Zheng Linmei, Tang Rong, Shi Lei, Zhou Zhongyi, Song Jie, Lu Zhicheng
Department of Obstetrics, Hainan General Hospital, Hainan Affiliated Hospital of Hainan Medical University, HuaXiu Road 19th, Haikou, 570311, Hainan, China.
Department of Hepatobiliary Surgery, Hainan General Hospital, Hainan Affiliated Hospital of Hainan Medical University, Haikou, 570311, China.
In Vitro Cell Dev Biol Anim. 2023 Feb;59(2):131-141. doi: 10.1007/s11626-023-00751-9. Epub 2023 Feb 27.
The maintenance of migration of trophoblast cells is beneficial to pregnancy, and its weakening can lead to preeclampsia (PE). CD142 is considered as a classical motility-promoting factor. Our research aimed to explore the role of CD142 in trophoblast cell migration and potential mechanism. Through fluorescence-activated cell sorting (FACS) and gene transduction assays, CD142 expression levels of mouse trophoblast cell lines were upregulated and downregulated respectively. Then, the migratory level was detected through Transwell assays in different groups of trophoblast cells. The corresponding chemokines were screened by ELISA in different sorted trophoblast cells. Based on gene overexpression and knockdown assays, the production mode of identified valuable chemokine was analyzed by detecting gene and protein expression in trophoblast cells. Finally, the contribution of autophagy response to specific chemokine regulated by CD142 was explored by combining different groups of cells and autophagy regulators. Our results showed that both CD142 positive sorting and CD142 overexpression promoted the migratory ability of trophoblast cells, and trophoblast cells with the highest level of CD142 had the strongest migratory ability. In addition, CD142 cells contained the highest level of IL-8. Consistently, CD142 overexpression promoted IL-8 protein expression in trophoblast cells while CD142 silencing was contrary. However, both CD142 overexpression and CD142 silencing did not affect IL-8 mRNA expression. Moreover, both CD142 and CD142-overexpressed cells showed higher BCL2 protein expression and poorer autophagic activity. Importantly, autophagy activation with TAT-Beclin1 recovered the increased IL-8 protein expression in CD142 cells. Obviously, the migratory ability of CD142 cells inhibited by TAT-Beclin1 was recovered by the addition of IL-8 recombinant factor. In conclusion, CD142 inhibits the degradation of IL-8 through the inhibition of BCL2-Beclin1-autophagy signal transduction, thereby promoting the migration of trophoblast cells.
滋养层细胞迁移的维持有利于妊娠，其减弱会导致先兆子痫（PE）。CD142被认为是一种经典的促进迁移因子。我们的研究旨在探讨CD142在滋养层细胞迁移中的作用及潜在机制。通过荧光激活细胞分选（FACS）和基因转导实验，分别上调和下调小鼠滋养层细胞系的CD142表达水平。然后，通过Transwell实验检测不同组滋养层细胞的迁移水平。在不同分选的滋养层细胞中通过ELISA筛选相应的趋化因子。基于基因过表达和敲低实验，通过检测滋养层细胞中的基因和蛋白表达来分析已鉴定的有价值趋化因子的产生模式。最后，通过结合不同组细胞和自噬调节剂来探讨自噬反应对CD142调节的特定趋化因子的贡献。我们的结果表明，CD142阳性分选和CD142过表达均促进了滋养层细胞的迁移能力，且CD142水平最高的滋养层细胞具有最强的迁移能力。此外，CD142细胞中IL-8水平最高。一致地，CD142过表达促进滋养层细胞中IL-8蛋白表达，而CD142沉默则相反。然而，CD142过表达和CD142沉默均不影响IL-8 mRNA表达。而且，CD142和CD142过表达细胞均显示出较高的BCL2蛋白表达和较差的自噬活性。重要的是，用TAT-Beclin1激活自噬可恢复CD142细胞中增加的IL-8蛋白表达。显然，添加IL-8重组因子可恢复被TAT-Beclin1抑制的CD142细胞的迁移能力。总之，CD142通过抑制BCL2-Beclin1-自噬信号转导来抑制IL-8的降解，从而促进滋养层细胞的迁移。
