Key Laboratory of Birth Defects and Related Diseases of Women and Children (Sichuan University), Ministry of Education, West China Second University Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R. China.
Int J Mol Med. 2019 Jul;44(1):281-290. doi: 10.3892/ijmm.2019.4175. Epub 2019 Apr 25.
Placental hypoxia serves a role in the early stages of normal pregnancy and is involved in the pathophysiology of preeclampsia. Previously, it was suggested that p57kinase inhibitory protein (KIP)2 regulates the cell cycle during embryogenesis and apoptosis. Recent evidence has indicated that p57KIP2 is increased in preeclamptic placentas and absence of p57KIP2 induces preeclampsia‑type symptoms in rats. However, effects of p57KIP2 on apoptosis under hypoxic conditions remain to be elucidated. In the present study, HTR‑8/SVneo trophoblasts were cultured under hypoxic conditions (2% O2). Knockdown using small interfering (si)RNA and overexpression of p57KIP2 were utilized to explore the biological function of p57KIP2 in apoptosis and cell function in vitro. Furthermore, expression of p57KIP2 and apoptosis were evaluated by western blotting, flow cytometry and TUNEL assays, and the response of trophoblasts to hypoxia and the role of p57KIP2 in trophoblast migration and invasion was assessed. The role of p57KIP2 in the JNK signaling pathway in HTR‑8/SVneo trophoblasts was further studies. In vitro, protein expression of p57KIP2 was increased in HTR‑8/SVneo cells exposed to 2% O2. Exogenous p57KIP2 overexpression significantly decreased the expression of pro‑apoptosis proteins, including p53, Bax and cleaved caspase3, under hypoxic conditions for 24 h. In addition, knockdown of p57KIP2 increased the response to apoptosis following hypoxia for 24 h. The present study revealed that overexpression of p57KIP2 decreased the levels of phosphorylated‑JNK. JNK inhibitor treatment combined with the overexpression of p57KIP2 significantly decreased the levels of apoptosis and increased cell invasion and migration. Taken together, p57KIP2 knockdown significantly increased apoptosis in HTR‑8/SVneo cells exposed to 2% O2, whereas overexpression of p57KIP2 had opposite effects, mediated by the JNK/stress activated protein kinase (SAPK) signaling pathway. The results indicated that hypoxia‑induced expression of p57KIP2 promoted trophoblast migration and invasion by mediating the JNK/SAPK signaling pathway, which is crucial during placentation. These results may provide a novel molecular mechanism to understand the involvement of p57KIP2 in the pathogenesis of preeclampsia.
胎盘缺氧在正常妊娠的早期阶段发挥作用，并参与子痫前期的病理生理学。先前有研究表明，p57 激酶抑制蛋白（KIP）2 在胚胎发生和细胞凋亡过程中调节细胞周期。最近的证据表明，p57KIP2 在子痫前期胎盘组织中增加，p57KIP2 的缺失会在大鼠中诱导子痫前期样症状。然而，p57KIP2 在缺氧条件下对细胞凋亡的影响仍有待阐明。在本研究中，将 HTR-8/SVneo 滋养层细胞在缺氧条件下（2%O2）培养。使用小干扰（si）RNA 敲低和 p57KIP2 的过表达来探索 p57KIP2 在体外细胞凋亡和细胞功能中的生物学功能。此外，通过蛋白质印迹、流式细胞术和 TUNEL 分析评估 p57KIP2 的表达和细胞凋亡，评估滋养层对缺氧的反应以及 p57KIP2 在滋养层迁移和侵袭中的作用。进一步研究了 p57KIP2 在 HTR-8/SVneo 滋养层细胞中 JNK 信号通路中的作用。在体外，暴露于 2%O2 的 HTR-8/SVneo 细胞中 p57KIP2 的蛋白表达增加。在缺氧 24 小时的情况下，外源性过表达 p57KIP2 显著降低了促凋亡蛋白，包括 p53、Bax 和 cleaved caspase3 的表达。此外，p57KIP2 敲低增加了缺氧 24 小时后的凋亡反应。本研究表明，p57KIP2 的过表达降低了磷酸化-JNK 的水平。JNK 抑制剂处理联合 p57KIP2 的过表达显著降低了凋亡水平，增加了细胞侵袭和迁移。综上所述，p57KIP2 敲低显著增加了暴露于 2%O2 的 HTR-8/SVneo 细胞中的细胞凋亡，而过表达 p57KIP2 则通过 JNK/应激激活蛋白激酶（SAPK）信号通路产生相反的作用。结果表明，缺氧诱导的 p57KIP2 表达通过调节 JNK/SAPK 信号通路促进滋养层迁移和侵袭，这在胎盘形成过程中至关重要。这些结果可能为理解 p57KIP2 参与子痫前期发病机制提供新的分子机制。
