巨噬细胞中诱导型一氧化氮合酶来源的 N O

PEG 10mRNA 的抑制作用最强,表明靶mRNA 的

二级结构对siRNA 的功能有很大影响。

本研究中我们构建针对PEG 10基因的siRNA 真核表达载体,并通过酶切鉴定和测序鉴定,说明我们构建的针对PEG 10的真核表达载体是正确有效的,可以用于后续PEG 10基因功能的研究,以及沉默PEG 10后抑制肝癌细胞生长,诱导肝癌细胞凋亡,从而为肝癌的基因治疗提供理论依据和实验工具。

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[编辑:刘红武]

收稿日期:2006202221;修回日期:2006204214基金项目:怀化市科技计划资助项目(0502)

作者单位:1.418000湖南怀化医学高等专科学校检验　

系;2.中国科学院生物物理研究所结构与分子生物学研究　中心作者简介:张申(1955-),男,学士,教授,主要从事自由　基生物学研究

巨噬细胞中诱导型一氧化氮合酶来源的NO 对共培养HL 60细胞凋亡的影响

张　申1,尹利华1,卫涛涛2

E ffects of Inducible Nitric Oxide Synthase Derived Nitric Oxide on Apoptosis of H L 60Cells Co 2cultured with RAW 264.7Macrophages

ZHAN G Shen 1,YIN Li 2hua 1,WEI Tao 2tao 2

1.De partment of L aboratory Medicine ,H uai hua Medical College ,H uai hua 418000,China;

2.Center f or S t ructural and Molecular B iology ,I nstitute of B iophysics ,Chinese A cadem y of Sciences

Abstract :Objective To study effects of inducible nitric oxide synthase (iNOS )2derived nitric oxide (NO )on apoptosis of HL 60cells co 2cultured with RAW 264.7macrophages.Methods Upon stimulation with

lipopolysaccharide (L PS )and interferon 2

γ(IFN 2γ),inducible nitric oxide synthase gene was expressed in RAW 264.7macrophages ,which caused the consequent generation of nitric oxide.Effects of nitric oxide on HL 60cells viability ,expression of bcl 22and bax protein ,activity of Caspase 23and cell apoptosis were evaluated with M T T assay ,Western blot analysis ,fluorescence analysis ,flow cytometry (FCM ),trans 2mission electron microscopy (TEM )and DNA agarose gel electrophoresis.R esults The results showed that iNOS 2derived nitric oxide caused oxidative damage of HL 60cells co 2cultured with RAW 264.7mac 2rophages ,and decreased cell viability ,and evidently reduced expression of bcl 22and increased expression of bax ,and induced activity of caspase 23and DNA f ragmentation.Conclusion The results suggested im 2portant effect of iNOS 2derived nitric oxide on apoptosis of cells in RAW 264.7macrophages.

K ey

w ords :

Inducible

nitric

oxide

synthase ;Nitric oxide ;Apoptosis ;Mac 2

rophage

摘　

要:目的　研究巨噬细胞中诱导型一氧化氮合酶(iN 2OS )来源的一氧化氮(NO )对共培养HL 60细胞凋亡的影响。

方法　以脂多糖(L PS )和γ2干扰素(IN F 2γ)诱导RAW 264.7巨噬细胞iNOS 基因的表达产生过量NO 为实验模型,通过