摘要
目的探讨FOXO3a-Bim信号通路在雷公藤内酯醇诱导膀胱癌T24细胞细胞凋亡的作用。方法培养膀胱癌T24细胞，将实验分为空白对照组，雷公藤内酯醇组（50nmo/L），MK2206组（50nmo/L雷公藤内酯醇＋5μmol／LMK2206），FOXO3a-siRNA组（50nmo/L雷公藤内酯醇＋100nmo/LFOXO3a-siRNA）和Bim-siRNA组（50nmol/L雷公藤内酯醇＋100nmol/L Bim-siRNA），用噻唑蓝（MTF）法检测124细胞生长抑制率，Annexin V／Propidiumiodide（PI）双染法检测细胞凋亡，Western印迹方法检测p-Akt、Akt、p-FOXO3a、FOXO3a、Bim、Bax、cleaved-caspase3表达。结果25、50、100、250nmo/L雷公藤内酯醇处理24h（17％±9％、24％±5％、43％±8％、61％±8％）、48h（20％±7％、34％±6％、56％±7％、74％±5％）和72h（32％±8％、41％±7％、69％±7％、84％±3％）后细胞生长抑制率均明显高于对照组（P〈0．05）。24、48、72h的Ic。分别为（113±10）、（91±8）、（68±5）nmo/L。24h（10％±4％、15％±5％、29％±8％、46％±8％）、48h（16％±5％、24％±6％、40％±7％、55％±9％）和72h（27％±4％、38％±5％、50％±9％、65％±8％）后细胞凋亡率均明显高于对照组（均P〈0．05）。25、50、100nmo/L雷公藤内酯醇诱导细胞凋亡、降低p-Akt、P-FOXO3a表达同时增高Bim、Bax、cleaved-caspase3表达。FOXO3a-siRNA及Bim-siRNA降低了雷公藤内酯醇诱导Bim、Bax、cleaved-caspase3高表达。雷公藤内酯醇组细胞抑制率（30％±8％）明显高于空白对照组（P〈0．05）。MK2206组（54％±6％）、FOXO3a-siRNA组（18％±7％）和Bim-siRNA组（11％±6％）细胞抑制率亦高于对照组，但与雷公藤内酯醇组相比，MK2206组细胞抑制率明显增高，FOXO3a-siRNA组和Bim．siRNA组细胞抑制率明显降低（P〈0．05）。结论雷公藤内酯醇通过激活FOXO3a-Bim信号通路诱导细胞凋亡，抑制124细胞生长。
Objective To investigate the role of FOXO3a-Bim signaling in triptolide induced bladder cancer T24 cells apoptosis. Methods T24 cells were used and divided into control group, triptolide group （50 nmol/L）, MK2206 group （ 50 nmol/L triptolide ＋ 5 μmol/L MK2206 ）, FOXO3a-siRNA group （50 nmol/L triptolide ＋ 100 nmol/L FOXO3a-siRNA）, Bim-siRNA group （50 nmol/L triptolide ＋ 100 nmol/L Bim-siRNA）. MTT assay was used to analyze the cells growth inhibition. Annexin V/PI staining was implemented to detect cell apoptosis rate, the expression of p-Akt, Akt, p-FOXO3a, FOXO3a, Bim, Bax. Cleaved-caspase 3 was analyzed by Western blot. Results After treatment with triptolide 25,50,100, 250 nmol/L, the cell growth inhibition rates at 24 hours（ 17% ± 9%, 24% ± 5% ,43% ± 8% ,61% ± 8% ）, 48 hours （20% ±7% ,34% ±6% ,56% ±7% ,74% ±5% ） and72 hours（32% ±8% ,41% ±7% ,69% ± 7% ,84% ± 3% ） were significantly higher than control group respectively. The ICs0 at 24, 48, 72 hourswere （ 113 ± 10）, （91 ± 8 ）, （68 ± 5 ） nmol/L ; the cell apoptosis rates at 24 hours （ 10% ± 4%, 15 % ± 5 %, 29% ±8%,46% ±8%）, 48 hours （16% ±5%,24% ±6%,40% ±7%,55% ±9%） and 72 hours （27% ±4% ,38% ± 5% ,50% ± 9% ,65% ± 8% ） were significantly increased （P 〈 0. 05）. Western blot showed that triptolide reduced the expression of p-Akt, p-FOXO3a and increased the expression of Bim, Bax, cleaved-caspase 3. The cell inhibition rate in Triptolide group （30% ± 8% ） was significantly higher than that in the control group （P 〈0. 05） and the rates in MK2206 group （54% ±6% ）, FOXO3a-siRNA group （ 18% ± 7% ） and Bim-siRNA group （ 11% ± 6% ） were also higher than the control group. Compared with the triptolide group, the inhibition rate in MK2206 group was significantly increased, but decreased in FOXO3a-siRNA group and Bim-siRNA group （P 〈 0.05 ）. Conclusion Triptolide induces 324 ceils apoptosis through FOXO3a-Bim signaling pathway.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2017年第15期1187-1190,共4页
National Medical Journal of China
基金
国家自然科学基金（81573657）
