目的绒毛外滋养细胞焦亡是否参与子痫前期的发生发展值得探讨。文章旨在构建H 2 O 2诱导滋养细胞焦亡模型,探讨绒毛外滋养细胞焦亡是否参与子痫前期的发生发展,为探究子痫前期发病机制提供新的方向。方法人滋养细胞HTR-8/SVneo培养于164...目的绒毛外滋养细胞焦亡是否参与子痫前期的发生发展值得探讨。文章旨在构建H 2 O 2诱导滋养细胞焦亡模型,探讨绒毛外滋养细胞焦亡是否参与子痫前期的发生发展,为探究子痫前期发病机制提供新的方向。方法人滋养细胞HTR-8/SVneo培养于1640+10%胎牛血清+1%抗生素中,培养12 h后,给予H 2 O 2(100、150、200、250μmol/L)处理细胞(2、4、6、12 h),对照为1640+10%胎牛血清+1%抗生素正常培养细胞。提取细胞总蛋白,Western blot检测焦亡相关分子蛋白水平的表达;RT-qPCR检测细胞中焦亡相关分子mRNA水平;倒置相差显微镜观察细胞形态学改变。结果当H 2 O 2为150μmol/L,处理人滋养细胞4 h时,焦亡相关分子NLRP3、caspase1、Cleaved caspase1、GSDMD及IL-1β的蛋白水平明显高于对照,随着作用时间的延长及浓度的增加,蛋白质表达水平被抑制;当H 2 O 2为150μmol/L,作用2 h时,焦亡经典通路中上游“启动”信号关键分子NLRP3及IL-1β的mRNA水平较对照明显升高(P<0.000);4 h时,焦亡经典通路中的关键分子GSDMD的mRNA水平及下游炎症因子IL-18的mRNA水平明显高于对照组(P<0.05)。通过焦亡相关分子mRNA水平的逆向验证,H 2 O 2诱导滋养细胞焦亡模型最佳条件为150μmol/L和4 h,在该条件下,光镜下可明显看到细胞肿胀、碎裂及质膜气泡形成等细胞焦亡典型改变。结论体外成功建立了氧化应激反应诱导滋养细胞焦亡模型,模拟了氧化应激反应诱导滋养细胞发生焦亡损伤的病生理过程,为后续子痫前期发病机制的研究提供实验基础。展开更多
Objective: To investigate the effect of klotho on invasion of Extrachorionic trophoblastic cell line HTR-8/SVneo cells. Methods: Klotho protein expression in placenta of preeclampsia and normal pregnant women was dete...Objective: To investigate the effect of klotho on invasion of Extrachorionic trophoblastic cell line HTR-8/SVneo cells. Methods: Klotho protein expression in placenta of preeclampsia and normal pregnant women was detected by immunohistochemistry. Experimental grouping:Normal control group, Klotho recombinant protein group, klotho SiRNA group. The abilities of cell were measured by Transwell assay. The protein expression were detected by Western blot. Results: The expression of klotho in placenta of preeclampsia was significantly lower than that in normal placenta. Treatment with Klotho recombinant protein improved the HTR-8/SVneo cell invasion abilities, and upregulated the expression of MMP-2 and MMP-9. The effects were re-versed by treatment with Klotho siRNA. Conclusion: Klotho regulate the invasion of trophoblast may via MMP-2 and MMP-9 expression. It provides a new clue for the study of the pathogenesis of preeclampsia.展开更多
OBJECTIVE:To elucidate the regulatory effects of salvianolic acid B(Sal B)on trophoblast cells in preeclampsia(PE).METHODS:3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide(MTT)assays were used to detect th...OBJECTIVE:To elucidate the regulatory effects of salvianolic acid B(Sal B)on trophoblast cells in preeclampsia(PE).METHODS:3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide(MTT)assays were used to detect the viability of human extravillous trophoblast HTR-8/Svneo cells induced by H_(2)O_(2)following treatment with different concentrations of Sal B.The levels of oxidative stressrelated molecules,including superoxide dismutase,glutathione-Px and malondialdehyde were detected using corresponding kits.Cell apoptosis was detected using a Terminal deoxynucleotidyl transferase(Td T)d UTP NickEnd Labeling(TUNEL)assay,and the expression of apoptosis-related proteins was detected using western blot analysis.In the present study,wound healing and Transwell assays were performed to measure the levels of cell invasion and migration.Western blot analysis was also used to detect the expression levels of epithelialmesenchymal transition-related proteins.The mechanisms underlying Sal B were further investigated using reverse transcription-quantitative real-time polymerase chain reaction(RT-q PCR)and western blot analysis,to determine the expression levels of matrix metallopeptidase 9(MMP-9)and phosphatidylinositol-4,5-bisphosphate 3-kinase(PI3K)/protein kinase B(Akt).RESULTS:Sal B increased the activity of HTR-8/Svneo cells,inhibited oxidative damage and promoted the invasion and migration of trophoblast cells induced by H_(2)O_(2).Furthermore,the expression levels of MMP-9 and members of the PI3K/Akt signaling pathway were significantly decreased.The pathway agonist,LY294002,and MMP-9 inhibitor,GM6001,reversed the effects of Sal B on H_(2)O_(2)-induced cells.CONCLUSIONS:Sal B promoted the invasion and migration of H_(2)O_(2)-induced HTR-8/Svneo trophoblast cells by upregulating MMP-9 via the PI3K/Akt signaling pathway.展开更多
文摘目的绒毛外滋养细胞焦亡是否参与子痫前期的发生发展值得探讨。文章旨在构建H 2 O 2诱导滋养细胞焦亡模型,探讨绒毛外滋养细胞焦亡是否参与子痫前期的发生发展,为探究子痫前期发病机制提供新的方向。方法人滋养细胞HTR-8/SVneo培养于1640+10%胎牛血清+1%抗生素中,培养12 h后,给予H 2 O 2(100、150、200、250μmol/L)处理细胞(2、4、6、12 h),对照为1640+10%胎牛血清+1%抗生素正常培养细胞。提取细胞总蛋白,Western blot检测焦亡相关分子蛋白水平的表达;RT-qPCR检测细胞中焦亡相关分子mRNA水平;倒置相差显微镜观察细胞形态学改变。结果当H 2 O 2为150μmol/L,处理人滋养细胞4 h时,焦亡相关分子NLRP3、caspase1、Cleaved caspase1、GSDMD及IL-1β的蛋白水平明显高于对照,随着作用时间的延长及浓度的增加,蛋白质表达水平被抑制;当H 2 O 2为150μmol/L,作用2 h时,焦亡经典通路中上游“启动”信号关键分子NLRP3及IL-1β的mRNA水平较对照明显升高(P<0.000);4 h时,焦亡经典通路中的关键分子GSDMD的mRNA水平及下游炎症因子IL-18的mRNA水平明显高于对照组(P<0.05)。通过焦亡相关分子mRNA水平的逆向验证,H 2 O 2诱导滋养细胞焦亡模型最佳条件为150μmol/L和4 h,在该条件下,光镜下可明显看到细胞肿胀、碎裂及质膜气泡形成等细胞焦亡典型改变。结论体外成功建立了氧化应激反应诱导滋养细胞焦亡模型,模拟了氧化应激反应诱导滋养细胞发生焦亡损伤的病生理过程,为后续子痫前期发病机制的研究提供实验基础。
文摘Objective: To investigate the effect of klotho on invasion of Extrachorionic trophoblastic cell line HTR-8/SVneo cells. Methods: Klotho protein expression in placenta of preeclampsia and normal pregnant women was detected by immunohistochemistry. Experimental grouping:Normal control group, Klotho recombinant protein group, klotho SiRNA group. The abilities of cell were measured by Transwell assay. The protein expression were detected by Western blot. Results: The expression of klotho in placenta of preeclampsia was significantly lower than that in normal placenta. Treatment with Klotho recombinant protein improved the HTR-8/SVneo cell invasion abilities, and upregulated the expression of MMP-2 and MMP-9. The effects were re-versed by treatment with Klotho siRNA. Conclusion: Klotho regulate the invasion of trophoblast may via MMP-2 and MMP-9 expression. It provides a new clue for the study of the pathogenesis of preeclampsia.
文摘OBJECTIVE:To elucidate the regulatory effects of salvianolic acid B(Sal B)on trophoblast cells in preeclampsia(PE).METHODS:3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide(MTT)assays were used to detect the viability of human extravillous trophoblast HTR-8/Svneo cells induced by H_(2)O_(2)following treatment with different concentrations of Sal B.The levels of oxidative stressrelated molecules,including superoxide dismutase,glutathione-Px and malondialdehyde were detected using corresponding kits.Cell apoptosis was detected using a Terminal deoxynucleotidyl transferase(Td T)d UTP NickEnd Labeling(TUNEL)assay,and the expression of apoptosis-related proteins was detected using western blot analysis.In the present study,wound healing and Transwell assays were performed to measure the levels of cell invasion and migration.Western blot analysis was also used to detect the expression levels of epithelialmesenchymal transition-related proteins.The mechanisms underlying Sal B were further investigated using reverse transcription-quantitative real-time polymerase chain reaction(RT-q PCR)and western blot analysis,to determine the expression levels of matrix metallopeptidase 9(MMP-9)and phosphatidylinositol-4,5-bisphosphate 3-kinase(PI3K)/protein kinase B(Akt).RESULTS:Sal B increased the activity of HTR-8/Svneo cells,inhibited oxidative damage and promoted the invasion and migration of trophoblast cells induced by H_(2)O_(2).Furthermore,the expression levels of MMP-9 and members of the PI3K/Akt signaling pathway were significantly decreased.The pathway agonist,LY294002,and MMP-9 inhibitor,GM6001,reversed the effects of Sal B on H_(2)O_(2)-induced cells.CONCLUSIONS:Sal B promoted the invasion and migration of H_(2)O_(2)-induced HTR-8/Svneo trophoblast cells by upregulating MMP-9 via the PI3K/Akt signaling pathway.
