Objective: To study whether miR-21 targets and inhibits tumor suppressor gene PTEN can promote prostate cancer cell proliferation and invasion. Methods: Prostate cancer cell lines PC-3 were cultured and divided into n...Objective: To study whether miR-21 targets and inhibits tumor suppressor gene PTEN can promote prostate cancer cell proliferation and invasion. Methods: Prostate cancer cell lines PC-3 were cultured and divided into negative control group (NC group), miR-21 group, pcDNA3.1 group, miR-21+pcDNA3.1 group and miR-21+PTEN group that were transfected with different miR and plasmid, respectively. After 12 h and 24 h of transfection, the cell viability and invasive cell number were determined; after 24 h of transfection, Bcl-2, Survivin, MMP2, MMP9, PTEN, PI3K, and AKT expression in cells were determined. Results: After 12 h and 24 h of transfection, OD value and invasive cell number of miR-21 group were significantly higher than those of NC group; after 24 h of transfection, Bcl-2, Survivin, MMP2, MMP9, PI3K and AKT expression levels were significantly higher than those of NC group while PTEN expression level was significantly lower than that of NC group; after 12 h and 24 h of transfection, OD value and invasive cell number of miR-21+pcDNA3.1 group were significantly higher than those of pcDNA3.1 group, and the OD value and invasive cell number of 1niR-21+PTEN group were significantly lower than those of miR-21+pcDNA3.1 group; after 24 h of transfection, Bcl-2, Survivin, MMP2 and MMP9 content of miR-21+pcDNA3.1 group were significantly higher than those of pcDNA3.1 group, and Bcl-2, Survivin, MMP2 and MMP9 content of miR-21+PTEN group were significantly lower than those of miR-21+pcDNA3.1 group. Conclusions: miR-21 can target and inhibit tumor suppressor gene PTEN expression to promote prostate cancer cell proliferation and invasion.展开更多
In this study, we reported the effect of the ATP binding site competitive inhibitor Torin1 on activated α2-macroglobulin (α2M*)-induced cell proliferation and activation of mTORC1 and mTORC2 signaling in prostate ca...In this study, we reported the effect of the ATP binding site competitive inhibitor Torin1 on activated α2-macroglobulin (α2M*)-induced cell proliferation and activation of mTORC1 and mTORC2 signaling in prostate cancer cells. Torin1 significantly inhibited α2M*-induced cellproliferation as measured by protein and DNA synthesis. Translational activity, a major cellular response in malignant cells,is coordinately regulated by the mTORC1-S6-kinaseand mTORC1-4EBP1 axes. Torin1 significantly inhibited α2M*- and insulin-induced activation of mTORC1 as determined by phosphorylation of S6-kinaseat Thr389 and 4EBP1 at Thr37/46 compared to untreated cells employing Raptor immunoprecipitates. Torin1 also significantly inhibited α2M*- and insulin-induced upregulation of p-AktT308 and p-AktS473 in prostate cancer cells. The effect was comparable to that of insulin employed as a positive control. Finally, Torin1 inhibited α2M*- and insulin-induced activation of mTORC2 kinase assayas measured by phosphorylation of Akt at Ser473 inRictor immunoprecipitates of prostate cancer cells.展开更多
目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响...目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响;流式细胞术检测细胞凋亡率;Western印迹检测PPⅠ对磷酸化细胞外调节蛋白激酶1/2(p-ERK1/2)、细胞外调节蛋白激酶1/2(ERK1/2)、核转录因子kappa B(NF-κB)/p65以及DNA甲基转移酶1(DNMT1)蛋白表达的影响,并加入ERK1/2抑制剂(PD98059)后检测PPⅠ对NF-κB/p65表达的影响。结果:MTT结果显示,PPⅠ能抑制PC3细胞的体外增殖,与对照组相比,PC3细胞从给药浓度0.4μmol/L(0.85±0.05 vs 1.00±0.00,P<0.01)开始明显下降,且呈时间和剂量依赖性。流式细胞术检测显示,PPⅠ能明显诱导PC3细胞早期凋亡,与对照组相比,PC3细胞早期凋亡率从给药浓度0.8μmol/L(13.83±2.97 vs 4.83±0.95)开始明显增加(P均<0.01),且呈剂量依赖性;Western印迹显示,PPⅠ以时间依赖性上调p-ERK1/2和ERK1/2蛋白的激活与表达,与对照组相比,给药时间从2 h(1.73±0.17 vs 1.00±0.00,P<0.01)开始,p-ERK1/2明显被激活表达,PPⅠ以剂量依赖性下调NF-κB/p65、DNMT1蛋白的表达,与对照组相比,给药浓度分别从1.6μmol/L(0.67±0.11 vs 1.00,P<0.01)与1.2μmol/L(0.63±0.06 vs 1.00±0.00,P<0.01)开始,NF-κB/p65、DNMT1表达明显下调;抑制ERK1/2磷酸化能逆转重楼皂苷Ⅰ对NF-κB/p65蛋白表达的下调,与PPⅠ组相比,PD98059+PPⅠ组NF-κB/p65蛋白表达(0.86±0.18 vs 0.43±0.09,P<0.05)明显上调。结论:PPⅠ可能通过介导ERK1/2通路抑制NF-κB/p65和DNMT1蛋白表达,诱导PC3细胞早期凋亡,进而抑制细胞增殖。展开更多
基金supported by Science and Technology Plan Project of Beijing(No:Z151100004015194)
文摘Objective: To study whether miR-21 targets and inhibits tumor suppressor gene PTEN can promote prostate cancer cell proliferation and invasion. Methods: Prostate cancer cell lines PC-3 were cultured and divided into negative control group (NC group), miR-21 group, pcDNA3.1 group, miR-21+pcDNA3.1 group and miR-21+PTEN group that were transfected with different miR and plasmid, respectively. After 12 h and 24 h of transfection, the cell viability and invasive cell number were determined; after 24 h of transfection, Bcl-2, Survivin, MMP2, MMP9, PTEN, PI3K, and AKT expression in cells were determined. Results: After 12 h and 24 h of transfection, OD value and invasive cell number of miR-21 group were significantly higher than those of NC group; after 24 h of transfection, Bcl-2, Survivin, MMP2, MMP9, PI3K and AKT expression levels were significantly higher than those of NC group while PTEN expression level was significantly lower than that of NC group; after 12 h and 24 h of transfection, OD value and invasive cell number of miR-21+pcDNA3.1 group were significantly higher than those of pcDNA3.1 group, and the OD value and invasive cell number of 1niR-21+PTEN group were significantly lower than those of miR-21+pcDNA3.1 group; after 24 h of transfection, Bcl-2, Survivin, MMP2 and MMP9 content of miR-21+pcDNA3.1 group were significantly higher than those of pcDNA3.1 group, and Bcl-2, Survivin, MMP2 and MMP9 content of miR-21+PTEN group were significantly lower than those of miR-21+pcDNA3.1 group. Conclusions: miR-21 can target and inhibit tumor suppressor gene PTEN expression to promote prostate cancer cell proliferation and invasion.
文摘In this study, we reported the effect of the ATP binding site competitive inhibitor Torin1 on activated α2-macroglobulin (α2M*)-induced cell proliferation and activation of mTORC1 and mTORC2 signaling in prostate cancer cells. Torin1 significantly inhibited α2M*-induced cellproliferation as measured by protein and DNA synthesis. Translational activity, a major cellular response in malignant cells,is coordinately regulated by the mTORC1-S6-kinaseand mTORC1-4EBP1 axes. Torin1 significantly inhibited α2M*- and insulin-induced activation of mTORC1 as determined by phosphorylation of S6-kinaseat Thr389 and 4EBP1 at Thr37/46 compared to untreated cells employing Raptor immunoprecipitates. Torin1 also significantly inhibited α2M*- and insulin-induced upregulation of p-AktT308 and p-AktS473 in prostate cancer cells. The effect was comparable to that of insulin employed as a positive control. Finally, Torin1 inhibited α2M*- and insulin-induced activation of mTORC2 kinase assayas measured by phosphorylation of Akt at Ser473 inRictor immunoprecipitates of prostate cancer cells.
文摘目的:研究重楼皂苷Ⅰ(PPⅠ)对去势抵抗性人前列腺癌PC3细胞生长的抑制作用及其分子机制。方法:体外培养PC3细胞,予不同浓度(0.4、0.8、1.2、1.6、2.0、2.4μmol/L)的PPⅠ处理24、48、72 h,另设对照组,MTT法观察PPⅠ对PC3细胞增殖的影响;流式细胞术检测细胞凋亡率;Western印迹检测PPⅠ对磷酸化细胞外调节蛋白激酶1/2(p-ERK1/2)、细胞外调节蛋白激酶1/2(ERK1/2)、核转录因子kappa B(NF-κB)/p65以及DNA甲基转移酶1(DNMT1)蛋白表达的影响,并加入ERK1/2抑制剂(PD98059)后检测PPⅠ对NF-κB/p65表达的影响。结果:MTT结果显示,PPⅠ能抑制PC3细胞的体外增殖,与对照组相比,PC3细胞从给药浓度0.4μmol/L(0.85±0.05 vs 1.00±0.00,P<0.01)开始明显下降,且呈时间和剂量依赖性。流式细胞术检测显示,PPⅠ能明显诱导PC3细胞早期凋亡,与对照组相比,PC3细胞早期凋亡率从给药浓度0.8μmol/L(13.83±2.97 vs 4.83±0.95)开始明显增加(P均<0.01),且呈剂量依赖性;Western印迹显示,PPⅠ以时间依赖性上调p-ERK1/2和ERK1/2蛋白的激活与表达,与对照组相比,给药时间从2 h(1.73±0.17 vs 1.00±0.00,P<0.01)开始,p-ERK1/2明显被激活表达,PPⅠ以剂量依赖性下调NF-κB/p65、DNMT1蛋白的表达,与对照组相比,给药浓度分别从1.6μmol/L(0.67±0.11 vs 1.00,P<0.01)与1.2μmol/L(0.63±0.06 vs 1.00±0.00,P<0.01)开始,NF-κB/p65、DNMT1表达明显下调;抑制ERK1/2磷酸化能逆转重楼皂苷Ⅰ对NF-κB/p65蛋白表达的下调,与PPⅠ组相比,PD98059+PPⅠ组NF-κB/p65蛋白表达(0.86±0.18 vs 0.43±0.09,P<0.05)明显上调。结论:PPⅠ可能通过介导ERK1/2通路抑制NF-κB/p65和DNMT1蛋白表达,诱导PC3细胞早期凋亡,进而抑制细胞增殖。
