长链非编码RNA-lncRNA 121610异常表达导致子宫内膜异位症内膜容受性受损的分子机制

Endometriosis (EM) with infertility is very prevalent in women at child bearing age. This is largely attributed to damaged endometrial receptivity in patients, however, the inner molecular mechanism is unclear. Our previous study showed that the expressions of molecular biomarkers in endometrial receptivity, such as leukemia inhibitory factor (LIF) were significantly lower in EM patients with infertility compared to controls. The structure of endometrium was damaged. Lon non-coding RNA 121610 was found to be down-regulated in the eutopic endometrium in patients from transcriptome screening. Knockdown of lncRNA 121610 inhibited the expression of LIF. The level of lncRNA 121610 in the cytoplasm of endometrial cells was significantly up-regulated after progesterone treatment. And antagonism of the progesterone receptor (PR) diminished the stimulatory effects. Has-miR-580-3p is a potential target of lncRNA 121610. Knockdown of this miRNA promoted the expression of LIF. Accordingly, we propose that progesterone can promote the expression of lncRNA 121610 through progesterone receptor (PR); LncRNA 121610 can sponge has-miR-580-3p by competing endogenous RNAs (ceRNA) mechanism which modulates the expression of LIF and αvβ3 Integrin, and then participate in the development of endometrial receptivity. To validate the presumption, the project intends to systematically study the up- and downstream signaling pathway by which lncRNA 121610 participates in the development of endometrial receptivity using cell and mouse model, together with clinical samples. We try to reveal the reason why abnormal expression of lncRNA 121610 causes the damage to endometrial receptivity in endometriosis with infertility. Results from the project will provide reference to the understanding of the etiology of endometriosis with infertility.

子宫内膜异位症（EM）合并不孕症发病率高，这与其内膜容受性受损密切相关，但分子机制尚不清楚。我们前期研究发现患者内膜形态受损，内膜容受性标记分子LIF等表达下调；表达谱筛选显示长链非编码RNA lncRNA 121610在患者中表达也显著降低；而下调该lncRNA抑制LIF表达；孕激素可显著上调该lncRNA的胞浆表达，PR受体拮抗剂则抑制这种效应；has-miR-580-3p是其潜在靶标，下调该miRNA促进LIF表达。据此，我们推测孕激素通过自身受体PR促进lncRNA 121610的表达，进而通过ceRNA机制吸附has-miR-580-3p，调控LIF和αvβ3 Integrin的表达，参与内膜容受性的建立。本项目拟利用细胞、小鼠和临床样本，系统研究lncRNA 121610参与内膜容受性建立的上下游分子机制，揭示其异常表达导致EM患者内膜容受性受损，从而参与EM不孕症发生的机制。

子宫内膜异位症（EMs）是一种雌激素依赖性女性疾病，是子宫内膜腺体细胞和基质细胞易位至宫腔之外的部位而引发的，以不孕和疼痛为典型症状。蜕膜化是子宫内膜的一种特殊分化过程，是内膜胚胎着床和建立妊娠的基础。前期芯片数据中筛选出分泌中期EMs组与正常组内膜组织中差异表达的lncRNA并进行验证发现 Lnc121610在EMs中呈低表达。随着蜕膜化的进展，lnc121610的表达逐渐升高。在诱导蜕膜化Day2、Day4和Day6时，lnc121610敲低组的蜕膜化标志基因PRL和IGFBP1的表达水平降低。随着BMP2浓度升高，PRL和IGFBP1表达水平明显升高，lnc121610表达水平明显升高。敲低/过表达lnc121610后，则PRL和IGFBP1的表达水平降低/升高。lnc121610敲低组分化期SMAD1、p-SMAD1、p-SMAD1/5/8蛋白表达明显降低。此外，通过分析子宫内膜的单细胞数据，发现TWIST2在蜕膜组织中异常表达，且敲降TWIST2后的结果提示，蜕膜化进程受阻。Chip-qPCR和双荧光素酶报告验证TWIST2与PR可能一起调控蜕膜化相关基因的转录，并且TWIST2可能增强PR对其下游分子的调控能力。本课题系统研究 lnc121610参与HESC细胞蜕膜化的分子机制。lnc121610的敲低可抑制HESC细胞蜕膜化表型的建立；外源性BMP2可影响HESC细胞中lnc121610的表达；lnc121610的敲低可抑制BMP2经典作用通路上的关键分子p-SMAD1/5/8蛋白的表达；TWIST2与PR互作，且TWIST2增强PR对下游CEBPB、IGFBP1的转录调控。阐明lncRNA在子宫内膜蜕膜化调控中的重要作用以及与EMs的关联，为内异症发病机制的解析和治疗靶点的筛选提供参考。