Protein Rad23, a nucleotide excision repair factor, mainly involves in repairing the DNA damage from environment, such as UV light. The function of Rad23 protein involved in DNA damage repair from many environmental f...Protein Rad23, a nucleotide excision repair factor, mainly involves in repairing the DNA damage from environment, such as UV light. The function of Rad23 protein involved in DNA damage repair from many environmental factors has been studied extensively, but it is not clear from ultraviolet irradiation. To further investigate the photo-protective function of Rad23 protein on HeLa cells damaged from UV light irradiation, firstly, HeLa cells were irradiated by UV light and incubated with the fusion protein of pCold-Rad23, then the cell viability and apoptosis rate were detected by MTT and Hoechst33342/Pl fluorescent staining, respectively. The results show that the recombinant Rad23 protein can protect the HeLa cells from UV irradiation, and inhibit the apoptosis of HeLa cell by UV irradiation.展开更多
To compare the cytotoxicity on HeLa cells induced by nanosized and microsized tellurium powders, HeLa cells were exposed to different concentrations of tellurium powders （0, 50, 100, 150 and 200 μg/mL） for 12 h. In...To compare the cytotoxicity on HeLa cells induced by nanosized and microsized tellurium powders, HeLa cells were exposed to different concentrations of tellurium powders （0, 50, 100, 150 and 200 μg/mL） for 12 h. In this study, detection of a series of biomarkers, including reactive oxygen species （ROS）, glutathione （GSH）, 8-hydroxy-2＇- deoxyguanosine （8-OHdG）, in addition to DNA and protein crosslink （DPC） and MTT assay, were conducted to evaluate the cytotoxicity. It is indicated that compared with the control group, there was no significant difference in the induced cytotoxicity at concentrations lower than 50 μg/mL for both nanosized and microsized tellurium powders. While there appears a significant difference in the induced cytotoxicity for nanosized tellurium powders when the concentration is higher than 100 μg/mL as well as for microsized tellurium powders when the concentration is higher than 200 μg/mL. Moreover, it is found that the cytotoxicity induced on HeLa cells exhibits a certain dose-effect relationship with the concentration of tellurium powders. A conclusion has been reached that the toxicity on HeLa cells can be induced by both nanosized and microsized tellurium powders, and the toxicity of the nanosized tellurium powders is significantly greater than the microsized one.展开更多
背景与目的:Ⅰ型磷酸酶抑制亚基-1(inhibitor-1 of protein phosphataseⅠ,PPI1)依赖于35位苏氨酸的磷酸化而发挥抑制作用。本研究目的在于观察PPI1野生型和持续活化型突变体的表达对宫颈癌细胞株的凋亡及凋亡刺激敏感性的影响,并初步...背景与目的:Ⅰ型磷酸酶抑制亚基-1(inhibitor-1 of protein phosphataseⅠ,PPI1)依赖于35位苏氨酸的磷酸化而发挥抑制作用。本研究目的在于观察PPI1野生型和持续活化型突变体的表达对宫颈癌细胞株的凋亡及凋亡刺激敏感性的影响,并初步探讨其可能的作用机制。方法:通过Lipofectamine介导,用PPI1野生型和持续活化型突变体表达质粒分别转染HeLa细胞,以未转染的及转染空载体的HeLa细胞为对照组,用RT-PCR和Western blot鉴定目的蛋白的表达;通过克隆形成实验观察PPI1对细胞克隆形成能力的影响,用Hoechst33258荧光染色、亚二倍体分析法分析PPI1对HeLa细胞凋亡的影响,用流式细胞术分析转染PPI1后的HeLa细胞对肿瘤坏死因子(tumor-necrosis factor,TNF)凋亡刺激的敏感性,用Western blot分析PPI1对凋亡相关蛋白表达及MAPK信号转导通路的影响。结果:PPI1野生型和持续活化型突变体在HeLa细胞中能够有效表达。荧光染色和亚二倍体分析均表明PPI1持续活化型突变体的表达使细胞凋亡增多,并能明显抑制HeLa细胞的克隆形成能力。Western blot分析表明,PPI1持续活化型突变体可使促凋亡分子P53、P27、BAK表达上调,而抗凋亡分子Bcl-2和Bcl-xL的表达下调。流式细胞术分析表明PPI1持续活化型突变体表达的HeLa细胞在TNF-α刺激后细胞凋亡率为19.06%,而对照组为2.67%和1.89%。Westernblot分析表明,TNF刺激后,在JNK信号转导通路中,该组细胞JNK的磷酸化程度比对照组明显增强,而且持续的时间也较长;对于p38信号转导通路,则在TNF刺激30min后,p38出现磷酸化,而其他组均无。结论:PPI1持续活化型突变体的表达可诱导宫颈癌细胞株的凋亡,并能明显增强HeLa细胞对TNF凋亡刺激的敏感性,其中涉及到JNK、p38信号转导通路的活化增强。展开更多
基金Project(31171176)supported by the National Natural Science Foundation of China
文摘Protein Rad23, a nucleotide excision repair factor, mainly involves in repairing the DNA damage from environment, such as UV light. The function of Rad23 protein involved in DNA damage repair from many environmental factors has been studied extensively, but it is not clear from ultraviolet irradiation. To further investigate the photo-protective function of Rad23 protein on HeLa cells damaged from UV light irradiation, firstly, HeLa cells were irradiated by UV light and incubated with the fusion protein of pCold-Rad23, then the cell viability and apoptosis rate were detected by MTT and Hoechst33342/Pl fluorescent staining, respectively. The results show that the recombinant Rad23 protein can protect the HeLa cells from UV irradiation, and inhibit the apoptosis of HeLa cell by UV irradiation.
文摘To compare the cytotoxicity on HeLa cells induced by nanosized and microsized tellurium powders, HeLa cells were exposed to different concentrations of tellurium powders （0, 50, 100, 150 and 200 μg/mL） for 12 h. In this study, detection of a series of biomarkers, including reactive oxygen species （ROS）, glutathione （GSH）, 8-hydroxy-2＇- deoxyguanosine （8-OHdG）, in addition to DNA and protein crosslink （DPC） and MTT assay, were conducted to evaluate the cytotoxicity. It is indicated that compared with the control group, there was no significant difference in the induced cytotoxicity at concentrations lower than 50 μg/mL for both nanosized and microsized tellurium powders. While there appears a significant difference in the induced cytotoxicity for nanosized tellurium powders when the concentration is higher than 100 μg/mL as well as for microsized tellurium powders when the concentration is higher than 200 μg/mL. Moreover, it is found that the cytotoxicity induced on HeLa cells exhibits a certain dose-effect relationship with the concentration of tellurium powders. A conclusion has been reached that the toxicity on HeLa cells can be induced by both nanosized and microsized tellurium powders, and the toxicity of the nanosized tellurium powders is significantly greater than the microsized one.
文摘背景与目的:Ⅰ型磷酸酶抑制亚基-1(inhibitor-1 of protein phosphataseⅠ,PPI1)依赖于35位苏氨酸的磷酸化而发挥抑制作用。本研究目的在于观察PPI1野生型和持续活化型突变体的表达对宫颈癌细胞株的凋亡及凋亡刺激敏感性的影响,并初步探讨其可能的作用机制。方法:通过Lipofectamine介导,用PPI1野生型和持续活化型突变体表达质粒分别转染HeLa细胞,以未转染的及转染空载体的HeLa细胞为对照组,用RT-PCR和Western blot鉴定目的蛋白的表达;通过克隆形成实验观察PPI1对细胞克隆形成能力的影响,用Hoechst33258荧光染色、亚二倍体分析法分析PPI1对HeLa细胞凋亡的影响,用流式细胞术分析转染PPI1后的HeLa细胞对肿瘤坏死因子(tumor-necrosis factor,TNF)凋亡刺激的敏感性,用Western blot分析PPI1对凋亡相关蛋白表达及MAPK信号转导通路的影响。结果:PPI1野生型和持续活化型突变体在HeLa细胞中能够有效表达。荧光染色和亚二倍体分析均表明PPI1持续活化型突变体的表达使细胞凋亡增多,并能明显抑制HeLa细胞的克隆形成能力。Western blot分析表明,PPI1持续活化型突变体可使促凋亡分子P53、P27、BAK表达上调,而抗凋亡分子Bcl-2和Bcl-xL的表达下调。流式细胞术分析表明PPI1持续活化型突变体表达的HeLa细胞在TNF-α刺激后细胞凋亡率为19.06%,而对照组为2.67%和1.89%。Westernblot分析表明,TNF刺激后,在JNK信号转导通路中,该组细胞JNK的磷酸化程度比对照组明显增强,而且持续的时间也较长;对于p38信号转导通路,则在TNF刺激30min后,p38出现磷酸化,而其他组均无。结论:PPI1持续活化型突变体的表达可诱导宫颈癌细胞株的凋亡,并能明显增强HeLa细胞对TNF凋亡刺激的敏感性,其中涉及到JNK、p38信号转导通路的活化增强。
