Mubritinib_HNMR_15426_MedChemExpress
雷公藤甲素通过抑制细胞铁死亡减轻博来霉素诱发的小鼠肺纤维化
雷公藤甲素通过抑制细胞铁死亡减轻博来霉素诱发的小鼠肺纤维化*张致琴1， 吴洁2， 秦艺璐2， 梁舒2， 左淑飞2， 张超2，高晓2， 范文强2， 张志强1△（1新乡医学院第一附属医院，河南 新乡 453100；2新乡市中心医院，河南 新乡 453000）［摘要］ 目的：探讨雷公藤甲素（TPL ）对博来霉素（BLM ）诱导肺纤维化小鼠的改善效果，揭示铁死亡参与TPL 改善肺纤维化的潜在机制。
方法：选择SPF 级小鼠，随机分为3组：对照组、模型组（BLM 处理组）和实验组（BLM+TPL 处理组），每组10只。
Day 0，模型组和实验组小鼠气管灌注BLM 水溶液（50 μL ，5 mg/kg ），对照组小鼠接受等体积的生理盐水。
Day 1起，实验组小鼠开始灌胃TPL 悬液（200 μL ，0.25 mg/kg ），每3 d 一次，共7次，对照组和模型组分别接受等体积生理盐水。
Masson 染色检测肺组织纤维化，TUNEL 染色检测肺组织细胞凋亡；体外CCK8法检测细胞活力，流式细胞术及活死染色检测细胞凋亡，免疫荧光法检测细胞脂质过氧化，Western blot 检测肺组织及细胞中目的蛋白表达。
结果：TPL 处理通过下调肺组织中I 型胶原（Col I ）和α-平滑肌肌动蛋白（α-SMA ）的表达（P <0.05），显著降低BLM 诱导小鼠肺部纤维化程度（P <0.01）；同时，TPL 处理可显著上调肺组织中谷胱甘肽过氧化酶4（GPX4）和铁死亡抑制蛋白1（FSP1）表达（P <0.05），下调转铁蛋白受体1（TfR1）表达（P <0.05），从而抑制BLM 诱导的肺部细胞铁死亡和凋亡（P <0.01）。
体外研究结果表明，TPL 处理可显著抑制肺上皮细胞凋亡（P <0.01），进一步检测发现TPL 处理可显著降低BLM 诱导的肺泡上皮细胞内脂质过氧化（P <0.01），显著上调细胞内GPX4和FSP1蛋白表达（P <0.01），下调TfR1蛋白表达（P <0.05），通过抑制细胞铁死亡来显著降低BLM 诱导的肺上皮细胞凋亡率。
茶多酚对人脂肪来源间充质干细胞成骨分化的影响
茶多酚对人脂肪来源间充质干细胞成骨分化的影响王华1，齐玉成-杨云芳-赵艺洋2,王慧1，陈培1，杨旭芳1(1.牡丹江医学院，黑龙江牡丹江157011；2.南方医科大学第一临床医学院，广东广州510515)摘要：目的探讨茶多酚(Epigallocatechin-3-gallate,EGCG)对人脂肪间充质干细胞(human adipose-derived mesenchy^-mal stem cells,hADSCs)成骨分化的影响。
方法利用胶原酶消化法和贴壁筛选法从人脂肪组织中分离、培养及扩增hADSCs,倒置显微镜下观察各代hADSCs的形态学特点;利用流式细胞术检测各代hADSCs免疫学表型;取P3代细胞进行成骨诱导分化，实验分三组，即未诱导组、常规成骨诱导组与EGCG组(常规成骨诱导+5^mol/L EGCG),14d后，镜下观察细胞形态学改变及碱性磷酸酶(ALP)染色。
结果体外分离、培养的hADSCs形态均一;流式细胞术结果显示hADSCs具备间充质干细胞的免疫学表型，即CD44、CD73、CD105阳性;成骨诱导14d后部分细胞由长梭形变成多角形，细胞呈现聚集趋势；ALP染色显示EGCG组呈强阳性。
结论成功的从脂肪组织中分离培养出了hADSCs,EGCG能加强其成骨分化能力，这将为骨质疏松症的临床药物开发提供新的思路，亦为组织工程骨的构建提供丰富可靠的种子细胞来源。
关键词：EGCG；人脂肪来源间充质干细胞;成骨分化中图分类号：R595.2文献标识码：A文章编号：1001-7550(2021)01-0001-04Effect of EGCG on osteogenic differentiation of human adipose-derived mesenchymal stem cellsWANG Hua et al(Mudanjiang Medical University,Mudanjiang157011,China)Abstract:Objective To explore the effect of tea polyphenol EGCG on the osteogenic differentiation of human adipose-derived mesenchymal stem cells(hADSCs) .Methods To isolate,culture and amplify hADSCs from human adipose tissue by collagenase digestion and adherent screening methods, the morphological characteristics of each passage of hADSCs were observed under an inverted microscope.The immunophenotype of each generation of hADSCs was detected by flow-cytometry.P3passage cells were taken for osteogenic induction and differentiation,and were divided into three groups:non-induced group, conventional osteogenic induction group and EGCG group(conventional osteogenic induction with+5Rmol/L EGCG).After14days,morphological changes and alkaline phosphatase(ALP)staining were observed under the microscope.Results The morphology of hADSCs isolated and cultured in vitro was uni-form.The results of flow cytometry showed that hADSCs had the immunophenotype of mesenchymal stem cells,such as CD44,CD73and CD105.After14days of osteogenic induction,some cells changed from long spindle shape to polygonal shape,and the cells showed aggregation trend.ALP staining showed strong positive in EGCG group.Conclusion hADSCs have been successfully isolated and cultured from adipose tissue.EGCG can enhance the osteogenic differentiation ability of hADSCs,which will provide a new idea for the clinical drug development of osteoporosis and provide an abundant and reliable source of seed cells for the construction of tissue-engineered bone.Key words:EGCG;human adipose-derived mesenchymal stem cells；osteogenic differentiation随着人口老龄化,骨质疏松症已成为影响人们生活质量的主要因素之一⑷。
黄芩素通过调节HIF-1α
黄芩素通过调节HIF -1α/VEGF 信号通路抑制类风湿关节炎大鼠的炎症反应和病理性血管生成*杜红丽1，张晨宇1，赵清2△［1河南中医药大学第五临床医学院（郑州人民医院）风湿免疫科，河南郑州450053；2河南大学淮河医院风湿免疫科，河南开封475099］［摘要］目的：探讨黄芩素（BA ）调节缺氧诱导因子1α（HIF -1α）/血管内皮生长因子（VEGF ）信号通路对类风湿关节炎（RA ）大鼠炎症反应和病理性血管生成的影响。
方法：按照随机数字表法将SD 大鼠分为对照（control ）组、模型（model ）组、低剂量（10mg/kg ）BA （BA -L ）组、高剂量（30mg/kg ）BA （BA -H ）组、雷公藤多苷片（TWP ；6.25mg/kg ）组和BA -H+HIF -1α激动剂二甲基草酰甘氨酸（DMOG ；40mg/kg ）组，每组12只。
除control 组外，其它组大鼠均采用II 型胶原蛋白-完全弗氏佐剂法诱导RA 大鼠模型。
第2次免疫24h 后开始给药处理，每天给药一次，持续4周。
检测大鼠在给药第0、7、14和28天时的足趾肿胀度，计算关节炎指数；计算大鼠胸腺和脾脏指数；HE 染色检测大鼠踝关节滑膜组织病理损伤；ELISA 法检测大鼠踝关节滑膜组织中肿瘤坏死因子α（TNF -α）和白细胞介素6（IL -6）水平；免疫组化检测大鼠踝关节滑膜组织中VEGF 和VEGF 受体2（又称激酶插入域受体，KDR ）表达；Western blot 检测各组大鼠踝关节滑膜组织中HIF -1α和VEGF 蛋白表达。
结果：与control 组比较，model 组大鼠踝关节滑膜组织病理损伤严重，足趾肿胀度、关节炎指数、胸腺和脾脏指数，以及滑膜组织TNF -α、IL -6、VEGF 、KDR 、HIF -1α和VEGF 水平均显著升高（P <0.05）；与model 组比较，BA -L 组、BA -H 组和TWP 组对应指标变化趋势与上述相反（P <0.05）；BA -H 组与TWP 组比较，上述指标变化差异无统计学意义（P >0.05）；DMOG 减弱了BA -H 对RA 大鼠炎症反应和病理性血管生成的抑制作用。
PPAR-γ在巨噬细胞炎症调控中的作用及机制的研究进展
158CARCINO GENESIS ，TERATO GENESIS &MUTA GENESISPPAR-γ在巨噬细胞炎症调控中的作用及机制的研究进展涂永梅1，3，彭洁1，#，龙子1，#，孔德钦1，陈宇豪2，覃梓瀚2，刘瑞1，李文丽1，*，于卫华1，*(1．空军军医大学军事预防医学院毒理学教研室，陕西省自由基生物学重点实验室，特殊作业环境危害评估与防治教育部重点实验室，陕西西安710032；2．空军军医大学基础医学院学员大队，陕西西安710032；3．陕西中医药大学公共卫生学院，陕西咸阳712046)收稿日期：2021-09-18；修订日期：2022-03-07基金项目：国家自然科学基金(NSFC-32171231，NSFC-31800706)作者信息：涂永梅，E-mail ：*****************。
#并列第一作者，彭洁，E-mail ：***************；龙子，E-mail ：luoze0317@ 。
*通信作者，李文丽，E-mail ：****************.cn ；于卫华，E-mail ：*****************.cn【摘要】巨噬细胞是机体内主要的炎症效应细胞，一般可分化为促炎(M1)型和抗炎(M2)型两大类，针对巨噬细胞极化调控成为炎症相关疾病治疗的新靶点。
研究表明，过氧化物酶体增殖物激活受体-γ(PPAR-γ)作为一个配体激活的核转录因子，可抑制M1型巨噬细胞促炎信号通路的启动，并促进M2型巨噬细胞抗炎信号的表达。
本文主要综述了PPAR-γ在巨噬细胞抗炎症反应中的关键作用，以及PPAR-γ激动剂在脓毒症、肠道炎症、代谢性炎症和自身免疫性炎症疾病治疗中的应用，以期为相关基础研究和临床用药提供指导。
【关键词】PPAR-γ；巨噬细胞；促炎反应；抗炎反应；氧化还原；炎症调控中图分类号：364.5文献标志码：A文章编号：1004-616X(2022)02-0158-04doi ：10.3969/j.issn.1004-616x.2022.02.016过氧化物酶体增殖物激活受体-γ(peroxisome proliferator-activated receptor-γ，PPAR-γ)属于核受体转录因子超家族的关键蛋白，主要通过配体结合途径激活，参与调控脂肪细胞分化和脂肪酸代谢，在肥胖和糖尿病等疾病进展中发挥重要作用[1]。
2_型糖尿病肾病患者血浆D_二聚体水平的检测及临床意义分析
· 医学检验 ·糖尿病新世界 2024年1月糖尿病新世界 DIABETES NEW WORLD2型糖尿病肾病患者血浆D 二聚体水平的检测及临床意义分析陈君颖，游含宇，叶玉琼厦门市中医院检验科，福建厦门 361009[摘要] 目的 评估血浆D 二聚体水平的检测应用在2型糖尿病肾病患者中的作用和临床意义。
方法 回顾性分析厦门市中医院在2021年1月—2022年1月期间收治的120例2型糖尿病肾病患者的全部病历资料，按照尿微量白蛋白排泄率（Urinary Albumin Ejection Rate ， UACR ）予以分组，分为正常蛋白尿组（n =40，UACR<20 μg/min ）、微量蛋白尿组（n =40，UACR 在20~200 μg/min 之间）、大量尿蛋白组（n =40，UACR>200 μg/min ），每组40例。
选取同期健康体检者40例作为对照组，全部均接受血浆D 二聚体水平检查，分析效果与临床意义。
结果 大量尿蛋白组的D 二聚体高于微量蛋白尿组、正常蛋白尿组、对照组，且4组D 二聚体水平对比，差异有统计学意义（P 均<0.05）。
血浆D 二聚体与2型糖尿病肾病患者蛋白尿水平呈正相关（r =0.253）。
结论 2型糖尿病肾病患者蛋白尿水平越高，血浆D 二聚体水平越高，血浆D 二聚体可以作为检测糖尿病肾病患者疾病进展的重要参考指标。
[关键词] 血浆D 二聚体水平；2型糖尿病肾病；临床意义[中图分类号] R587 [文献标识码] A [文章编号] 1672-4062（2024）01（b ）-0052-04Detection and Clinical Significance of Plasma D-dimer Level in Patients with Type 2 Diabetic NephropathyCHEN Junying, YOU Hanyu, YE YuqiongDepartment of Laboratory, Xiamen Hospital of Traditional Chinese Medicine, Xiamen, Fujian Province, 361009 China [Abstract ] Objective To evaluate the role and clinical significance of plasma D-dimer level detection in patients with type 2 diabetic nephropathy. Methods All the medical records of 120 patients with type 2 diabetic nephropathy admitted to Xiamen Hospital of Traditional Chinese Medicine from January 2021 to January 2022 were retrospectively analyzed. According to the urinary albumin ejection rate (UACR), the patients were divided into normal albuminuria group (n =40, UACR<20 μg/min), microalbuminuria group (n =40, UACR>200 μg/min) and large urinary protein group (n =40, UACR>200 μg/min), with 40 cases in each group. 40 healthy subjects in the same period were selected as the control group, and all of them were examined by plasma D-dimer level to analyze the effect and clinical significance. Results The level of D-dimer in the large urinary protein group was higher than that in the microalbuminuria group, the normal albuminuria group and the control group, and comparison of D-dimer levels in 4 groups the differences were statistically significant (all P <0.05). Plasma D-dimer was positively correlated with albuminuria level in type 2 diabetic nephropathy patients (r =0.253). Conclusion The higher the level of proteinuria in type 2 diabetic nephropa⁃thy patients, the higher the level of plasma D-dimer, which can be used as an important reference index to detect the disease progression in diabetic nephropathy patients.[Key words ] Plasma D-dimer level; Type 2 diabetic nephropathy; Clinical significance糖尿病肾病是一种严重的糖尿病微血管病变，其发病率呈逐年上升趋势[1]。
苯环喹溴铵药理毒理研究进展
苯环喹溴铵药理毒理研究进展刘红1王宝辉1王宇1张俊毅1王祥艳1孙艾楠1程颜彬1赵李宏2（1白求恩医科大学制药厂，吉林长春130012；2吉林省成大方圆医药连锁有限公司，吉林长春130041）【摘要】变应性鼻炎和慢性阻塞性肺病是当前两种发病率高、发病人群广泛、病程缠绵且危害显著的呼吸系统疾病。
作为有针对性的新一代治疗药物，苯环喹溴铵是我国自主研发的具有自主知识产权的国家一类新药，具有广阔的应用前景。
【关键词】苯环喹溴铵；药理；毒理；研究进展Research progress of pharmacological effects and toxicological informationof bencycloquidium bromideLIU Hong1 WANG Bao-hui1WANG Y u1 ZHANG Jun-yi1 W ANG Xiang-yan1 SUN Ai-nan1CHENG Y an-bin1 ZHAO Li-hong2(1 Pharmaceutical Factory Norman Bethune University of Medical Science，Jilin Changchun 130012；2 Cheng DaFang Y uan Pharmaceutical Co,Ltd of Jilin Province，Jilin Changchun 130041)【Abtract】At present, allergic rhinitis and chronic obstructive pulmonary disease are the two kinds of respiratory diseases, with the features of high incidence、widespread population incidence、long course and remarkable harm. As a new generation of targeted therapeutic drug, bencycloquidium bromide is an independent intellectual property rights and national class of drug that would get broad application prospect. 【KeyWords】bencycloquidium bromide；pharmacological effects；toxicological information；research progress变应性鼻炎（allergic rhinitis，AR）又称为过敏性鼻炎，是特应性个体接触致敏源后导致的，包含IgE介导的炎症介质释放和多种免疫活性细胞因子、细胞因子参与的鼻黏膜慢性反应性疾病。
重组贻贝粘蛋白的表征及功效评价
生物技术进展 2023 年 第 13 卷 第 4 期 596 ~ 603Current Biotechnology ISSN 2095‑2341研究论文Articles重组贻贝粘蛋白的表征及功效评价李敏 ， 魏文培 ， 乔莎 ， 郝东 ， 周浩 ， 赵硕文 ， 张立峰 ， 侯增淼 *西安德诺海思医疗科技有限公司，西安 710000摘要：为了推进重组贻贝粘蛋白在医疗、化妆品领域的应用，对大肠杆菌规模化发酵及纯化生产获得的重组贻贝粘蛋白进行了表征及功效评价。
经Edman 降解法、基质辅助激光解吸电离飞行时间质谱、PITC 法、非还原型SDS -聚丙烯酰胺凝胶电泳法、凝胶法、改良的Arnow 法对重组贻贝粘蛋白进行氨基酸N 端测序、相对分子量分析、氨基酸组成分析、蛋白纯度分析、内毒素含量测定、多巴含量测定；通过细胞迁移、斑马鱼尾鳍修复效果对重组贻贝粘蛋白进行功效评价。
结果显示，获得的重组贻贝粘蛋白与理论的一级结构一致，蛋白纯度达95%以上，内毒素<10 EU ·mg -1,多巴含量大于5%；重组贻贝粘蛋白浓度为60 μg ·mL -1时能够显著促进细胞增殖的活性（P <0.01）；斑马鱼尾鳍面积样品组与模型对照组相比极显著增加（P <0.001）。
研究结果表明，重组贻贝粘蛋白具有显著的促细胞迁移和修复愈合的功效，具备作为生物医学材料的潜质。
关键词：贻贝粘蛋白；基因重组；生物材料；表征；功效评价DOI ：10.19586/j.20952341.2023.0021 中图分类号：S985.3+1 文献标志码：ACharacterization and Efficacy Evaluation of Recombinant Mussel Adhesive ProteinLI Min ， WEI Wenpei ， QIAO Sha ， HAO Dong ， ZHOU Hao ， ZHAO Shuowen ， ZHANG Lifeng ，HOU Zengmiao *Xi'an DeNovo Hith Medical Technology Co.， Ltd ， Xi'an 710000， ChinaAbstract ：In order to promote the application of recombinant mussel adhesive protein in the medical and cosmetics field ， the recombi⁃nant mussel adhesive protein obtained from scale fermentation and purification of Escherichia coli was characterized and its efficacy was evaluated. Amino acid N -terminal sequencing ， relative molecular weight analysis ， amino acid composition analysis ， protein purityanalysis ， endotoxin content ， dihydroxyphenylalanine （DOPA ） content of recombinant mussel adhesive protein were determined by the following methods ： Edman degradation ， matrix -assisted laser desorption ionization time -of -flight mass spectrometry （MALDI -TOF -MS ）， phenyl -isothiocyanate （PITC ）， nonreductive SDS -polyacrylamide gel electrophoresis （SDS -PAGE ）， gel method ， modified Ar⁃now. The efficacy of recombinant mussel adhesive protein was evaluated by cell migration and repairing effect of zebrafish tail fin. Re⁃sults showed that the obtained recombinant mussel adhesive protein was confirmed to be consistent with the theoretical primary structure ， protein purity of more than 95%， endotoxin <10 EU ·mg -1， DOPA content above 5%. When the recombinant mussel adhesive protein concentration was 60 μg ·mL -1， the effect of promoting cell proliferation was the most obvious ， and it had very significant activity （P <0.01）. The caudal fin area of zebrafish in sample group was significantly increased compared with model control group （P <0.001）. The results indicated that recombinant mussel adhesive protein can promote cell migration and repair healing and has the potential to be used as biomedical materials.Key words ：mussel adhesive protein ； gene recombination ； biological materials ； representation ； efficacy evaluation贻贝粘蛋白（mussel adhesive protein ， MAP ）也称作贻贝足丝蛋白（mussel foot protein ，Mfps ），收稿日期：2023⁃02⁃24； 接受日期：2023⁃03⁃31联系方式：李敏 E -mail:*******************;*通信作者 侯增淼 E -mail:***********************.cn李敏，等：重组贻贝粘蛋白的表征及功效评价是海洋贝类——紫贻贝（Mytilus galloprovincalis）、厚壳贻贝（Mytilus coruscus）、翡翠贻贝（Perna viri⁃dis）等分泌的一种特殊的蛋白质，贻贝中含有多种贻贝粘蛋白，包括贻贝粘蛋白（Mfp 1～6）、前胶原蛋白（precollagens）和基质蛋白（matrix proteins）等［1］。
211232350_莪术醇通过调节lncRNA_NR2F1-AS1
莪术醇通过调节lncRNA NR2F1-AS1/miR-145-5p表达抑制前列腺癌细胞增殖及转移汪洋杨君①（咸宁市中心医院，湖北科技学院附属第一医院泌尿外科，咸宁 437100）中图分类号R737.25 文献标志码 A 文章编号1000-484X（2023）04-0745-05［摘要］目的：探讨莪术醇对前列腺癌细胞增殖及转移的影响及其可能作用机制。
方法：不同浓度的莪术醇处理人前列腺癌细胞LNCap，采用脂质体转染法将si-NC、si-lncRNA NR2F1-AS1分别转染至LNCap细胞，将pcDNA、pcDNA-lncRNANR2F1-AS1分别转染至LNCap细胞后加入100 μg/ml莪术醇处理；CCK-8法、平板克隆形成实验、Transwell实验分别检测细胞增殖、克隆形成、迁移及侵袭；qRT-PCR检测lncRNA NR2F1-AS1、miR-145-5p表达量；双荧光素酶报告实验验证lncRNA NR2F1-AS1与miR-145-5p的靶向关系。
结果：莪术醇可降低细胞存活率，并可降低lncRNA NR2F1-AS1表达量，克隆形成数、迁移及侵袭细胞数减少（P<0.05），而miR-145-5p表达量升高（P<0.05）；lncRNA NR2F1-AS1可靶向调控miR-145-5p的表达（P< 0.05）；转染si-lncRNA NR2F1-AS1后细胞存活率降低（P<0.05），miR-145-5p表达量升高（P<0.05），克隆形成数、迁移及侵袭细胞数减少（P<0.05）；转染pcDNA-lncRNA NR2F1-AS1可降低莪术醇对LNCap细胞增殖及转移的作用。
结论：莪术醇可通过调节lncRNA NR2F1-AS1/miR-145-5p表达，抑制前列腺癌细胞增殖、克隆形成、迁移及侵袭。
［关键词］莪术醇；前列腺癌；lncRNA NR2F1-AS1；miR-145-5p；细胞增殖；迁移；侵袭Curcumol inhibits proliferation and metastasis of prostate cancer cells by regulating expression of lncRNA NR2F1-AS1/miR-145-5pWANG Yang， YANG Jun. Department of Urology， Xianning Central Hospital， the First Affiliated Hospital of Hubei University of Science and Technology， Xianning 437100， China［Abstract］Objective：To explore the effect of curcumol on the proliferation and metastasis of prostate cancer cells and its pos‐sible mechanism. Methods：Different concentrations of curcumol were used to treat human prostate cancer cells LNCap. si-NC， si-lnc-RNA NR2F1-AS1， pcDNA and pcDNA-lncRNA NR2F1-AS1 were transfected into LNCap cells by liposome transfection method re‐spectively. CCK-8 method， plate colony formation experiment， Transwell experiment were used to detect cell proliferation， colony for‐mation， migration and invasion. qRT-PCR was used to detect the expression of lncRNA NR2F1-AS1 and miR-145-5p. The dual lucifer‐ase report experiment was used to verify the targeting relationship between lncRNA NR2F1-AS1 and miR-145-5p. Results：Curcurmol decreased the cell survival rate and the expression of lncRNA NR2F1-AS1， and the number of colony formation， migration and inva‐sion cells were decreased （P<0.05）， while the expression of miR-145-5p was increased （P<0.05）. lncRNA NR2F1-AS1 could target the expression of miR-145-5p （P<0.05）， while the cell survival rate was decreased （P<0.05）， and the numbers of clone formation，migration and invasion cells were decreased （P<0.05）. pcDNA-lncRNA NR2F1-AS1 transfected could reduce the effect of curcumol on the proliferation and metastasis of LNCap cells. Conclusion：Curcumol can inhibit the proliferation， clone formation， migration and invasion of prostate cancer cells by regulating the expression of lncRNA NR2F1-AS1/miR-145-5p.［Key words］Curcumol；Prostate cancer；lncRNA NR2F1-AS1；miR-145-5p；Cell proliferation；Migration；Invasion前列腺癌是常见的一种恶性肿瘤，中药或从天然植物中提取的活性成分具有抗前列腺癌的作用，其主要通过多靶点、多途径等发挥抗肿瘤作用［1-4］。
肿瘤环境下脂肪间充质干细胞旁分泌因子促进结肠癌细胞侵袭
肿瘤环境下脂肪间充质干细胞旁分泌因子促进结肠癌细胞侵袭陈冬梅;刘淑丹;马会明;刘晓明;梁雪云;魏军【摘要】Objective To investigate the underpinning mechanisms by which human adipose-derived mesenchymal stem cells ( hAMSCs) promote cancer metastasis remain elusive. Methods Colon cancer cells were co-cultured with AMSCs using a Transwell model. The capacity of invasion of HCT116 cells in the indicated conditions was de-tected by a Transwell invasive assay. The transcripts of EMT-associated genes in HCT116 treated with indicated conditions were determined by a qRT-PCR assay. Concentration of indicated paracrine factors and cytokines of the culture medium was ascertained by an ELISA. Results The results showed that AMSCs could promote colon canc-er cells to a mesenchymal-like phenotype in a contact-dependent manner. Reciprocally, colon cancer cells were a-ble to induce AMSCs to produce metastasis-related factors and cytokines, such as fibroblast growth factor 10 ( FGF10 ) , vascular endothelial growth factor C ( VEGFC ) , tumor necrosis factor-α( TNF-α) and interleukin 10 (IL-10) in part through a mechanism of an activation of tumor microenvironment. Intriguingly, an inhibition of contact with AMSCs led a reduced capacity of invasion of colon cancer cells in vitro. Conclusion These findings thus suggest that the crosstalk between the microenvironment of cancer cells and paracrine factors of AMSCs has an implication in colon cancer malignancy. This study thus uncovers a novel paracrine factors mediated-crosstalk be-tween colon cancer cells andAMSCs in colon cancer malignancy.%目的确定肿瘤环境下人类脂肪间充质干细胞( hAMSCs)的旁分泌特征变化与肿瘤细胞侵袭的关系. 方法体外分离培养AMSCs,收集培养上清液制备条件培养基,用于培养HCT116细胞. 利用Transwell 培养板共培养AM-SCs与HCT116 细胞. 通过检测穿透人工基底胶( Matrigel胶)的能力比较两种培养条件下HCT116细胞侵袭能力的差异. 实时荧光定量 PCR 检测 HCT116 细胞上皮间质转化( EMT)相关基因的表达变化. ELISA法检测共培养后AM-SCs基因表达和旁分泌因子表达变化. 结果结肠癌细胞系HCT116与AMSCs 的Transwell共培养系统中,HCT116细胞的侵袭能力较AMSCs条件培养液培养显著增强. 同时结肠癌细胞系HCT116构成的肿瘤微环境也影响了AMSCs旁分泌因子表达的变化,其中成纤维生长因子10 ( FGF10 )、血管内皮生长因子C( VEGFC)、肿瘤坏死因子α( TNF-α)和白细胞介素10(IL-10)的表达较正常培养条件下显著上调. 结论肿瘤微环境影响下的AMSCs旁分泌因子表达改变,这些变化能够增强结肠肿瘤细胞的侵袭能力.【期刊名称】《安徽医科大学学报》【年(卷),期】2016(051)001【总页数】6页(P36-41)【关键词】转移性;旁分泌因子;脂肪间充质干细胞;结肠癌【作者】陈冬梅;刘淑丹;马会明;刘晓明;梁雪云;魏军【作者单位】宁夏医科大学总医院干细胞研究所,银川 750004;宁夏医科大学总医院干细胞研究所,银川 750004;宁夏医科大学教育部生育力保持重点实验室,银川750004;宁夏医科大学总医院干细胞研究所,银川 750004;宁夏医科大学总医院干细胞研究所,银川 750004;宁夏医科大学总医院干细胞研究所,银川 750004【正文语种】中文【中图分类】R73-37肥胖已成为发达国家和部分发展中国家共同面对的人口健康问题。
益生菌对阿尔茨海默病作用的研究进展
益生菌对阿尔茨海默病作用的研究进展发布时间：2021-12-14T06:08:15.523Z 来源：《中国结合医学杂志》2021年12期作者：宋鑫萍1,2，李盛钰2，金清1[导读] 阿尔茨海默病已成为威胁全球老年人生命健康的主要疾病之一，患者数量逐年攀升，其护理的经济成本高，给全球经济造成重大挑战。
近年来研究显示，益生菌在适量使用时作为有益于宿主健康的微生物，在防治阿尔茨海默病方面具有积极影响，其作用机制可能通过调节肠道菌群，影响神经免疫系统，调控神经活性物质以及代谢产物，通过肠-脑轴影响该病发生和发展。
宋鑫萍1,2，李盛钰2，金清11.延边大学农学院，吉林延吉 1330022.吉林省农业科学院农产品加工研究所，吉林长春 130033摘要：阿尔茨海默病已成为威胁全球老年人生命健康的主要疾病之一，患者数量逐年攀升，其护理的经济成本高，给全球经济造成重大挑战。
近年来研究显示，益生菌在适量使用时作为有益于宿主健康的微生物，在防治阿尔茨海默病方面具有积极影响，其作用机制可能通过调节肠道菌群，影响神经免疫系统，调控神经活性物质以及代谢产物，通过肠-脑轴影响该病发生和发展。
本文综述了近几年来国内外益生菌对阿尔茨海默病的作用进展，以及其预防和治疗阿尔茨海默病的潜在作用机制。
关键词：益生菌；阿尔茨海默病；肠道菌群；机制Recent Progress in Research on Probiotics Effect on Alzheimer’s DiseaseSONG Xinping1,2，LI Shengyu2，JI Qing1*(1.College of Agricultural, Yanbian University, Yanji 133002，China)(2.Institute of Agro-food Technology, Jilin Academy of Agricultural Sciences, Chanchun 130033, China)Abstract：Alzheimer’s disease has become one of the major diseases threatening the life and health of the global elderly. The number of patients is increasing year by year, and the economic cost of nursing is high, which poses a major challenge to the global economy. In recent years, studies have shown that probiotics, as microorganisms beneficial to the health of the host, have a positive impact on the prevention and treatment of Alzheimer’s disease. Its mechanism may be through regulating intestinal flora, affecting the nervous immune system, regulating the neuroactive substances and metabolites, and affecting the occurrence and development of the disease through thegut- brain axis. This paper reviews the progress of probiotics on Alzheimer’s disease at home and abroad in recent years, as well as its potential mechanism of prevention and treatment.Key words：probiotics; Alzheimer’s disease; gut microbiota; mechanism阿尔茨海默病（Alzheimer’s disease, AD），系中枢神经系统退行性疾病，属于老年期痴呆常见类型，临床特征主要包括：记忆力减退、认知功能障碍、行为改变、焦虑和抑郁等。
氰基硼氢化钠还原胺化京尼平合成拟生物碱与活性
氰基硼氢化钠还原胺化京尼平合成拟生物碱与活性秦杰琛 1），曾小娟 1），张韶湘 1），张晓梅 2），刘鑫洋 1），邹 澄 1），赵 庆 2）（1）昆明医科大学药学院暨云南省天然药物药理重点实验室，云南 昆明 650500；2）云南中医药大学中药学院，云南 昆明 650500）[ 摘要 ] 目的 以京尼平苷为原料通过还原胺化反应合成拟生物碱的方法。
方法 京尼平与胺类化合物在氰基硼氢化钠存在下进行还原反应：京尼平与芳基乙胺的甲醇溶液混合后，加入过量氰基硼氢化钠，放置室温下反应3d，产物经石油醚-异丙醇-二乙胺，石油醚-乙酸乙酯等洗脱分离。
结果 合成共得到9个拟生物碱并对部分拟生物碱进行活性筛选，找到治疗Ⅱ型糖尿病的PTP1B 抑制剂。
结论 部分受试化合物对PTP1B 有抑制作用。
一系列活性衍生物的获得为化合物结构及其生物活性间的构效关系研究打下了基础，有利于寻找具有更高活性的PTP1B 抑制剂。
[ 关键词 ] 京尼平； 拟生物碱； 还原胺化； 抗PTP1B 活性[ 中图分类号 ] R284.1 [ 文献标志码 ] A [ 文章编号 ] 2095 − 610X （2021）02 − 0018 − 05Reductive Amination of Genipin with NaBH 3CN toSynthesize Alkaloid-likes and BioactivityQIN Jie-chen 1），ZENG Xiao-juan 1），ZHANG Shao-xiang 1），ZHANG Xiao-mei 2），LIU Xin-yang 1），ZOU Cheng 1），ZHAO Qing 2）（1） School of Pharmaceutical Science & Yunnan Key Laboratory of Pharmacology for Natural Products ，Kunming Medical University ，Kunming Yunnan 650500; 2） Faculty of Pharmacy ，Yunnan University of Chinese Medicine ，Kunming Yunnan 650500，China ）［Abstract ］ Objective To explore a method for the synthesis of alkaloid-likes from Genipin by reductive amination is reported. Methods The reduction of Genipin and amines in the presence of sodium cyanoborohydride：after the methanol solution of Genipin and arylethylamine was mixed，excessive sodium cyanoborohydride was added and the reaction was kept at room temperature for 3 days. The product was eluted and separated on silica gel by petroleum ether-isopropyl alcohol-diethylamine and petroleum ether-ethyl acetate. Results Nine alkaloid-likes were synthesized. Some alkaloid-likes were screened for inhibition activity of PTP1B enzyme for Ⅱ diabetes treatment. Conclusions All of the tested compounds have a certain inhibitory effect on PTP1B. The acquisition of a series of active derivatives has laid a foundation for the study of the structure-activity relationship between the compounds and their bioactivities，so as to facilitate the search for more active PTP1B inhibitors.［Key words ］ Genipin；Alkaloid-likes；Reductive amination；Inhibition activity against PTP1B目前，通过发现先导化合物是现代新药研发的重要出发点，研究者对具有特定生物活性的先导化合物，利用生物、化学方法进行结构修饰，从而减少化合物毒副作用，提高化合物的活性，增强其生物利用度，最终找到一些作用效果明显，副作用少的新药应用于临床治疗。
锝[99Tc]亚甲基二膦酸盐注射液对类风湿性关节炎的治疗作用及对心理弹性、生活质量的影响
![锝[99Tc]亚甲基二膦酸盐注射液对类风湿性关节炎的治疗作用及对心理弹性、生活质量的影响](https://img.taocdn.com/s3/m/88327f25dcccda38376baf1ffc4ffe473368fd06.png)
DOI：10.19368/ki.2096-1782.2023.15.001锝[99Tc]亚甲基二膦酸盐注射液对类风湿性关节炎的治疗作用及对心理弹性、生活质量的影响聂浩劫1，陈思思1，胡宇萍1，吴开明1，徐永亮1，郭慧萍1，孙平21.黑龙江中医药大学附属第一医院核医学科，黑龙江哈尔滨150040；2.牡丹江医学院解剖教研室，黑龙江牡丹江157011[摘要]目的探讨锝[99Tc]亚甲基二膦酸盐注射液对类风湿性关节炎的治疗作用及对心理弹性、生活质量的影响。
方法选取2021年1—12月黑龙江中医药大学附属第一医院类风湿性关节炎患者100例为研究对象，按照随机数表法分为对照组和观察组，各50例。
对照组采用传统治疗方案（甲氨蝶呤片+来氟米特），观察组在对照组基础上加用锝[99Tc]亚甲基二膦酸盐注射液进行治疗。
统计分析两组临床治疗效果、心理弹性、生活质量。
结果用药后，观察组压痛关节计数、肿胀关节计数、疼痛评分、患者对疾病总体状况的自我评分、第三方医生对患者目前疾病总体状况的评分、类风湿因子、血沉、C反应蛋白水平均低于对照组，差异有统计学意义（P<0.05）；用药后，观察组乐观性、力量性、韧性评分均高于对照组，差异有统计学意义（P<0.05）。
观察组生活质量评分为（91.45±5.52）分，高于对照组，差异有统计学意义（t=12.143，P<0.05）。
结论锝[99Tc]亚甲基二膦酸盐注射液对类风湿性关节炎的治疗作用显著，能够改善患者的心理弹性、生活质量。
[关键词]类风湿性关节炎；锝[99Tc]亚甲基二膦酸盐注射液；治疗作用；心理弹性；生活质量[中图分类号]R593.22 [文献标识码]A [文章编号]2096-1782（2023）08(a)-0001-04Therapeutic Effects of Technetium [99Tc] Methylene diphosphate Injection on Rheumatoid Arthritis and Effects on Psychological Resilience and Qual⁃ity of LifeNIE Haojie1, CHEN Sisi1, HU Yuping1, WU Kaiming1, XU Yongliang1, GUO Huiping1, SUN Ping21.Department of Nuclear Medicine, the First Affiliated Hospital of Heilongjiang University of Traditional Chinese Medicine, Harbin, Heilongjiang Province, 150040 China;2.Department of Anatomy, Mudanjiang Medical College, Mu⁃danjiang, Heilongjiang Province, 157011 China[Abstract] Objective To investigate the therapeutic effect of technetium [99Tc] methylene diphosphate injection on rheumatoid arthritis and its effect on psychological resilience and quality of life. Methods 100 patients with rheuma⁃toid arthritis in the First Affiliated Hospital of Heilongjiang University of Traditional Chinese Medicine from January to December 2021 were selected as the study objects. They were divided into a control group and an observation group, according to the random number table, each with 50 cases. The control group was treated with traditional treat⁃ment program (methotrexate tablets+leflunomide), and the observation group was treated with technetium [99Tc] methy⁃lene diphosphate injection on on the basis of the control group. The clinical treatment effect, psychological resilience, and quality of life of the two groups were statistically analyzed. Results After medication, the patients in the observa⁃tion group had lower pressure-pain joint counts, swollen joint counts, VAS scores, patients' self-ratings of the overall [基金项目]黑龙江省卫生健康委科研课题（2020-300）；黑龙江省医药卫生科研项目（20220101020737）。
细胞蛇的研究进展
2007年,英国牛津大学的刘骥陇等在研究果蝇U 小体和P 小体(U 小体和P 小体是真核生物细胞质中的无膜细胞器)的功能关系时,用4种针对Cup (P 小体中的一种蛋白质)的抗体,对雌性果蝇的卵巢组织进行免疫组织化学染色,染色结果除了预期标记上的P 小体外,还标记出了长条形的丝状结构[1]。
这种结构的形状和数量与纤毛很相似,导致当时以为在果蝇中找到了有纤毛的新细胞类型。
但后来的一系列实验表明,该结构与纤毛没有关系,于是将其命名为“细胞蛇”。
最初是抗Cup 抗体不纯产生假象,意外发现的细胞蛇,而采用亲和层析纯化后的抗Cup 抗体无法再DOI:10.16605/ki.1007-7847.2020.10.0258细胞蛇的研究进展收稿日期:2020-10-22;修回日期:2020-11-19;网络首发日期:2021-07-27基金项目:宁夏自然科学基金项目(2020AAC03179);国家自然科学基金资助项目(31560329)作者简介:李欣玲(1999—),女,广西贵港人,学生;*通信作者:俞晓丽(1984—),女,宁夏银川人,博士,副教授,主要从事干细胞与生殖生物学研究,E-mail:********************。
李欣玲,张樱馨,李进兰,潘文鑫,王彦凤,杨丽蓉,王通,俞晓丽*(宁夏医科大学生育力保持教育部重点实验室临床医学院基础医学院,中国宁夏银川750000)摘要:细胞蛇是近年来细胞生物学研究的热门方向之一,由于其在细胞的增殖、代谢和发育上具有一定的生物学功能,因此,对一些疾病如癌症等的临床诊断或治疗具有一定的指导意义。
细胞蛇是由三磷酸胞苷合成酶(cytidine triphosphate synthetase,CTPS)聚合而成的无膜细胞器,其形成过程及功能在不同类型的细胞中不尽相同。
例如:细胞蛇能促进癌细胞增殖,并使患者病情恶化;过表达的细胞蛇可抑制神经干细胞增殖,影响大脑皮层发育;在卵泡细胞中,细胞蛇相当于CTPS 的存储库,在卵子发生过程起到促进细胞增殖和代谢的作用。
15523666_迷迭香酸对哮喘小鼠氧化性肺损伤的保护作用
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摘要为评价迷迭香酸对哮 喘 小 鼠 模 型 氧 化 性 肺 损 伤 的 保 护 作 用"本 研 究 用 卵 清 蛋 白 %,(;'致 敏$激 发 雌 性 &;B&&S小鼠建立哮喘模型"并用 ,(; 和 Z"," 联合激发小鼠作为氧化肺损伤阳性对照模型#在最后一次滴鼻激 发 "$O 后 "取 支 气 管 肺 泡 灌 洗 液 %&;B['进 行 细 胞 计 数 并 测 定 活 性 氧 %9,D'$超 氧 化 物 歧 化 酶 %D,''和 谷 胱 甘 肽 过 氧化物酶%1DZ?CQ'水平"取左侧肺脏固定做 Z= 染色#结果显示"迷迭香酸可明显减少 &;B[ 中 细 胞 总 数 和 嗜 酸 性粒细胞数目"显著抑制肺组织和 &;B[ 中 9,D的产生"升高 D,' 和 1DZ?CQ水 平"改 善 肺 组 织 病 理 变 化# 本 试 验 结 果 表 明 "迷 迭 香 酸 对 氧 化 肺 损 伤 起 到 明 显 的 保 护 作 用 # 关 键 词 迷 迭 香 酸 *哮 喘 *氧 化 性 肺 损 伤 *抗 氧 化 中 图 分 类 号 9"0434 文 献 标 识 码 ; 文 章 编 号 !+6!?6":+%"8!6'!"?:+48?8+
小檗碱通过激活Nrf2-HO-1GPX4通路抑制小鼠海马神经元HT22细胞的铁死亡
细胞死亡的类型根据细胞形态可以分为3种：自噬、凋亡以及细胞坏死［1］。
铁死亡是区别于以上3种细胞形态的新的细胞形态，它与铁离子水平相关，是由于脂质过氧化物产生而发生的［2］。
它的实质是氧化损伤，即主要为依赖铁离子的脂质过氧化物产生过量并积累，随后产生线粒体变化［3］。
线粒体功能障碍和异常的能量代谢是许多急性和慢性神经退行性疾病的常见上游介质。
研究发现，多种疾病与铁死亡的关联紧密，如阿尔茨海默病、帕金森病等神经退行性疾病［4-7］。
神经退行性疾病是由神经元结构和功能的渐进性丧失或神经元死亡引起的疾病［8］。
大多数神经退行性疾病的发病机制尚不清楚，但越来越多的证据表明，神经退行性疾病的发病机制与兴奋毒素和氧化应激有Berberine inhibits erastin-induced ferroptosis of mouse hippocampal neuronal cells possibly by activating the Nrf2-HO-1/GPX4pathwayHUANG Qingyang 1,JI Dongdong 1,TIAN Xiuyun 1,MA Linyan 2,SUN Xiaojin 21Department of Clinical Medicine,2Department of Pharmacy,Bengbu Medical College,Biochemical Drugs Engineering and Technological Research Center of Anhui Province,Bengbu 233030,China摘要：目的探讨小檗碱对于Erastin 诱导小鼠海马神经元HT22细胞的铁死亡的保护作用及其可能机制。
方法以HT22小鼠海马神经元细胞为研究对象，分为对照组、Erastin 模型组、Erastin+30μmol/L BBR 组、Erastin+60μmol/L BBR 组。
牙源性间充质干细胞对成骨前体细胞成骨分化的影响
现物医学 Progress in Modern Biomedicine VoL21 N C K2 JAN.2021•205 •doi: 10.13241/ki.pmb.2021.02.002牙源性间充质干细胞对成骨前体细胞成骨分化的影响*李慧马博余曰月A郑明珠宋慧妍(首都医科大学附属北京世纪坛医院口腔科北京100038)摘要目的:探讨牙源性间充质干细胞对成骨前体细胞成骨分化的影响。
方法:将小鼠成骨前体细胞M C3T3-E1分为两组,观察组为牙源性间充质干细胞与M C3T3-E1细胞共培养，对照组为单一M C3T3-E1细胞培养。
采用C C K-8法检测细胞增殖水平，采用酶联免疫法检测碱性嶙酸酶(Alkaline phosphatase,A L P)活性并进行茜素红染色，采用q R T-P C R、Western blot检测A L P与骨桥素(osteopontin,O P N)m R N A与蛋白表达水平=结果：细胞共培养丨d与3d后，观察组的细胞增殖指数、A L P活性显著高于对照组(f><0.05)。
与对照组相比，观察组的矿化结节显著增加，经茜素红染色呈红褐色。
细胞共培养1d与3d后，观察组的A L P、O P N m R N A与蛋白相对表达水平显著高于对照组(P<0.05)。
结论：牙源性间充质干细胞能促进成骨前体细胞的A L P、0P N表达，提高A L P活性，增加细胞增殖能力，诱发矿化,从而促进成骨分化。
关键词：牙源性间充质干细胞;成骨前体细胞;碱性磷酸酶;骨桥素；成骨分化中图分类号:R-33;R331.2;R781.4 文献标识码:A文章编号：1673-6273(2021 >02-205-05Effects of Odontogenic Mesenchymal Stem Cells on OsteogenicDifferentiation of Osteoblast Precursor Cells*LI Hui, MA Bo, YU Ri-yue^, ZHENG Ming-zhu, SONG Hui-yan(Department o f S tomatology, Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China) ABSTRACT Objective: 丁o investigate the effects of odontogenic mesenchymal stem cells on osteogenic differentiation o f osteoblast precursor cells. Methods: The MC3T3-E1 mouse osteoblast precursor cells were divided into two groups. The observation group were co-cultured with odontogenic mesenchymal stem cells and MC3T3-E1 cells. The control group were cultured with single MC3T3-E1 cell. CCK-8method were used to detect cell proliferation level, enzyme-linked immunoassay were used to detect Alkaline phosphatase (ALP) activity and alizarin red staining, and qRT-PCR and Western blot were used to detect ALP and osteopontin (osteopontin, OPN) mRNA and protein expression levels. Results: After the cells were co-cultured for 1and 3 days, the cell proliferation index and ALP activity in the observation group were significantly higher than those in the control group (P<0.05). Compared with the control group, the mineralized nodules in the observation group increased significantly, and they were reddish brown after stained with alizarin red. After co-culture of cells for 1and 3 days, the relative expression levels o f ALP, OPN mRNA and protein in the observation group were significantly higher than those in the control group (P<0.05). Conclusion: Dental derived mesenchymal stem cells can promote the expression o f ALP and OPN o f osteoblast precursor cells, increase ALP activity, increase cell proliferation ability, induce mineralization, and thus promote osteogenic differentiation.Key words: Odontogenic mesenchymal stem cells; Osteoblast precursor cells; Alkaline phosphatase; Osteopontin; Osteogenic differentiationChinese Library Classification(CLC): R-33; R331.2; R781.4 Document code: AArticle ID: 1673-6273(2021)02-205-05.X j C. —.刖目牙周病在临床上比较常见，也为一种慢性感染性疾病，可导致牙齿支持组织的破坏，最终造成牙周袋形成与牙槽骨吸收，使得患者牙齿丧失m。
Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes(NI
Sibirny 169, Elaine C.M. Silva-Zacarin 170, Hans-Uwe Simon 171, Cristiano Simone 172, Anne Simonsen 173, Mark A. Smith 174, Katharina Spanel-Borowski 175, Vickram Srinivas 168,Meredith Steeves 34, Harald Stenmark 173, Per E. Stromhaug 176, Carlos S. Subauste 177,Seiichiro Sugimoto 178, David Sulzer 179, Toshihiko Suzuki 180, Michele S. Swanson 181, Ira Tabas 182, Fumihiko Takeshita 183, Nicholas J. Talbot 184, Zsolt Tallóczy 179, KeijiTanaka 95, Kozo Tanaka 185, Isei Tanida 186, Graham S. Taylor 187, J. Paul Taylor 188, Alexei Terman 189, Gianluca Tettamanti 190, Craig B. Thompson 102, Michael Thumm 191, Aviva M.Tolkovsky 192, Sharon A. Tooze 193, Ray Truant 194, Lesya V. Tumanovska 195, YasuoUchiyama 196, Takashi Ueno 96, Néstor L. Uzcátegui 197, Ida van der Klei 89, Eva C.Vaquero 198, Tibor Vellai 199, Michael W. Vogel 200, Hong-Gang Wang 201, Paul Webster 202,John W. Wiley 203, Zhijun Xi 204, Gutian Xiao 205, Joachim Yahalom 206, Jin-Ming Yang 207,George Yap 208, Xiao-Ming Yin 209, Tamotsu Yoshimori 139, Li Yu 107, Zhenyu Yue 210,Michisuke Yuzaki 211, Olga Zabirnyk 212, Xiaoxiang Zheng 213, Xiongwei Zhu 174, and Russell L. Deter 2141 Life Sciences Institute, and Departments of Molecular, Cellular and Developmental Biology and Biological Chemistry; University of Michigan; Ann Arbor, Michigan USA2 Department of Biochemistry and Food Science; Hebrew University; Rehovot, Israel3 Department of Molecular Cell Biology; Catholic University of Leuven; Leuven, Belgium4 Creighton University School of Medicine; Department of Biomedical Sciences;Omaha, Nebraska USA5 Department of Biology; College of Sciences; University of Texas at San Antonio;San Antonio, Texas USA6 Department of Pathology & Laboratory Medicine; University of Cincinnati College of Medicine; Cincinnati, Ohio USA7 Department of Chemical and Biological Sciences; Japan Women’s University; Tokyo, Japan8 Department of Cancer Biology; University of Massachusetts Medical School;Worcester, Massachusetts USA9 Department of Pharmaceutical Sciences; University of Connecticut; Storrs,Connecticut USA 10 Telethon Institute of Genetics and Medicine; Napoli, Italy 11 Department of Biological Sciences; University of Toledo; Toledo, Ohio USA 12 Department of Genetics, Development and Cell Biology,and Plant Sciences Institute; Iowa State University; Ames, Iowa USA 13 Center for Research on Biology and Pathology of Aging; University of Pisa; Pisa, Italy 14 Basic Medical Sciences; Western University of Health Sciences; Pomona, California USA 15 Centre d’études d’agents Pathogènes et Biotechnologies pour la Santé;CNRS UM1, UM2; Institut de Biologie; Montpellier, France 16 Department of Microbiology andImmunology; Indiana University School of Medicine; Indianapolis, Indiana USA 17 Buck Institute for Age Research; Novato, California USA 18 Department of Biological Sciences; University of Pittsburgh;Pittsburgh, Pennsylvania USA 19 Cell Biology Program; Hospital for Sick Children; Toronto, Ontario,Canada 20 Division of Pharmacology; Faculty of Health Sciences; Linköping University; Linköping, Sweden 21 Department of Medicine I; Division of Oncology; Institute of Cancer Research; Medical University of Vienna; Vienna, Austria 22 UMR5095; CNRS; Université de Bordeaux 2; Bordeaux, France 23 Department of Microbiology; Instituto de Fermentaciones Industriales; Madrid, Spain 24 Dulbecco Telethon Institute—IRCCS Santa Lucia Foundation and Department of Biology; University of Rome Tor Vergata; Rome, Italy 25 Department of Immunology; Peking University; Center for Human Disease Genomics; Beijing, China 26 Department of Pharmacology; Emory University School of Medicine; Atlanta, Georgia USA 27 Division of Pulmonary and Critical Care Medicine; Brigham & Womens Hospital; Harvard Medical School; Boston,Massachusetts USA 28 Department of Pathology and Center for Neuroscience; University of Pittsburgh;Pittsburgh, Pennsylvania USA 29 National Creative Research Initiatives Center for Cell Growth Regulation;Department of Biological Sciences; Korea Advanced Institute of Science and Technology; Republic of Korea 30 Département de Biologie Cellulaire et de Morphologie; Université de Lausanne; Lausanne, Switzerland 31 Safar Center for Resuscitation Research; Pittsburgh, Pennsylvania USA 32 Department of Chemistry and Biochemistry and the Molecular Biology Institute; University of California, Los Angeles; Los Angeles,California USA 33 Laboratoire de Génétique Moléculaire des Champignons; Institut de Biochimie et de Génétique Cellulaires; CNRS; Université de Bordeaux 2; Bordeaux, France 34 Department of CancerBiology; The Scripps Research Institute; Jupiter, Florida USA 35 INSERM U756, and the Université Paris-Sud 11; Châtenay-Malabry, France 36 Laboratorio de Biología Celular y Molecular-Instituto de Histología NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author Manuscripty Embriología; Universidad Nacional de Cuyo-CONICET; Mendoza, Argentina 37 Departamento deMorfología y Biología Celular; Universidad de Oviedo; Oviedo, Spain 38 Keck Graduate Institute of Applied Sciences; Claremont, California USA 39 Department of Anatomy and Structural Biology and ofDevelopmental and Molecular Biology; Marion Bessin Liver Research Center; Albert Einstein College of Medicine; Bronx, New York USA 40 Department of Pathology; University of California, San Francisco; San Francisco, California USA 41 Laboratorio Nazionale Consorzio Interuniversitario Biotecnologie; Trieste,Italy 42 Department of Genome Science; University of Cincinnati; Cincinnati, Ohio USA 43 Medical Oncology Branch; National Cancer Institute/Navy Medical Oncology; Bethesda, Maryland USA 44Department of Molecular Genetics and Microbiology; University of New Mexico Health Science Center;Albuquerque, New Mexico USA 45 Department of Biochemistry & Molecular Biology; and ARC Centre of Excellence in Structural and Functional Microbial Genomics; Monash University; Clayton Campus;Melbourne, Victoria, Australia 46 Department of Biology; University of Tor Vergata; via della Ricerca Scientifica; Rome, Italy 47 Department of Physiology; Tufts University; Boston, Massachusetts USA 48Department of Neurology; Massachusetts General Hospital; Charlestown, Massachusetts USA 49Department of Molecular, Cellular & Developmental Biology; Yale University; New Haven, Connecticut 50 Departments of Medicine, Pharmacology and Pathology; Comprehensive Cancer Center; Case Western Reserve University and University Hospitals of Cleveland; Cleveland, Ohio USA 51 Immunotec Research Ltd.; Quebec, Canada 52 Virologie et Immunologie Moleculaires; INRA UR892; Jouy-en-Josas, France 53 Department of Anatomy and Cell Biology; University of Florida College of Medicine; Gainesville, Florida USA 54 Department of Biochemistry; University of Tuebingen; Tuebingen, Germany 55 Baylor College of Medicine; Houston, Texas USA 56 Department of Biological Chemistry; The Weizmann Institute of Science;Rehovot, Israel 57 Department of Biological and Environmental Sciences; University of Helsinki; Helsinki,Finland 58 Departments of Biochemistry and Molecular Biology; Apoptosis and Genomics Research Group of the Hungarian Academy of Sciences; Research Center for Molecular Medicine; University of Debrecen;Debrecen, Hungary 59 Cellular Neurobiology Laboratory; The Salk Institute for Biological Studies; San Diego, California USA 60 Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED); Departamento Bioquímica y Biología Molecular y Genética; E.U. Enfermería; Universidad de Extremadura; Cáceres, Spain 61 Department of Neuro-Oncology; MD Anderson Cancer Center;University of Texas; Houston, Texas USA 62 Department of Frontier Veterinary Medicine; KagoshimaUniversity; Kagoshima, Japan 63 Department of Zoology and Animal Biology; University of Geneva; Geneva,Switzerland 64 Gladstone Institute of Neurological Disease and Department of Neurology; University of California; San Francisco, California USA 65 Department of Pharmacology and Toxicology and Massey Cancer Center; Virginia Commonwealth University; Richmond, Virginia USA 66 Biochemistry and Medical Genetics; Manitoba Institute of Cell Biology; Winnipeg, Manitoba, Canada 67 Institute for Biochemistry and Molecular Biology II; Heinrich Heine University; Düsseldorf, Germany 68 Department of Cell Biology;Harvard Medical School; Boston, Massachusetts USA 69 Department of Developmental Genetics and Gene Control; Institute of Genetics; The University of Nottingham; Queen’s Medical Centre; Nottingham UK 70Genome Sciences Center; British Columbia Cancer Agency; Vancouver, British Columbia, Canada 71BioScience Center; San Diego State University, San Diego; San Diego, California USA 72 Deparment of Internal Medicine; Heinrich Heine Universität; Düsseldorf, Germany 73 Department of Immunology; Duke University Medical Center; Durham, North Carolina USA 74 Department of Pharmacology; University of Colorado Health Sciences Center; Aurora, Colorado USA 75 Division of Cardiology; University of Texas Southwestern Medical Center; Dallas, Texas USA 76 Apoptosis Department and Centre for Genotoxic Stress Research; Institute of Cancer Biology; Danish Cancer Society; Copenhagen, Denmark 77 Cancer Institute,The Second Affiliated Hospital; Zhejiang University School of Medicine; Hangzhou, Zhejiang, China 78Department of Life Science; National Taiwan University; Taipei, Taiwan 79 Department of Immunobiology;Yale University School of Medicine; New Haven, Connecticut USA 80 Department of Microbiology and Molecular Genetics; Medical College of Wisconsin; Milwaukee, Wisconsin USA 81 Memorial Sloan-Kettering Cancer Center; New York, New York USA 82 Department of Pharmacology; University of Medicine and Dentistry of New Jersey—Robert Wood Johnson Medical School; Piscataway, New Jersey USA 83NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptBiochemistry Department; Institute of Medical Biology; University of Tromsø; Tromsø, Norway 84Department of Molecular Microbiology and Immunology; University of Southern California Keck Medical School; Los Angeles, California USA 85 Department of Applied Biological Chemistry; Niigata University;Niigata, Japan 86 Department of Molecular Biology; University of Texas Southwestern Medical Center;Dallas, Texas USA 87 Department of Laboratory Medicine and Pathology; University of Minnesota Cancer Center; Minneapolis, Minnesota USA 88 Department of Medicine & Pharmacology; Wayne State University School of Medicine; Detroit, Michigan USA 89 Molecular Cell Biology; Groningen Biomolecular Sciences and Biotechnology Institute (GBB); University of Groningen; Haren, The Netherlands 90 Division of Pulmonary, Allergy and Critical Care Medicine; University of Pittsburgh Medical Center; Pittsburgh,Pennsylvania USA 91 Department of Molecular Genetics; Weizmann Institute of Science; Rehovot, Israel 92 Department of Radiation Oncology; University Hospitals of Cleveland; Cleveland, Ohio USA 93Department of Biotechnology; University of Tokyo; Tokyo, Japan 94 Department of Cell Biology; Centro de Investigación Príncipe Felipe; Valencia, Spain 95 Laboratory of Frontier Science; Tokyo Metropolitan Institute of Medical Science; Tokyo, Japan 96 Department of Biochemistry; Juntendo University School of Medicine; Tokyo, Japan 97 Department of Neurosurgery; The University of Texas MD Anderson Cancer Center; Houston, Texas USA 98 Department of Anatomy, Cell and Developmental Biology; Eötvös Loránd University; Budapest, Hungary 99 INSERM U848; Institut Gustave Roussy, and the Université Paris-Sud 11;Villejuif, France 100 Division of Developmental Biology; Cincinnati Children’s Hospital ResearchFoundation; Cincinnati, Ohio USA 101 Molecular and Cellular Oncology; MD Anderson Cancer Center;Houston, Texas USA 102 Abramson Family Cancer Research Institute; University of Pennsylvania School of Medicine; Philadelphia, Pennsylvania USA 103 Laval University Cancer Research Center; Québec,Canada 104 Department of Biology; Eastern Michigan University; Ypsilanti, Michigan USA 105 Institute of Health Sciences; Shanghai Jiao Tong University School of Medicine & Shanghai Institutes for Biological Sciences; Chinese Academy of Sciences; Shanghai, China 106 Department of Microbiology and Immunology;National Cheng Kung University; Taiwan 107 Molecular Development Section; Laboratory of Immunology;National Institute of Allergy and Infectious Diseases; National Institutes of Health; Bethesda, Maryland USA 108 Departments of Internal Medicine and Microbiology; University of Texas Southwestern Medical Center;Dallas, Texas USA 109 University of Michigan Medical School; Department of Pathology; Ann Arbor,Michigan USA 110 Neurodegeneration Research Laboratory; National Neuroscience Institute; Singapore 111 Biotechnology Institute; Zhejiang University; Hangzhou, China 112 Department of Anatomy; Chang Gung University; Taiwan 113 Department of Experimental Therapeutics; University of Texas MD Anderson Cancer Center; Houston, Texas USA 114 Departamento de Bioquímica y Biología Molecular; InstitutoUniversitario de Oncología; Universidad de Oviedo; Oviedo, Spain 115 Department of Radiation Oncology;Vanderbilt University; Nashville, Tennessee USA 116 Ben May Department for Cancer Research; University of Chicago; Gordon Center for Integrative Sciences; Chicago, Illinois USA 117 Drug Research andEvaluation; Istituto Superiore di Sanita; Rome, Italy 118 Department of Pharmacology; University of Antwerp; Wilrijk, Antwerp, Belgium 119 Faculty of Agriculture; Kyushu University; Fukuoka, Japan 120GSF—National Research Center for Environment and Health; Munich, Germany 121 Department of Medical Biochemistry; Academic Medical Center; Amsterdam, The Netherlands 122 Biology Department; Queens College; City University of New York; Flushing, New York USA 123 Research Unit for Tropical Diseases de Duve Institute and Laboratory of Biochemistry; Université Catholique de Louvain; Brussels, Belgium 124Department of Biological Chemistry; University of Padua; Padua, Italy 125 Department of Healthcare Sciences; The University College of Antwerp; Antwerp, Belgium 126 Department of Physiology and Cell Biology; Tokyo Medical and Dental University; Tokyo, Japan 127 INSERM ERI 21, and the Laboratoire de Pathologie Clinique et Expérimentale; IFR-50; Nice, France 128 Department of Cell Biology; University Medical Centre Utrecht; Utrecht, The Netherlands 129 Plymouth Marine Laboratory; Plymouth, UK 130Institute of Physiology; Center for Neuroscience and Cell Biology; Coimbra, Portugal 131 Division of Life Science; Graduate School of Science and Engineering; Saitama University; Saitama, Japan 132 Department of Physiological Sciences; Warsaw Agricultural University; Warsaw, Poland 133 Laboratory of ViralImmunobiology; The Rockefeller University; New York, New York USA 134 Novartis Institutes for BiomedicalNIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptResearch; Cambridge, Massachusetts USA 135 Fungal Patho-Biology Group; Temasek Life Sciences Laboratory; National University of Singapore; Singapore 136 Department of Genetics, Cell Biology &Development; University of Minnesota; Minneapolis, Minnesota USA 137 Department of Neuromuscular Research; National Institute of Neuroscience; National Center of Neurology and Psychiatry (NCNP);Kodaira, Tokyo, Japan 138 Nathan Kline Institute; New York University School of Medicine; Orangeburg,New York USA 139 Department of Cellular Regulation; Research Institute for Microbial Diseases; Osaka University; Osaka, Japan 140 Institute for Biochemistry and Molecular Biology II; Heinrich HeineUniversity, Düsseldorf; Düsseldorf, Germany 141 Department of Microbiology and Immunology; University of Tokyo; Tokyo, Japan 142 Departments of Radiation Oncology, Biochemistry and Environmental Health Sciences, and the Case Comprehensive Cancer Center; Case Western Reserve University; Cleveland, Ohio USA 143 Department of Molecular, Cellular and Developmental Biology; University of Michigan; Ann Arbor,Michigan USA 144 Department of Oncology; Chaim-Sheba Medical Center; Ramat-Gan, Israel 145Department of Oral and Craniofacial Biology; Louisiana State University Health Sciences Center School of Dentistry; New Orleans, Louisiana USA 146 Department of Cell Biology and Biophysics; Panepistimiopolis,Athens, Greece 147 Inflammatory Bowel Disease Research Group; Addenbrooke’s Hospital; University of Cambridge; Cambridge, UK 148 Department of Pediatrics; University of Pittsburgh School of Medicine and Children’s Hospital of Pittsburgh; Pittsburgh, Pennsylvania USA 149 Department of Biology; University of Tor Vergata; via della Ricerca Scientifica; Rome, Italy 150 Department of Neurobiochemistry; Tel-Aviv University; Ramat-Aviv, Tel-Aviv, Israel 151 Department of Biochemistry & Molecular Biology; Monash University; Clayton Campus; Melbourne, Victoria, Australia 152 Department of Molecular Biology;University of Tuebingen; Tuebingen, Germany 153 Arthritis and Rheumatism Branch; National Institute of Arthritis and Musculoskeletal and Skin Diseases; National Institutes of Health; Bethesda, Maryland USA 154 Institute of Cellular and Molecular Anatomy; Anatomie III; Clinic of the JWG-University; Frankfurt,Germany 155 Ophthalmology and Neurosciences Division; Medical University of South Carolina;Charleston, South Carolina USA 156 Department of Medical Genetics; Cambridge Institute for Medical Research; Cambridge, United Kingdom 157 Tumour Cell Death Laboratory; Beatson Institute for Cancer Research; Glasgow, Scotland UK 158 Department of Cell Biology and Molecular Medicine; Cardiovascular Research Institute; University of Medicine and Dentistry of New Jersey—New Jersey Medical School; Newark,New Jersey USA 159 Department of Diagnostic and Therapeutic Sciences; Meikai University School ofDentistry; Sakado, Japan 160 Division of Applied Life Sciences; Kyoto University; Kyoto, and JST; CREST;Kyoto, Japan 161 Department of Biomedical Science; University of Padova, Padova; and Dulbecco Telethon Institute at Venetian Institute of Molecular Medicine; Padova, Italy 162 Department of Microbiology and Immunology; University of Tokyo; Tokyo, Japan 163 Laboratorio Nazionale Consorzio Interuniversitario Biotecnologie; Trieste, Italy 164 Department of Cell Biology; Institute for Cancer Research; Rikshospitalet-Radiumhospitalet HF and Department of Molecular Biosciences; University of Oslo; Oslo, Norway 165Department of Animal Science; University of Wyoming; Laramie, Wyoming USA 166 Cancer Center;Massachusetts General Hospital; Charlestown, Massachusetts USA 167 Department of Pathology;University of Alabama at Birmingham; Birmingham, Alabama USA 168 Department of Orthopaedic Surgery;Thomas Jefferson University; Philadelphia, Pennsylvania USA 169 Institute of Cell Biology; NationalAcademy of Sciences of Ukraine; Lviv, Ukraine USA 170 Universidade Federal de Sao Carlos; Sorocaba,Brasil 171 Department of Pharmacology; University of Bern; Bern, Switzerland 172 Department ofTranslational Pharmacology; Consorzio Mario Negri Sud; Santa Maria Imbaro, Italy 173 Centre for Cancer Biomedicine; University of Oslo, and Department of Biochemistry; The Norwegian Radium Hospital;Montebello, Oslo, Norway 174 Department of Pathology; Case Western Reserve University; Cleveland, Ohio USA 175 Institute of Anatomy; University of Leipzig; Leipzig, Germany 176 Division of Biology; University of Missouri; Columbia, Missouri USA 177 Departments of Ophthalmology and Medicine; Case Western Reserve University School of Medicine; Institute of Pathology; Cleveland, Ohio USA 178 Department of Neurology; National Hospital Organization; Miyazaki Higashi Hospital; Miyazaki, Japan 179 Departments of Neurology and Psychiatry; Columbia University; New York, New York USA 180 Department ofMicrobiology; University of the Ryukyus; Okinawa, Japan 181 Department of Microbiology andNIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptImmunology; University of Michigan; Ann Arbor, Michigan USA 182 Department of Medicine; Columbia University; New York, New York USA 183 Department of Molecular Biodefense Research; Yokohama City University Graduate School of Medicine; Yokohama, Japan 184 School of Biosciences; University of Exeter;Exeter UK 185 Institute of Development, Aging and Cancer; Center for Research Strategy and Support;Tohoku University; Miyagi, Japan 186 Department of Biochemistry and Cell Biology; National Institute of Infectious Diseases; Tokyo, Japan 187 CRUK Institute for Cancer Studies; University of Birmingham;Birmingham, UK 188 Department of Neurology; University of Pennsylvania School of Medicine;Philadelphia, Pennsylvania USA 189 Division of Geriatric Medicine; Faculty of Health Sciences; Linköping University; Linköping, Sweden 190 Department of Structural and Functional Biology; University of Insubria;Varese, Italy 191 Zentrum Biochemie und Molekulare Zellbiologie; Georg-August-Universitaet; Goettingen,Germany 192 Department of Biochemistry; University of Cambridge; Cambridge UK 193 Secretory Pathways Laboratory; Cancer Research UK London Research Institute; London, England UK 194Biochemistry and Biomedical Sciences; McMaster University; Hamilton, Ontario, Canada 195 Department of General and Molecular Pathophysiology; Bogomoletz Institute of Physiology; Kiev, Ukraine 196Deptartment of Cell Biology and Neurosciences; Osaka University Graduate School of Medicine; Osaka,Japan 197 Escuela de Bioanálisis; Universidad Central de Venezuela; Caracas, Venezuela 198 Hospital Clínic; Division of Gastroenterology; Barcelona, Catalonia, Spain 199 Department of Genetics; Eötvös Loránd University; Budapest, Hungary 200 Maryland Psychiatric Research Center; Baltimore, Maryland USA 201 H. Lee Moffitt Cancer Center & Research Institute; Tampa, Florida USA 202 Ahmanson Center for Advanced Electron Microscopy & Imaging; House Ear Institute; Los Angeles, California USA 203Department of Internal Medicine; University of Michigan; Ann Arbor, Michigan USA 204 Department of Urology; Peking University First Hospital; Beijing, China 205 Department of Microbiology and Molecular Genetics; University of Pittsburgh School of Medicine; Pittsburgh, Pennsylvania USA 206 Department of Radiation Oncology; Memorial Sloan-Kettering Cancer Center; New York, New York USA 207 The Cancer Institute of New Jersey; University of Medicine and Dentistry of New Jersey—Robert Wood Johnson Medical School; New Brunswick, New Jersey USA 208 Department of Medicine; University of Medicine and Dentistry of New Jersey—New Jersey Medical School, Newark, New Jersey USA 209 Department of Pathology;University of Pittsburgh School of Medicine; Pittsburgh, Pennsylvania USA 210 Department of Neurology and of Neuroscience; Mount Sinai School of Medicine; New York, New York USA 211 Department of Neurophysiology; Keio University School of Medicine; Tokyo, Japan 212 Metabolism and CancerSusceptibility Section; Laboratory of Comparative Carcinogenesis; Center for Cancer Research; NCI—Frederick; National Institutes of Health; Frederick, Maryland USA 213 Department of BiomedicalEngineering; Zhejiang University; Zhejiang, China 214 Department of Obstetrics and Gynecology; Baylor College of Medicine; Houston, Texas USA AbstractResearch in autophagy continues to accelerate,1 and as a result many new scientists are entering thefield. Accordingly, it is important to establish a standard set of criteria for monitoringmacroautophagy in different organisms. Recent reviews have described the range of assays that havebeen used for this purpose.2,3 There are many useful and convenient methods that can be used tomonitor macroautophagy in yeast, but relatively few in other model systems, and there is muchconfusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A keypoint that needs to be emphasized is that there is a difference between measurements that monitorthe numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus,a block in macroautophagy that results in autophagosome accumulation needs to be differentiatedfrom fully functional autophagy that includes delivery to, and degradation within, lysosomes (in mosthigher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for theselection and interpretation of the methods that can be used by investigators who are attempting toexamine macroautophagy and related processes, as well as by reviewers who need to provide realisticand reasonable critiques of papers that investigate these processes. This set of guidelines is not meantNIH-PA Author Manuscript NIH-PA Author ManuscriptNIH-PA Author Manuscriptto be a formulaic set of rules, because the appropriate assays depend in part on the question beingasked and the system being used. In addition, we emphasize that no individual assay is guaranteedto be the most appropriate one in every situation, and we strongly recommend the use of multipleassays to verify an autophagic response.Keywords autolysosome; autophagosome; flux; lysosome; phagophore; stress; vacuole At the first Keystone Symposium on Autophagy in Health and Disease, one of the researchers in the audience, after listening to several comments detailing inadequacies in documenting autophagy, asked the question “What are the essential criteria for demonstrating autophagy?”This is a reasonable question, particularly considering that each of us may have his/her own opinion regarding the answer. Unfortunately, this presents something of a “moving target” for researchers who may think they have met those criteria, only to find out that the reviewer of their paper has different ideas. Conversely, as a reviewer, it is tiresome to raise the same objections repeatedly, wondering why researchers have not fulfilled some of the basic requirements for establishing the occurrence of an autophagic process. In addition, drugs that potentially modulate autophagy are increasingly being used in clinical trials, and screens are being carried out for new drugs that can modulate autophagy for therapeutic purposes. Clearly it is important to determine whether these drugs are truly affecting autophagy based on a set of accepted criteria. Accordingly, we describe here a basic set of contemporary guidelines that can be used by researchers to plan and interpret their experiments, by clinicians to decide which avenue of treatment is appropriate, and by both authors and reviewers to justify or criticize an experimental approach.Several fundamental points must be kept in mind as we establish guidelines for the selectionof appropriate methods to monitor autophagy. Importantly, there are no absolute criteria fordetermining the autophagic status that apply to every situation. This is because some assaysare inappropriate, problematic or may not work at all in particular cells, tissues or organisms.2 In addition, these guidelines may evolve as new methodologies are developed and currentassays of the process are superseded. Nonetheless, it is useful to establish guidelines foracceptable assays that can reliably monitor autophagy in many experimental systems. It isimportant to note that in this set of guidelines the term “autophagy” generally refers tomacroautophagy; other autophagy-related processes are specifically designated whenappropriate.An important point is that autophagy is a dynamic, multi-step process that can be modulatedat several steps, both positively and negatively. In this respect, the autophagic pathway is notdifferent from other cellular pathways. An accumulation of autophagosomes (be they measuredby electron microscopy (EM) image analysis, as fluorescent GFP-LC3 dots, or as LC3lipidation on a western blot), could, for example, reflect either increased autophagosomeformation due to increases in autophagic activity, or to reduced turnover of autophagosomes(Fig. 1). The latter can occur by inhibiting their maturation to amphisomes or autolysosomes,which happens if there are defects in fusion with endosomes or lysosomes, respectively, orfollowing inefficient degradation of the cargo once fusion has occurred.4 For the purposes ofthis review, the autophagic compartments are referred to as the sequestering(preautophagosomal) phagophore,5 the autophagosome,6 the amphisome (generated by fusionof autophagosomes with endosomes, also referred to as an acidic late autophagosome 7)8 andthe autolysosome (generated by fusion of autophagosomes or amphisomes with a lysosome,also referred to as an autophagolysosome).6 We note that the use of the term “phagophore” inthis review has no implied meaning in regard to the origin of the autophagosomal membrane.NIH-PA Author ManuscriptNIH-PA Author ManuscriptNIH-PA Author Manuscript。