miR-29调控TGF-β/Smad信号通路在宫腔粘连中的作用机制研究

Intrauterine adhesions (IUA) as the terminations of aberrant fibrosis after endometrium injury，was a urgently clinical puzzle for seriously threatening to female reproductive health. miRNA was concerned about the important role in regulation of fibrosis pathway in multiple organ fibrosis disease. Earlier stage of project terms detected that: the high expression of TGF-β, Smad2 and Smad3 in IUA endometria positively correlating with the severity of the IUA; the level of miR-29 having a negative correlation with protein expression of TGF-β, Smad2 and Smad3 in IUA. This fact preliminary confirmed miR-29 might control and regulate the TGF-β/Smad signaling pathways and involve in the occurrence of IUA. On this basis ,the investigation design to: 1) Respectively construct plasmid with mimicker and inhibitor of miR-29, transfection endometrial stroma cells, test the fibrosis; gradually increasing TGF-β stimulation of endometrial stromal cells in time, up-regulation of miR-29 level, test its role in reversing the process of fibrosis, and to further clarify the signaling pathways in regulation of fibrosis; confirm that TGF-β is the target genes of miR-29 by being co-transfected TGF-β 3'UTR-psiCHECK-2 and miR-29 with endometrial stromal cells; 2) Regulate miR-29 level in vivo of IUA animal models, elucidate its role in the process of IUA fibrosis, provide a new theoretical foundation to clarify the pathogenesis of IUA, and provide new targets for the treatment.

宫腔粘连（IUA）是子宫内膜损伤后发生的纤维化病变，严重威胁女性生殖健康，是临床亟待解决难题。miR-29以其在纤维化疾病中的调节作用而备受关注。项目组前期发现：IUA子宫内膜中TGF-β、Smad2和Smad3高表达，与IUA分期正相关；miR-29与TGF-β、Smad2及Smad3表达均呈负相关，初步证实miR-29与TGF-β/Smad通路相关，并可能参与IUA的发生。本研究拟：1) miR-29 mimic和miR-29 inhibitor分别转染子宫内膜间质细胞，检测纤维化进程；渐进延长TGF-β刺激细胞时间，上调miR-29，检测其逆转纤维化；TGF-β 3'UTR-psiCHECK-2和miR-29 mimic共转染子宫内膜间质细胞，证实TGF-β是miR-29靶基因；2)调控IUA模型miR-29水平，探讨可能作用机制。为阐明IUA发病机制、探寻治疗新方向提供理论基础。

宫腔粘连(IUA)是子宫内膜基底层损伤后发生的纤维化病变，可引起月经减少、不孕不育及妊娠相关并发症等，其分子发病机制不清、治疗效果欠佳，严重威胁女性生殖健康。miR-29 和TGF-β/Smad信号通路以其参与了多个器官纤维化疾病的调节作用而备受关注。我们团队前期研究发现IUA 子宫内膜中TGF-β、Smad2 和Smad3 高表达，而miR-29 低表达，初步证实miR-29与TGF-β/Smad 通路相关，并可能参与了IUA 的发生。为进一步探讨其作用机制，我们又做了以下研究：①检测了人IUA子宫内膜中TGF-β1、Smad2、Smad3J及miR-29a,结果显示前3者呈高表达且随分期逐渐增加，miR-29a则呈低表达；②分离培养了人原代子宫内膜基质细胞(HESCs)，建立了TGF-β1诱导的HESCs纤维化模型，转染miR-29，检测提示能够抑制纤维化，使细胞中COL1A1、α-SMA在mRNA及蛋白水平表达下降；③建立大鼠IUA模型，预防性转染agomir-29b，提示可有效抑制子宫内膜纤维化。从人体、动物和细胞上证实了miR-29 能通过抑制TGF-β/Smad信号通路以减轻IUA，为临床IUA的防治提供新靶点和治疗方法。.另外我们在临床雌激素治疗和手术预防宫腔再粘连的技巧方面也进行了探索研究，发现使用INTERCEED防粘连膜可以使IUA的复发率减少和减轻，并提高子宫内膜容受性；IUA早期应用雌激素治疗可以减少粘连程度，有一定预防作用；全反式维甲酸ATRA可以通过抑制TGF-β1/smad4信号通路抑制子宫内膜纤维化。这些研究为临床治疗IUA提供了可靠的科学依据和新的治疗方法。项目按计划顺利完成，共发表文章21篇，其中SCI 6篇；培养人才9人，其中博士研究生4人、硕士研究生5人。