PTBP1促进Circ_0029692的生成调节Twist 1形成反馈环路调控子痫前期滋养细胞EMT的机制

The mechanism of decreased invasiveness in preeclampsia trophoblast cells is not clear. Our team found RNA-binding protein PTBP1 exhibited low expression in PE placenta tissue, and its expression level was positively correlated with the expression of Vimentin and N-cadherin; Chip detection results proved the down-regulated expression of circ_0029692; Bioinformatics Software predicted there were binding sites between PTBP1 and both end flanking sequences of circ_0029692. Whether PTBP1 promotes the generation of circ_0029692, whether circ_0029692 combines with miR-135b-5p and miR-372-3p thus altering regulation of its target genes, and whether Twist1 facilitates the transcriptional regulation on PTBP1 and form a feedback loop, consequently regulating epithelial to mesenchymal transition remained to be expounded. This project explicitly showed that PTBP1 could promote the generation of circ_0029692; Circ_0029692 could combine with miR-372-3p and miR-135b-5p thus altering its negative regulation on TWIST1, N-cadherin and Vimentin; Twist1 could facilitate the transcriptional regulation on PTBP1, forming feedback loop and controling EMT of PE trophoblast cells. The project will provide a new approach for further research on the pathogenesis of PE.

子痫前期滋养细胞侵袭力降低的机制不清。课题组发现RNA结合蛋白PTBP1在PE胎盘组织中低表达，与N-cadherin和Vimentin表达正相关；芯片检测发现circ_00296922表达下调；软件预测到circ_0029692 两端侧翼序列存在PTBP1 的结合位点。PTBP1是否能促进circ_0029692生成；circ_0029692能否结合miR-372-3p和miR-135b-5p，改变其对靶基因的调控；Twist1能否转录调控PTBP1形成反馈环路，调控EMT？有待阐明。项目拟明确PTBP1促进circ_0029692生成；circ_0029692结合miR-372-3p和miR-135b-5p，改变其对Twist1、N-cadherin和Vimentin的负性调控；Twist1转录调控PTBP1形成反馈环路，共同调控EMT的机制。为深入研究PE的发病机制提供新思路。

子痫前期为妊娠期特有疾病，全球死亡率高达3-8%，其发病率、死亡率较高，严重威胁母、儿生命安全。滋养细胞侵袭能力不足，胎盘螺旋动脉重塑失败为子痫前期主要发病机制。环状RNA作为一类非编码RNA，其稳定性高，特异性强，适合作为疾病诊断的预测分子。环状RNA在疾病发生发展的病理生理过程中，发挥着重要的作用，有潜力作为一种新的临床治疗方案。. 本项目以环状RNA通过调节滋养细胞侵袭能力参与子痫前期的发生发展为切入点展开研究。发现一个新的环状RNA（hsa_circ_0029692），其在子痫前期患者胎盘中表达水平显著下调，体外实验发现明显促进滋养细胞EMT、迁移和侵袭能力。体内外实验证实，RNA结合蛋白PTBP1促进hsa_circ_0029692的剪接生成，hsa_circ_0029692与twist1竞争性结合miR-372-3p，从而改变twist1表达水平，twist1作为一种转录因子，实验结果证明twist1能够促进PTBP1的转录，上调PTBP1的表达，由此形成反馈环路：在子痫前期中，低水平表达的PTBP1，剪接生成hsa_circ_0029692水平降低，通过miR-372-3p介导twist1表达水平下调，最终导致滋养细胞EMT、迁移和侵袭能力减弱，而twist1的表达下调反馈调节PTBP1转录水平降低，引发子痫前期的发病。实验还发现，反馈环路中PTBP1和hsa_circ_0029692的表达水平下调，导致hsa_circ_0029692通过miR-135b-5p介导结合竞争的vimentin表达下调，由于twist1、vimentin均对滋养细胞EMT、迁移和侵袭有显著调控作用，由此证实：PTBP1剪接hsa_circ_0029692，通过miR-372-3p（/miR-135b-5p）介导调控twist1（/vimentin），形成正反馈环路调节滋养细胞EMT、迁移和侵袭，诱导子痫前期的发生。. 本研究发现一条新的反馈信号通路PTBP1/circ_0029692/miR-372-3p(miR-135b-5p)/Twist1，调控子痫前期细胞滋养细胞EMT、迁移和侵袭的作用机制，为揭示子痫前期发生发展的新机制，以及防治子痫前期，提供新的理论与实验依据，对子痫前期的早期诊断或临床治疗策略的建立提供帮助。