[1]杨晓姗,林雅军,魏洁,等.重组人p66Shc腺病毒抑制HeLa细胞增殖的机制研究[J].第三军医大学学报,2014,36(19):1991-1995.
 Yang Xiaoshan,Lin Yajun,Wei Jie,et al.Recombinant adenovirus of human p66Shc inhibits proliferation in HeLa cells[J].J Third Mil Med Univ,2014,36(19):1991-1995.
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
36卷
期数:
2014年第19期
页码:
1991-1995
栏目:
论著
出版日期:
2014-10-15

文章信息/Info

Title:
Recombinant adenovirus of human p66Shc inhibits proliferation in HeLa cells
作者:
杨晓姗林雅军魏洁胡刚徐蓉许晓东马桂蕾孙洪范
北京协和医学院中国医学科学院生物医学工程研究所,天津市生物医学材料重点实验室;卫生部北京医院:卫生部北京老年医学研究所,卫生部老年医学重点实验室,血液科
Author(s):
Yang Xiaoshan Lin Yajun Wei Jie Hu Gang Xu Rong Xu Xiaodong Ma Guilei Sun Hongfan

Institute of Biomedical Engineering, Tianjin Key Laboratory of Biomaterial Research, Peking Union Medical College & Chinese Academy of Medical Sciences, Tianjin, 300192; Beijing Institute of Geriatrics, Key Laboratory of Geriatrics,Department of Hematology, Beijing Hospital, Ministry of Health, Beijing, 100730, China

关键词:
p66ShcHeLa细胞活性氧p53
Keywords:
p66Shc HeLa cells reactive oxygen species p53
分类号:
R73-362; R730.23; R737.33
文献标志码:
A
摘要:

目的      探讨重组人p66Shc腺病毒(AdenoⅩ-p66Shc)对宫颈癌HeLa细胞增殖的抑制作用及机制。       方法        MTT法检测细胞活力,DCFH-DA荧光探针检测活性氧(ROS)生成;ELISA检测8-羟基脱氧鸟苷(8-OHdG)含量;Annexin Ⅴ-FITC/PI荧光染色检测细胞凋亡率;Western blot检测相关蛋白表达;免疫共沉淀检测p66Shc与p53之间相互作用。       结果        重组人p66Shc腺病毒呈剂量依赖性地抑制HeLa细胞增殖(P<0.05),而N-乙酰半胱氨酸能够阻断重组人p66Shc腺病毒的作用。p66Shc能引起细胞ROS水平显著上升(P<0.05),同时伴随8-OHdG含量的升高(P<0.05);p66Shc能引起p53蛋白和CyclinB1蛋白表达及p53磷酸化修饰(p-p53)显著升高(P<0.05),但免疫共沉淀实验结果显示p66Shc与p53并非直接结合,提示p66Shc通过其他信号分子间接调控p53的表达及活性。       结论        重组人p66Shc腺病毒通过使HeLa细胞ROS水平上升,引起DNA氧化损伤,从而诱导p53表达及其磷酸化修饰升高抑制HeLa细胞增殖。

Abstract:

Objective        To determine the inhibitory effect of a recombinant adenovirus of human p66Shc (AdenoⅩ-p66Shc) on the proliferation of human cervical cancer HeLa cells and investigate the possible underlying mechanism.       Methods        The Hela cells were infected with AdenoⅩ-LacZ and AdenoⅩ-p66Shc respectively. The cells without infection served as blank control. Cell viability was analyzed by MTT assay. Reactive oxygen species (ROS) was measured by DCFH-DA fluorescent probes and flow cytometry. The levels of 8-OHdG were detected by ELISA assay. Cell apoptosis was detected by Annexin Ⅴ-FITC/PI staining. Western blotting was used to observe the expression of associated proteins. Co-immunoprecipitation (Co-IP) was used to investigate the possible interaction between p66Shc and p53.       Results        MTT assay showed that the proliferation of HeLa cells was significantly suppressed (P<0.05) by AdenoⅩ-p66Shc in a dose-dependent manner, and its inhibition could be blocked by N-acetyl-L-cysteine. Meanwhile, the levels of ROS and 8-OHdG were significantly increased after AdenoⅩ-p66Shc infection (P<0.05). The expression of p53, Cyclin B1 and p-p53 was up-regulated with the over-expression of p66Shc, but the result of Co-IP demonstrated that p66Shc and p53 were not directly combined, suggesting the expression and activity of p53 was regulated by other molecular activated by p66Shc.       Conclusion        AdenoⅩ-p66Shc induces ROS generation and DNA oxidative damage, and up-regulates the expression of p53 and p-p53, and thereby inhibits the proliferation of HeLa cells.

参考文献/References:

杨晓姗, 林雅军, 魏洁, 等. 重组人p66Shc腺病毒抑制HeLa细胞增殖的机制研究[J].第三军医大学学报,2014,36(19):1991-1995.

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更新日期/Last Update: 2014-09-29