Generalized Bloch Spheres for m-Qubit States
博士入学PPT模板
Results
2.2. Overexpressed of PTBP1 promotes migration of lung cancer cells
Results
2.3. Knockdown of PTBP1 inhibits levels of EMT-related proteins in lung cancer cells
Background
Seven alternative splicing (AS) subgroups: • Exon skipping accounts for nearly 40% of AS events; • alternative 3′ splice site (3′SS) selection (18.4%) and 5′SS
3. Dewei Niu, ******, Shanze Yi, Feng Wang*. Gene cloning, protein expression and functional analysis of a type 3 metallothionein gene from Sonneratia alba with biosorption potential. Polish Journal of Environmental Studies, Accepted. PJOES-00647-2017-02.
A
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A. PTBP1 expression was elevated in LUAD tissues (N=515) compared with normal lung tissues (N=59) according to TCGA database (p<0.01); B. Kaplan-Meier plots of patients with LUAD according to high (N=127) and low (N=375) PTBP1 expression from the TCGA database and compared by paired t-test, p<0.01.
外泌体在结直肠癌中的作用
外泌体在结直肠癌中的作用包久兵;史良会【摘要】外泌体是细胞释放的细胞外囊泡,包含mRNA、miRNA、蛋白质和部分特定区域的DNA等,参与细胞间通讯,并涉及许多生物学和病理学过程.来源于结直肠癌(CRC)细胞的外泌体与肿瘤的发生、肿瘤细胞的存活、增殖、侵袭和转移有关.本文介绍了外泌体及其纯化,并对CRC来源的外泌体种类、作用及其机制进行了综述.【期刊名称】《沈阳医学院学报》【年(卷),期】2018(020)004【总页数】4页(P361-364)【关键词】外泌体;结直肠癌;侵袭与转移【作者】包久兵;史良会【作者单位】皖南医学院研究生院,安徽芜湖 241001;皖南医学院弋矶山医院胃肠外科【正文语种】中文【中图分类】R735.3结直肠癌(CRC)是我国最常见的恶性肿瘤之一,在肿瘤导致的死亡中居第5位[1],预计到2030年,全球CRC的发病率将增加60%[2]。
对CRC侵袭及转移的分子机制及肿瘤细胞与外界信息交流的机制的了解,将有助于进一步预防和治疗CRC,其中外泌体介导的运输形式发挥着重要作用。
癌细胞能释放多种囊泡,这些囊泡可以通过体液,如外周血、唾液、尿液和腹水等进行转移[3],某些特殊的细胞囊泡称为“外泌体(exosomes)”,其与癌症的进展相关[4]。
1 外泌体1.1 外泌体的产生外泌体是在内化过程中由质膜产生的。
首先,细胞通过内吞作用形成早期内涵体(early endosomes,EE),其逐渐变为晚期内涵体或多泡体(MVBs),MVBs膜内陷形成腔内囊泡(ILVs),MVBs与溶酶体膜融合可降解蛋白质,并且释放ILVs进入溶酶体,或者MVBs与质膜融合,ILVs被释放到细胞外环境中,称为外泌体[5]。
外泌体融合了mRNA、miRNA、蛋白质和部分特定区域的DNA等。
外泌体的大小在30~100 nm[6],40~100 nm[7],50~150 nm[8]不等。
然而,外泌体的生物起源机制尚不十分清楚,还需要进行更深入的研究。
【精品】翻译综合
一个抑制肿瘤的连续模型-------艾丽斯H伯杰,阿尔弗雷德G. Knudson 与皮埃尔保罗潘多尔菲今年,也就是2011 年,标志着视网膜母细胞瘤的统计分析的第四十周年,首次提供了证据表明,肿瘤的发生,可以由两个突变发起。
这项工作提供了“二次打击”的假说,为解释隐性抑癌基因(TSGs)在显性遗传的癌症易感性综合征中的作用奠定了基础。
然而,四十年后,我们已经知道,即使是部分失活的肿瘤抑制基因也可以致使肿瘤的发生。
在这里,我们分析这方面的证据,并提出了一个关于肿瘤抑制基因功能的连续模型来全方位的解释肿瘤抑制基因在癌症过程中的突变。
虽然在1900 年之前癌症的遗传倾向已经被人认知,但是,是在19 世纪曾一度被忽视的孟德尔的遗传规律被重新发现之后,癌症的遗传倾向才更趋于合理化。
到那时,人们也知道,肿瘤细胞中的染色体模式是不正常的。
接下来对癌症遗传学的理解做出贡献的人是波威利,他提出,一些染色体可能刺激细胞分裂,其他的一些染色体 a 可能会抑制细胞分裂,但他的想法长期被忽视。
现在我们知道,这两种类型的基因,都是存在的。
在这次研究中,我们总结了后一种类型基因的研究历史,抑癌基因(TSGs),以及能够支持完全和部分失活的肿瘤抑制基因在癌症的发病中的作用的证据。
我们将抑制肿瘤的连续模型与经典的“二次打击”假说相结合,用来说明肿瘤抑制基因微妙的剂量效应,同时我们也讨论的“二次打击”假说的例外,如“专性的单倍剂量不足”,指出部分损失的抑癌基因比完全损失的更具致癌性。
这个连续模型突出了微妙的调控肿瘤抑制基因表达或活动的重要性,如微RNA(miRNA)的监管和调控。
最后,我们讨论了这种模式在┲⒌恼锒虾椭瘟乒 讨械挠跋臁!岸 未蚧鳌奔偎?第一个能够表明基因的异常可以导致癌症的发生的证据源自1960 年费城慢性粒细胞白血病细胞的染色体的发现。
后来,在1973 年,人们发现这个染色体是是第9 号和第22 号染色体异位的结果,并在1977 年,在急性早幼粒细胞白血病患者中第15 号和第17 号染色体易位被识别出来。
用温和的胰蛋白酶消化法分离小胶质细胞
High-Yield Isolation of MurineMicroglia by Mild TrypsinizationJOSEP SAURA,1*JOSEP MARIA TUSELL,2AND JOAN SERRATOSA1 1Department of Pharmacology and Toxicology,Institut d’Investigacions Biome`diques deBarcelona,IIBB-CSIC,Barcelona,Spain2Department of Neurochemistry,Institut d’Investigacions Biome`diques de Barcelona,IIBB-CSIC,Barcelona,SpainKEY WORDS in vitro;trypsin;CD11b;M-CSF;lipopolysaccharide;interferon␥ABSTRACT Microglia can be isolated with high purity but low yield by shaking off loosely adherent cells from mixed glial cultures.Here we describe a new technique for isolating microglia with an average yield close to2,000,000microglial cells/mouse pup, more thanfive times higher than that of the shaking method.Confluent mixed glial cultures are subjected to mild trypsinization(0.05–0.12%)in the presence of0.2–0.5mM EDTA and0.5–0.8mM Ca2ϩ.This results in the detachment of an intact layer of cells containing virtually all the astrocytes,leaving undisturbed a population offirmly at-tached cells identified asϾ98%microglia.These almost pure microglial preparations can be kept in culture for weeks and show proliferation and phagocytosis.Treatment with macrophage colony-stimulating factor and lipopolysaccharide,alone or in the presence of interferon␥,induces typical microglial responses in terms of proliferation, morphological changes,nuclear factor-B translocation,NO,and tumor necrosis␣release and phagocytosis.This method allows for the preparation of highly enriched mouse or rat microglial cultures with ease and reproducibility.Because of its high yield, it can be especially convenient when high amounts of microglial protein/mRNA are required or in cases in which the starting material is limited,such as microglial cultures from transgenic animals.The Supplementary Video Clip1referred to in this article can be found at the GLIA website(/jpages/0894-1491/suppmat/ 2003/v44.html©2003Wiley-Liss,Inc.INTRODUCTIONMicroglial cells,the resident immune cell population in the CNS,are derived from cells of mesodermal origin that enter the CNS during development.In the healthy adult brain,microglial cells are small,highly ramified cells with a low-profile phenotype.As a response to alterations in the environment,particularly neuronal damage,microglial cells exhibit marked morphological changes,proliferate,become phagocytic,and upregu-late the expression of a large number of molecules such as cytokines,adhesion molecules,membrane receptors, and transcription factors.This process,called micro-glial activation,is a physiological response aimed at protecting the affected neural tissue.However,due to their capacity to produce highly neurotoxic species, chronically activated microglial cells may participate in the pathogenesis of neurodegenerative disorders such as Alzheimer’s disease.Many features of microglial activation can be repro-duced in culture.Isolation of microglial cells for cultur-ing can be obtained by several methods,including iso-lation from CNS tissue by Percoll gradient(Ford et al., 1995),isolation from primary cultures by nutritional deprivation(Hao et al.,1991)or by collectingfloating cells(Ganter et al.,1992),and preparation of microglia-enriched primary cultures with horse serum(Colton et al.,1991).By far the most popular protocol is the shak-ing method described simultaneously by Giulian and Grant sponsor:the Spanish Ministerio de Ciencia y Tecnologı´a;Grant number: SAF2001-2240.*Correspondence to:Dr.Josep Saura,Department of Pharmacology and Tox-icology,IIBB-CSIC Rossello´161,6a planta,08036Barcelona,Spain.E-mail:jsafat@iibb.csic.esReceived19February2003;Accepted14April2003DOI10.1002/glia.10274GLIA44:183–189(2003)©2003Wiley-Liss,Inc.Baker(1986)and Frei et al.(1986).With this method, microglial cells are separated from confluent primary mixed glial cultures from newborn rodent cerebral cor-tex by agitation on a rotary shaker.This method allows for the preparation of highly enriched(Ͼ95%)micro-glial cultures;unfortunately,the amount of microglial cells obtained is low.In the present study,we describe a new method to isolate microglial cells from primary mixed glial cul-tures of rodent brain by a mild trypsinization protocol. This method is simple,reproducible,and allows for the preparation of microglial cultures of high purity(Ͼ98%)with a much higher yield than the shaking method.Microglial cells obtained with this method are functional as assessed by their capacity to proliferate, phagocyte,change morphology,release NO and tumor necrosis factor␣(TNF␣),or translocate NF-B in re-sponse to specific stimuli.MATERIALS AND METHODSTrypsin-EDTA solution(0.25%trypsin,1mM EDTA in HBSS;25200-072),Dulbecco’s modified Eagle medi-um-F-12nutrient mixture(DMEM-F12;31330-038), macrophage serum-free medium(M-SFM;12065-074), fetal bovine serum(FBS),and penicillin-streptomycin were from Invitrogen.Deoxyribonuclease I,trypsin in-hibitor,mouse anti-␣smooth muscle actin(␣SMA; clone1A4),biotin-labeled tomato lectin,extravidin per-oxidase,Harris hematoxylin,macrophage colony-stim-ulating factor(M-CSF;M-9170),sulfanilamide,phos-phoric acid,N-1-naphtylenediamine,bisbenzimide (Hoechst no.33258),and lipopolysaccharide(LPS; L-2654)from Escherichia coli were from Sigma.Inter-feron␥(IFN␥;585-IF)was from R&D Systems.Rabbit anticow glialfibrillary acidic protein(GFAP)was from Dako.Rat antimouse CD11b(clone5C6)was from Se-rotec.Goat anti-p65was from Santa Cruz Biotechnol-ogy.Biotinylated antirabbit IgG was from Pierce.Bio-tinylated antimouse IgG and biotinylated antirat IgG were from Vector Laboratories.FluoSpheres carboxy-late microspheres(F-8826)and Alexa Fluor488goat antimouse were from Molecular Probes.Fluorescein (FITC)-conjugated streptavidin was from Chemicon In-ternational.5-bromo-2Ј-deoxyuridine(BrdU)and Mowiol were from Calbiochem.BrdU staining kit (HCS24)was from Oncogene.Mouse TNF␣enzyme-linked immunosorbent assay(ELISA)kit(EM-TNFA) was from Endogen.Cell CulturesMixed glial cultures were prepared from cerebral cortices of1-day-old Bl/C57mice(Charles River, France)according to the method of Giulian and Baker (1986).After mechanical and chemical dissociation, cortical cells were seeded in DMEM-F12with10%FBS at a density of250,000cells/ml(ϭ62,500cells/cm2)and cultured at37°C in humidified5%CO2/95%air.Me-dium was replaced every4–5days and confluency was achieved after10–12days in vitro(DIV).Microglial cultures were prepared by two methods: mild trypsinization and shaking.For the mild trypsinization method,various parameters,such as trypsin,Ca2ϩ,and EDTA concentrations,were studied and the optimal protocol obtained is described below. For comparison,the shaking method of Giulian and Baker(1986)was applied as follows.Mouse primary mixed glial cultures were prepared on25cm2flasks at different seeding densities in the60,000–200,000cells/ cm2range.The highest microglia yield was obtained when cells were plated at100,000–120,000cells/cm2. Microglial cells were obtained by shaking theflasks overnight at200rpm.Floating cells were pelleted and subcultured at400,000cells/ml(ϭ100,000cells/cm2) on mixed glial-conditioned medium.Immunocytochemistry and Lectin Staining Cells werefixed with4%paraformaldehyde(60min, 22°C)for tomato lectin,GFAP,or CD11b immunode-tection,and with methanol(8min,Ϫ20°C)for␣SMA, phosphorylated H3histone,and p65immunodetection. When immunocytochemistry was revealed with diami-nobenzidine(DAB),a standard protocol was used (Casal et al.,2001).Dilutions of primary antibodies/ lectins were anti-GFAP,1:1,000;anti-CD11b,1:3,000; anti-␣SMA,1:1000;biotinylated tomato lectin,1:500. Dilutions of secondary biotinylated antibodies were an-tirabbit,1:1,000;antirat,1:300;and antimouse,1:200. For immunofluorescence,cells were cultured on glass coverslips,a mouse anti-p65(1g/ml)was the primary antibody and Alexa Fluor488-labeled goat antimouse antibody(1:1,000)was the secondary antibody.Cover-slips were mounted with Mowiol and stored at4°C.Cell CountsAfter immunocytochemistry,nuclei were counter-stained with Harris hematoxylin.Underϫ20objective, 15fields of0.135mm2were photographed per well. Total cells,immunopositive and immunonegative cells were counted for GFAP,tomato lectin,CD11b,and ␣SMA.Nitrite AssayNitric oxide production was assessed by the Griess reaction,a colorimetric assay that detects nitrite (NO2Ϫ),a stable reaction product of NO and molecular oxygen.Briefly,100l of conditioned medium were incubated with100l of Griess reagent at22°C for10 min.The optical density of the samples was measured at540nm.The nitrite concentration was determined from a sodium nitrite standard curve.184SAURA ET AL.TNF ␣The amount of TNF ␣released in 100l of the con-ditioned medium was determined using an ELISA kit specific for mouse TNF ␣.ELISA measurements were performed using the standard and instructions sup-plied by the manufacturer.BrdU IncorporationCells were labeled with 10M BrdU for 2h and fixed for 10min in 70%ethanol.After blocking endogenous peroxidases with 3%hydrogen peroxide,the protocol supplied by the manufacturer was followed.PhagocytosisCells were cultured on glass coverslips and incubated for 10,30,or 90min at 37°C in the presence of 2m diameter FluoSpheres at 0.01%solid mass.Cells were then washed with PBS,fixed in methanol (8min,Ϫ20°C),labeled with biotinylated tomato lectin 1:500,and visualized with FITC-conjugated streptavidin (1:100).After 5-min incubation with Hoechst-33258(2.5g/ml in PBS),coverslips were mounted with Mowiol.Statistical AnalysisResults are expressed as mean ϮSD from at least three independent cultures.Statistical analysis of TNF ␣release in treated cells vs.their respective con-trols was performed using Student’s t -test.Statistical analysis of nitrite production in treated cells vs.their respective controls was performed using one-way ANOVA and the Turkey posthoc test.RESULTSThe starting point of this study was the serendipi-tous observation that incubation of mixed glial cultures with a trypsin solution (0.25%trypsin,1mM EDTA in HBSS;named henceforth trypsin 0.25%)diluted 1:4in DMEM-F12resulted in the detachment of an upper layer of cells in one piece,whereas a number of cells remained attached to the bottom of the well (Fig.1and Video Clip 1(available at the Glia Web site at:http://Fig. 1.Photographic sequence of isolation of microglia by mild trypsinization of mixed glial cultures.A shows a confluent murine mixed glial culture at DIV20prior to mild trypsinization.The same field in A is shown in B and C 23and 24min,respectively,since the beginning of trypsinization performed as indicated in text.Note the progressive and rapid detachment of the mainly astrocytic layer of cells and the presence of a population of microglial-looking cells at-tached to the surface of the well.D shows the isolated microglial cells after trypsinization is terminated (30min).The process is also illus-trated as a supplementary material Video Clip 1found at the GLIA website ( /jpages /0894-1491/suppmat /2003/v44.html ).Magnification bar,40m.Video Clip 1.This movie shows the temporal sequence of mild trypsinization of a murine mixed glial culture.It illustrates the detachment of a layer of cells containing virtually all the astrocytes and some microglia.A highly enriched micro-glial population remains attached to the bottom of the well.The sequence lasted 30min and consists of 23individual photographs.185MURINE MICROGLIA CULTURES BY TRYPSINIZATION/jpages/0894-1491/suppmat/2003/v44.html)).The detachment started in the pe-riphery of the well after approximately 15min in tryp-sin solution and was typically completed after 25–35min.The effect was inhibited in the presence of serum (7%)or soybean trypsin inhibitor (0.3mg/mg trypsin),indicating that trypsin was indeed responsible for the detachment.A similar detachment of an intact layer of cells was observed when trypsin 0.25%was diluted 1:3,1:2,or 1:1in DMEM-F12but not with undiluted tryp-sin 0.25%,in which case all the cells detached,individ-ually or in small clumps.Interestingly,whereas tryp-sin 0.25%diluted 1:4in PBS containing 1mM CaCl 2induced the effect seen with trypsin 0.25%:DMEM-F121:4,trypsin 0.25%diluted 1:4in Ca 2ϩ-free PBS did not.Note that the Ca 2ϩconcentration in DMEM:F12is 1mM.These results indicate that the presence of Ca 2ϩis necessary for the trypsin-induced separation of a de-tached layer of cells from a population of attached cells.Twenty-four hours after trypsinization,the isolated cells were stained for various cellular markers.A great majority of cells (98.4–99.2%;range of four experi-ments)were positive for CD11b (Fig.2),and the same figure was obtained with tomato lectin histochemistry.In contrast,0.8–1.7%were positive for ␣SMA,which is present in pericytes and astrocytes but not in micro-glia,and 0.2–0.5%for GFAP,an astroglial marker.Cells negative for CD11b and tomato lectin were large,flat,and polygonal,and this was the morphology of ␣SMA-positive cells.These observations indicate that a large majority (Ͼ98%)of cells isolated by trypsiniza-tion were microglia.After isolation by trypsinization,adherent microglial cells could be grown in various culture media.For short-term culture (Ͻ3days),DMEM-F12or M-SFM with or without 10%FBS could be used but resulted in poor viability for long-term culture.DMEM-F12with 10%FBS conditioned by mixed glial cultures allowed cells to be cultured for at least 2weeks.Except for experiments requiring no serum,this was the medium selected for all the experiments.Cell density of cultures obtained by trypsinization was 31,160Ϯ8,524cells/cm 2(n ϭ7experiments).Since the plating density was 62,500cells/cm 2,this represents a 50%yield.Considering that on average we obtain 3,500,000cortical cells/mouse pup,this results in a yield of 1,750,000microglial cells/pup.In contrast,with the shaking method,225,000Ϯ92,000microglial cells were obtained from one 25cm 2flask initially plated with 2,500,000cells (9%yield or 315,000micro-glial cells/pup).Cells obtained by shaking were mainly microglia (Ͼ98%),as estimated by CD11b and tomato lectin staining.Time-course experiments revealed that the age of the mixed glial culture determines the yield of the trypsinization method (Fig.3).A high yield was ob-tained between DIV15and DIV35,peaking at DIV20to DIV25,whereas nonconfluent cultures (DIV10)re-sulted on a much lower yield.As shown in Figure 3,purity of the microglial cultures was low when obtained from mixed glial cultures at DIV10(78%),peaked be-tween DIV15and DIV30(Ͼ97%),and declined at DIV35(94%).Since confluency is reached before (or after)when primary glial cultures are plated at a higher (or lower)density,the graphs in Figure 3canbeFig.3.Microglial cell density and purity as a function of the age of the murine primary mixed glial culture.Bars show cell density and solid line %microglia 24h after trypsinization of mixed glial cultures of 10–35DIV.Preparation of microglial cultures by trypsinization of mixed glial cultures of DIV15–30results in both high density (Ͼ20,000cells/cm 2)and high purity (Ͼ97%).Data were obtained from three independent experiments and bars showSD.d trypsinization of murine primary cortical mixed glial cultures results in the isolation of highly enriched microglial cultures.Images show staining for CD11b in mixed glial cultures (A )or 24h after mild trypsinization (B ).After trypsinization,virtually all cellsare positive for CD11b.Note also the ramified morphology and high abundance of microglial cells in mixed glial cultures.Magnification bar,50m.186SAURA ET AL.somewhat shifted to the left or right depending on the seeding cell density.From these experiments,the following protocol was established for24-well plates.First,prepare primary mixed glial cortical cultures as described e them between DIV15and DIV30for isolation of micro-glia.It is important that mixed glial cultures have been confluent for at least3days.Second,wash mixed glial cells for1min in DMEM:F12to eliminate serum.Keep the conditioned medium for step 5.Third,incubate cells at37°C with500l per well of trypsin0.25%: DMEM-F121:3until intact layer is detached.This step takes20–45min.However,longer incubations(up to 6h)do not affect microglial yield or viability.Fourth, add500l of DMEM-F12with10%FBS for trypsin inactivation.Fifth,aspirate medium containing the layer of detached cells and replace with mixed glial-conditioned medium from step2.This protocol has been successfully used for isolating microglial cells from mixed glial cultures of rat and mouse growing on uncoated or polylysine-coated6-, 24-,or48-well plates or glass coverslips.After step5, isolated microglial cells can be recovered by a5-min incubation with trypsin0.25%with vigorous pipetting and replated.The morphology of microglial cells obtained by this method was typically elongated,either bipolar or unipolar.Cells with ameboid morphology were also abundant,whereas round refringent cells often seen in mixed glial cultures were extremely rare.Treatment with IFN␥(10ng/ml),and especially with LPS(1g/ ml)or LPSϩIFN␥,increased the number of cells with ameboid morphology.In contrast,M-CSF(200ng/ml) reduced the proportion of ameboid cells and induced an even more elongated morphology.When cultured in M-SFM for72h,microglial cells isolated by this method showed a low proliferation rate (Fig.4A).In these conditions,a24-h exposure to M-CSF(200ng/ml)strongly increased the number of pro-liferating,BrdU-positive cells(Fig.4B).We also as-sessed the ability of various factors to induce NF-B nuclear translocation in these cells.By immunocyto-chemistry,the NF-B subunit p65was localized in control cells throughout the cytoplasm with a weaker signal in the nucleus(Fig.4C).Six hours after treat-ment with LPS(1g/ml;Fig.4D)or LPSϩIFN␥(10 ng/ml;not shown),p65immunoreactivity was predom-inantly localized in the nucleus.In order to test the capacity of microglial cells iso-lated by this method to release NO,nitrite concentra-tion was estimated in the conditioned medium24h after treatment with various factors.Nitrite levels in control medium were low(4.5Ϯ0.9M)and were not affected by IFN␥(10ng/ml)or M-CSF(200ng/ml).In contrast,LPS(1g/ml)induced a modest but signifi-cant release of NO(8.2Ϯ1.4M;PϽ0.05),which was markedly potentiated by IFN␥(19.1Ϯ2.4;PϽ0.001). We also studied whether trypsinization-isolated micro-glial cells were able to release TNF␣upon stimulation. TNF␣levels in medium conditioned by control cells were barely detectable(8Ϯ20pg/ml),whereas a sig-nificant increase in TNF␣concentration was observed in the conditioned medium of microglial cells treated for6h with LPSϩIFN␥(1,277Ϯ174pg/ml;PϽ0.001).Trypsinization-isolated microglial cells were incu-bated withfluorescein-labeled latex beads in order to ascertain their phagocytic capacity.Since latex beads are easily seen by phase-contrast microscopy,we could follow the phagocytic process“live.”Beadsfloating in the medium had a Brownian movement even when located on top of a microglial cell.This movement abruptly stopped,indicating that the cell had captured the particle.This was followed by a rapid transport (5–10min)of the latex bead to a perinuclear location. After90min,microglial cells were packed with latex beads(Ͼ200beads per cell)concentrated around the nucleus.In contrast,flat,large,polygonal cells resem-bling contaminating␣SMA-positive cells did not phagocyte latex beads.Exposure to LPS(1g/ml,24h) enhanced microglial phagocytic capacity as shown by the increased number of beadsincorporated after10or 30min(Fig.4E and F).DISCUSSIONTrypsinization of primary mixed glial cultures fol-lowed by replating at low density is a common practice for preparing highly enriched astroglial cultures. Trypsinization is done at0.05–0.25%in calcium-free medium generally in the presence of0.5mM EDTA and results in the rapid detachment of all cells in the cul-ture.In the present study,we have observed that trypsinization at0.05–0.12%in the presence of0.2–0.5 mM EDTA did not induce the individual detachment of all cells but the detachment of an intact layer of cells provided Ca2ϩwas present in the medium at concen-trations between0.52and0.84mM.Since this Ca2ϩconcentration is higher than the chelating capacity of the EDTA present,our hypothesis is that the nonse-questered free Ca2ϩpartially inhibits trypsin and this results in the observed partial trypsinization.The detachment of an intact cell layer left a popula-tion of cellsfirmly attached to the bottom of the well. With histochemical markers,these cells were identified as predominantly(Ͼ98%)microglia.Therefore,this trypsinization step can be used as a new method to isolate microglia from mixed glial cultures.The process resembles an accelerated form of that occurring when confluent mixed glial cultures are nutritionally de-prived,which results in a progressive retraction of astrocytes and the appearance of increasing number of ameboid microglial cells(Hao et al.,1991).The method here presented for the isolation of mi-croglial cells is simple,reproducible,and versatile.It can be used for isolating mouse or rat microglia from mixed glial cultures growing in a variety of supports.If needed,microglial cells can be recovered after isolation and replated at a different density or in a different187MURINE MICROGLIA CULTURES BY TRYPSINIZATIONsupport.A major advantage of this method with re-spect to the shaking method is its higher yield.Thus,with the optimal conditions for the shaking method,we obtain a yield of 315,000microglial cells/pup,which is in the range of that reported for the shaking method (Sawada et al.,1990;Hassan et al.,1991;Abromson-Leeman et al.,1993;Fischer et al.,1993;Pinteaux et al.,2002)or other methods (Colton et al.,1991;Slepko and Levi,1996).In contrast,with the novel trypsiniza-tion method,an average yield of 1,750,000microglial cells/pup was obtained.Therefore,the trypsinization method provides a greater than fivefold increase in microglial yield when compared with the shaking method.A critical point for validating the trypsinization pro-tocol as an alternative method for preparing microglial cultures was to show that microglial cells obtained were functionally equivalent to microglial cells isolated by shaking.To this end,we studied their response to two activating stimuli such as M-CSF and LPS (ϩIFN ␥).In microglial cells isolated by the shaking method,M-CSF,a hematopoietic cytokine,induces pro-liferation (Suzumura et al.,1990;Casal et al.,2001)and morphological elongation (Sawada et al.,1990;Suzumura et al.,1991).We have reproduced both ob-servations in microglial cells isolated by trypsinization.On the other hand,treatment of microglial cells iso-lated by shaking with the bacterial endotoxinLPSFig.4.Cells isolated by mild trypsinization show typical microglial responses.In A and B ,isolated microglial cells were allowed to incor-porate BrdU as indicated in text.A:Control cells.B:Cells treated with M-CSF (200ng/ml,24h).M-CSF induces a marked increase in the number of proliferating,BrdU-positive cells as well as an elonga-tion of microglial cells.Arrows with (ϩ)and (Ϫ)point to examples of BrdU-positive and –negative microglial cells,respectively.Magnifica-tion bar,50m.In C and D ,the NF-B subunit p65was immunode-tected in isolated microglia.C:Control cells.D:Cells treated for 6h with LPS (1g/ml).In control cells p65is localized throughout the cytoplasm,whereas LPS induces the nuclear translocation of theprotein.Magnification bar,30m.In E and F ,microglial cells were exposed to fluorescein-labeled latex beads for 30min.E:Control cells.F:Microglial cells treated with LPS (1g/ml)for 24h.Control micro-glial cells internalize latex beads showing the phagocytic activity of these cells in basal conditions.Pretreatment with LPS enhances microglial phagocytic activity as shown by the increased amount of internalized latex beads.LPS also induces transformation into an ameboid morphology.Arrows show examples of fluorescent latex beads.Microglial cells are labeled with tomato lectin.Magnification bar,30m.188SAURA ET AL.alone or in the presence of the proinflammatory cyto-kine IFN␥results in an in vitro model of microglial activation with a characteristic ameboid morphology (Suzumura et al.,1991;Wollmer et al.,2001),NF-B activation(Heyen et al.,2000;Wollmer et al.,2001), NO release(Boje and Arora,1992;Chao et al.,1992b), TNF␣release(Sawada et al.,1989;Chao et al.,1992a), and increased phagocytosis(Peterson et al.,1993).All these signs of activation were also observed with mi-croglia obtained by trypsinization.Altogether,these findings demonstrate that cells isolated by this novel protocol not only express microglial markers but also behave like microglia.In summary,a novel technique for isolating micro-glial cells by trypsinization is described.The reproduc-ibility and ease of the method and the purity of the microglial cultures obtained are as high as,if not higher than,those of the existing methods.Trypsiniza-tion-isolated microglia responses to M-CSF,LPS,and IFN␥are characteristic of microglial cells.The higher yield of the new method may allow for the design of experiments requiring high amounts of microglial pro-tein/mRNA or in which the starting material is limited. It may become particularly useful for the preparation of microglial cultures from transgenic mice.ACKNOWLEDGMENTSThe authors thank Dr.V.Petegnief and Dr.C.Sola` for critically reading of the article.Joseph Saura is the recipient of a Ramo´n y Cajal contract from the Spanish Ministerio de Ciencia y Tecnologı´a.REFERENCESAbromson-Leeman S,Hayashi M,Martin C,Sobel R,al Sabbagh A, Weiner H,Dorf ME.1993.T cell responses to myelin basic protein in experimental autoimmune encephalomyelitis-resistant BALB/c mice.J Neuroimmunol45:89–101.Boje KM,Arora PK.1992.Microglial-produced nitric oxide and reac-tive nitrogen oxides mediate neuronal cell death.Brain Res587: 250–256.Casal C,Tusell JM,Serratosa J.2001.Role of calmodulin in the differentiation/activation of microglial cells.Brain Res902:101–107.Chao CC,Hu S,Close K,Choi CS,Molitor TW,Novick WJ,Peterson PK.1992a.Cytokine release from microglia:differential inhibition by pentoxifylline and dexamethasone.J Inf Dis166:847–853. Chao CC,Hu S,Molitor TW,Shaskan EG,Peterson PK.1992b. Activated microglia mediate neuronal cell injury via a nitric oxide mechanism.J Immunol149:2736–2741.Colton CA,Yao J,Taffs RE,Keri JE,Oster-Granite ML.1991.Abnor-mal production of interleukin-1by microglia from trisomy16mice. Neurosci Lett132:270–274.Fischer HG,Nitzgen B,Germann T,Degitz K,Daubener W,Hadding U.1993.Differentiation driven by granulocyte-macrophage colony-stimulating factor endows microglia with interferon-gamma-inde-pendent antigen presentation function.J Neuroimmunol42:87–95. Ford AL,Goodsall AL,Hickey WF,Sedgwick JD.1995.Normal adult ramified microglia separated from other central nervous system macrophages byflow cytometric sorting.Phenotypic differences de-fined and direct ex vivo antigen presentation to myelin basic pro-tein-reactive CD4ϩT cells compared.J Immunol154:4309–4321. Frei K,Bodmer S,Schwerdel C,Fontana A.1986.Astrocyte-derived interleukin3as a growth factor for microglia cells and peritoneal macrophages.J Immunol137:3521–3527.Ganter S,Northoff H,Mannel D,Gebicke-Harter PJ.1992.Growth control of cultured microglia.J Neurosci Res33:218–230.Giulian D,Baker TJ.1986.Characterization of ameboid microglia isolated from developing mammalian brain.J Neurosci6:2163–2178.Hao C,Richardson A,Fedoroff S.1991.Macrophage-like cells origi-nate from neuroepithelium in culture:characterization and proper-ties of the macrophage-like cells.Intl J Dev Neurosci9:1–14. Hassan NF,Rifat S,Campbell DE,McCawley LJ,Douglas SD.1991. Isolation andflow cytometric characterization of newborn mouse brain-derived microglia maintained in vitro.J Leukocyte Biol50: 86–92.Heyen JR,Ye S,Finck BN,Johnson RW.2000.Interleukin(IL)-10 inhibits IL-6production in microglia by preventing activation of NF-kappaB.Mol Brain Res77:138–147.Peterson PK,Gekker G,Hu S,Chao CC.1993.Intracellular survival and multiplication of Toxoplasma gondii in astrocytes.J Inf Dis 168:1472–1478.Pinteaux E,Parker LC,Rothwell NJ,Luheshi GN.2002.Expression of interleukin-1receptors and their role in interleukin-1actions in murine microglial cells.J Neurochem83:754–763.Sawada M,Kondo N,Suzumura A,Marunouchi T.1989.Production of tumor necrosis factor-alpha by microglia and astrocytes in culture. Brain Res491:394–397.Sawada M,Suzumura A,Yamamoto H,Marunouchi T.1990.Activa-tion and proliferation of the isolated microglia by colony stimulating factor-1and possible involvement of protein kinase C.Brain Res 509:119–124.Slepko N,Levi G.1996.Progressive activation of adult microglial cells in vitro.Glia16:241–246.Suzumura A,Sawada M,Yamamoto H,Marunouchi T.1990.Effects of colony stimulating factors on isolated microglia in vitro.J Neu-roimmunol30:111–120.Suzumura A,Marunouchi T,Yamamoto H.1991.Morphological transformation of microglia in vitro.Brain Res545:301–306. Wollmer MA,Lucius R,Wilms H,Held-Feindt J,Sievers J,Mentlein R.2001.ATP and adenosine induce ramification of microglia in vitro.J Neuroimmunol115:19–27.189MURINE MICROGLIA CULTURES BY TRYPSINIZATION。
核酸适配体
S 钾离子能够使富含鸟嘌呤的核酸适配体形成特异的 Gquadruplex 二级构型并对此结构 有一定的稳定(wěndìng)作用, 因此可以通过监测核酸适配体二级结构信号的变化实现对 钾离子的检测
Lindner等还通过aptamer芯片系统成功地从混合蛋白质中识 别出专一性的蛋白,而且利用凝血酶aptamer证明了在同一 芯片上同时检测两种蛋白方法的可行性。总之,寡核苷酸 aptamer作为低分子量的分子受体,它在芯片上能专一性地 检测蛋白质而且很稳定,以它为阵列来捕获蛋白质将为蛋 白质组学研究的发展起到重共四要十六页作用。
共四十六页
亲和介质 分离 (jièzhì)
S 一些具有亲和表面的介质也用于适配体的筛选,如琼脂糖、 纤维素及具有亲和表面的小珠或小柱等。
S 如J.Colin Cox等人利用链霉亲和素标记的磁珠完成了溶菌 酶适配体的自动化筛选。具体(jùtǐ)流程为:通过链酶亲和素 与生物素的相互作用,将生物素化的靶蛋白固定在磁珠上。 随后特异结合序列的分离,RT-PCR扩增和转录都通过设定 的程序自动完成,最后筛选得到的序列克隆到载体中进行 测序鉴定。通过这种自动化筛选工作台,Cox等只用了不到 两天的时间就完成了12轮的筛选。
S Macugen是一种选择性血管内皮生产因子 (vascular endothelial growth factor,VEGF) 拮抗剂。
共四十六页
核酸适配体的化学本质(běnzhì)与识 别机理
S 核酸适配体的化学(huàxué)本质是核酸,它与配体的结合是基 于单链核酸结构和空间构象的多样性。在靶分子存在的条 件下,它可通过链内某些互补碱基间的配对以及静电作用、 氢键作用等自身发生适应性折叠形成发卡(hairpin)、假结 (pseudoknot)、凸环(stem loop)、G2四分体(G2quartet)等稳 定的三维空间结构。这样形成的适配体结构与靶分子之间 有较大的接触面积,能与靶物质的紧密结合,具有高亲和 力和高特异性。
依泽麦布抑制膀胱癌细胞增殖、迁移及其机制
-基础研.*依泽麦布抑制膀胱癌细胞增殖、迁移及其机制熊康平,彭天辰,熊连祎,肖宇,鞠林高,王刚,王行环(武汉大学中南医院泌尿外科,湖北武汉430071)Ezetimibe inhibits the proliferation and migration of bladder cancer cells and the mechanismXIONG Kangping,PENG Tianchen,XIONG Yaoyi,XIAO Yu,JU Lingao,WANG Gang,WANG Xinghuan (Department of Urology,Zhongnan Hospital of Wuhan University,Wuhan430071,China)ABSTRACT:Objective To explore the effects of ezetimibe on the proliferation,migration and cell cycle of bladder cancer cells and to investigate the underlying mechanism.Methods Afer bladder cancer cell lines T24and5637were treated wth different concentrations of ezetimibe(0,5,1020,40,60,80and100mmol/L)for48hours,the proliferation,migration and cell cycle distribution were detected wth MTT assay,wound-healing assay and flow cytometry,respectively.The expressions of CDK2,CDK4,1-catenin and vimentin were detected with Western blot.Results Afer exposure to ezetimibe,the proliferation ofbladdercancerce l sT24and5637weresignifican(lyinhibi(edinaconcen(raion-dependen(manner.Thece l cyclewasarres-ted at G0/G1phase,and cell migration was decreased.Western blot showed that ezetimibe downregulated the protein expressions of CDK4,CDK6,1-catenin and vimentin in bladder cancer cells.Conclusion Ezetimibe can inhibit the proliferation and m1grat1onofbladdercancerce l sand1nduceG0/G1ce l cyclearrest.KEY WORDS:ezetimibe;bladder cancer;proliferation;cell cycle distribution;migration ability摘要:/的探究依泽麦布(Ezetimibe)对膀胱癌细胞增殖、迁移以及周期分布的影响及其可能机制)12将膀胱癌细胞系T24和5637分别用不同浓度(0、5、10、20、40、60、80、100mmol/L)的依泽麦布处理48h后,采用+哩兰(MTT)实验测试依泽麦布对膀胱癌细胞增殖能力的影响;运用划痕实验检测依泽麦布对细胞迁移能力的影响;应用流式细胞仪来检测依泽麦布处理后细胞周期的分布情况;Western blot检测CDK2和CDK4以及上1连接素(p-catenin)和波形蛋白(vimentin)的表达情况。
地牡宁神口服液联合可乐定透皮贴治疗儿童抽动-秽语综合征效果与安全性研究
地牡宁神口服液联合可乐定透皮贴治疗儿童抽动-秽语综合征效果与安全性研究*文伟① 肖珊① 罗丽② 【摘要】 目的:探讨地牡宁神口服液联合可乐定透皮贴治疗儿童抽动-秽语综合征(TS)患儿的效果。
方法:选取2021年1月—2022年7月赣州市妇幼保健院收治的60例TS患儿,按随机数字表法分为两组,各30例。
对照组给予可乐定透皮贴治疗,观察组加用地牡宁神口服液治疗,均持续6个月。
对比两组临床疗效、症状改善时间、血清学指标及安全性。
结果:观察组治疗总有效率96.67%,高于对照组的73.33%(P<0.05)。
观察组运动性抽动改善时间(1.89±0.34)个月,发声性抽动改善时间(1.95±0.28)个月,综合性损伤症状改善时间(2.04±0.55)个月,均早于对照组的(2.46±0.57)、(2.48±0.61)、(2.78±0.81)个月,差异均有统计学意义(P<0.05)。
治疗前两组免疫功能、血清学指标比较,差异均无统计学意义(P>0.05);治疗后观察组血清免疫球蛋白G(IgG)水平为(6.05±0.47)g/L,免疫球蛋白A(IgA)水平为(0.49±0.10)g/L,免疫球蛋白M(IgM)水平为(0.51±0.15)g/L,白细胞介素-12(IL-12)水平为(101.15±8.64)pg/mL,肿瘤坏死因子-α(TNF-α)水平为(177.44±13.60)pg/mL,均低于对照组的(6.52±0.55)g/L、(0.68±0.13)g/L、(0.67±0.14)g/L、(112.74±9.55)pg/L、(202.49±14.68)pg/mL,差异均有统计学意义(P<0.05)。
两组不良反应比较,差异无统计学意义(P>0.05)。
异氰酸酯聚氨酯英文
Carbohydrate Polymers 101 (2014) 1043–1060Contents lists available at ScienceDirectCarbohydratePolymersj o u r n a l h o m e p a g e :w w w.e l s e v i e r.c o m /l o c a t e /c a r b p olReviewFunctionalized bacterial cellulose derivatives and nanocompositesWeili Hu,Shiyan Chen ∗,Jingxuan Yang,Zhe Li,Huaping Wang ∗State Key Laboratory for Modification of Chemical Fibers and Polymer Materials,Key Laboratory of High-performance Fibers and Products,Ministry of Education College of Materials Science and Engineering,Donghua University,Shanghai 201620,Chinaa r t i c l ei n f oArticle history:Received 9August 2013Received in revised form 23September 2013Accepted 29September 2013Available online 6 October 2013Keywords:Bacterial cellulose Modification Nanocomposites Functionalizationa b s t r a c tBacterial cellulose (BC)is a fascinating and renewable natural nanomaterial characterized by favor-able properties such as remarkable mechanical properties,porosity,water absorbency,moldability,biodegradability and excellent biological affinity.Intensive research and exploration in the past few decades on BC nanomaterials mainly focused on their biosynthetic process to achieve the low-cost preparation and application in medical,food,advanced acoustic diaphragms,and other fields.These investigations have led to the emergence of more diverse potential applications exploiting the function-ality of BC nanomaterials.This review gives a summary of construction strategies including biosynthetic modification,chemical modification,and different in situ and ex situ patterns of functionalization for the preparation of advanced BC-based functional nanomaterials.The major studies being directed toward elaborate designs of highly functionalized material systems for many-faceted prospective applications.Simple biosynthetic or chemical modification on BC surface can improve its compatibility with differ-ent matrix and expand its utilization in nano-related applications.Moreover,based on the construction strategies of functional nanomaterial system,different guest substrates including small molecules,inor-ganic nanoparticles or nanowires,and polymers can be incorporated onto the surfaces of BC nanofibers to prepare various functional nanocomposites with outstanding properties,or significantly improved physicochemical,catalytic,optoelectronic,as well as magnetic properties.We focus on the preparation methods,formation mechanisms,and unique performances of the different BC derivatives or BC-based nanocomposites.The special applications of the advanced BC-based functional nanomaterials,such as sensors,photocatalytic nanomaterials,optoelectronic devices,and magnetically responsive membranes are also critically and comprehensively reviewed.Crown Copyright © 2013 Published by Elsevier Ltd. All rights reserved.Contents 1.Introduction ..........................................................................................................................................10442.Construction strategies of BC-based functional nanomaterials .....................................................................................10452.1.Biosynthetic modification .. (1045)2.1.1.Altered BC structure ................................................................................................................10452.1.2.Nanocomposites .. (1046)2.2.Chemical surface modification ...............................................................................................................10462.3.In situ formation of nanostructures . (1047)2.3.1.In situ formation of nanostructures through reduction reaction ..................................................................10482.3.2.In situ formation of nanostructures through precipitation reaction...............................................................10482.3.3.In situ formation of nanostructures through sol-gel reaction (1048)2.4.Ex situ introduction of components..........................................................................................................10492.5.Other combined strategies ...................................................................................................................10493.Applications of BC-based functional nanomaterials .................................................................................................10533.1.Sensors........................................................................................................................................10533.2.Photocatalytic nanomaterials . (1053)∗Corresponding authors.Tel.:+862167792950;fax:+862167792726.E-mail addresses:chensy@ (S.Chen),wanghp@ (H.Wang).0144-8617/$–see front matter.Crown Copyright © 2013 Published by Elsevier Ltd. All rights reserved./10.1016/j.carbpol.2013.09.1021044W.Hu et al./Carbohydrate Polymers101 (2014) 1043–10603.3.Optoelectronics (1054)3.3.1.Electrically conductivefilms (1054)3.3.2.Optically transparentfilms (1055)3.3.3.Flexible displays (1056)3.3.4.Photoluminescent and photochromicfilms (1056)3.4.Magnetically responsivefilms (1057)4.Concluding remarks (1058)Acknowledgements (1059)References (1059)1.IntroductionIt is well known that cellulose is a very important and fasci-nating biopolymer and an almost inexhaustible and sustainable natural polymeric raw material,which is of special importance both in industries and in daily lives.In the past decade,the design and development of renewable resources and innovative prod-ucts for science,medicine and technology have led to a global revival of interdisciplinary research and utilization of this abun-dant natural polymer.Formed by repeated connection of glucose building blocks,cellulose possesses abundant surface hydroxyl groups forming plentiful inter-and intra-molecular hydrogen bonds,characterized by its hydrophilicity,chirality,biodegrad-ability,and broad chemical-modifying capacity(Klemm,Heublein, Fink,&Bohn,2005).The properties of cellulose largely depend on the specific assembling and supramolecular order controlled by the origin and treatment of cellulose(Eichhorn et al.,2010).Bacterial cellulose(BC)has the same molecular formula as plant cellulose, but with unique and sophisticated three-dimensional porous net-work structures.Intrinsically originated from the unique structure, BC demonstrates a serious of distinguished structural features and properties such as high purity,high degree of polymerization(up to8000),high crystallinity(of70–80%),high water content to99%, and high mechanical stability,which is quite different from the natural cellulose(Barud et al.,2011).These specific parameters are determined by the biofabrication approach of BC(Fig.1)and the controllable shape and supramolecular structure through the alter-ation of cultivation conditions during fermentation.These amazing physicochemical properties have attracted significant interest from both research scientists and industrialists.So far,BC has wide appli-cation in variousfields including medical,food,advanced acoustic diaphragms,and so on(Klemm et al.,2011).These research and exploration have led to the emergence of more diverse potential applications exploiting the functionality of BC nanomaterials.In view of expanding the scope of BC applications,it is important to take full advantage of the unique structure and properties of BC nanomaterials to develop novel BC-based nanomaterials with ground-breaking new features.Various modification methods have been explored to open up possibilities for endowing BC with new functionalities(Klemm et al.,2011).Simple biosynthetic or chemical modification on BC surface can improve its compatibility with different matrixes and expand its utilization in nano-related applications.Another notable feature of BC is its high aspect ratio and abundant active functional hydroxyl groups,which makes it suitable for combination with different nanostructures by provid-ing powerful interaction of BC with surrounding species,such as inorganic and polymeric nanoparticles and nanowires(Huang& Gu,2011).This novel concept breaks new ground to make optimum use of the specific chemical properties of the guest substrates,in association with the unique features of renewable BC resources. As a promising template in the synthesis of a great variety of nanostructures with designed properties and functionalities,BC can play the role of reducing agent,structure-directing agent and stabilizer(Yang,Xie,Deng,Bian,&Hong,2012).The polymeric or inorganic ingredients can be incorporated and cooperatively inter-act with BC nanofibers to attain some functional products.Such new high-value materials are the subject of continuing research and are commercially interesting in terms of new products from the inexhaustible and sustainable natural polymeric raw material.In the last few years,growing worldwide activity can be observed regarding extensive scientific investigation and increas-ing efforts for the practical use of the BC materials.There is an increasing annual publication activity on BC(also known as micro-bial cellulose or bacterial nanocellulose)since2000as presented in Fig.2.In recent years,the investigation and utilization of BC in functional materials have been the focus of research,and a growing number of works have been included in thisfield.Functional BC-based nanomaterials are especially an attractive topic because they enable the creation of materials with improved or new properties by mixing multiple constituents and exploiting synergistic effects, such as electronic,optical,magnetic,catalytic properties and bioac-tivity.With a special property or several remarkable functions, functional BC-based nanomaterials are a type of high value-added materials possessing potential applications in specificfields.In this review,recent developments on BC-based advanced functional nanomaterials including modified BC nanomaterials,functional BC-based nanocomposites and their applications will be discussed and reviewed.A variety of surface functionalization through biosyn-thetic or chemical modification will be considered,which can improve the functionality of BC nanomaterials and expandits Fig.1.The illustration of biofabrication process of BC.W.Hu et al./Carbohydrate Polymers101 (2014) 1043–10601045Fig.2.The illustration of the annual number of publications on BC since2000 (SciFinder Scholar search system,search term“bacterial cellulose”).potential applicationfields.Various approaches to the preparation of functional BC-based nanocomposites by incorporating different guest substrates including small molecules,inorganic nanoparti-cles or nanowires,and polymers on the surfaces of BC nanofibers are summarized,which mainly focus on the preparation methods,per-formances and some formation mechanisms of specific functional nanomaterials.2.Construction strategies of BC-based functional nanomaterialsAlthough BC nanomaterial has unique physical and chemical characteristics,its high degree of crystallinity and sole functional group lead to its poor dissolubility and processability,thereby limiting the applicationfields.BC possesses an abundance of hydroxyl groups on the surface,where modification can be easily achieved.It can be modified to achieve alternative functional groups and patterns of functionalization using in situ and ex situ modification methods as shown in Fig.3.The properties of BC derivatives are primarily determined by the type of the functional groups.In particular,modified BC with more than one functional group possessing different surface characteristics such as lipophilic–hydrophilic properties,magnetic and optical properties combined with a controlled pre-set functionalization pattern is in the center of interest.2.1.Biosynthetic modificationThe shape and supramolecular structure of BC can be controlled by the change of cultivation conditions such as the type of strain, carbon source,and additives(Heßler&Klemm,2009;Klemm et al., 2006).Studies have demonstrated the potential for manipulating the biogenesis of BC in order to produce modified BC nanofibers with a controlled composition,morphology,and properties.The inclusion of additives in the nutrient media components during biosynthesis can influence the assembly and microstructure of BC including the crystallinity,crystalline polymorphism,crystal-lite size and ribbon width.The presence of additives in the media may interfere with the bacterial cells or bind directly to the cel-lulose during production,thereby affecting the yield,structure, morphology and physical properties of BC.The in situ generation of composites can also be effectively regulated during biosynthesis by the inclusion of additives and dispersed particles.2.1.1.Altered BC structureAs a remarkable benefit of BC,the ultrastructure and morphol-ogy of BC can be altered by introducing additives not specifically required for bacterial cell growth in the media.The effects of the water-soluble agents in culture media on the aggregation and crystallization of BC microfibrils were intensively studied.Adding various water-soluble chemical reagents can modify the microfib-rillar features of the cellulosic ribbons.Adding nalidixic acid or chloramphenicol produced ribbons with an apparently larger width,probably because several ribbons from a cluster of cells whose dividing process was inhibited,combined or intertwined. While adding dithiothreitol produced ribbons with only45%width of the control ribbons on average(Yamanaka&Sugiyama,2000). Fig.3.Schematic illustration of the generalized synthetic routes to modified BC nanomaterials(Chen et al.,2010;Geng et al.,2011;Heßler&Klemm,2009;Hu,Chen,Xu, et al.,2011;Ifuku et al.,2009;Oshima et al.,2008;Shen et al.,2009;Yamanaka&Sugiyama,2000).1046W.Hu et al./Carbohydrate Polymers101 (2014) 1043–1060The pore system and water content of BC can be controlled in situ by the incorporation of water soluble polymers such as carboxymethylcellulose(CMC),hydroxypropylmethyl cellu-lose(HPMC),methylcellulose(MC),poly(vinyl alcohol)(PVA)and polyethylene glycol(PEG)to the culture medium(Seifert,Hesse, Kabrelian,&Klemm,2004).In the presence of these additives,the pore system and elasticity of BC,as well as the water adsorption and water holding capacity can be controlled.It has been claimed the addition of HPMC,CMC and MC can cause decreased crystallinity and crystal size,as well as greater thermal stability and pore size (Chen,Chen,Huang,&Lin,2011).Along with the formation of porelike network structure,the water retention ability and the ion absorption capacity increase.The functionalized BC produced with CMC also showed good adsorption performances for copper and lead ions(Chen et al.,2009).Nevertheless the presence of PVA in the culture medium results in a reduced water absorption ability and a slightly higher copper ion capacity in comparison with original BC.The addition of-cyclodextrin or PEG400causes a remarkable pore size increase(Heßler&Klemm,2009).Surprisingly,these co-substrates act as removable auxiliaries not incorporated in the BC samples.Other polymers such as Tween80,urea,fluorescent bright-ener and sodium alginate(NaAlg)have also been incorporated into the BC fermentation medium,with the observed differences in pore size,degree of polymerization,crystallinity,fiber widths and mechanical strength(Huang,Chen,Lin,Hsu,&Chen,2010; Ruka,Simon,&Dean,2013;Zhou,Sun,Hu,Li,&Yang,2007). The results revealed that the addition of urea can increase the mechanical strength.The bundle widths of BC produced withfluo-rescent brightener increased and the cellulose network void grew. BC produced with NaAlg added has a lower crystallinity,a smaller crystalline size and an enhanced yield.2.1.2.NanocompositesIn particular,the additives can be in situ incorporated into the growing BCfibrils to create a novel type of nanocomposites,which represents a very specific and important modification method of BC.This in situ technique can combine the properties of altered supramolecular structure of BC with those of the incorporated com-ponents.So far,researchers have put different efforts to obtain BC nanocomposites by applying various additives such as organic com-pounds,polymers and inorganic substances(Klemm et al.,2011).The biological fermentation of BC in presence of cationic starch leads to the formation of double-network BC composites by incor-poration of the starch derivative in the BC network(Heßler& Klemm,2009).This double-stage structure consists of an opaque upper and a transparent under part.As shown in Fig.4a,in case of the upper layer,the additive adsorbs at the cellulosefibers and causes an irregular distribution of the pore sizes.The under layer indicates the incorporation of the starch into the solvate shells of the BC prepolymer,forming an exciting skinnyfilm structure (Fig.4b).Other characteristic examples include the additives of poly(ethylene oxide)(Brown&Laborie,2008),PVA(Gea,Bilotti, Reynolds,Soykeabkeaw,&Peijs,2010)and starch(Grande et al., 2009)in the media for the formation of nanocomposites with the incorporation of these additives into the network of BC.Along with the increase of the additive content,the cellulose crystallized into smaller nanofibers,which further bonded together into bundles. The BC nanofibers were well dispersed in the composites and the nanocomposites typically show significantly improved mechanical properties.The inorganic additives can drastically modify the performance of BC.BC/multiwalled carbon nanotubes(MWCNTs)composites can be obtained in the presence of MWCNTs in an agitated cul-ture(Park,Kim,Kwon,Hong,&Jin,2009;Yan,Chen,Wang,Wang, &Jiang,2008).Interestingly,a core–shell structure model was demonstrated,with the packed MWNTs attached nascent sub-elementaryfibrils as the core of the cellulose assemblies as shown in Fig.4c.While in the static culture method,band-like assemblies with sharp bends and rigidness were produced in the presence of MWCNTs.Similarly,the crystallinity index,crystallite size,and cellulose I␣content also changed,which may be attributed to the interaction between the hydroxyl groups of treated MWCNTs and the sub-elementary BCfibrils,interfering with the aggre-gation and crystallization of BC microfibrils.By adding silica or titanium precursor into the static growth medium,the SiO2and TiO2nanoparticles about several tens of nanometers in size can be incorporated onto BC microfibrils(Geng et al.,2011;Yano,Maeda, Nakajima,Hagiwara,&Sawaguchi,2008)(Fig.4d).It is inferred that the basic proteins in the outer membrane of bacterium cell act as the catalyst for the hydrolysis and condensation of inorganic precur-sor,the surface of both outer membrane and BC nanofibers renders the nucleation and growth sites for inorganic nanoparticles.The space-temporal effect endows bacteria the delicate control ability over formation of the nanocomposites.2.2.Chemical surface modificationAlthough the biosynthetic modification of BC is a kind of green sustainable technology,the strict microbial fermentation environ-ment restricts the introduction of some additives.Other questions involving the interaction mechanism between the additives and microfibrils growth,as well as the structure control of BC nanofibers still need to be addressed.While chemically modified methods are not limited by the required types of pared to the biosynthesis method,its more clearly defined objectives and prin-ciples make it a feasible modified method for BC material.Since the chemical composition of BC is similar to plantfibers,it can also be carboxymethylated,acetylated,phosphorylated,and mod-ified by other graft copolymerization and crosslinking reaction to obtain a series of BC derivatives.The introduction of new functional groups to the BC structure can endow BC with various features such as hydrophobicity,ions adsorption capacity and optical proper-ties while maintaining the unique three-dimensional nano network and excellent mechanical properties of BC.In most cases,the modification of BC focuses on the improve-ment of its applicability and performance in different application fields.Several methods have been employed to achieve BC derivatives with improved metal ions absorption capacity.The functionalized diethylenetriamine-BC(EABC),amidoximated BC (Am-BC)and phosphorylated BC have been prepared as new adsor-bents for metal ions(Chen,Shen,Yu,Hu,&Wang,2010;Oshima, Kondo,Ohto,Inoue,&Baba,2008;Shen et al.,2009).The experi-mental data showed that the microporous network structure of BC was maintained after the modification and these novel adsorbent showed good adsorption performances for different metal ions.To anchor metallic ions on BC nanofibers,the carboxylate groups have been introduced onto the BC nanofiber surface using the2,2,6,6-tetramethylpiperidine-1-oxyradical(TEMPO)-mediated oxidation system(Ifuku,Tsuji,Morimoto,Saimoto,&Yano,2009).The oxida-tion can proceed under mild aqueous conditions maintaining the crystallinity and crystal size of BC nanofibers.In order to improve the dispersability and compatibility in dif-ferent solvents or matrices that are suitable in the production of nanocomposites,the acetylation of BC based on a non-swelling reaction mechanism was reported recently.BC could be partially acetylated by thefibrous acetylation method to modify its physical properties,while preserving the microfibrillar morphology(Kim, Nishiyama,&Kuga,2002).The hydrophobicity of the acetylated surface is advantageous for maintaining a large surface area on drying from water and would make the microfibrils compatible with other hydrophobic materials.While most of the chemicallyW.Hu et al./Carbohydrate Polymers101 (2014) 1043–10601047Fig.4.SEM images of freeze-dried upper part of BC/starch composite(a),under part of BC/starch composite(b),middle layer of BC/MWCNTs composites(c),and BC/SiO2 nanocomposites(d)(Geng et al.,2011;Heßler&Klemm,2009;Park et al.,2009).modified techniques require tedious solvent exchanges that strongly diminish the environmental benefit of the use of BC.A major thrust of recent BC modification research focused on the design of more economical and environmentally friendly method to obtain novel BC derivatives.Thus,a solvent-free derivatization system appears as an important goal to get BC derivative prod-ucts with hydrophobic surfaces with a minimum environment impact.In line with the solvent-free BC derivatization concept,BC preserving the microfibrillar morphology has been partially acety-lated using acetic anhydride in the presence of iodine as a catalyst (Hu,Chen,Xu,&Wang,2011).Another example is the reported vapor-phase technique which has been applied recently for the surface esterification of BC microfibrils with the help of gaseous trifluoroacetic acid mixed with acetylating agents(Berlioz,Molina-Boisseau,Nishiyama,&Heux,2009).Experimental results have shown the acetylation proceeded from the surface to the interior crystalline core of BC nanfibers.Hence,for moderate degree of sub-stitution,the surface was fully grafted whereas the cellulose core remained unmodified and the originalfibrous morphology was maintained.The obtained acetylated BC membrane shows more hydrophobic surface and good mechanical properties as shown in Fig.5,which is in favor of enhancing the hydrophobic non-polar polymeric matrix.2.3.In situ formation of nanostructuresBC has unique micro-nano porous three-dimensional network, which can facilitate the penetration of various metal ions into the interior.It also possesses a great deal of hydroxyl and ether bonds,forming the effective reactive sites to anchor metallic ions on the surface of the nanofibers.Then a variety of inorganic nanoparticles or nanowires can be formed through precipitation, oxidation–reduction and sol–gel reaction as shown in Fig.6.Dif-ferent from the doped nanoparticles into BC matrix,the size and morphology of the nanoparticles can be regulated by adjusting the structure of BC template and the in situ preparation condi-tions.At the same time,the nanospace in the BCfibers can behave as an effective nanoreactor to prevent the unwanted agglomera-tion phenomenon,ensuring the effective dispersion of the formed nanoparticles in the BC matrix.In the process of in situ preparation of BC-based nanocompos-ites,the second components such as metal and polymer in the form of particles orfibers can be introduced into the BC matrix,while retaining the unique three-dimensional nanoporous network of BC.BC can be viewed as a soft template to control the synthesis of desired nano-materials or nano-structure with specific size and shape.This can obtain a variety of novel functional materialswith Fig.5.(a)FE-SEM image of vacuum-dried acetylated BC,and the inset shows the profile of water droplets on the membrane surface.(b)Tensile stress–strain behaviors of air dried BC and acetylated BC(Hu,Chen,Xu,et al.,2011).1048W.Hu et al./Carbohydrate Polymers 101 (2014) 1043–1060Fig.6.Schematic diagram of the in situ preparation of nanoparticles/BC nanocomposites.unique features and outstanding ing BC as a tem-plate in the in situ synthesis of nano-materials has the following advantages:BC is a kind of environment-friendly and renewable material which can be produced from a wide range of raw mate-rials.The shape,structure and properties can be easily adjusted during the biosynthesis,pretreatment and chemical modification process.Furthermore,the BC matrix can be removed by calcination to obtain the pure inorganic nanoparticles and nanowires structure.Therefore,BC with controllable structure and pore size can provide restrictive environment to ensure a variety of nanostruc-tures effectively embedded in the matrix.The method does not require severe conditions,and is simple and easy to implement,which can obtain the nano-particles with narrow size distribution.So far,this method has been applied to synthesize different nano-materials such as metal,semiconductor and electrically conducting polymers through different preparation methods.2.3.1.In situ formation of nanostructures through reduction reactionThere have been some reports regarding the application of BC as a soft template to in situ form different metal nanoparticles by the reduction reaction as shown in Table 1.The size and mor-phology of the formed nanoparticles can be controlled by using different reducing agents including sodium borohydride (NaBH 4),triethanolamine,hydrazine (NH 2NH 2),hydroxylamine (NH 2OH),ascorbic acid,polyethylenimine (PEI)and so on (Barud et al.,2008;Yang,Xie,Hong,Cao,&Yang,2012;Zhang et al.,2010).These reducing agents can serve as an assistant material for stabilizing nanoparticles,preventing their aggregation.In addition,BC itself can be used as a reductant to produce metal nanoparticles (Yang,Xie,Deng,et al.,2012),without introducing additional reducing agent or stabilizing agent,thus avoiding the secondary pollutants and guaranteeing the feasibility of its applications in medical and catalytic field.2.3.2.In situ formation of nanostructures through precipitation reactionWell-separated nanofibrils of BC create an extensive surface area forming active sites for metal ion adsorption,and the sub-sequent introduction of precipitating agent will react with the immobilized metal ions to generate the initial nuclei of metal or oxide.Then the nuclei continue to grow,thereby further forming functional inorganic nanoparticles as shown in Fig.7.The growth of the nanoparticles was readily controlled by repeated alternating dipping of BC membranes in the metal ion and precipitating agent solution followed by a rinse step.So it is feasible for BC to serve as an excellent matrix in the synthesis of nanoparticles through the in situ precipitation reaction.CdS and CdSe nanoparticles have been synthesized and stabi-lized on BC nanofibers using in situ precipitation method (Li et al.,2009a;Yang et al.,2012a ).At first,hydroxyl and ether groups of BC anchored Cd 2+or thioglycolic acid capped Cd 2+,then the anchored Cd 2+reacted with S 2−or Se 2−to generate CdS or CdSe nanoparti-cles on the BC nanofibers as shown in Fig.8.The results indicatedthat nanoparticles with the diameter of 20–30nm deposited on BC nanofibres are well-dispersed in the BC nanofibre-network.AgCl nanoparticles with a size of several tenths of nanometers have been in situ synthesized in the three-dimensional non-woven network of BC nanofibrils (Hu et al.,2009).The growth of the nanoparticles was readily obtained by repeated alternating dipping of BC mem-branes in the solution of silver nitrate or sodium chloride followed by a rinse step.The nanopore is essential for introduction of silver ions and reaction with Cl −to form AgCl particles into BC fibers and removal of the excess chemicals from BC fibers.BC can serve as an excellent matrix in the in situ synthesis of the Fe 3O 4nanoparticles through the coprecipitation reaction (Zhang et al.,2011).The ultrafine network architecture of BC gives a good tunnel for Fe 2+/Fe 3+adsorption and ensures the effec-tive anchoring of absorbed ions onto the BC nanofibers through ion–dipole interactions.After being rinsed with distilled water to remove the unanchored ions,the obtained ions/BC complexes were immersed into the excessive NaOH solution.The interaction between Fe 2+/Fe 3+and OH −can induce the formation of Fe 3O 4nanoparticles with the diameter of 80–100nm as shown in Fig.9.During the in situ formation process of different nanostruc-tures,the size and size distribution of the formed nanoparticles are controllable by adjusting synthetic parameters such as the con-centration of metal ions.When the concentration of metal ions is too high,the BC nanofibers fails to fully immoblize and dis-perse the large number of metal ions due to the limited hydroxyl reactive sites shown in Fig.10.Meanwhile,the presence of excess metal ions would result in the agglomeration of the nanoparticles due to the higher aggregation speed than the orientation speed in the particle growth process.Therefore the concentration of the reaction solution needs to be reduced appropriately to ensure the metal ions and nanoparticles effectively dispersed and fixed onto the surface of the nanofibers.However,when the concentra-tion is too low,the loading amount of the nanoparticles will be significantly reduced,which would degrade the performance of the final products.The exploration of optimal reaction conditions would be particularly important to achieve the effective dispersion of nanoparticles with sufficient loading amount,thereby obtain-ing the functional nanocomposite materials with excellent optical,electrical,magnetic and antibacterial properties.2.3.3.In situ formation of nanostructures through sol-gel reactionTaking advantage of the ultrafine nanofibrous structure,as well as abundant porous channels inside the network,BC nanomateri-als can be used as the templates to in situ prepare metal oxides through the sol–gel reaction.The schematic diagram of the in situ preparation process is described in Fig.11.When immersing BC into the precursor solution,the inorganic ions can be immobilized onto the BC nanofibers.Then it can be solidified to form the gel through hydrolysis and condensation,and subsequently heated to form the desired oxides.BC template can stabilize and disperse the formed oxides nanoparticles through van der Waals forces and hydrogen bonding interaction,prompting generated nanoparticles distributed along the surface of nanofibers.The BC template can。
遗传育种相关名词中英文对照
遗传育种相关名词中英文对照中英文对照的分子育种相关名词 3"untranslated region (3"UTR) 3"非翻译区 5"untranslated region (5; UTR) 5"非翻译区 A chromosome A 染色体 AATAAA 多腺苷酸化信号aberration 崎变 abiogenesis 非生源说 accessory chromosome 副染色体 accessory nucleus 副核 accessory protein 辅助蛋白 accident variance 偶然变异 Ac-Ds system Ac-Ds 系统 acentric chromosome 无着丝粒染色体acentric fragment 无着丝粒片段 acentric ring 无着丝粒环 achromatin 非染色质 acquired character 获得性状acrocentric chromosome 近端着丝粒染色体 acrosyndesis 端部联会 activating transcription factor 转录激活因子activator 激活剂 activator element 激活单元 activator protein( AP)激活蛋白 activator-dissociation system Ac-Ds 激活解离系统 active chromatin 活性染色质 activesite 活性部位 adaptation 适应 adaptive peak 适应高峰adaptive surface 适应面 addition 附加物 addition haploid 附加单倍体 addition line 附加系 additiveeffect 加性效应 additive gene 加性基因 additive genetic variance 加性遗传方差additive recombination 插人重组additive resistance 累加抗性 adenosine 腺昔adenosine diphosphate (ADP )腺昔二鱗酸adenosine triphosphate( ATP)腺昔三憐酸adjacent segregation 相邻分离A- form DNA A 型 DNAakinetic chromosome 无着丝粒染色体akinetic fragment 无着丝粒片断alien addition monosomic 外源单体生物alien chromosome substitution 外源染色体代换alien species 外源种 alien-addition cell hybrid 异源附加细胞杂种 alkylating agent 焼化剂 allele 等位基因allele center 等位基因中心 allele linkage analysis 等位基因连锁分析 allele specific oligonucleotide(ASO)等位基因特异的寡核苷酸 allelic complement 等位(基因)互补 allelic diversity 等位(基因)多样化 allelic exclusion 等位基因排斥 allelic inactivation 等位(基因)失活 allelic interaction 等位(基因)相互作用allelic recombination 等位(基因)重组 allelicreplacement 等位(基因)置换 allelic series 等位(基因)系列 allelic variation 等位(基因)变异 allelism 等位性 allelotype 等位(基因)型 allodiploid 异源二倍体 allohaploid 异源单倍体 allopatric speciation 异域种alloploidy 异源倍性 allopolyhaploid 异源多倍单倍体allopolyploid 异源多倍体 allosyndesis 异源联会allotetraploid 异源四倍体 alloheteroploid 异源异倍体alternation of generation 世代交替 alternative transcription 可变转录 alternative transcription initiation 可变转录起始 Alu repetitive sequence, Alu family Alu 重复序列,Alu 家族ambiguous codon 多义密码子 ambisense genome 双义基因组 ambisense RNA 双义 RNA aminoacyl-tRNA binding site 氨酰基 tRNA 接合位点 aminoacyl-tRNA synthetase 氨酰基 tRNA 连接酶 amixis 无融合amorph 无效等位基因amphidiploid 双二倍体amphipolyploid 双多倍体amplicon 扩增子amplification 扩增 amplification primer 扩增引物analysis of variance 方差分析 anaphase (分裂)后期anaphase bridge (分裂)后期桥anchor cell 锚状细胞 androgamete 雄配子aneuhaploid 非整倍单倍体aneuploid 非整倍体 animal genetics 动物遗传学annealing 复性 antibody 抗体anticoding strand 反编码链anticodon 反密码子anticodon arm 反密码子臂anticodon loop 反密码子环 antiparallel 反向平行antirepressor 抗阻抑物antisense RNA 反义 RNAantisense strand 反义链 apogamogony 无融合结实apogamy 无配子生殖apomixis 无融合生殖 arm ratio (染色体)臂比artificial gene 人工基因 artificial selection 人工选择 asexual hybridization 无性杂交 asexual propagation 无性繁殖 asexual reproduction 无性生殖assortative mating 选型交配 asynapsis 不联会 asynaptic gene 不联会基因atavism 返祖 atelocentric chromosome 非端着丝粒染色体 attached X chromosome 并连 X 染色体 attachmentsite 附着位点 attenuation 衰减 attenuator 衰减子autarchic gene 自效基因auto-alloploid 同源异源体 autoallopolyploid 同源异源多倍体 autobivalent 同源二阶染色体 auto-diploid 同源二倍体;自体融合二倍体 autodiploidization 同源二倍化autoduplication 自体复制 autogenesis 自然发生autogenomatic 同源染色体组 autoheteroploidy 同源异倍性autonomous transposable element 自主转座单元autonomously replicating sequence(ARS)自主复制序列autoparthenogenesis 自发单性生殖 autopolyhaploid 同源多倍单倍体 autopolyploid 同源多倍体 autoradiogram 放射自显影图 autosyndetic pairing 同源配对 autotetraploid 同源四倍体 autozygote 同合子 auxotroph 营养缺陷体 B chromosome B 染色体 B1,first backcross generation 回交第一代 B2,second backcross generation 回交第二代back mutation 回复突变 backcross 回交backcross hybrid 回交杂种 backcross parent 回交亲本 backcross ratio 回交比率 background genotype 背景基因型 bacterial artification chromosome( BAC )细菌人工染色体Bacterial genetics 细菌遗传学 Bacteriophage 噬菌体balanced lethal 平衡致死 balanced lethal gene 平衡致死基因 balanced linkage 平衡连锁 balanced load 平衡负荷balanced polymorphism 平衡多态现象 balanced rearrangements 平衡重组balanced tertiary trisomic 平衡三级三体balanced translocation 平衡异位balancing selection 平衡选择band analysis 谱带分析 banding pattern (染色体)带型basal transcription apparatus 基础转录装置 base analog 碱基类似物base analogue 类減基base content 减基含量base exchange 碱基交换 base pairing mistake 碱基配对错误 base pairing rules 碱基配对法则 base substitution 减基置换 base transition 减基转换 base transversion 减基颠换 base-pair region 碱基配对区base-pair substitution 碱基配对替换 basic number of chromosome 染色体基数 behavioral genetics 行为遗传学behavioral isolation 行为隔离 bidirectionalreplication 双向复制 bimodal distribution 双峰分布binary fission 二分裂binding protein 结合蛋白binding site 结合部位 binucleate phase 双核期biochemical genetics 生化遗传学 biochemical mutant 生化突变体biochemical polymorphism 生化多态性 bioethics 生物伦理学 biogenesis 生源说 bioinformatics 生物信息学biological diversity 生物多样性 biometrical genetics 生物统计遗传学(简称生统遗传学) bisexual reproduction 两性生殖 bisexuality 两性现象 bivalent 二价体 blending inheritance 混合遗传 blot transfer apparatus 印迹转移装置 blotting membrane 印迹膜 bottle neck effect 瓶颈效应 branch migration 分支迁移 breed variety 品种breeding 育种,培育;繁殖,生育 breeding by crossing 杂交育种法 breeding by separation 分隔育种法 breeding coefficient 繁殖率 breeding habit 繁殖习性 breeding migration 生殖回游,繁殖回游 breeding period 生殖期breeding place 繁殖地 breeding population 繁殖种群breeding potential 繁殖能力,育种潜能 breeding range繁殖幅度 breeding season 繁殖季节 breeding size 繁殖个体数 breeding system 繁殖系统 breeding true 纯育breeding value 育种值 broad heritability 广义遗传率bulk selection 集团选择 C0,acentric 无着丝粒的Cl,monocentric 单着丝粒 C2, dicentric 双着丝粒的C3,tricentric 三着丝粒的 candidate gene 候选基因candidate-gene approach 候选基因法 Canpbenmodel 坎贝尔模型carytype 染色体组型,核型 catabolite activator protein 分解活化蛋白catabolite repression 分解代谢产物阻遏catastrophism 灾变说 cell clone 细胞克隆 cell cycle 细胞周期 cell determination 细胞决定 cell division 细胞分裂 cell division cycle gene(CDC gene) 细胞分裂周期基因 ceU division lag 细胞分裂延迟 cell fate 细胞命运cell fusion 细胞融合 cell genetics 细胞的遗传学 cell hybridization 细胞杂交 cell sorter 细胞分类器 cell strain 细胞株 cell-cell communication 细胞间通信center of variation 变异中心 centimorgan(cM) 厘摩central dogma 中心法则 central tendency 集中趋势centromere DNA 着丝粒 DNA centromere interference 着丝粒干扰centromere 着丝粒 centromeric exchange ( CME)着丝粒交换centromeric inactivation 着丝粒失活 centromeric sequence( CEN sequence)中心粒序列 character divergence 性状趋异chemical genetics 化学遗传学chemigenomics 化学基因组学chiasma centralization 交叉中化chiasma terminalization 交叉端化chimera 异源嵌合体Chi-square (x2) test 卡方检验 chondriogene 线粒体基因 chorionic villus sampling 绒毛膜取样 chromatid abemition 染色单体畸变chromatid break 染色单体断裂chromatid bridge 染色单体桥chromatid interchange 染色单体互换 chromatid interference 染色单体干涉 chromatid segregation 染色单体分离chromatid tetrad 四分染色单体chromatid translocation 染色单体异位chromatin agglutination 染色质凝聚chromosomal aberration 染色体崎变chromosomal assignment 染色体定位chromosomal banding 染色体显带chromosomal disorder 染色体病chromosomal elimination 染色体消减 chromosomal inheritance 染色体遗传chromosomal interference 染色体干扰chromosomal location 染色体定位chromosomal locus 染色体位点 chromosomal mutation 染色体突变chromosomal pattern 染色体型chromosomal polymorphism 染色体多态性 chromosomal rearrangement 染色体质量排chromosomal reproduction 染色体增殖chromosomal RNA 染色体 RNAchromosomal shift 染色体变迁,染色体移位chromosome aberration 染色体畸变 chromosome arm 染色体臂chromosome association 染色体联合chromosome banding pattern 染色体带型chromosome behavior 染色体动态chromosome blotting 染色体印迹chromosome breakage 染色体断裂chromosome bridge 染色体桥 chromosome coiling 染色体螺旋chromosome condensation 染色体浓缩chromosome constriction 染色体缢痕chromosome cycle 染色体周期chromosome damage 染色体损伤chromosome deletion 染色体缺失chromosome disjunction 染色体分离chromosome doubling 染色体加倍chromosome duplication 染色体复制chromosome elimination 染色体丢失 chromosome engineering 染色体工程chromosome evolution 染色体进化 chromosome exchange 染色体交换chromosome fusion 染色体融合 chromosome gap 染色体间隙chromosome hopping 染色体跳移chromosome interchange 染色体交换chromosome interference 染色体干涉chromosome jumping 染色体跳查chromosome knob 染色体结 chromosome loop 染色体环chromosome lose 染色体丢失chromosome map 染色体图 chromosome mapping 染色体作图chromosome matrix 染色体基质chromosome mutation 染色体突变 chromosome non-disjunction 染色体不分离 chromosome paring 染色体配对chromosome polymorphism 染色体多态性 chromosome puff 染色体疏松 chromosome rearrangement 染色体质量排chromosome reduplication 染色体再加倍 chromosome repeat 染色体质量叠 chromosome scaffold 染色体支架chromosome segregation 染色体分离 chromosome set 染色体组chromosome stickiness 染色体粘性chromosome theory of heredity 染色体遗传学说chromosome theory of inheritance 染色体遗传学说chromosome thread 染色体丝chromosome walking 染色体步查chromosome-mediated gene transfer 染色体中介基因转移 chromosomology 染色体学 CIB method CIB 法;性连锁致死突变出现频率检测法 circular DNA 环林 DNA cis conformation 顺式构象 cis dominance 顺式显性 cis-heterogenote 顺式杂基因子 cis-regulatory element 顺式调节兀件 cis-trans test 顺反测验cladogram 进化树 cloning vector 克隆载体 C-meiosis C 减数分裂C-metaphase C 中期C-mitosis C 有丝分裂 code degeneracy 密码简并coding capacity 编码容量 coding ratio 密码比 coding recognition site 密码识别位置 coding region 编码区coding sequence 编码序列 coding site 编码位置 coding strand 密码链 coding triplet 编码三联体 codominance 共显性 codon bias 密码子偏倚 codon type 密码子型coefficient of consanguinity 近亲系数 coefficient of genetic determination 遗传决定系数 coefficient of hybridity 杂种系数 coefficient of inbreeding 近交系数coefficient of migration 迁移系数 coefficient of relationship 亲缘系数 coefficient of variability 变异系数 coevolution 协同进化 coinducer 协诱导物 cold sensitive mutant 冷敏感突变体colineartiy 共线性combining ability 配合力comparative genomics 比较基因组学competence 感受态competent cell 感受态细胞competing groups 竞争类群 competition advantage 竞争优势competitive exclusion principle 竞争排斥原理complementary DNA (cDNA)互补 DNAcomplementary gene 互补基因 complementation test 互补测验complete linkage 完全连锁 complete selection 完全选择 complotype 补体单元型 composite transposon 复合转座子 conditional gene 条件基因 conditional lethal 条件致死conditional mutation 条件突变 consanguinity 近亲consensus sequence 共有序列 conservative transposition 保守转座 constitutive heterochromatin 组成型染色质continuous variation 连续变异convergent evolution 趋同进化cooperativity 协同性 coordinately controlled genes 协同控制基因 core promoter element 核心启动子 core sequence 核心序列 co-repressor 协阻抑物correlation coefficient 相关系数 cosegregation 共分离 cosuppression 共抑制cotranfection 共转染cotranscript 共转录物 cotranscriptional processing 共转录过程 cotransduction 共转导cotransformation 共转化 cotranslational secrection 共翻译分泌counterselection 反选择coupling phase 互引相 covalently closed circular DNA(cccDNA)共价闭合环状 DNAcovariation 相关变异criss-cross inheritance 交叉遗传 cross 杂交crossability 杂交性crossbred 杂种cross-campatibility 杂交亲和性 cioss-infertility 杂交不育性 crossing over 交换crossing-over map 交换图crossing-over value 交换值crossover products 交换产物 crossover rates 交换率crossover reducer 交换抑制因子crossover suppressor 交换抑制因子crossover unit 交换单位 crossover value 值crossover-type gamete 交换型配子C-value paradox C 值悖论 cybrid 胞质杂种 cyclin 细胞周期蛋白cytidme 胞苷 cytochimera 细胞嵌合体cytogenetics 细胞遗传学 cytohet 胞质杂合子cytologic 细胞学的cytological map 细胞学图cytoplasm 细胞质cytoplasmic genome 胞质基因组 cytoplasmic heredity 细胞质遗传 cytqplasmic incompatibility 细胞质不亲和性cytoplasmic inheritance 细胞质遗传cytoplasmic male sterility 细胞质雄性不育cytoplasmic mutation 细胞质突变 cytofdasmic segregation 细胞质分离cytoskeleton 细胞骨架Darwin 达尔文 Darwinian fitness 达尔文适合度Darwinism 达尔文学说 daughter cell 子细胞 daughter chromatid 子染色体 daughter chromosome 子染色体deformylase 去甲酰酶 degenerate code 简并密码degenerate primer 简并引物 degenerate sequence 简并序列 degenerated codon 简并密码子degeneration 退化 degree of dominance 显性度delayed inheritance 延迟遗传 deletant 缺失体deletion 缺失。
常用免疫学名词解释
附录:常用免疫学名词解释Aabsorption吸收应用特异性抗原与溶液中的抗体结合,形成不溶性复合物而除去抗体,例如用此法处理血清,即称为吸收血清。
用作吸收的抗原称为吸收剂。
accessory cell辅佐细胞特异性免疫应答需要的细胞,但不是实际介导的,通常用于描述抗原呈递细胞(APC)。
acquired immunity获得性免疫机体在生活过程中所获得的免疫力,称为获得性免疫,它与先天性免疫或天然免疫相反。
获得性免疫可分为:自动免疫,被动免疫,体液免疫与细胞免疫。
参看适应性免疫(adaptive immunity),过继性免疫(adoptive immunity),免疫耐受(immune tolerance)。
acquired immunodeficiency Sydrom(AIDS)获得性免疫缺陷综合征(艾滋病)由人类免疫缺陷病毒(HIV)所致的免疫缺陷病。
HIV感染主要引起T淋巴细胞CD4亚群的极度减少。
患者表现为迟发型超敏反应降低或消失,对机会感染菌极其易感,易发生某些少见的,如Kaposi氏肉瘤或Burkitt氏淋巴瘤。
HIV也可引起B 淋巴细胞多克隆性扩增,导致高丙种球蛋白血症。
尽管血清中免疫球蛋白量明显增加,但对抗原不能发生免疫反应。
这种综合征发生于“危险”人群,包括同性恋的男子,滥用静脉药物者,血液或血液制品的接受者,以及某些来自中非或加勒比海的人群。
在“危险”人群的异性伙伴中和AIDS母亲的婴儿中也已发现了这种综合症。
active immunization主动免疫(作用)抗原进入机体起免疫应答(自动免疫)adaptation tolerance适应性耐受生物在长期进化过程,宿主与寄生物在相互反应中,宿主的防御能力选择性地被减弱。
adaptative immunity适应免疫机体与抗原接触而发生的免疫力(包括主动体液免疫与主动细胞免疫)。
adherent cell粘附细胞在体外能粘附于表面的细胞。
2013-Probucol prevents blood–brain barrier dysfunction in wild-type mice普罗布考 血脑屏障功能障碍
Probucol prevents blood–brain barrier dysfunction in wild-type mice induced by saturated fat or cholesterol feedingRyusuke Takechi,*†Susan Galloway,*Menuka M Pallebage-Gamarallage,*Virginie Lam,*Satvinder S Dhaliwal*†and John C Mamo*†*Faculty of Health Sciences,School of Public Health,Curtin Health Innovation Research Institute Biosciences Research Precinct,Curtin University,Bentley,and†Centre for Metabolic Fitness,Australian Technology Network,Perth,WA,AustraliaSUMMARY1.Dysfunction of the blood–brain barrier(BBB)is an early pathological feature of vascular dementia and Alzheimer’s disease(AD)and is triggered by inflammatory stimuli.Probu-col is a lipid-lowering agent with potent anti-oxidant proper-ties once commonly used for the treatment of cardiovascular disease.Probucol therapy was found to stabilize cognitive symptoms in elderly AD patients,whereas in amyloid trans-genic mice probucol was shown to attenuate amyloidosis. However,the mechanisms underlying the effects of probucol have note been determined.2.In the present study we investigated whether probucol can prevent BBB disturbances induced by chronic ingestion of proinflammatory diets enriched with either20%(w/w)sat-urated fats(SFA)or1%(w/w)cholesterol.Mice were fed the diets for12weeks before they were killed and BBB integrity was measured.3.Mice maintained on either the SFA-or cholesterol-sup-plemented diets were found to have a30-and sevenfold greater likelihood of BBB dysfunction,respectively,as deter-mined by the parenchymal extravasation of plasma-derived immunoglobulins and endogenous lipoprotein enrichment with b-amyloid.In contrast,mice fed the SFA-or cholesterol-enriched diets that also contained1%(w/w)probucol showed no evidence of BBB disturbance.The parenchymal expression of glialfibrillary acidic protein,a marker of cerebrovascular inflammation,was significantly greater in mice fed the SFA-enriched diet.Plasma lipid,b-amyloid and apolipoprotein B levels were not increased by feeding of the SFA-or cholesterol-enriched diets.However,mice fed the SFA-or cholesterol-enriched diets did exhibit increased plasma non-esterified fatty acid levels that were not reduced by probucol.4.The data suggest that probucol prevents disturbances of BBB induced by chronic ingestion of diets enriched in SFA or cholesterol by suppressing inflammatory pathways rather than by modulating plasma lipid homeostasis.Key words:blood–brain barrier,cholesterol,neuroinflam-mation,probucol,saturated fatty acid,vascular dementia.INTRODUCTIONDisturbed blood–brain barrier(BBB)function,astroglial cell acti-vation and endothelial degeneration are common early pathologi-cal features of vascular dementia(VaD)and Alzheimer’s disease (AD).1–3Plasma proteins that are normally excluded from the brain parenchyma are found to increasingly penetrate as disease progresses,potentially creating a vicious inflammatory cycle.4,5 Elevated plasma cholesterol,high blood pressure and exposure to environmental toxins,such as those derived from smoking,are risk factors for VaD and AD.6,7However,in AD cerebrovascular dysfunction is also exacerbated because of the formation of pro-teinaceous deposits that are enriched in b-amyloid(A b). Several lines of evidence suggest that the risk of developing VaD and AD is modulated by diet.7,8Population studies have shown that the intake of saturated fats(SFA)and cholesterol is a positive risk factor,whereas polyunsaturated oils,particularly the x-3and x-6fatty acids confer protection.8–10Direct evidence of a dietary lipid–cerebrovascular link also comes from animal stud-ies.Wild-type mice or rabbits maintained on hyperlipidaemic ath-erogenic diets exhibit compromised BBB function and the extravasation of plasma immunoglobulins;8,11,12in mice geneti-cally engineered to overexpress A b,atherogenic SFA-or choles-terol-supplemented diets significantly accelerate amyloid deposition,whereas docosahexaenoic fatty acid-enriched diets suppresses cerebral A b.13,14The mechanisms by which dietary SFA and cholesterol influ-ence inflammatory pathways that compromise cerebrovascular integrity are currently unknown.Both lipids stimulate synthesis and secretion of pro-atherogenic apolipoprotein(apo)B lipopro-teins from lipogenic organs and this may occur commensurate with the enrichment of these macromolecules with A b.15–18 Thesefindings suggest that exaggerated exposure to plasma lipids,apoB,lipoproteins or A b associated with these macromolecules induce cerebrovascular disturbances.Consistent with this hypothesis,studies in three strains of amyloid transgenic mice with different susceptibility to disease have shown that the secretion into the blood of apoB–lipoprotein–A b was strongly associated with disease onset and progression.19In human cada-Correspondence:Professor John Mamo,Faculty of Health Sciences,School of Public Health,Curtin Health Innovation Research Institute Bio-sciences Research Precinct,Curtin University,GPO Box U1987,Perth,WA6845,Australia.Email:J.Mamo@.auReceived10July2012;revision24October2012;accepted15Novem-ber2012.©2012The AuthorsClinical and Experimental Pharmacology and Physiology©2012Wiley Publishing Asia Pty LtdClinical and Experimental Pharmacology and Physiology(2013)40,45–52doi:10.1111/1440-1681.12032ver specimens,immunoreactive apoB was clearly evident in amy-loid plaque20and,similarly,significant parenchymal colocaliza-tion of apoB with early diffuse amyloid plaque has been reported in transgenic amyloid mice.21Probucol is a cholesterol-lowering agent once commonly used to reduce the risk of coronary artery disease.22,23However,in addition to its lipid-lowering actions,probucol has been shown to have effects that could confer significant cerebrovascular benefits. For example,probucol has a suppressive effect on lipoprotein biogenesis and,by extension,may limit A b secretion,24thereafter reducing BBB exposure to circulating lipoprotein–A b.A syner-gistic effect of probucol on A b kinetics is the possibility of enhanced clearance of lipoprotein–A b via receptor-mediated path-ways.25Alternatively and independent of lipoprotein–A b homeo-stasis,probucol is an exceedingly potent anti-oxidant that can prevent membrane degradation under conditions of oxidative stress.22,23In addition to protein and lipid products,mitochon-drial hyperactivity is a hallmark feature of cerebrovascular and neuronal atrophy.26Notionally,probucol could attenuate cerebro-vascular and neuronal proinflammatory pathways.In the present study,the putative efficacy of probucol in maintaining cerebro-vascular integrity was explored in established murine models of dietary-induced BBB dysfunction.8,12,18,27METHODSAnimalsSix-week-old female C57BL/6J mice were purchased from the Animal Resources Centre(Murdoch,WA,Australia).Mice were randomly allocated to receive either a low-fat(LF)control diet or the LF diet enriched with SFA(20%w/w)or cholesterol(1%w/ w).Another two groups of mice were simultaneously fed either the SFA-or cholesterol-enriched diet and treated with probucol. The feed preparations were made by Specialty Feeds(Perth,WA, Australia).The LF control diet was standard AIN93M rodent chow containing<4%(w/w)fat as polyunsaturates and was free of cholesterol(for details of the composition of the different diets,see Table S1available as Supplementary Material to this paper).The SFA in the SFA-enriched diet was derived primarily from cocoa butter.To achieve a daily dose of probucol of approximately30mg/day,the drug was incorporated into the chow at a concentration of1%w/w because mice consume approximately3g chow daily.24,28Mice were maintained in an accredited animal holding facility with regulated temperature,air pressure and lighting(12h light–dark cycle).Mice had ad libitum access to feed and water.Three months after the commencement of the dietary intervention,eight mice from each group were killed by cardiac exsanguination under complete anaesthesia(pentobarbitone).All experimental procedures in this study were approved by a National Health and Medical Research Council of Australia-accredited Animal Ethics Committee(Curtin University approval no.R34/08).Collection of tissue and plasma samplesFollowing3months dietary intervention,brain tissue and plasma samples were collected as described previously.12Briefly,mice were anaesthetized with pentobarbitone and blood samples were obtained by cardiac puncture.Plasma was separated by low-speed centrifuga-tion(2500g for10min at5°C)and stored immediately atÀ80°C. Brains were carefully removed and washed in chilled phosphate-buf-fered saline(PBS).For immunofluorescent microscopy,the right hemisphere was segmented andfixed in4%paraformaldehyde for 24h followed by cryoprotection in20%sucrose solution for3days at4°C.Tissues were then frozen in isopentane–dry ice and stored at À80°C.The left hemisphere wasfixed in4%paraformaldehyde for 24h and processed using a Leica TP1020tissue processor(Leica, Wetzlar,Germany)before being embedded in paraffin.Immunofluorescent detection of IgG and apoB in cerebral tissueThe integrity of the BBB was assessed by the immunofluorescent detection of plasma IgG and apoB immunoreactivity in the brain parenchyma,as described previously.8,12,21,29Briefly,18l m cryosection specimens were prepared from the right hemisphere of each mouse.For IgG detection,sections were incubated with Alexa488-conjugated polyclonal goat anti-mouse IgG antibody (1:100;Invitrogen,Carlsbad,CA,USA)for2h at room tem-perature.Subsequently,the sections were imaged using an invertedfluorescent microscope(AxioVert200M;Zeiss,Oberko-chen,Germany)and AXIOVISION software version4.6(Zeiss).The apoB lipoproteins were detected by overnight incubation of the tissue samples with polyclonal rabbit anti-apoB antibody (1:200;Abcam,Cambridge,UK).The primary antibody was then visualized with Alexa488-conjugated goat anti-rabbit IgG (Invitrogen).Negative controls were included for all immunofluorescence experiments,in which the primary antibody was replaced with buffer or an irrelevant serum.Fluorescent staining was not observed for any negative controls.Immunofluorescent detection of glialfibrillary acidic protein As a marker of the cerebral inflammatory response following die-tary and drug intervention,activated glial cells were measured using semiquantitative three-dimensional(3D)immunofluorescent microscopy.Cryosections(20l m)were prepared and blocked with anti-goat serum for30min before rabbit anti-mouse glial fibrillary acidic protein(GFAP)was applied to the sections (1:250;Abcam)for20h at4°C.After a thorough wash with PBS,the sections were incubated with Alexa480-conjugated goat anti-rabbit IgG(1:100;Invitrogen)for1h at room temperature and nuclei were counterstained with4′,6′-diamidino-2-phenylin-dole.Sections were subsequently imaged under an invertedfluo-rescent microscope(AxioVert200M;Zeiss)using AXIOVISION software version4.6(Zeiss).Fluorescent image capture and semiquantitative measurementsAllfluorescent images were captured using the AxioVert200M (Zeiss)fluorescent microscope coupled to an MRm digital camera (Zeiss)and managed by AXIOVISION software(Zeiss).Three-dimensional images were taken using the ApoTome optical sec-tioning technique(Zeiss).Quantification was determined within46R Takechi et al.the cortex excluding the hippocampus(CTX)and in the hippocampal formation(HPF)in the approximate stereotaxic areas of 1.7mm interaual and 2.1mm Bregma,as described previously.12For each mouse,a minimum of three cryosection specimens was prepared from the right hemisphere.For each specimen,up to seven3D ApoTome images were taken at random within each designated region of the brain.For quantitative measurement of IgG and apoB,images were captured at9200magnification (4309322l m).Each3D image consisted of between6and13 Z-stack images and the distance between the Z-stack slices was 1.225l m optimized by Nyquist theory(29oversampling in axial direction).The optical density sum for the protein of inter-est was determined in three dimensions(138891040pixel two-dimensional planes)using the automated optical density mea-surement tool(AXIOVISION;Zeiss).The total optical density sum of the entire3D image was then divided by the number of Z-stack images and is expressed as per volume unit. Immunohistochemical staining of CD68Immunoreactivity to CD68was investigated in the present study as a marker of activated microglia.Paraffin-embedded sections (20l m)were deparaffinized in xylene and rehydrated.After blocking with3%hydrogen peroxide in methanol for30min and 20%goat serum in PBS for30min,sections were incubated with rabbit anti-mouse CD68(1:150;Abcam)for20h at4°C.The sections were subsequently incubated with horseradish peroxidase (HRP)-conjugated anti-rabbit IgG for45min at20°C(1:500; Dako,Glostrup,Denmark).Positive immunostaining was visual-ized using a liquid diaminobenzidine plus substrate chromogen kit(DAKO).All sections were counterstained with Harris’hae-matoxylin.Determination of plasma S100calcium-binding protein B and A b levelsPlasma S100calcium-binding protein B(S100B)levels were measured by ELISA(CosmoBio,Tokyo,Japan)according to the manufacturer’s instructions.Briefly,20l L plasma samples or 20l L S100B standards(0,98,197,394,1575,3150and 6300pg/mL)were incubated overnight at4°C in96-well micro-plates coated with the primary antibody.Thereafter,plates were incubated with conjugated secondary antibody for2h,followed by2h incubation with streptavidin–HRP.Finally,samples were incubated with substrate solution for20min and the reaction was terminated with stopping solution.Optical absorbance was mea-sured at490nm.Plasma concentrations of mouse A b(1–40)and A b(1–42)were determined by ELISA(Invitrogen)according to the manufac-turer’s instructions.Briefly,100l L of fourfold-diluted plasma samples or A b standards(A b(1–40):0,1,2.5,5.0,10.0,25.0, 50.0,100pmol/L;A b(1–42):0,0.1,0.5, 1.0, 2.0, 5.0,10.0, 20.0pmol/L)were dispensed into wells,incubated overnight at 4°C and then washed thoroughly with washing solution provided with the kits.The HRP-conjugated A b antibody was then added and samples were incubated for1h at room temperature. According to the instructions provided with the kits,TMB solution was added and samples were left for30min in the dark before the reaction was terminated by the addition of stopping solution.Optical absorbance was measured at450nm.Plasma lipidsPlasma triglyceride and total cholesterol levels were measured using colorimetric assays(Randox,Crumlin,UK),according to the instructions provided by the manufacturer with minor modifi-cations.Briefly,2l L plasma samples or standards were loaded to96-well microplates,200l L reaction solution was then added and samples were incubated for5min at37°C.Optical absor-bance was read at550nm.Plasma non-esterified fatty acids(NEFA)were measured using a colorimetric assay(Wako,Tokyo,Japan)according manufac-turer’s instructions.Briefly,7l L plasma samples or standards were loaded onto96-well microplates,300l L Reagent1was added and samples were incubated for3min at37°C before the addition of150l L Reagent2and a further incubation for 4.5min at37°C.Optical absorbance was read at550nm. Measurement of plasma apoBA method for the quantification of plasma apoB100and apoB48 was established by modifying previously published methods for human apoB using sodium dodecyl sulphate(SDS)–polyacryl-amide gel electrophoresis and western blotting.30Briefly,2l L plasma samples were mixed with25l L NuPAGE sample buffer (Invitrogen),6l L of25%2-mercaptoethanol and67l L water. Then,samples(20l L)were loaded onto3–8%gradient Tris-ace-tate SDS-NuPAGE gels(Invitrogen)and electrophoresed at 150V for approximately1.5h.Isolated apoB proteins were then electrotransferred to polyvinylidene difluoride membranes at 40V for1h.Following30min blocking with2%skim milk in Tris-buffered saline Tween-20(TBST),membranes were incu-bated with anti-apoB antibody(1:100;Abcam)for2h. Subsequently,membranes were incubated with HRP-conjugated anti-rabbit IgG(1:30000;Dako)for45min.Enhanced chemi-luminescence(ECL)was performed using an Amersham ECL analysis system and visualized on ECL hyperfilm(Amersham, Amersham,UK).Thefilms were scanned and the pixel intensity of each band was analysed using IMAGEJ(National Institutes of Health,Bethesda,MD,USA).Levels of apoB100and apoB48 were quantified on the basis of the estimated pixel intensity of samples relative to the pixel intensity of the apoB standard run on the same gel.Statistical analysisData are the meanÆSEM of eight mice per group.Data were analysed using one-way ANOVA with Tukey’s or Dunnett’s post hoc tests and statistical significance was set at P<0.05(two-tailed).In addition to quantitative analysis of parenchymal IgG extravasation,for each of the randomly selected‘fields of view’quantitatively analysed,the‘presence’or‘absence’of leakage was compared with that in the LF control diet-fed group using binary logistic regression analysis.Findings are expressed as an odds ratio(OR)for leakage within each treatment arm,with OR>1indicating a greater likelihood of leakage for the treatment compared with control.Effects of probucol on BBB dysfunction47RESULTSMice fed the SFA-or cholesterol-enriched diets for12weeks exhibited significant perivascular abundance of IgG and of apoB lipoproteins compared with age-matched mice maintained on the LF cholesterol-free diet.The distribution of IgG and apoB was significantly greater in the CTX than the HPF(Figs1,2a,b).In contrast,the abundance of IgG and apoB lipoproteins in the CTX and HPF of probucol-treated mice fed the SFA-or cholesterol-enriched diets was comparable to that in the LF control diet-fed group(Figs1,2a,b).An OR for BBB dysfunction was determined by binary logistic regression analysis based on the parenchymal abundance of IgG and expressed relative to the LF diet-fed control group.As indi-cated in Fig.2c,mice maintained on the SFA-or cholesterol-enriched diets had a30-and sevenfold greater likelihood of BBB disturbances,respectively.Consistent with the parenchymal abun-dance of IgG and apoB(Figs1,2a,b),probucol treatment reduced the ORs in mice fed the SFA-or cholesterol-enriched diets simi-lar to those in the control group.Plasma concentrations of S100B are commonly used surrogate markers of BBB dysfunction and astrocytic inflammation.27,31,32 In the present study,plasma S100B levels were twofold greater in mice fed the SFA-or cholesterol-enriched diets compared with control(Fig.3).Conversely,plasma concentrations of S100B were not increased in probucol-treated mice fed the SFA-or cho-lesterol-enriched diets compared with control.Consistently,substantial activation of parenchymal glial cells was detected by3D immunofluorescent microscopy in mice fed the SFA-or cholesterol-enriched diets(Fig.4a).Semiquantitative data revealed significantly greater GFAP levels in mice fed the SFA-or cholesterol-enriched diets(Fig.4b).Consistent with these observations,reactive microglia(as evidenced by positive CD68staining)were seen in brains from mice fed the SFA-enriched diet(Fig.5).In contrast,GFAP abundance in pro-bucol-treated mice fed the SFA-or cholesterol-enriched diets was comparable to that in control mice.The SFA-and cholesterol-enriched diets were well tolerated, with no significant changes in plasma cholesterol,triglyceride, A b(1–40),A b(1–42)or apoB levels(Table1).Weight gain was also similar between the different groups(data not shown).None-theless,cholesterol was slightly increased in the cholesterol-fed group and probucol treatment significantly reduced total choles-terol in mice fed either the SFA-or cholesterol-enriched diet,as well as apoB levels in the latter group(Table1).Plasma concen-trations of NEFA were markedly increased in mice fed the SFA-or cholesterol-enriched diets,and probucol treatment had no sig-nificant effect on these increases in NEFA levels(Table 1).(a)(b)Fig.1Cerebral parenchymal abundance of IgG and apolipoprotein(apo)B.The integrity of the blood–brain barrier was assessed by three-dimensional immunofluorescent microscopy of perivascular leakage of plasma IgG and apoB.Mice were fed a control(Ctrl)diet or diets enriched with1%(w/w)cho-lesterol(Chol)or20%(w/w)saturated fatty acids(SFA),with or without concomitant probucol(approximately30mg/day;Prob)treatment,for 12weeks.(a)Representative images of perivascular IgG leakage are shown at lower(original magnification9200;bar,100l m)and higher magnifica-tion in three dimensions(x,y and z axes=80,70and14l m).Green indicates IgG staining,blue indicates nuclear(4′,6′-diamidino-2-phenylindole)stain-ing.(b)Two-dimensional images of plasma apoB leakage shown at lower(original magnification9200)and higher(x,y and z axes=80,70,8l m) magnification.Red indicates apoB staining.48R Takechi et al.DISCUSSIONThe present study used an established dietary lipid-induced model of BBB dysfunction to explore the putative effect of pro-bucol,a lipid-lowering agent with potent anti-oxidant activity. The high fat-fed C57BL/6J model is now an established model of BBB dysfunction and is widely used.8,12,18,27,33We confirmed the significant cerebral extravasation of plasma IgG and apoB in mice fed for12weeks with diets enriched with SFA or supple-mented with cholesterol and,in addition,demonstrated complete suppression of this phenomenon following the combined adminis-tration of1%(w/w)probucol at in the diet.The profound effi-cacy of probucol in this model was unlikely to be a consequence of the substantial reduction in plasma cholesterol in the SFA-or cholesterol-fed mice because the untreated SFA-and cholesterol-fed groups had comparable plasma cholesterol levels to those seen in the LF control group.Instead,plasma concentrations of NEFA were markedly increased in the SFA-and cholesterol-fed mice and this may have been a significant contributing factor to BBB dysfunction.Probucol had no significant effect on plasma NEFA levels in mice fed diets supplemented with SFA or choles-(a)(b)(c)Fig.2Quantitative and qualitative analyses of parenchymal IgG and apolipoprotein(apo)B extravasation.Mice were fed a control diet or diets enriched with1%(w/w)cholesterol(Chol)or20%(w/w)saturated fatty acids(SFA),with or without concomitant probucol(approximately30mg/day;Prob)treatment,for12weeks.The integrity of the blood–brain barrier wasassessed by using immunomicroscopic quantitative analysis and qualitative binarylogistic regression analysis.(a,b)Three-dimensional semiquantitative results of cerebral IgG(a)and apoB(b)extravasation expressed per volume unit. Data was collected from the cerebral cortex(CTX)and hippocampal formation(HPF).A minimum of three sections was tested from each mouse and each group contained eight mice.Data are the meanÆSEM.Columns with different symbols differ significantly(P<0.05,one-way ANOVA Tukey’s test).(h),control;(&),Chol;(),Chol+Prob;(),SFA;(),SFA+Prob.(c)The presence or absence of leakage in the randomly selectedof view was compared with the control group using binary logistic regression analysis and is expressed as an odds ratio for leakage within each treatment arm.Values>1indicate a greater likelihood of leakage for the treatment compared with control.Fig.3Plasma levels of S100calcium-binding protein B(S100B),a sur-rogate marker of cerebrovascular integrity,in mice fed a control diet or diets enriched with1%(w/w)cholesterol(Chol)or20%(w/w)saturated fatty acids(SFA),with or without concomitant probucol(approximately 30mg/day;Prob)treatment,for12weeks.Data are the meanÆSEM (n=8).Columns with different letters differ significantly(P<0.05,one-way ANOVA with Tukey’s test).(a)(b)Fig.4Cerebral parenchymal distribution of activated glial cells in response to perivascular inflammation.Activated glial cells were detected by three-dimensional immunofluorescent staining for glialfibrillary acidic protein(GFAP)in the cortex of mice fed a control diet or diets enriched with1%(w/w)cholesterol(Chol)or20%(w/w)saturated fatty acids (SFA),with or without concomitant probucol(approximately30mg/day; Prob)treatment,for12weeks.(a)Representative extended focus images. Bar,100l m.Green indicates GFAP staining,whereas nuclei are stained blue.Arrowheads indicate blood vessels.(b)Activated glial cells were detected and analysed semiquantitatively.Data are the meanÆSEM. Columns with different symbols differ significantly(P<0.05,one-way ANOVA with Tukey’s test).A minimum of three sections was tested from each mouse and each group contained eight mice.Effects of probucol on BBB dysfunction49terol,indicating that the bene ficial effects of probucol on BBB function were either downstream of vascular exposure to NEFA or that damage occurred via other mechanisms.The signi ficant blood-to-brain delivery,retention and accumu-lation of apoB lipoprotein shown in the present and previous studies,12primarily within the CTX of SFA-and cholesterol-fed mice,is consistent with the distribution of apoB and amyloid pla-que in brain specimens from individuals with VaD and/or AD.20The pattern of parenchymal abundance of apoB –lipoprotein –A b in wild-type mice maintained on pro-atherogenic diets also repli-cates the pattern of BBB dysfunction and of apoB –lipoprotein –A b distribution in established murine models of AD.12,21,29,34Therefore,genetically unmanipulated mice with dietary-induced cerebrovascular in flammation are a physiologically useful model in which to explore compounds that may be bene ficial in main-taining BBB function.Dietary lipotoxicity is a term used to broadly describe pro-cesses leading to end-organ damage following excess exposure to lipids.First identi fied in the context of fat-induced insulin resis-tance,the process has been implicated in endothelial dysfunction,atherosclerosis,organ failure and autoimmune and in flammatory disorders.35Animal feeding studies have shown that SFA enriched diets increase protein oxidation and lipid peroxida-tion.36,37Exogenous fatty acid supplementation results in signi fi-cant shifts in neuronal phospholipids and lipid raft composition,35key regulators of in flammation.However,signi ficant differences in the cytotoxic effects of fatty acids have been reported,with longer-chain SFAs being the most potent and the unsaturated fatty acids being cytoprotective.38–40Morgan suggests that one mechanism underlying the toxicity of SFA is a consequence of disturbances in protein processing and endoplasmic reticulum (ER)dysfunction.41Other mechanisms underlying SFA-induced changes in BBB function include stimulation of NADPH oxidase-derived reactive oxygen species generated by activated microglial cells or modulation of mammalian target of rapamycin (mTOR),a key signal transduction protein that regulates vascular endothelial fenestration.42Patil et al.43concluded that dietary pal-mitic acid induced region-speci fic cerebral damage because of the higher fatty acid-metabolizing capacity of cortical astroglia.Conversely,cell culture studies suggest that incubation with longer-chain unsaturated fatty acids has an antagonistic effect on in flammatory pathways induced by SFAs.44Studies by Ghribi et al.11found that,like SFA,dietary choles-terol results in BBB dysfunction in New Zealand white rabbits.Similarly,we found that modest dietary supplementation with cholesterol (1%w/w)disturbs BBB function 45but,unlike the studies in rabbits,the mice were not dyslipidaemic.Cell culture studies suggest several mechanisms by which dietary cholesterol may be proin flammatory and some of these appear to be analo-gous to the effects of dietary SFA.For example,Yao and Tabas reported that excess cholesterol causes ER and mitochondrial stress that can lead to apoptosis.46Mitochondrial activityorFig.5Immunohistochemical analysis of microglial activation.Activated microglia were detected by immunohistochemical staining for CD68in mice fed either the control low-fat chow or a saturated fatty acid (SFA)-enriched diet.Immunoreactivity for CD68appears as brown staining and is indicated by the arrows.Cell nuclei were stained with Harris ’haematoxylin (blue).Bar,100l m.Table 1Plasma concentrations of lipids and b -amyloid in the different treatment groups Treatment group TC (mmol/L)TG (mmol/L)NEFA (mEq/L)ApoB (l g/mL)A b (1–40)(pg/mL)A b (1–42)(pg/mL)Control 6.89Æ0.580.34Æ0.020.38Æ0.0998.5Æ11.742.64Æ3.0616.66Æ2.61Chol8.65Æ0.240.43Æ0.050.69Æ0.11*105.9Æ8.239.72Æ1.9817.48Æ1.40Chol +Prob 2.80Æ0.44**0.28Æ0.040.80Æ0.13*64.1Æ5.5*42.53Æ1.2119.47Æ0.31SFA7.09Æ0.510.32Æ0.040.70Æ0.04*77.3Æ7.543.30Æ2.9221.94Æ1.11SFA +Prob3.45Æ0.39**0.37Æ0.040.74Æ0.07*83.2Æ7.039.88Æ2.2717.13Æ2.47Data are the mean ÆSEM (n =8mice per group).*P <0.05,**P <0.01compared with control (one-way ANOVA followed by Dunnett ’s test).Mice were fed a control diet or diets enriched with 1%(w/w)cholesterol (Chol)or 20%(w/w)saturated fatty acids (SFA),with or without concomitant probucol (approximately 30mg/day;Prob)treatment,for 12weeks.TC,total cholesterol;TG,triglyceride;NEFA,non-esteri fied fatty acids;ApoB,apolipoprotein B;A b ,b -amyloid.50R Takechi et al.。
消化系统英文版共12页
The stomach is divided into the fundic, cardiac, body, and pyloric regions. The stomach has three layers of muscle: an outer longitudinal layer, a middle circular layer, and an inner oblique layer. The inner lining consists of four layers: the serosa, the muscularis, the submucosa, and the mucosa. The mucosa is densely packed with gastric glands, which contain cells that produce digestive enzymes, hydrochloric acid, and mucus. The stomach is a multi-functional organ, it stores food, breaks it down and then empties it into the small intestine.
The digestive system is responsible for breaking down food, absorbing nutrients from the food, and ridding the body of waste products from the food. It is roughly nine meters long and begins at the mouth and ends at the anus.
从内质网生化鲜明囊泡至中过氧化物酶
Biochemically Distinct Vesiclesfrom the Endoplasmic Reticulum Fuse to Form PeroxisomesAdabella van der Zand,1,*Ju¨rgen Gent,1,3Ineke Braakman,1,2and Henk F.Tabak1,21Cellular Protein Chemistry,Faculty of Science,Utrecht University,NL-3584CH Utrecht,The Netherlands2These authors contributed equally to this work3Present address:Institute for Life Sciences and Chemistry,Hogeschool Utrecht,NL-3572JE Utrecht,The Netherlands *Correspondence:a.vanderzand@uu.nlDOI10.1016/j.cell.2012.01.054SUMMARYAs a rule,organelles in eukaryotic cells can derive only from pre-existing organelles.Peroxisomes are unique because they acquire their lipids and membrane proteins from the endoplasmic reticulum (ER),whereas they import their matrix proteins directly from the cytosol.We have discovered that peroxisomes are formed via heterotypic fusion of at least two biochemically distinct preperoxisomal vesicle pools that arise from the ER.These vesicles each carry half a peroxisomal translocon complex. Their fusion initiates assembly of the full peroxisomal translocon and subsequent uptake of enzymes from the cytosol.Ourfindings demonstrate a remarkable mechanism to maintain biochemical identity of organelles by transporting crucial components via different routes to theirfinal destination. INTRODUCTIONCompartmentalization of the eukaryotic cell is one of the major transitions in the evolution of life.The multiplication of these compartments(organelles)in dividing cells reflects aspects of their evolutionary past.Autonomous organelles such as mito-chondria and chloroplasts form via proliferation of pre-existing organelles.They contain their own protein import machineries indicative of their endosymbiotic origin(reviewed by Nunnari and Walter,1996;Warren and Wickner,1996).In contrast, organelles of the secretory pathway,such as the Golgi complex and plasma membrane,rely on the endoplasmic reticulum(ER) for their formation and protein import.Peroxisomes are unusual in this respect because their biogen-esis requires both these assembly lines:(1)the ER provides lipids and peroxisomal membrane proteins(PMPs)and yields a perox-isomal precompartment(Hoepfner et al.,2005;Kragt et al.,2005; Tam et al.,2005;Kim et al.,2006;Motley and Hettema,2007;van der Zand et al.,2010),and(2)the cytosol provides the matrix proteins,which are imported via the peroxisomal translocon (Hazra et al.,2002;Agne et al.,2003).Together these processes define the beginning and end of the peroxisomal biogenesis pathway.This atypical assembly line has led to controversial discussions,particularly in the recent literature(Ma et al.,2011; Nuttall et al.,2011).We have now taken a fundamental step forward and demonstrate that these two routes do not operate independently of each other;both the ER and the peroxisomal translocon play an essential role in peroxisome biogenesis. Real-time imaging of live cells has given many insights into the spatial and temporal organization of membrane and PMPflow from the ER to peroxisomes.The nature of the membrane carriers between both compartments however remained unre-solved.To dissect the events leading to the formation of new peroxisomes we studied the interactions between PMPs as they traffic from the ER to peroxisomes.We discovered that PMPs leave the ER via different routes.This results in formation of vesicular carriers that upon heterotypic fusion combine their PMP content.From this moment onward an active peroxisomal translocon is assembled,which only then can begin with the import of enzymes from the cytoplasm.RESULTSAssembly of PMP Complexes during Peroxisome Biogenesis:Experimental Set-UpThe peroxisomal translocon translocates enzymes carrying a peroxisomal targeting signal(PTS1/PTS2)from the cytosol into the peroxisomal lumen(reviewed by Ruckta¨schel et al., 2011).It is functionally divided into two halves:the docking complex formed by the PMPs Pex13p and Pex14p,and a RING finger complex composed of the PMPs Pex2p,Pex10p,and Pex12p(Agne et al.,2003).We used bimolecularfluorescence complementation(BiFC),also called split-GFP(Hu et al.,2002; Nyfeler and Hauri,2007;Kerppola,2008)to follow the assembly of docking and RINGfinger PMPs into functional peroxisomal translocon complexes in living yeast cells(Figure1A).PEX genes were genomically fused at their30ends to either VN(aa1–173), the N-terminal half of Venusfluorescent protein,or to VC(aa 155–238),the C-terminal half of Venusfluorescent protein.As a result the tagged PMPs were expressed from their endogenous promoter in place of the wild-type untagged PMP.Because Venusfluorescent protein half-molecules are notfluorescent, Cell149,397–409,April13,2012ª2012Elsevier Inc.397wild-type haploid yeast cells expressing PEX-VC or PEX-VN were nonfluorescent.To visualize peroxisomes,we introduced a fluo-rescently-tagged matrix protein marker CFP-PTS1(red)into thehaploid cells expressing PEX-VN .The VN-and VC-tagged PMPs were fully functional as they mediated peroxisomal import of CFP-PTS1(data notshown).Figure 1.PMP Interactions at Different Stages of Peroxisome Formation(A)Experimental set-up of the spit-GFP assay combined with cell mating.Haploid cells expressing PEX x -VN and CFP-PTS1were mated in various combinations with haploid cells of the opposite mating type expressing PEX y -VC ,upon which their contents fuse.Cells were mated and followed for up to 72hr for restoration of Venus fluorescence by live-cell microscopy.The import of CFP-PTS1(red)into peroxisomes containing the PMP complexes (green)was monitored (yellow).(B)Wild-type haploid yeast cells expressing PEX14-VC (AZY357)or PEX10-VC (AZY425)were mated in various combinations with wild-type cells coexpressing the peroxisomal marker protein CFP-PTS1(red)and either PEX13-VN (AZY355)or PEX2-VN (AZY424).Scale bar,5m m.(C)Summary of the split-GFP/mating results in the different wild-type and PEX mutant cells:(À)no Venus fluorescence;(+)Venus fluorescence.See also Figures S1–S4.398Cell 149,397–409,April 13,2012ª2012Elsevier Inc.Haploid cells expressing PEX-VN and CFP-PTS1were mated in various combinations with haploid cells of the opposite mating type expressing PEX-VC,upon which their cytoplasmic contents including the two nuclei fuse.As a consequence each diploid zygote now expressed tagged as well as untagged versions of the PMPs under investigation.Mated cells were in-spected at24hr intervals up to72hr forfluorescence comple-mentation.When PMPs interacted,the fused VN and VC halves were brought together,associated,and formed thefluorescent Venus signal(green).We used colocalization with CFP-PTS1 (red)to confirm the peroxisomal localization of the PMP complexes.Because mature peroxisomes do not fuse(Motley and Het-tema,2007),only the newly synthesized pool of VN-or VC-tagged PMPs produced bimolecularfluorescent complexes. These newly formed peroxisomes have thus been synthesized after mating and import the constitutively expressed matrix protein marker CFP-PTS1.The split-GFP assay combined with cell mating thus allowed us to follow the assembly of newly synthesized PMPs with time at specific cellular locations.To examine the interaction between PMPs in peroxisomes and validate our approach,we used wild-type cellsfirst.Genes en-coding Pex2p,Pex10p,Pex13p,and Pex14p were genomically tagged with the N-or C-terminal half of Venusfluorescent protein respectively(Figures1B and1C).We foundfluorescence complementation of the Venus fragments for all combinations of PMPs tested(Figures1B and1C).The reconstitutedfluores-cence(green)colocalized with import-competent peroxisomes (CFP-PTS1)(red)demonstrating the functional assembly of various PMP complexes in the peroxisomal membrane(Fig-ure1B).We concluded that all tagged PMPs showed functional interactions and that they localized properly to peroxisomes. As controls we used Pex1p and Pex6p.Although Pex1p and Pex6p associate with the peroxisomal translocon(Rosenkranz et al.,2006)we did not detect any direct interactions with the translocon using this assay(Figure S1available online).We did howeverfind interactions between Pex1p and Pex6p(Figure S1) as was reported before(Faber et al.,1998).These data confirmed that the split-GFP assay was highly specific in vivo. We then determined when during peroxisome biogenesis newly synthesized Pexp-VN and Pexp-VC start to interact.We used a collection of PEX deletion mutants to identify genes that blocked peroxisome biogenesis at distinct stages.Of the PMPs Pex3p has been the most extensively studied,and the trafficking of newly synthesized Pex3p-YFP is well documented (Hoepfner et al.,2005;Kragt et al.,2005;Tam et al.,2005).Wild-type cells and several peroxisome-deficient strains(D pex1, D pex6,D pex10,D pex13,D pex15,D pex19)coexpressed the peroxisomal membrane protein PEX3-YFP(green),the peroxi-somal matrix protein marker CFP-PTS1(red),or the ER marker SEC63-CFP(red)(Figure S2).In D pex19cells PMP export from the ER is blocked(Lam et al.,2010;van der Zand et al.,2010; Agrawal et al.,2011)and consequently Pex3p-YFP was trapped in the ER.It represented thefirst block in peroxisome biogenesis. Cells that lacked components of the AAA+complex or its membrane receptor Pex15p(Birschmann et al.,2003)(D pex1, D pex6,or D pex15)signified the next stage in peroxisome biogenesis,where Pex3p-YFP localized to one dot per cell.In all these mutants peroxisomes were absent and CFP-PTS1 localized to the cytosol.Cells that lacked components of the docking(D pex13)or RING finger(D pex10)complex showed more Pex3p-YFP labeled puncta per cell,and were therefore a progression from the single dot stage.In these two mutants a functional peroxisomal trans-locon cannot be formed;consequently CFP-PTS1remained mislocalized to the cytosol.In wild-type cells CFP-PTS1was effi-ciently sequestered into peroxisomes and colocalized with Pex3p-YFP.PMP Complex Formation in the ER MembraneWe used cells lacking PEX3or PEX19(D pex3or D pex19)to examine the interaction between PMPs in the ER.In these cells PMPs are inserted into the ER membrane but cannot leave this compartment(van der Zand et al.,2010).Haploid D pex3cells expressing PEX14-VC or PEX10-VC were mated in various combinations with haploid D pex3cells ex-pressing CFP-PTS1and either PEX13-VN or PEX2-VN.Because peroxisomes are absent in D pex3cells,the peroxisomal marker CFP-PTS1(red)localized to the cytosol(Figure S3).Cells were mated and inspected at24hr intervals up to72hr.Fluorescence complementation of the Venus fragments(green)was only found between Pex13p and Pex14p(docking complex),and between Pex2p and Pex10p(RINGfinger complex)(Figures1C and S3). Because PMPs cannot exit the ER in D pex3cells,the data implied that the docking and RINGfinger subcomplexes were assembled already in the ER membrane.Contrary to peroxi-somes,however,the full peroxisomal translocon did not assemble in the ER,as we did not detect anyfluorescence complementation between Pex2p-Pex14p and Pex10p-Pex13p in the72hr time course.The cytosolic localization of CFP-PTS1in the D pex3cells was therefore not only a reflection of the absence of mature peroxisomes,but also of an incom-pletely assembled peroxisomal translocon(Hazra et al.,2002). We concluded that the full peroxisomal translocon is not assem-bled in the ER.Identical results were obtained with D pex19cells (Figure1C).PMP Complex Formation in Preperoxisomal VesiclesIn D pex1or D pex6cells,PMPs not only reside in the ER but also in immature vesicles that are not yet capable of importing PTS1/PTS2-containing matrix enzymes(Figure S2).The afore-mentioned split-GFP matings were repeated in either D pex1or D pex6cells(Figures1C and S4).Again both mutant cells were mated and inspected at24hr intervals up to72hr.Fluorescence complementation of the Venus fragments(green)was found only between Pex13p and Pex14p(docking complex),and between Pex2p and Pex10p(RINGfinger complex),indicating the pres-ence of the docking-and RINGfinger subcomplexes in preper-oxisomal vesicles.Like in the ER the full peroxisomal translocon was not assem-bled in the preperoxisomal vesicles,as we did not detect any fluorescence complementation between Pex2p-Pex14p and Pex10p-Pex13p in the72hr time course.We concluded that also in the D pex1and D pex6cells a functional peroxisomal translocon did not assemble,as shown by the cytosolic localiza-tion of CFP-PTS1.Cell149,397–409,April13,2012ª2012Elsevier Inc.399The Docking and RING Finger Complexes Are Kept in Distinct Subcellular Structures Early during Peroxisome BiogenesisThe failure to assemble the peroxisomal translocon in D pex3,D pex19,D pex1,and D pex6cells can be explained in two ways:(1)the two half-translocons leave the ER in one compart-ment but their physical separation is retained as it is in the ER,or (2)the half-translocons traffic in different membrane carriers that need to fuse to complete their functional assembly.To distin-guish between these two possibilities we biochemically isolated organellar fractions from wild-type,D pex19,D pex1,and D pex6cells by buoyant-density centrifugation and followed the behavior of 15markers (Figures 2and S5).Samples were analyzed by western blot with indicated antibodies.In wild-type cells,the PMPs comigrated with the peroxisomal matrix proteins thiolase (Pot1p),catalase (Cta1p),and CFP-PTS1.In D pex19cells PMPs failed to exit the ER and as a result the peak fractions shifted to a lower density that coequilibrated with the ER marker Sec63p.In D pex1and D pex6cells,PMPs started to accumulate in different low-density fractions.Surpris-ingly the RING finger PMPs (Pex2p,Pex10p,and Pex12p),Pex11p,and Pex15p were not present in the same low-density fractions as the docking PMPs (Pex13p,Pex14p)and Pex5p.The docking PMPs equilibrated at higher densities (V2)than the RING finger PMPs (V1),implying their presence in distinct vesicular structures (Figures 2A and S5).Consistent with the presence of only peroxisomal half-translocons in the preperoxi-somal vesicles,V1and V2vesicles did not contain detectable levels of matrix proteins (Figure 2C).We noted a substantial pool of PMPs comigrated with the ER marker in D pex1and D pex6cells.A likely explanation for this is the depletion of perox-isomal budding factors from the ER membrane,because we found the majority of Pex3p in both V1and V2vesicle fractions and only limited amounts in the ER in D pex1and D pex6cells.These data suggest that the peroxisomal half-translocons leave the ER via separate low-density membrane carriers,where they cannot support matrix protein import.To confirm the exis-tence of two biochemically distinct vesicle pools,we performed coimmunoprecipitation experiments between the docking PMP Pex13p and the RING finger PMP Pex2p (Figure 3A).PEX13-CFP and 3HA-PEX2were integrated into the genome and ex-pressed from the GAL1promoter to obtain comparable protein levels in the various PEX mutants.Wild-type cells served as a positive control,where Pex2p coprecipitated with Pex13p in peroxisomes.As a negative control,we used D pex19cells where ER exit was blocked,and the amount of Pex2p coprecipitating with Pex13p was comparable to background levels (Figure S6A).In the D pex1and D pex6mutants,when Pex2p and Pex13p reside in different preperoxisomal vesicles,the amount of Pex2p coprecipitating with Pex13p was reduced by more than 50%when compared to wild-type cells.Although Pex2p and Pex13p also reside in the ER in these mutants,these PMPs do not associate in the ER membrane as demonstrated in the D pex19cells.We failed to detect interactions with nonperoxiso-mal proteins such as Kar2p (Figure S6B)or Sec63p (data not shown).We also tested by microscopy for colocalization between flu-orescently (YFP [green]or CFP [red])tagged docking (Pex13p,Pex14p)and RING finger (Pex2p,Pex10p)PMPs in wild-type,D pex1,or D pex6cells (Figures 3B and 3C).PMPs were chromo-somally tagged to create endogenously expressed C-terminal fusions with either CFP or YFP.In cells that coexpressed only the docking PMPs or the RING finger PMPs the fluorescent signals always overlapped,regardless of genetic background.When we coexpressed docking and RING finger PMPs (Pex2p-Pex14p or Pex10p-Pex13p)in D pex1and D pex6cells,the fluorescently labeled structures were juxtaposed and the amount of colocalization was strongly reduced (<30%)compared to wild-type cells.It suggests that the accumulated preperoxisomal vesicles in D pex1and D pex6cells represent topologically distinct compart-ments.We concluded that in D pex1and D pex6cells the two half-translocons were physically segregated in different membrane carriers that precluded their assembly into a full translocon.The small but significant amounts of colocalization and coimmu-noprecipitation we found is in agreement with previously pub-lished data of secretory cargoes that are sorted into different exit routes from the ER (Castillon et al.,2009).WepostulateFigure 2.Subcellular Localization of PMPs in Wild-Type and PEX Mutant Cells(A–C)Western blot analysis.(A)PMP cargo proteins and the ER protein Sec63p.(B)Pex1p,Pex6p,and Pex3p.(C)Peroxisomal matrix proteins thio-lase [Pot1p],catalase [Cta1p],and CFP-PTS1)of peak fractions (3,8,16,and 21)taken from sucrose step-gradients after buoyant-density centrifugation.Fractions of wild-type,D pex19,D pex1,or D pex6homogenates are shown.PMPs were grouped according to their functional relationships.The biochemical identity of the peak fraction is annotated by peroxisomes (P),ER,preperoxisomal vesicles (V1and V2).Arrows denote specific protein bands.Anti-GFP antibodies were used to detect XFP-tagged PMPs.Pex11p*does not represent the total cellular pool of Pex11p,but rather phosphorylated Pex11p,which specifically associates with the ER and not peroxisomes (Knoblach and Rachubinski,2010).See also Figure S5.400Cell 149,397–409,April 13,2012ª2012Elsevier Inc.Figure3.RING Finger and Docking PMPs Reside in Different Preperoxisomal Vesicles(A)Protein complexes were isolated from wild-type,D pex19,D pex1,and D pex6cells expressing3HA-PEX2and PEX13-CFP using immunoprecipitation with rabbit polyclonal anti-GFP-antibody coupled to protein A Sepharose beads followed by western blot analysis.Pex13p and Pex2p immunoprecipitations were quantified using ImageJ.The amount of Pex2p coprecipitating with Pex13p is expressed as a ratio(see Supplemental Information for details).Lanes represent equal amounts of protein.*Background bands.(B)Colocalization analysis between endogenously expressed CFP(red)-and YFP(green)-tagged versions of docking(Pex13p and Pex14p)and/or RINGfinger PMPs(Pex2p,Pex10p)in wild-type,D pex1,and D pex6cells.Scale bar,5m m.See also Figure S7A.(C)Quantification of the percentage of colocalization of the data shown in(B)was done using ImageJ software(JACoP plugin).Error bars represent the SD of three independent experiments.(D)Fluorescence pulse-chase analysis of Pex2p-YFP(green)and Pex13p-CFP(red)in wild-type cells(AZY615).Scale bar,5m m.See also Figures S6C and S7C.(E)Quantification of the percentage of colocalization between Pex2p-YFP and Pex13p-CFP during a complete and representativefluorescent pulse-chase experiment(Figure3D)compared to D pex1and D pex6cells(Figure3B).Quantifications as in(C).See also Figures S6and S7.Cell149,397–409,April13,2012ª2012Elsevier Inc.401402Cell149,397–409,April13,2012ª2012Elsevier Inc.that the observed biochemical associations andfluorescent co-localization between the docking and RINGfinger PMPs in D pex1or D pex6cells must have occurred after budding because we failed to detect interactions between these PMPs in the ER. We usedfluorescence pulse-chase experiments to demon-strate that the physical separation of the two half-translocons in the ER and in the preperoxisomal structures occurred in wild-type cells too(Figures3D,3E and S6C).Colocalization between the pool of newly synthesized RINGfinger PMP (Pex2p-YFP or YFP-Pex12p)and the docking PMP Pex13p-CFP was measured with time.PMPs were tagged with YFP (green)and CFP(red)respectively,put under control of the GAL1promoter and integrated into the yeast genome.Diploid cells were used,so that for every galactose-inducible PEX locus also a wild-type endogenous(chromosomal)copy existed to ensure that cells contained peroxisomes.The inducible GAL1promoter was used to produce a limited wave of PMP synthesis.Before induction nofluorescence signal was detected(Figure3D:0min).A15min pulse in galactose induced synthesis of the XFP-tagged PMPs simultaneously, whereas further synthesis was stopped by repression of the GAL1promoter with glucose.We showed before that the amount of PMP-XFP that is synthesized corresponds well with endogenous levels and that overproduction is prevented(Hoepf-ner et al.,2005;van der Zand et al.,2010).At early time points Pex2p and Pex13p existed as separate fluorescent puncta(Figure3D:45min)that failed to colocalize. At later time points these puncta started to coalesce and coloc-alize(Figure3D:180min).Similar results were found for Pex12p and Pex13p(Figure S6C).Thus the RINGfinger PMPs were kept at distinct cellular locations from the docking PMP Pex13p during the early stages of peroxisome biogenesis in wild-type cells.In contrast in cells where we coinduced either the docking PMPs Pex13p-CFP and Pex14p-YFP,or the RINGfinger PMPs Pex2p-CFP and YFP-Pex12p(Figure S6C),thefluorescence signals colocalized from the earliest time point(90min)and re-mained colocalized for the duration of the chase(180min),sug-gesting that the individual peroxisomal half-translocon complexes assemble early during their biogenesis and remain together while they traffic from the ER to peroxisomes.The percentage colocalization between Pex2p and Pex13p during early time points of the pulse-chase(45–60min)(Fig-ure3E)compared very well to the amount of colocalization when expressed in the D pex1or D bined, our data imply that during peroxisome biogenesis the docking and RINGfinger half-translocons exist in distinct subcellular structures.We propose that the docking and RINGfinger PMPs leave the ER in different vesicles.Peroxisomes Are Formed via the Heterotypic Fusionof Preperoxisomal VesiclesA crucial question to resolve concerns thefinal stages of perox-isome formation.Two options arise:(1)model1,in which vesic-ular carriers fuse among each other and develop into new mature peroxisomes,thereby adding new peroxisomes to the existing population,or(2)model2,in which vesicular carriers with PMPs fuse with pre-existing mature peroxisomes,which grow and divide to form additional organelles,a scenario proposed before(Motley and Hettema,2007).We appliedfluorescence pulse-chase assays combined with cell fusion to follow the fate of pulse-labeled preperoxisomal vesicles with time(Figure4A).To label preperoxisomal vesicles or peroxisomes we had to switch markers.We found that the RINGfinger PMPs were characterized by a short half-life (<5hr).This made them ineffective markers for organelle fusion in our assay,where we followed pulse-labeled protein over a24hr time course.To this end we replaced the RINGfinger PMPs with Pex11p as a marker.Pex11p behaved identically to the RINGfinger PMPs(Figure S7).To assay fusion,PEX11and PEX13were tagged with YFP and CFP respectively and their expression controlled by the induc-ible galactose(GAL1)promoter.Integration plasmids were used,so that for every galactose-inducible PEX locus also a wild-type endogenous(chromosomal)copy existed.To induce synthesis of Pex13p-CFP or Pex11p-YFP in haploid D pex1or D pex6cells,respectively,cells were grown in galactose for 2hr.To stop further synthesis and allow the pool of newly synthesized Pex11p-YFP or Pex13p-CFP to be imported into preperoxisomal vesicles,cells were grown for a further2hr in glucose.Cells then were allowed to mate under conditions that prevented any further synthesis of Pex11p-YFP and Pex13p-CFP.We inspected cells at5hr,10hr,and24hr for colocaliza-tion of Pex11p-YFP(green)and Pex13p-CFP(red).To test whether labeled preperoxisomal vesicles fused together(Figure4B,top),we mated haploid D pex1cells express-ing PEX13-CFP with D pex6cells expressing PEX11-YFP.After mating,Pex11p and Pex13p colocalized in the same puncta. The PMPs that had accumulated in different precompartments during the pulse-chase protocol hence came together in the same compartment in the diploids.In a control experiment we confirmed that only the preperox-isomal vesicles are fusogenic(Figure4B).We mated haploidFigure4.Heterotypic Fusion of Preperoxisomal Vesicles(A)Experimental set-up of(pre-)peroxisomal fusion assay.Galactose-inducible andfluorescently tagged PMPs were used to label specific organelles in the cell. Cells were mated and inspected at5hr,10hr,or24hr for colocalization by live-cellfluorescence microscopy.Pex11p-YFP(green)and Pex13p-CFP(red)were used to pulse-label preperoxisomal vesicles(V1and V2)or peroxisomes(P).Their colocalization was used as a measure for organelle fusion.(B)Haploid D pex1cells expressing PEX13-CFP(AZY496)were mated with D pex6cells expressing PEX11-YFP(AZY495)(top)or haploid wild-type cells ex-pressing PEX13-CFP(AZY399)were mated with wild-type cells expressing PEX11-YFP(AZY400)(bottom).Scale bar,5m m.(C)Haploid D pex1cells expressing either PEX11-YFP(AZY557)or PEX13-CFP were mated(AZY496),and in parallel we mated haploid D pex6cells expressing either PEX11-YFP(AZY495)or PEX13-CFP(AZY556).Scale bar,5m m.(D)Colocalization analysis of TomatoRed-tagged(blue)Pex3p,Pex1p and Pex6p with CFP-tagged(red)docking PMP Pex13p and YFP-tagged(green)RING finger PMP Pex10p in wild-type,D pex1,and D pex6cells.Scale bar,5m m.(E)Quantification of the percentage of colocalization of the data shown in(D)was done using ImageJ software(JACoP plugin).Error bars represent the SD of at least three independent experiments.Cell149,397–409,April13,2012ª2012Elsevier Inc.403wild-type cells expressing PEX13-CFP with wild-type cells ex-pressing PEX11-YFP to test whether pulse-labeled peroxisomes fused or not.We never detected any colocalization between Pex11p and Pex13p in the newly formed diploid.It suggests that existing mature peroxisomes cannot fuse,as was demon-strated before(Motley and Hettema,2007).The idea that different preperoxisomal vesicles fuse to form peroxisomes was described before(Titorenko et al.,2000).The authors reported that heterotypic fusion of biochemically purified preperoxisomal vesicles was dependent on both NSF-like (AAA+)proteins Pex1p and Pex6p(Titorenko and Rachubinski, 2000).To demonstrate that indeed Pex1p and Pex6p were both required for heterotypic fusion of preperoxisomal vesicles we investigated whether or not Pex11p-YFP(green)and Pex13p-CFP(red)colocalized in diploids derived from either D pex1haploid cells or D pex6haploid cells(Figure4C). Haploid D pex1cells expressing either PEX11-YFP or PEX13-CFP were mated,and in parallel,haploid D pex6cells expressing either PEX11-YFP or PEX13-CFP were mated.We never de-tected any colocalization between Pex11p and Pex13p in diploids derived from both matings,suggesting that Pex1p and Pex6p were necessary to mediate the heterotypic fusion of pre-peroxisomal vesicles.To further dissect the function of both Pex1p and Pex6p and the budding factor Pex3p we assayed their subcellular localiza-tion biochemically and by live-cell imaging in several mutants. Buoyant-density centrifugation(Figure2B)demonstrated that Pex1p,Pex6p,and Pex3p comigrated only in the peroxisomal and ER fractions.In D pex1or D pex6cells they showed a differen-tial distribution,whereby Pex3p equilibrated in both preperoxi-somal vesicle fractions(V1and V2),Pex1p was restricted to V2 only whereas Pex6p was specific to V1.We confirmed these results byfluorescence microscopy in wild-type,D pex1or D pex6cells that coexpressed the docking PMP Pex13p-CFP(red)and the RINGfinger PMP Pex10p-YFP (green)together with either TomatoRed-tagged(blue)Pex3p, Pex1p,or Pex6p(Figures4D and4E).In wild-type cells Pex10p and Pex13p colocalized with Pex3p,Pex1p,or Pex6p. In D pex1and D pex6cells,Pex3p was evenly distributed over the Pex10p and Pex13p marked puncta,whereas Pex6p specif-ically associated with Pex10p,and Pex1p with Pex13p.We concluded that Pex3p was a shared component of both preperoxisomal vesicle populations,whereas Pex1p and Pex6p each specifically associated with V2or V1,respectively. Heterotypic Fusion of Preperoxisomal Vesicles Results in Formation of the Active Peroxisomal Translocon Because the split-GFP mating assay allowed monitoring of both PMP complex formation and the import competence of the newly formed peroxisomal translocons,we used it to demon-strate that the preperoxisomal structures accumulated in D pex1and D pex6cells indeed were productive intermediates (Figure5).Haploid D pex1cells expressing PEX14-VC or PEX10-VC were mated with haploid D pex6cells expressing CFP-PTS1and either PEX2-VN or PEX13-VN.In the absence of peroxisomes in haploid D pex6yeast cells and in early zygotes,CFP-PTS1(red) is localized to the cytosol(Figure5).After mating D pex1with D pex6cells,each diploid received from its mating partner a VN-and a VC-tagged PMP and a corresponding wild-type copy of Pex1p or Pex6p allowing fusion of preperoxisomal vesi-cles(Figure4)and subsequent formation of peroxisomes in the diploids.Cells were mated and inspected at12hr and48hr after mating forfluorescence complementation.At12hr CFP-PTS1 import had not commenced and we found a mixed population of zygotes,some showedfluorescence complementation of Venus fragments(green),indicative of assembly of the full peroxisomal translocon,others did not.Thus,as a result of the contribution of Pex1p and Pex6p,preperoxisomal vesicles fused in the diploid zygote,thereby bringing the two half-trans-locons together.Import of CFP-PTS1into the reconstituted peroxisomal translocon complexes took more time.When we next inspected the cells again at24hr some Venus-positive zygotes showed a redistribution of cytosolic CFP-PTS1to peroxisomes(puncta).At48hr all zygotes contained peroxi-somal CFP-PTS1,which colocalized with the Venus labeled structures.These data can be explained in two ways:preperoxisomal vesicles fuse and form new import-competent peroxisomes,or preperoxisomal vesicles fuse with the CFP-PTS1-labeled perox-isomes that are formed by complementation of PEX1and PEX6. Preperoxisomal Vesicles Mature into New Peroxisomes To test whether vesicular carriers can fuse with peroxisomes,we usedfluorescently tagged(CFP or YFP)PMPs to label specific organelles in the cell(Figure6A).Like before,PEX11and PEX13were fused to YFP and CFP,respectively,and their expression controlled by the inducible galactose(GAL1) promoter.Wild-type cells were used for labeling peroxisomes whereas D pex1or D pex6cells were used to label preperoxiso-mal vesicles.The haploid cells were grown in galactose for2hr to induce expression.To stop further synthesis and allow the pool of newly synthesized Pex11p or Pex13p to be imported into perox-isomes or preperoxisomal vesicles,cells were grown for a further 2hr in glucose.Cells then were allowed to mate under condi-tions that prevented any further synthesis of Pex11p and Pex13p.Cells were inspected at5hr,10hr,and24hr. Pex11p-YFP(green)and Pex13p-CFP(red)never colocalized (Figure6A).These data suggest that preperoxisomal vesicles do not fuse with peroxisomes.The formation of new import-competent peroxisomes from fusion of preperoxisomal vesicles therefore explains the colocalization we found between the Venus-reconstituted peroxisomal translocon complexes and CFP-PTS1in Figure5.We next used split-GFP experiments to demonstrate that new peroxisomes are formed by maturation of preperoxisomal vesicles(Figure6B).Wild-type haploid cells expressing PEX14-VC or PEX10-VC were mated with wild-type haploid cells expressing CFP-PTS1and either PEX2-VN or PEX13-VN as described for Figure1.In this instance,however,a galac-tose-inducible copy of the peroxisomal CFP-PTS1marker was used to pulse-label a pre-existing population of peroxi-somes before cells were allowed to mate.To label the popula-tion of existing peroxisomes before mating,haploid cells were404Cell149,397–409,April13,2012ª2012Elsevier Inc.。
伯氏疏螺旋体 英文
伯氏疏螺旋体英文Title: Borrelia burgdorferi: An Insight into the Lyme Disease PathogenBorrelia burgdorferi, also known as Lyme disease spirochete, is a bacterial species belonging to the genus Borrelia within the spirochete family. It is a Gram-negative, microaerophilic organism that possesses a unique helical shape, characterized by 3 to 10 sparse spirals under microscopic examination. These spirochetes arefurther distinguished by their flagella, visible under electron microscopy, with 7 to 15 flagella at each end.The significance of Borrelia burgdorferi lies in its role as the causative agent of Lyme disease, a tick-borne infection that affects humans and other animals. Lyme disease is a multisystemic illness that can manifest with a range of symptoms, including skin rashes, arthritic pain, neurological problems, and cardiac abnormalities. The spirochete is transmitted to humans through the bite of infected ticks, primarily the Ixodes species.The lifecycle of Borrelia burgdorferi is intricately linked to its arthropod hosts. It spends part of its lifecycle within the tick, undergoing multiple stages of development before being transmitted to a new host. Once inside the host, the spirochete can disseminate throughout the body, invading various tissues and organs.The pathogenesis of Lyme disease involves complex interactions between Borrelia burgdorferi and the host's immune system. The spirochete possesses various virulence factors that enable it to evade immune clearance andpersist within the host. These factors include adhesinsthat facilitate attachment to host cells, proteases that degrade host proteins, and antigenic variation that allows the spirochete to evade immune recognition.The diagnosis of Lyme disease can be challenging due to the variable presentation of symptoms and the absence of a single, reliable diagnostic test. However, a combination of clinical symptoms, serological testing, and, in some cases, molecular detection methods can aid in the diagnosis. Treatment typically involves the administration ofantibiotics, which can effectively eliminate the spirochete in the early stages of infection.The ecology and epidemiology of Borrelia burgdorferi are also fascinating aspects of its biology. The spirochete is endemic in certain geographical regions, particularly those with high tick populations. Climate change and other environmental factors have been implicated in the expansion of tick habitats and, consequently, the increasing incidence of Lyme disease.Moreover, Borrelia burgdorferi exhibits genetic diversity, with multiple strains and subspecies identified. This diversity contributes to the variable clinical manifestations of Lyme disease and poses challenges for vaccine development and therapeutic strategies.Research into Borrelia burgdorferi and Lyme disease continues to evolve, with scientists seeking to better understand the spirochete's biology, pathogenesis, andhost-parasite interactions. This knowledge is crucial for developing more effective diagnostic tools, treatment options, and, ultimately, prevention strategies againstthis debilitating disease.In conclusion, Borrelia burgdorferi, the causativeagent of Lyme disease, is a complex and fascinatingorganism that holds significant implications for public health. Its unique biology and ability to evade immune clearance make it a challenging pathogen to combat. However, through continued research and innovation, we may one day find the key to preventing and effectively treating this debilitating disease.。
Reproduction numbers and sub-threshold endemic equilibria for compartmental models of disease trans
Reproduction numbers and sub-threshold endemicequilibria for compartmental models of disease transmissionP.van den Driesschea,1,James Watmough b,*,2aDepartment of Mathematics and Statistics,University of Victoria,Victoria,BC,Canada V8W 3P4b Department of Mathematics and Statistics,University of New Brunswick,Fredericton,NB,Canada E3B 5A3Received 26April 2001;received in revised form 27June 2001;accepted 27June 2001Dedicated to the memory of John JacquezAbstractA precise definition of the basic reproduction number,R 0,is presented for a general compartmental disease transmission model based on a system of ordinary differential equations.It is shown that,if R 0<1,then the disease free equilibrium is locally asymptotically stable;whereas if R 0>1,then it is unstable.Thus,R 0is a threshold parameter for the model.An analysis of the local centre manifold yields a simple criterion for the existence and stability of super-and sub-threshold endemic equilibria for R 0near one.This criterion,together with the definition of R 0,is illustrated by treatment,multigroup,staged progression,multistrain and vector–host models and can be applied to more complex models.The results are significant for disease control.Ó2002Elsevier Science Inc.All rights reserved.Keywords:Basic reproduction number;Sub-threshold equilibrium;Disease transmission model;Disease control1.IntroductionOne of the most important concerns about any infectious disease is its ability to invade a population.Many epidemiological models have a disease free equilibrium (DFE)at whichtheMathematical Biosciences 180(2002)29–48/locate/mbs*Corresponding author.Tel.:+1-5064587323;fax:+1-5064534705.E-mail addresses:pvdd@math.uvic.ca (P.van den Driessche),watmough@unb.ca (J.Watmough).URL:http://www.math.unb.ca/$watmough.1Research supported in part by an NSERC Research Grant,the University of Victoria Committee on faculty research and travel and MITACS.2Research supported by an NSERC Postdoctoral Fellowship tenured at the University of Victoria.0025-5564/02/$-see front matter Ó2002Elsevier Science Inc.All rights reserved.PII:S0025-5564(02)00108-630P.van den Driessche,J.Watmough/Mathematical Biosciences180(2002)29–48population remains in the absence of disease.These models usually have a threshold parameter, known as the basic reproduction number,R0,such that if R0<1,then the DFE is locally as-ymptotically stable,and the disease cannot invade the population,but if R0>1,then the DFE is unstable and invasion is always possible(see the survey paper by Hethcote[1]).Diekmann et al.[2]define R0as the spectral radius of the next generation matrix.We write down in detail a general compartmental disease transmission model suited to heterogeneous populations that can be modelled by a system of ordinary differential equations.We derive an expression for the next generation matrix for this model and examine the threshold R0¼1in detail.The model is suited to a heterogeneous population in which the vital and epidemiological parameters for an individual may depend on such factors as the stage of the disease,spatial position,age or behaviour.However,we assume that the population can be broken into homo-geneous subpopulations,or compartments,such that individuals in a given compartment are indistinguishable from one another.That is,the parameters may vary from compartment to compartment,but are identical for all individuals within a given compartment.We also assume that the parameters do not depend on the length of time an individual has spent in a compart-ment.The model is based on a system of ordinary equations describing the evolution of the number of individuals in each compartment.In addition to showing that R0is a threshold parameter for the local stability of the DFE, we apply centre manifold theory to determine the existence and stability of endemic equilib-ria near the threshold.We show that some models may have unstable endemic equilibria near the DFE for R0<1.This suggests that even though the DFE is locally stable,the disease may persist.The model is developed in Section2.The basic reproduction number is defined and shown to bea threshold parameter in Section3,and the definition is illustrated by several examples in Section4.The analysis of the centre manifold is presented in Section5.The epidemiological ramifications of the results are presented in Section6.2.A general compartmental epidemic model for a heterogeneous populationConsider a heterogeneous population whose individuals are distinguishable by age,behaviour, spatial position and/or stage of disease,but can be grouped into n homogeneous compartments.A general epidemic model for such a population is developed in this section.Let x¼ðx1;...;x nÞt, with each x i P0,be the number of individuals in each compartment.For clarity we sort the compartments so that thefirst m compartments correspond to infected individuals.The distinc-tion between infected and uninfected compartments must be determined from the epidemiological interpretation of the model and cannot be deduced from the structure of the equations alone,as we shall discuss below.It is plausible that more than one interpretation is possible for some models.A simple epidemic model illustrating this is given in Section4.1.The basic reproduction number can not be determined from the structure of the mathematical model alone,but depends on the definition of infected and uninfected compartments.We define X s to be the set of all disease free states.That isX s¼f x P0j x i¼0;i¼1;...;m g:In order to compute R0,it is important to distinguish new infections from all other changes inpopulation.Let F iðxÞbe the rate of appearance of new infections in compartment i,Vþi ðxÞbe therate of transfer of individuals into compartment i by all other means,and VÀi ðxÞbe the rate oftransfer of individuals out of compartment i.It is assumed that each function is continuously differentiable at least twice in each variable.The disease transmission model consists of non-negative initial conditions together with the following system of equations:_x i¼f iðxÞ¼F iðxÞÀV iðxÞ;i¼1;...;n;ð1Þwhere V i¼VÀi ÀVþiand the functions satisfy assumptions(A1)–(A5)described below.Sinceeach function represents a directed transfer of individuals,they are all non-negative.Thus,(A1)if x P0,then F i;Vþi ;VÀiP0for i¼1;...;n.If a compartment is empty,then there can be no transfer of individuals out of the compartment by death,infection,nor any other means.Thus,(A2)if x i¼0then VÀi ¼0.In particular,if x2X s then VÀi¼0for i¼1;...;m.Consider the disease transmission model given by(1)with f iðxÞ,i¼1;...;n,satisfying con-ditions(A1)and(A2).If x i¼0,then f iðxÞP0and hence,the non-negative cone(x i P0, i¼1;...;n)is forward invariant.By Theorems1.1.8and1.1.9of Wiggins[3,p.37]for each non-negative initial condition there is a unique,non-negative solution.The next condition arises from the simple fact that the incidence of infection for uninfected compartments is zero.(A3)F i¼0if i>m.To ensure that the disease free subspace is invariant,we assume that if the population is free of disease then the population will remain free of disease.That is,there is no(density independent) immigration of infectives.This condition is stated as follows:(A4)if x2X s then F iðxÞ¼0and VþiðxÞ¼0for i¼1;...;m.The remaining condition is based on the derivatives of f near a DFE.For our purposes,we define a DFE of(1)to be a(locally asymptotically)stable equilibrium solution of the disease free model,i.e.,(1)restricted to X s.Note that we need not assume that the model has a unique DFE. Consider a population near the DFE x0.If the population remains near the DFE(i.e.,if the introduction of a few infective individuals does not result in an epidemic)then the population will return to the DFE according to the linearized system_x¼Dfðx0ÞðxÀx0Þ;ð2Þwhere Dfðx0Þis the derivative½o f i=o x j evaluated at the DFE,x0(i.e.,the Jacobian matrix).Here, and in what follows,some derivatives are one sided,since x0is on the domain boundary.We restrict our attention to systems in which the DFE is stable in the absence of new infection.That is, (A5)If FðxÞis set to zero,then all eigenvalues of Dfðx0Þhave negative real parts.P.van den Driessche,J.Watmough/Mathematical Biosciences180(2002)29–4831The conditions listed above allow us to partition the matrix Df ðx 0Þas shown by the following lemma.Lemma 1.If x 0is a DFE of (1)and f i ðx Þsatisfies (A1)–(A5),then the derivatives D F ðx 0Þand D V ðx 0Þare partitioned asD F ðx 0Þ¼F 000 ;D V ðx 0Þ¼V 0J 3J 4;where F and V are the m Âm matrices defined byF ¼o F i o x j ðx 0Þ !and V ¼o V i o x jðx 0Þ !with 16i ;j 6m :Further ,F is non-negative ,V is a non-singular M-matrix and all eigenvalues of J 4have positive real part .Proof.Let x 02X s be a DFE.By (A3)and (A4),ðo F i =o x j Þðx 0Þ¼0if either i >m or j >m .Similarly,by (A2)and (A4),if x 2X s then V i ðx Þ¼0for i 6m .Hence,ðo V i =o x j Þðx 0Þ¼0for i 6m and j >m .This shows the stated partition and zero blocks.The non-negativity of F follows from (A1)and (A4).Let f e j g be the Euclidean basis vectors.That is,e j is the j th column of the n Ân identity matrix.Then,for j ¼1;...;m ,o V i o x jðx 0Þ¼lim h !0þV i ðx 0þhe j ÞÀV i ðx 0Þh :To show that V is a non-singular M-matrix,note that if x 0is a DFE,then by (A2)and (A4),V i ðx 0Þ¼0for i ¼1;...;m ,and if i ¼j ,then the i th component of x 0þhe j ¼0and V i ðx 0þhe j Þ60,by (A1)and (A2).Hence,o V i =o x j 0for i m and j ¼i and V has the Z sign pattern (see Appendix A).Additionally,by (A5),all eigenvalues of V have positive real parts.These two conditions imply that V is a non-singular M-matrix [4,p.135(G 20)].Condition (A5)also implies that the eigenvalues of J 4have positive real part.Ã3.The basic reproduction numberThe basic reproduction number,denoted R 0,is ‘the expected number of secondary cases produced,in a completely susceptible population,by a typical infective individual’[2];see also [5,p.17].If R 0<1,then on average an infected individual produces less than one new infected individual over the course of its infectious period,and the infection cannot grow.Conversely,if R 0>1,then each infected individual produces,on average,more than one new infection,and the disease can invade the population.For the case of a single infected compartment,R 0is simply the product of the infection rate and the mean duration of the infection.However,for more complicated models with several infected compartments this simple heuristic definition of R 0is32P.van den Driessche,J.Watmough /Mathematical Biosciences 180(2002)29–48insufficient.A more general basic reproduction number can be defined as the number of new infections produced by a typical infective individual in a population at a DFE.To determine the fate of a‘typical’infective individual introduced into the population,we consider the dynamics of the linearized system(2)with reinfection turned off.That is,the system _x¼ÀD Vðx0ÞðxÀx0Þ:ð3ÞBy(A5),the DFE is locally asymptotically stable in this system.Thus,(3)can be used to de-termine the fate of a small number of infected individuals introduced to a disease free population.Let wi ð0Þbe the number of infected individuals initially in compartment i and letwðtÞ¼w1ðtÞ;...;w mðtÞðÞt be the number of these initially infected individuals remaining in the infected compartments after t time units.That is the vector w is thefirst m components of x.The partitioning of D Vðx0Þimplies that wðtÞsatisfies w0ðtÞ¼ÀV wðtÞ,which has the unique solution wðtÞ¼eÀVt wð0Þ.By Lemma1,V is a non-singular M-matrix and is,therefore,invertible and all of its eigenvalues have positive real parts.Thus,integrating F wðtÞfrom zero to infinity gives the expected number of new infections produced by the initially infected individuals as the vector FVÀ1wð0Þ.Since F is non-negative and V is a non-singular M-matrix,VÀ1is non-negative[4,p.137 (N38)],as is FVÀ1.To interpret the entries of FVÀ1and develop a meaningful definition of R0,consider the fate of an infected individual introduced into compartment k of a disease free population.The(j;k)entry of VÀ1is the average length of time this individual spends in compartment j during its lifetime, assuming that the population remains near the DFE and barring reinfection.The(i;j)entry of F is the rate at which infected individuals in compartment j produce new infections in compartment i. Hence,the(i;k)entry of the product FVÀ1is the expected number of new infections in com-partment i produced by the infected individual originally introduced into compartment k.Fol-lowing Diekmann et al.[2],we call FVÀ1the next generation matrix for the model and set R0¼qðFVÀ1Þ;ð4Þwhere qðAÞdenotes the spectral radius of a matrix A.The DFE,x0,is locally asymptotically stable if all the eigenvalues of the matrix Dfðx0Þhave negative real parts and unstable if any eigenvalue of Dfðx0Þhas a positive real part.By Lemma1, the eigenvalues of Dfðx0Þcan be partitioned into two sets corresponding to the infected and uninfected compartments.These two sets are the eigenvalues of FÀV and those ofÀJ4.Again by Lemma1,the eigenvalues ofÀJ4all have negative real part,thus the stability of the DFE is determined by the eigenvalues of FÀV.The following theorem states that R0is a threshold parameter for the stability of the DFE.Theorem2.Consider the disease transmission model given by(1)with fðxÞsatisfying conditions (A1)–(A5).If x0is a DFE of the model,then x0is locally asymptotically stable if R0<1,but un-stable if R0>1,where R0is defined by(4).Proof.Let J1¼FÀV.Since V is a non-singular M-matrix and F is non-negative,ÀJ1¼VÀF has the Z sign pattern(see Appendix A).Thus,sðJ1Þ<0()ÀJ1is a non-singular M-matrix;P.van den Driessche,J.Watmough/Mathematical Biosciences180(2002)29–483334P.van den Driessche,J.Watmough/Mathematical Biosciences180(2002)29–48where sðJ1Þdenotes the maximum real part of all the eigenvalues of the matrix J1(the spectral abscissa of J1).Since FVÀ1is non-negative,ÀJ1VÀ1¼IÀFVÀ1also has the Z sign pattern.Ap-plying Lemma5of Appendix A,with H¼V and B¼ÀJ1¼VÀF,we have ÀJ1is a non-singular M-matrix()IÀFVÀ1is a non-singular M-matrix:Finally,since FVÀ1is non-negative,all eigenvalues of FVÀ1have magnitude less than or equal to qðFVÀ1Þ.Thus,IÀFVÀ1is a non-singular M-matrix;()qðFVÀ1Þ<1:Hence,sðJ1Þ<0if and only if R0<1.Similarly,it follows thatsðJ1Þ¼0()ÀJ1is a singular M-matrix;()IÀFVÀ1is a singular M-matrix;()qðFVÀ1Þ¼1:The second equivalence follows from Lemma6of Appendix A,with H¼V and K¼F.The remainder of the equivalences follow as with the non-singular case.Hence,sðJ1Þ¼0if and only if R0¼1.It follows that sðJ1Þ>0if and only if R0>1.ÃA similar result can be found in the recent book by Diekmann and Heesterbeek[6,Theorem6.13].This result is known for the special case in which J1is irreducible and V is a positive di-agonal matrix[7–10].The special case in which V has positive diagonal and negative subdiagonal elements is proven in Hyman et al.[11,Appendix B];however,our approach is much simpler(see Section4.3).4.Examples4.1.Treatment modelThe decomposition of fðxÞinto the components F and V is illustrated using a simple treat-ment model.The model is based on the tuberculosis model of Castillo-Chavez and Feng[12,Eq.(1.1)],but also includes treatment failure used in their more elaborate two-strain model[12,Eq.(2.1)].A similar tuberculosis model with two treated compartments is proposed by Blower et al.[13].The population is divided into four compartments,namely,individuals susceptible to tu-berculosis(S),exposed individuals(E),infectious individuals(I)and treated individuals(T).The dynamics are illustrated in Fig.1.Susceptible and treated individuals enter the exposed com-partment at rates b1I=N and b2I=N,respectively,where N¼EþIþSþT.Exposed individuals progress to the infectious compartment at the rate m.All newborns are susceptible,and all indi-viduals die at the rate d>0.Thus,the core of the model is an SEI model using standard inci-dence.The treatment rates are r1for exposed individuals and r2for infectious individuals. However,only a fraction q of the treatments of infectious individuals are successful.Unsuc-cessfully treated infectious individuals re-enter the exposed compartment(p¼1Àq).The diseasetransmission model consists of the following differential equations together with non-negative initial conditions:_E¼b1SI=Nþb2TI=NÀðdþmþr1ÞEþpr2I;ð5aÞ_I¼m EÀðdþr2ÞI;ð5bÞ_S¼bðNÞÀdSÀb1SI=N;ð5cÞ_T¼ÀdTþr1Eþqr2IÀb2TI=N:ð5dÞProgression from E to I and failure of treatment are not considered to be new infections,but rather the progression of an infected individual through the various compartments.Hence,F¼b1SI=Nþb2TI=NB B@1C CA and V¼ðdþmþr1ÞEÀpr2IÀm Eþðdþr2ÞIÀbðNÞþdSþb1SI=NdTÀr1EÀqr2Iþb2TI=NB B@1C CA:ð6ÞThe infected compartments are E and I,giving m¼2.An equilibrium solution with E¼I¼0has the form x0¼ð0;0;S0;0Þt,where S0is any positive solution of bðS0Þ¼dS0.This will be a DFE if and only if b0ðS0Þ<d.Without loss of generality,assume S0¼1is a DFE.Then,F¼0b100;V¼dþmþr1Àpr2Àm dþr2;givingVÀ1¼1ðdþmþr1Þðdþr2ÞÀm pr2dþr2pr2m dþmþr1and R0¼b1m=ððdþmþr1Þðdþr2ÞÀm pr2Þ.A heuristic derivation of the(2;1)entry of VÀ1and R0are as follows:a fraction h1¼m=ðdþmþr1Þof exposed individuals progress to compartment I,a fraction h2¼pr2=ðdþr2Þof infectious individuals re-enter compartment E.Hence,a fractionh1of exposed individuals pass through compartment I at least once,a fraction h21h2passthroughat least twice,and a fraction h k 1h k À12pass through at least k times,spending an average of s ¼1=ðd þr 2Þtime units in compartment I on each pass.Thus,an individual introduced into com-partment E spends,on average,s ðh 1þh 21h 2þÁÁÁÞ¼s h 1=ð1Àh 1h 2Þ¼m =ððd þm þr 1Þðd þr 2ÞÀm pr 2Þtime units in compartment I over its expected lifetime.Multiplying this by b 1gives R 0.The model without treatment (r 1¼r 2¼0)is an SEI model with R 0¼b 1m =ðd ðd þm ÞÞ.The interpretation of R 0for this case is simpler.Only a fraction m =ðd þm Þof exposed individuals progress from compartment E to compartment I ,and individuals entering compartment I spend,on average,1=d time units there.Although conditions (A1)–(A5)do not restrict the decomposition of f i ðx Þto a single choice for F i ,only one such choice is epidemiologically correct.Different choices for the function F lead to different values for the spectral radius of FV À1,as shown in Table 1.In column (a),treatment failure is considered to be a new infection and in column (b),both treatment failure and pro-gression to infectiousness are considered new infections.In each case the condition q ðFV À1Þ<1yields the same portion of parameter space.Thus,q ðFV À1Þis a threshold parameter in both cases.The difference between the numbers lies in the epidemiological interpretation rather than the mathematical analysis.For example,in column (a),the infection rate is b 1þpr 2and an exposed individual is expected to spend m =ððd þm þr 1Þðd þr 2ÞÞtime units in compartment I .However,this reasoning is biologically flawed since treatment failure does not give rise to a newly infected individual.Table 1Decomposition of f leading to alternative thresholds(a)(b)Fb 1SI =N þb 2TI =N þpr 2I 0000B B @1C C A b 1SI =N þb 2TI =N þpr 2I m E 000B B @1C C A Vðd þm þr 1ÞE Àm E þðd þr 2ÞI Àb ðN ÞþdS þb 1SI =N dT Àr 1E Àqr 2I þb 2TI =N 0B B @1C C A ðd þm þr 1ÞE ðd þr 2ÞI Àb ðN ÞþdS þb 1SI =N dT Àr 1E Àqr 2I þb 2TI =N 0B B @1C C A F0b 1þpr 200 0b 1þpr 2m 0 V d þm þr 10Àm d þr 2d þm þr 100d þr 2 q (FV À1)b 1m þpr 2mðd þm þr 1Þðd þr 2Þffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffib 1m þpr 2mðd þm þr 1Þðd þr 2Þs 36P.van den Driessche,J.Watmough /Mathematical Biosciences 180(2002)29–484.2.Multigroup modelIn the epidemiological literature,the term‘multigroup’usually refers to the division of a het-erogeneous population into several homogeneous groups based on individual behaviour(e.g., [14]).Each group is then subdivided into epidemiological compartments.The majority of mul-tigroup models in the literature are used for sexually transmitted diseases,such as HIV/AIDS or gonorrhea,where behaviour is an important factor in the probability of contracting the disease [7,8,14,15].As an example,we use an m-group SIRS-vaccination model of Hethcote[7,14]with a generalized incidence term.The sample model includes several SI multigroup models of HIV/ AIDS as special cases[8,15].The model equations are as follows:_I i ¼X mj¼1b ijðxÞS i I jÀðd iþc iþ iÞI i;ð7aÞ_S i ¼ð1Àp iÞb iÀðd iþh iÞS iþr i R iÀX mj¼1b ijðxÞS i I j;ð7bÞ_Ri¼p i b iþc i I iþh i S iÀðd iþr iÞR i;ð7cÞfor i¼1;...;m,where x¼ðI1;...;I m;S1;...;S m;R1;...;R mÞt.Susceptible and removed individu-als die at the rate d i>0,whereas infected individuals die at the faster rate d iþ i.Infected in-dividuals recover with temporary immunity from re-infection at the rate c i,and immunity lasts an expected1=r i time units.All newborns are susceptible,and a constant fraction b i are born into each group.A fraction p i of newborns are vaccinated at birth.Thereafter,susceptible individuals are vaccinated at the rate h i.The incidence,b ijðxÞdepends on individual behaviour,which determines the amount of mixing between the different groups(see,e.g.,Jacquez et al.[16]). The DFE for this model isx0¼ð0;...;0;S01;...;S0m;R01;...;R0mÞt;whereS0 i ¼b i d ið1Àp iÞþr iðÞd iðd iþh iþr iÞ;R0 i ¼b iðh iþd i p iÞd iðd iþh iþr iÞ:Linearizing(7a)about x¼x0givesF¼S0i b ijðx0ÞÂÃandV¼½ðd iþc iþ iÞd ij ;where d ij is one if i¼j,but zero otherwise.Thus,FVÀ1¼S0i b ijðx0Þ=ðd iÂþc iþ iÞÃ:P.van den Driessche,J.Watmough/Mathematical Biosciences180(2002)29–4837For the special case with b ij separable,that is,b ijðxÞ¼a iðxÞk jðxÞ,F has rank one,and the basic reproduction number isR0¼X mi¼1S0ia iðx0Þk iðx0Þd iþc iþ i:ð8ÞThat is,the basic reproduction number of the disease is the sum of the‘reproduction numbers’for each group.4.3.Staged progression modelThe staged progression model[11,Section3and Appendix B]has a single uninfected com-partment,and infected individuals progress through several stages of the disease with changing infectivity.The model is applicable to many diseases,particularly HIV/AIDS,where transmission probabilities vary as the viral load in an infected individual changes.The model equations are as follows(see Fig.2):_I 1¼X mÀ1k¼1b k SI k=NÀðm1þd1ÞI1;ð9aÞ_Ii¼m iÀ1I iÀ1Àðm iþd iÞI i;i¼2;...;mÀ1;ð9bÞ_Im¼m mÀ1I mÀ1Àd m I m;ð9cÞ_S¼bÀbSÀX mÀ1k¼1b k SI k=N:ð9dÞThe model assumes standard incidence,death rates d i>0in each infectious stage,and thefinal stage has a zero infectivity due to morbidity.Infected individuals spend,on average,1=m i time units in stage i.The unique DFE has I i¼0,i¼1;...;m and S¼1.For simplicity,define m m¼0. Then F¼½F ij and V¼½V ij ,whereF ij¼b j i¼1;j6mÀ1;0otherwise;&ð10ÞV ij¼m iþd i j¼i;Àm j i¼1þj;0otherwise:8<:ð11ÞLet a ij be the(i;j)entry of VÀ1.Thena ij¼0i<j;1=ðm iþd iÞi¼j;Q iÀ1k¼jm kQ ik¼jðm kþd kÞj<i:8>>><>>>:ð12ÞThus,R0¼b1m1þd1þb2m1ðm1þd1Þðm2þd2Þþb3m1m2ðm1þd1Þðm2þd2Þðm3þd3ÞþÁÁÁþb mÀ1m1...m mÀ2ðm1þd1Þ...ðm mÀ1þd mÀ1Þ:ð13ÞThe i th term in R0represents the number of new infections produced by a typical individual during the time it spends in the i th infectious stage.More specifically,m iÀ1=ðm iÀ1þd iÀ1Þis the fraction of individuals reaching stage iÀ1that progress to stage i,and1=ðm iþd iÞis the average time an individual entering stage i spends in stage i.Hence,the i th term in R0is the product of the infectivity of individuals in stage i,the fraction of initially infected individuals surviving at least to stage i,and the average infectious period of an individual in stage i.4.4.Multistrain modelThe recent emergence of resistant viral and bacterial strains,and the effect of treatment on their proliferation is becoming increasingly important[12,13].One framework for studying such sys-tems is the multistrain model shown in Fig.3,which is a caricature of the more detailed treatment model of Castillo-Chavez and Feng[12,Section2]for tuberculosis and the coupled two-strain vector–host model of Feng and Velasco-Hern a ndez[17]for Dengue fever.The model has only a single susceptible compartment,but has two infectious compartments corresponding to the two infectious agents.Each strain is modelled as a simple SIS system.However,strain one may ‘super-infect’an individual infected with strain two,giving rise to a new infection incompartment。
心室致密化不全诊治进展PPT课件
临床特点
❖ 儿童多见,本病在成人发病率高于预期,有家族倾 向
❖ 临床表现无特异性,心肌致密化不全患者出现症状 的年龄和临床表现程度个体差异很大,从无症状到 严重心功能不全,或发生猝死
❖ 虽然心肌致密化不全是先天性心肌病,但心脏病症 状的出现常在成年
临床特点
❖ Demographic characteristics of 53 patients Male gender 25 (47.2%)
anterior and anterolateral
segments
超声诊断
超声诊断-三维超声
超声诊断
Two-dimensional apical 4 chamber and parasternal short axis images at the level of the ventricles show dilatation of both ventricles, multiple trabeculae and intertrabecular recesses in inferior, lateral, anterior walls, middle and apical portions of the septum and apex of the left ventricle. A mild pericardial effusion can be observed
❖ Duration 7 ±5 months( range 1–24) ❖ Prevalence in thБайду номын сангаас INCICh 4/10,000/5 years
临床特点
Clinical and electrocardiographic characteristics IN 53 CASES OF NVM ❖ Precordial pain 11 (21%) ❖ Syncope 5 (9.4%) ❖ NYHA Functional class I/II 40 (75%) ❖ NYHA Functional class III/IV 13 (25%) ❖ Familiar occurrence 16 (30.2%) ❖ Cardiac Rhythm
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ρU Λ
where
+ m ρU Λ = {U ̺Λ U | U ∈ U (2 )}
and ̺Λ is the diagonal matrix ̺Λ = diag{Λ}, Λ = {[λ1 , λ2 , . . . , λ2m ] | λi real, λi = 1, λi ≥ 0 } ;
U (2m ) is the unitary group in 2m -dimensions and Λ is the probability spectrum generating the State ρU Λ . This construction warrants positivity and normalisation. It is however not always (or, better, almost never) convenient in the discussion of physical situations.∗ On the other hand writing ̺ as a vector in H confronts us with the problem of deriving conditions for the expansion coefficients † that guarantee the expansion to yield a state. Formulated in this general way the problem has no obvious solution: positivity and normalisation conditions can derived by expressing the eigenvalues in terms of the expansion coefficients, i.e. finding the zeroes of the characteristic polynomial as functions of these parameters. As we know from Abel and Galois a solution by rational operations and radicals does not exist for quintic or higher degrees, i.e. for general 3-qubit and a fortiori for higher systems. For the 2-qubit explicit expressions are given by the Ferrari-Cardano formulae. In this paper I explicitly construct classes of states for all m whose spectra are determined by charactistic polynomials factorizing into polynomials of a given degree. The novel point in our considerations is the use of hermitian matrix representations of a Clifford algebra to construct bases in H. This particular choice of basis allows us to arrange the 22m − 1 real coordinates of a m-Qubit state in multidimensional arrays which are shown to ’transform’ as O (2m) tensors. This fact implies that the probability spectrum of a mQubit state depends only on O (2m)-invariants, a considerable simplification
{m}
· Γj 1
1
Introduction
For many purposes it is useful to consider m-qubit states as vectors in a R-linear Hilbert space H whose basis is a set {Bi , i = 1 . . . 22m } of 2m × 2m orthonormal trace (Bi · Bj ) = δij , hermitian matrices:
2m
k =0
‡
2m k
= 22m
Complete in the sense that all states factorizing in a specific way are contained in this
set.
3
independent products. Furthermore because of the commutation relations we have trace Γi1
22m
H={
k =1
bk Bk | bk real}.
(H )Βιβλιοθήκη A state is either represented by a hermitian, normalized matrix or an appropriate coordinate vector [b1 , b2 , . . . , b22m ] (a formulation in an appropriate
Generalized Bloch Spheres for m-Qubit States.
Klaus Dietz∗ Sektion Physik, LMU, Muenchen, Theresienstrasse 37, 80333 Muenchen, and Dipartimento di Scienze Fisiche ed Astronomiche dell’Universita di Palermo, via Archirafi 36, 90123 Palermo, Italy February 1, 2008
Permanent Address:Physics Department,University of Bonn, 53115 Bonn, Germany, email: dietz@th.physik.uni-bonn.de
∗
1
projective space would more adequate).In [2] [3] the generators of the quantum invariance group SU (2m ) are proposed as such a basis, a possibility which we shall discuss in the Summary. A straightforward solution for the parametrisation of a state ̺ (a density matrix) is to write the set of all states as {̺} =
∗
2
of the parameter dependencies of these eigenvalues, indeed. This simplification leads to a complete characterisation of complete‡ sets of states which allow for an explicit construction of a parameter domain. In this way I find the set of all states (vector-states) whose parameter domain is the Bloch 2m-sphere. Furthermore a set of (bivector)-states is proposed whose novel parameter domain generalizes the notion of a Bloch sphere. Beyond these two domains the Descartes rule for the positivity of polynomial roots can be used to derive admissible parameter domains.
{m}
(1)
.
From the anticommutation relations we see immediately that the products Γj1 ,j2,...,jk := ik−1 Γj1 · Γj2 · . . . · Γjk k = 2...m (2) (3)
are totally anti-symmetric in the indices [j1 . . . jk ]. The only symmetric object constructed from Clifford numbers is the unity I = Γ2 i as we see from the anticommutation relations. A product consists of at most 2m factors. Hence we have
arXiv:quant-ph/0601013v1 3 Jan 2006
Abstract m-Qubit states are embedded in Cl2m Clifford algebra. Their probability spectrum then depends on O (2m)- or O (2m + 1)-invariants respectively. Parameter domains for O (2m (+1))-vector and -tensor configurations, generalizing the notion of a Bloch sphere, are derived.