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目的:探讨子痫前期(PE)血清激活血管平滑肌细胞蛋白激酶C-α(PKC-α)及前胶原Ⅰ表达是否由磷脂酶C-γ1(PLC-γ1)信息分子介导。方法:原代培养脐静脉内皮细胞(HUVEC)与脐动脉平滑肌细胞(HUASMC),体外建立HUVEC与HUASMC共培养体系,用PE血清处理共培养HUASMC细胞。MTT法检测HUASMC细胞活力;Annexin VFITC-流式细胞仪检测HUASMC凋亡;Western blot法检测PLC-γ1(磷酸化)、PKC-α(包膜/胞浆比值)活性以及前胶原Ⅰ表达。siRNA沉默PLC-γ1表达后,观察PE血清对共培养HUVEC细胞PKC-α活性及前胶原Ⅰ表达的影响。结果:PE血清可促进共培养HUASMC细胞PLC-γ1磷酸化、PKC-α膜转位及前胶原Ⅰ的表达(P<0.05);siRNA沉默HUASMC细胞PLC-γ1蛋白表达可逆转PE血清引起的PKC-α膜转位及前胶原Ⅰ的表达(P<0.05)。结论:PE血清通过PLC-γ1-PKC-α信息途径促进HUASMC前胶原Ⅰ的表达,可能在胎盘血管病理过程中起着重要的作用。
Aims: To investigate whether protein kinase C-α (PKC-α) and procollagen Ⅰ expression in serum-activated vascular smooth muscle cells of preeclampsia (PE) is mediated by phospholipase C-γ1 (PLC-γ1) signaling molecule. Methods: Human umbilical vein endothelial cells (HUVEC) and umbilical artery smooth muscle cells (HUASMC) were cultured in primary culture. HUVEC and HUASMC co-culture system was established in vitro. HUASMC cells were co-cultured with PE serum. The viability of HUASMC was detected by MTT assay. The apoptosis of HUASMC was detected by Annexin VFITC-flow cytometry. The activity of PLC-γ1 (phosphorylation), PKC-α (envelope / cytoplasm) and the expression of procollagen Ⅰ were detected by Western blot. After silencing PLC-γ1 by siRNA, the effect of PE serum on PKC-α activity and procollagen Ⅰ expression in co-cultured HUVECs was observed. Results: PE serum could promote PLC-γ1 phosphorylation and PKC-α membrane translocation and procollagen Ⅰ expression in HUASMC cells co-cultured with siRNA (P <0.05). SiRNA silencing PLC-γ1 protein in HUASMC cells could reverse the PKC -α membrane translocation and procollagen Ⅰ expression (P <0.05). Conclusion: PE serum promotes the expression of procollagen Ⅰ in HUASMC through the PLC-γ1-PKC-α signaling pathway, which may play an important role in the pathological process of placental vascularization.