The effects of nanometer realgar uterine cervix cancer cell line SiHa cells and suspension on proliferation and apoptosis of human oncogenic genes HPV16E6/E7 were investigated. A ＂micro-jet effiux＂ strategy was used...The effects of nanometer realgar uterine cervix cancer cell line SiHa cells and suspension on proliferation and apoptosis of human oncogenic genes HPV16E6/E7 were investigated. A ＂micro-jet effiux＂ strategy was used for the preparation of nanometer realgar suspension. SiHa cells were treated with nanometer Realgar suspension in various concentrations （6.25, 12.5, 25 and 50 mg/L） for different durations （12, 24, 48 and 72 h）. The inhibitive effect of nanometer realgar suspension on growth of SiHa cells was detected by MTT method. Special morphological changes of apoptosis were observed by transmission electron microscopy （TEM） and DNA fragments electrophoresis. The apoptotic rate was quantified by flow cytometry （FCM）. The expression of HPV 16E6/E7 mRNA and protein was assayed by RT-PCR and Western blot respectively. The results showed after being treated with 25--50 mg/L nanometer realgar suspension for 48 h, the survival rate of SiHa cells was decreased, and apoptotic rate markedly increased in a time- and concentration-dependent manner. TEM and DNA electrophoresis revealed the special morphological changes of apoptosis. The apoptotic rate of SiHa cells treated with nanometer realgar suspension was significantly higher than in the control group （P〈0.01）, and G0/G1 phase arrest appeared following treatment with nanometer realgar suspension in 25 and 50 mg/L for 48 h. RT-PCR and Western blot assay indicated that nanometer realgar suspension reduced the HPV16E6/E7 gene expression. Nanometer realgar suspension could inhibit the proliferation and induce apoptosis of SiHa cells. The mechanism may be related to the down-regulation of the HPV16E6/E7 gene expression.展开更多
Introduction: Cervical cancer is primarily caused by the human papilloma virus (HPV), which transforms normal cervical cells into cancerous cells that are highly resistant to radiation and chemotherapy. Induction of a...Introduction: Cervical cancer is primarily caused by the human papilloma virus (HPV), which transforms normal cervical cells into cancerous cells that are highly resistant to radiation and chemotherapy. Induction of apoptosis in transformed cells is a key strategy in successfully treating HPV-induced cervical cancer. TRAIL (tumor necrosis factor related apoptosis-inducing ligand) has been shown to selectively induce apoptosis in cancer cells by binding to death receptors and activating extrinsic pathways for apoptosis. However, certain cervical cancers—such as the cultured cell line SiHa—are remarkably resistant to TRAIL. In this study, SiHa cells were sensitized to TRAIL by using sanguinarine—derived from the plant Sanguinaria Canadensis—which is known to induce oxidative stress and lead to the upregulation of receptors for TRAIL. Methods: Cultured SiHa cells were exposed to sub-lethal doses of sanguinarine in combination with TRAIL. Cell viability changes as well as the production of reactive oxygen species (ROS) were assessed. The induction of apoptosis was investigated by assays for caspase activation. Flow cytometry was performed to analyze expression of death receptors 4/5. Results: Treatment of SiHa cells with a combination of sanguinarine and TRAIL led to a significant reduction in cell viability. Significant increase in ROS was observed and caspase activation assays confirmed the induction of apoptosis. Conclusions: The observed synergistic effect of sanguinarine and TRAIL on SiHa cells is promising for the treatment of cervical, and possibly other, HPV-induced cancers. Oxidative stress caused by sanguinarine seems to play a central role in this synergy. The precise link between reactive oxygen species and the possible upregulation of death receptors needs further investigation. This knowledge will enable us to devise more effective treatments for those who suffer from this devastating disease.展开更多
Objective： To evaluate the effects of HPV16 E6/E7 siRNAs on cervical cancer SiHa cells. Methods： The expressions of the E6, E7, p53 and Rb genes were assayed by RT-PCR and Western-bloting respectively. The prolifera...Objective： To evaluate the effects of HPV16 E6/E7 siRNAs on cervical cancer SiHa cells. Methods： The expressions of the E6, E7, p53 and Rb genes were assayed by RT-PCR and Western-bloting respectively. The proliferation and apoptosis of the cells were evaluated by MTT and flow cytometry. Results： HPV 16 E6 and E7 oncogenes were selectivly downregulated by HPV 16 E6 and E7 siRNAs, which sustained at least 96 h by single dose siRNA. Furthermore, reduction of E6 and E7 oncogenes expression upregulated the expressions of P53 and RB protein and induced apoptosis in SiHa cells. Conclusion： Introduction of HPV16 E6/E7 siRNA might be a potentially potent and specific approach to inhibit proliferation and induce apoptosis of SiHa cervical cancer cells.展开更多
目的 :探讨硼替佐米对裸鼠人宫颈癌SiHa细胞移植瘤生长的作用及其与Janus激酶2(Janus kinase2,JAK2)、信号转导与转录激活因子3(signal transducer and activator of transcription 3,STAT3)表达的关系。方法:建立裸鼠人宫颈癌SiHa细胞...目的 :探讨硼替佐米对裸鼠人宫颈癌SiHa细胞移植瘤生长的作用及其与Janus激酶2(Janus kinase2,JAK2)、信号转导与转录激活因子3(signal transducer and activator of transcription 3,STAT3)表达的关系。方法:建立裸鼠人宫颈癌SiHa细胞移植瘤模型,成瘤后随机分为对照组和给药组,每组8只。观察裸鼠体质量变化及移植瘤体积。干预结束后处死裸鼠,剥取瘤体称重,计算抑瘤率。将肿瘤组织行HE染色作病理观察,RT-PCR检测肿瘤组织JAK2、STAT3基因表达,Western blot印迹法检测肿瘤组织中JAK2、STAT3总蛋白及磷酸化的JAK2、STAT3蛋白表达。结果:(1)干预前及干预2周后两组裸鼠体质量无明显差异,但在干预后3周及4周给药组裸鼠体质量小于对照组(均P<0.05)。(2)干预后各时间段给药组肿瘤体积小于对照组(均P<0.05),给药组抑瘤率为21.4%。(3)对照组中肿瘤细胞侵袭结缔组织、脂肪组织区域,给药组中肿瘤组织与结缔组织、脂肪组织之间边界完整、规则;(4)两组JAK2和STAT3基因表达的差异无统计学意义(P>0.05)。(5)两组JAK2和STAT3总蛋白表达量的差异均无统计学意义(P>0.05),但给药组磷酸化JAK2和STAT3蛋白表达量低于对照组(均P>0.01)。结论:硼替佐米能抑制宫颈癌SiHa细胞移植瘤的生长及侵袭,其机制可能通过抑制磷酸化JAK2和STAT3蛋白表达,降低JAK2/STAT3信号通路活性。展开更多
目的探究敲减Rho鸟苷三磷酸酶激活蛋白30(Rho GTPase-activating protein 30,ARHGAP30)后,宫颈癌Siha细胞增殖及凋亡的变化。方法设计特异性shARHGAP30引物并连接pLKO.1载体,转化到大肠杆菌感受态细胞中,再与慢病毒辅助质粒转入HEK-293...目的探究敲减Rho鸟苷三磷酸酶激活蛋白30(Rho GTPase-activating protein 30,ARHGAP30)后,宫颈癌Siha细胞增殖及凋亡的变化。方法设计特异性shARHGAP30引物并连接pLKO.1载体,转化到大肠杆菌感受态细胞中,再与慢病毒辅助质粒转入HEK-293T细胞,收集细胞上清获得的病毒过滤后感染Siha细胞,RT-qPCR和Western blot检测敲减效率,以及转染后Bax及Bcl-2的表达变化;CCK-8法检测敲减后细胞的增殖水平。结果成功构建敲减ARHGAP30基因的慢病毒质粒,并建立Siha稳转细胞,ARHGAP30在Siha细胞中的转录和翻译减少(P<0.01),Bax/Bcl-2明显降低(P<0.01),凋亡减少,细胞增殖水平升高(P<0.01)。结论ARHGAP30参与Siha细胞的增殖与凋亡,调控ARHGAP30基因或将干扰宫颈癌的发生和发展。展开更多
基金a grant from Hubei Chal-lenging Program of Science and Technology,China(No.2007AA301B38-3)
文摘The effects of nanometer realgar uterine cervix cancer cell line SiHa cells and suspension on proliferation and apoptosis of human oncogenic genes HPV16E6/E7 were investigated. A ＂micro-jet effiux＂ strategy was used for the preparation of nanometer realgar suspension. SiHa cells were treated with nanometer Realgar suspension in various concentrations （6.25, 12.5, 25 and 50 mg/L） for different durations （12, 24, 48 and 72 h）. The inhibitive effect of nanometer realgar suspension on growth of SiHa cells was detected by MTT method. Special morphological changes of apoptosis were observed by transmission electron microscopy （TEM） and DNA fragments electrophoresis. The apoptotic rate was quantified by flow cytometry （FCM）. The expression of HPV 16E6/E7 mRNA and protein was assayed by RT-PCR and Western blot respectively. The results showed after being treated with 25--50 mg/L nanometer realgar suspension for 48 h, the survival rate of SiHa cells was decreased, and apoptotic rate markedly increased in a time- and concentration-dependent manner. TEM and DNA electrophoresis revealed the special morphological changes of apoptosis. The apoptotic rate of SiHa cells treated with nanometer realgar suspension was significantly higher than in the control group （P〈0.01）, and G0/G1 phase arrest appeared following treatment with nanometer realgar suspension in 25 and 50 mg/L for 48 h. RT-PCR and Western blot assay indicated that nanometer realgar suspension reduced the HPV16E6/E7 gene expression. Nanometer realgar suspension could inhibit the proliferation and induce apoptosis of SiHa cells. The mechanism may be related to the down-regulation of the HPV16E6/E7 gene expression.
文摘Introduction: Cervical cancer is primarily caused by the human papilloma virus (HPV), which transforms normal cervical cells into cancerous cells that are highly resistant to radiation and chemotherapy. Induction of apoptosis in transformed cells is a key strategy in successfully treating HPV-induced cervical cancer. TRAIL (tumor necrosis factor related apoptosis-inducing ligand) has been shown to selectively induce apoptosis in cancer cells by binding to death receptors and activating extrinsic pathways for apoptosis. However, certain cervical cancers—such as the cultured cell line SiHa—are remarkably resistant to TRAIL. In this study, SiHa cells were sensitized to TRAIL by using sanguinarine—derived from the plant Sanguinaria Canadensis—which is known to induce oxidative stress and lead to the upregulation of receptors for TRAIL. Methods: Cultured SiHa cells were exposed to sub-lethal doses of sanguinarine in combination with TRAIL. Cell viability changes as well as the production of reactive oxygen species (ROS) were assessed. The induction of apoptosis was investigated by assays for caspase activation. Flow cytometry was performed to analyze expression of death receptors 4/5. Results: Treatment of SiHa cells with a combination of sanguinarine and TRAIL led to a significant reduction in cell viability. Significant increase in ROS was observed and caspase activation assays confirmed the induction of apoptosis. Conclusions: The observed synergistic effect of sanguinarine and TRAIL on SiHa cells is promising for the treatment of cervical, and possibly other, HPV-induced cancers. Oxidative stress caused by sanguinarine seems to play a central role in this synergy. The precise link between reactive oxygen species and the possible upregulation of death receptors needs further investigation. This knowledge will enable us to devise more effective treatments for those who suffer from this devastating disease.
基金supported by the grants from the National Natural Science Foundation of China(No.30660192)the Key Project of Education Committee of Jiangxi Province(No.2005-179).
文摘Objective： To evaluate the effects of HPV16 E6/E7 siRNAs on cervical cancer SiHa cells. Methods： The expressions of the E6, E7, p53 and Rb genes were assayed by RT-PCR and Western-bloting respectively. The proliferation and apoptosis of the cells were evaluated by MTT and flow cytometry. Results： HPV 16 E6 and E7 oncogenes were selectivly downregulated by HPV 16 E6 and E7 siRNAs, which sustained at least 96 h by single dose siRNA. Furthermore, reduction of E6 and E7 oncogenes expression upregulated the expressions of P53 and RB protein and induced apoptosis in SiHa cells. Conclusion： Introduction of HPV16 E6/E7 siRNA might be a potentially potent and specific approach to inhibit proliferation and induce apoptosis of SiHa cervical cancer cells.
文摘目的 :探讨硼替佐米对裸鼠人宫颈癌SiHa细胞移植瘤生长的作用及其与Janus激酶2(Janus kinase2,JAK2)、信号转导与转录激活因子3(signal transducer and activator of transcription 3,STAT3)表达的关系。方法:建立裸鼠人宫颈癌SiHa细胞移植瘤模型,成瘤后随机分为对照组和给药组,每组8只。观察裸鼠体质量变化及移植瘤体积。干预结束后处死裸鼠,剥取瘤体称重,计算抑瘤率。将肿瘤组织行HE染色作病理观察,RT-PCR检测肿瘤组织JAK2、STAT3基因表达,Western blot印迹法检测肿瘤组织中JAK2、STAT3总蛋白及磷酸化的JAK2、STAT3蛋白表达。结果:(1)干预前及干预2周后两组裸鼠体质量无明显差异,但在干预后3周及4周给药组裸鼠体质量小于对照组(均P<0.05)。(2)干预后各时间段给药组肿瘤体积小于对照组(均P<0.05),给药组抑瘤率为21.4%。(3)对照组中肿瘤细胞侵袭结缔组织、脂肪组织区域,给药组中肿瘤组织与结缔组织、脂肪组织之间边界完整、规则;(4)两组JAK2和STAT3基因表达的差异无统计学意义(P>0.05)。(5)两组JAK2和STAT3总蛋白表达量的差异均无统计学意义(P>0.05),但给药组磷酸化JAK2和STAT3蛋白表达量低于对照组(均P>0.01)。结论:硼替佐米能抑制宫颈癌SiHa细胞移植瘤的生长及侵袭,其机制可能通过抑制磷酸化JAK2和STAT3蛋白表达,降低JAK2/STAT3信号通路活性。
文摘目的探究敲减Rho鸟苷三磷酸酶激活蛋白30(Rho GTPase-activating protein 30,ARHGAP30)后,宫颈癌Siha细胞增殖及凋亡的变化。方法设计特异性shARHGAP30引物并连接pLKO.1载体,转化到大肠杆菌感受态细胞中,再与慢病毒辅助质粒转入HEK-293T细胞,收集细胞上清获得的病毒过滤后感染Siha细胞,RT-qPCR和Western blot检测敲减效率,以及转染后Bax及Bcl-2的表达变化;CCK-8法检测敲减后细胞的增殖水平。结果成功构建敲减ARHGAP30基因的慢病毒质粒,并建立Siha稳转细胞,ARHGAP30在Siha细胞中的转录和翻译减少(P<0.01),Bax/Bcl-2明显降低(P<0.01),凋亡减少,细胞增殖水平升高(P<0.01)。结论ARHGAP30参与Siha细胞的增殖与凋亡,调控ARHGAP30基因或将干扰宫颈癌的发生和发展。
