目的探索Jurkat T细胞中胞外调节蛋白激酶(extracellular regulated protein kinases,ERK)活性动力学以及基质刚度对ERK活性的影响。方法利用荧光共振能量转移(fluorescence resonance energy transfer,FRET)技术实时观测Jurkat细胞中ER...目的探索Jurkat T细胞中胞外调节蛋白激酶(extracellular regulated protein kinases,ERK)活性动力学以及基质刚度对ERK活性的影响。方法利用荧光共振能量转移(fluorescence resonance energy transfer,FRET)技术实时观测Jurkat细胞中ERK活性的变化,或细胞处于I型胶原基质胶中检测其影响。结果部分Jurkat细胞中存在ERK活性脉冲现象,频率约为3次/h,FRET振幅变化约为20%。在抗体激活T细胞抗原受体(T-cell receptor,TCR)的条件下,ERK脉冲依然存在,频率和振幅无显著变化。当细胞处于I型胶原水凝胶中,随着胶基质刚度增加,脉冲频率有所下调。结论Jurkat T细胞中存在自发的ERK活性脉冲现象,初步实验显示其频率受基质刚度影响。而该信号波动的生理意义和分子机制仍有待探索。展开更多
Objective: This study was designed to explore whether inhibition of the extracellular-regulated kinase (ERK) and phosphatidylinositol-3-kinase (PI3K) signaling pathways can inhibit the growth of xenografts of endometr...Objective: This study was designed to explore whether inhibition of the extracellular-regulated kinase (ERK) and phosphatidylinositol-3-kinase (PI3K) signaling pathways can inhibit the growth of xenografts of endometrial cancer cell lines with different estrogen receptors (ER) profiles in vivo and to provide preliminary laboratory basis for the probability of endometrial adenocarcinoma treatment with blockage of the two pathways, especially to endometrial cancer with low ER status. Methods: Human endometrial cancer Ishikawa bearing ER and HEC-1Awith low ER status cells were subcutaneously injected into BALB/c nude mice to establish endometrial cancer xenograft tumor models. The effects of PI3K/Akt inhibitor LY294002, MAPK/ERK1/2 inhibitor PD-98059 and their combinations on the growth of the xenograft tumors and apoptotic state of Ishikawa and HEC-1Acells were tested in vivo using the inhibitory rate, the terminal deoxynucleotidyl transferase-mediated nick-end labeling assay, H/E-stain. Western blot analysis was used to detect the alterations of activated ERK (P-ERK) and AKT (P-AKT) during this process. Results: LY294002, a PI3K/Akt pathway inhibitor, induced significant suppression in the growth of both Ishikawa and HEC-1Acell xenograft tumors, concomitant with increased apoptosis in xenografts as evidenced by TUNEL. A similar effect was also observed when the MAPK/ERK1/2 signaling pathway was inhibited by PD98059. Concurrent inhibition of the PI3K/Akt and MAPK/ERK1/2 pathways showed enhanced anti-tumor effects in vivo as indicated by increased apoptosis. At the same time, the levels of P-ERK and P-AKT in both xenograft tumors decreased, and their levels in combination group was the lowest. Conclusions: PD98059, LY294002 and their combinations showed remarkable inhibitory effects on xenograft tumors of endometrial carcinoma cell lines with different expression status of ER in vivo through blockage of PI3K/Akt and MAPK/ERK1/2 signaling pathways. This suggests that targeting these pathways may be an effective therapeutic strategy against endometrial carcinomas, especially for ER-negative cancers which show poor response to endocrinal therapy.展开更多
目的：探讨细胞外信号调节蛋白激酶（ERK）通路在慢性哮喘大鼠气道平滑肌细胞（ASMC）凋亡中的作用及其机制。方法：30只Wistar大鼠分为正常对照组（A组）和慢性哮喘组（B组），用原位末端标记法和Annexin—V FIT CPI双染色法观察ASMC...目的：探讨细胞外信号调节蛋白激酶（ERK）通路在慢性哮喘大鼠气道平滑肌细胞（ASMC）凋亡中的作用及其机制。方法：30只Wistar大鼠分为正常对照组（A组）和慢性哮喘组（B组），用原位末端标记法和Annexin—V FIT CPI双染色法观察ASMC凋亡，免疫组织化学法检测bcl-2和bax的表达，Western blot检测caspase-3蛋白的表达，并用ERK激动剂表皮生长因子（EGF）和抑制剂PD98059干预两组ASMC，观察上述指标的变化。结果：与正常对照组ASMC比较，慢性哮喘组ASMC凋亡指数、早期凋亡细胞百分率明显下降。经PD98059干预之后，慢性哮喘组ASMC的凋亡指数与早期凋亡细胞百分率、bax蛋白表达量和caspase-3蛋白含量明显增高，bcl-2蛋白表达量明显降低。经EGF干预之后，慢性哮喘组ASMC凋亡指数与早期凋亡细胞百分率进一步下降，而这一作用可以被PD98059所抑制。结论：慢性哮喘组大鼠ASMC内源性增殖活性增加的同时，伴有凋亡活性下降。ERK1／2参与慢性哮喘ASMC凋亡调控，其机制与bcl-2家族和caspace-3有关。展开更多
文摘目的探索Jurkat T细胞中胞外调节蛋白激酶(extracellular regulated protein kinases,ERK)活性动力学以及基质刚度对ERK活性的影响。方法利用荧光共振能量转移(fluorescence resonance energy transfer,FRET)技术实时观测Jurkat细胞中ERK活性的变化,或细胞处于I型胶原基质胶中检测其影响。结果部分Jurkat细胞中存在ERK活性脉冲现象,频率约为3次/h,FRET振幅变化约为20%。在抗体激活T细胞抗原受体(T-cell receptor,TCR)的条件下,ERK脉冲依然存在,频率和振幅无显著变化。当细胞处于I型胶原水凝胶中,随着胶基质刚度增加,脉冲频率有所下调。结论Jurkat T细胞中存在自发的ERK活性脉冲现象,初步实验显示其频率受基质刚度影响。而该信号波动的生理意义和分子机制仍有待探索。
文摘Objective: This study was designed to explore whether inhibition of the extracellular-regulated kinase (ERK) and phosphatidylinositol-3-kinase (PI3K) signaling pathways can inhibit the growth of xenografts of endometrial cancer cell lines with different estrogen receptors (ER) profiles in vivo and to provide preliminary laboratory basis for the probability of endometrial adenocarcinoma treatment with blockage of the two pathways, especially to endometrial cancer with low ER status. Methods: Human endometrial cancer Ishikawa bearing ER and HEC-1Awith low ER status cells were subcutaneously injected into BALB/c nude mice to establish endometrial cancer xenograft tumor models. The effects of PI3K/Akt inhibitor LY294002, MAPK/ERK1/2 inhibitor PD-98059 and their combinations on the growth of the xenograft tumors and apoptotic state of Ishikawa and HEC-1Acells were tested in vivo using the inhibitory rate, the terminal deoxynucleotidyl transferase-mediated nick-end labeling assay, H/E-stain. Western blot analysis was used to detect the alterations of activated ERK (P-ERK) and AKT (P-AKT) during this process. Results: LY294002, a PI3K/Akt pathway inhibitor, induced significant suppression in the growth of both Ishikawa and HEC-1Acell xenograft tumors, concomitant with increased apoptosis in xenografts as evidenced by TUNEL. A similar effect was also observed when the MAPK/ERK1/2 signaling pathway was inhibited by PD98059. Concurrent inhibition of the PI3K/Akt and MAPK/ERK1/2 pathways showed enhanced anti-tumor effects in vivo as indicated by increased apoptosis. At the same time, the levels of P-ERK and P-AKT in both xenograft tumors decreased, and their levels in combination group was the lowest. Conclusions: PD98059, LY294002 and their combinations showed remarkable inhibitory effects on xenograft tumors of endometrial carcinoma cell lines with different expression status of ER in vivo through blockage of PI3K/Akt and MAPK/ERK1/2 signaling pathways. This suggests that targeting these pathways may be an effective therapeutic strategy against endometrial carcinomas, especially for ER-negative cancers which show poor response to endocrinal therapy.
文摘目的：探讨细胞外信号调节蛋白激酶（ERK）通路在慢性哮喘大鼠气道平滑肌细胞（ASMC）凋亡中的作用及其机制。方法：30只Wistar大鼠分为正常对照组（A组）和慢性哮喘组（B组），用原位末端标记法和Annexin—V FIT CPI双染色法观察ASMC凋亡，免疫组织化学法检测bcl-2和bax的表达，Western blot检测caspase-3蛋白的表达，并用ERK激动剂表皮生长因子（EGF）和抑制剂PD98059干预两组ASMC，观察上述指标的变化。结果：与正常对照组ASMC比较，慢性哮喘组ASMC凋亡指数、早期凋亡细胞百分率明显下降。经PD98059干预之后，慢性哮喘组ASMC的凋亡指数与早期凋亡细胞百分率、bax蛋白表达量和caspase-3蛋白含量明显增高，bcl-2蛋白表达量明显降低。经EGF干预之后，慢性哮喘组ASMC凋亡指数与早期凋亡细胞百分率进一步下降，而这一作用可以被PD98059所抑制。结论：慢性哮喘组大鼠ASMC内源性增殖活性增加的同时，伴有凋亡活性下降。ERK1／2参与慢性哮喘ASMC凋亡调控，其机制与bcl-2家族和caspace-3有关。
