| Endometriosis(Ems)is a benign disease affecting approximately 15% of premenopausal women,whose main symptoms are chronic pelvic pain,dysmenorrhea and infertility.It is characterized by the growth of endometrial tissue outside the uterus.All of these symptoms of the disease may affect the patient's psychology,which often affects their social life.Endometrial stromal cells(ESCs)invaded the uterine cavity to form endometriosis tissue acts as an important pathological basis for this disease.Therefore,inhibiton of invasion of ESCs may be an effective treatment for Ems.In recent years,a large number of studies have reported that micro RNAs(miRNAs)play an important role in endometriosis,but little research has been done on ESCs invasion in endometriosis.Aim:The purpose of this experiment was to study the effect of micro RNAs on the invasion of endometrial stromal cells(ESCs),to explore the specific mechanism of miRNAs regulating the invasion of ESCs,and to clarify the miR-202 and k-Ras /ERK1 / 2 signaling pathways in ESCs.The specific mechanism of mutual regulation provides a new target for the prevention and treatment of endometriosis.Method:1 Analyzed the micro RNA high-throughput chip data published in the GEO database,we screened out the micro RNA in the ectopic endometrium and eutopic endometrium of the patients with ovarian endometriosis,and determined the target micro RNA for further study by consulting the analysis papers and analyzing the data of micro RNA microarray.The expression patterns of miRNAs in ectopic endometrium and eutopic endometrium from patients with Ems was analyzed by quantitative real-time quantitative PCR(q RT-PCR)in 30 ectopic endometrium from Ems patients and 15 normal endometrium from healthy persons.2 The effect of miR-202-5p on the invasion of ESCs was detected by transfecting miR-202-5p mimics into ESCs.3 The online software predicted the target gene of miR-202,and detected the target gene protein by transfecting miR-202 mimics into ESCs.The interaction between miR-202 and the target gene was verified by a dual luciferase assay.q RT-PCR was used to detect the expression of target gene in ectopic endometrial tissues of patients with Ems,and the relationship between miR-202 and target gene was further analyzed.4 si-k-Ras was transfected into ESCs,and the effects of knockdown of k-Ras on the invasion of ESCs were observed by Western blot,Transwell and wound healing assays,respectively.At the same time,ESCs were co-transfected with pc DNA-k-Ras and miR-202-5p mimics,and the effect of k-Ras overexpression on the function of miR-202-5p mimics in invasion of ESCs was measured.5 pc DNA-k-Ras was co-transfected with miR-202 mimics into ESCs,and the expression changes of p-Raf1,p-MEK and p-ERK were detected by Western blot.Result:1 Microarray analysis showed significant changes in the expression of multiple micro RNAs,some of which were similar to previous reports,indirectly confirming the credibility of our Microarray analysis;compared with the normal control group,miR-202 was significantly downregulated in endometrial tissue from patients with Ems.2 Overexpression of miR-202 inhibited the invasion and migration of ESCs by regulating the process of Epithelial-Mesenchymal Transition(EMT).3 Bioinformatics and dual luciferase assays confirmed that k-Ras is a target of miR-202.Overexpression of miR-202 inhibits its protein expression.Further studies showed that k-Ras was significantly elevated,and inversely correlated with miR-202 in endometrial tissue of patients with Ems.4 Further studies demonstrated that knockdown of k-Ras significantly inhibited the invasive ability of ESCs,whereas k-Ras overexpression weaken the inhibitory effects of miR-202 overexpression on ESCs invasive abilitiy.5 Western Blot assay showed that overexpression of miR-202 significantly inhibited the expression of phosphorylated-Raf1,MEK and ERK,while overexpression of kRas reversed the inhibitory effect of miR-202 overexpression.It is suggested that miR-202-5p inhibits the invasion and migration of ESCs by inhibiting the kRas/Raf1/MEK/ERK signaling pathway.Conclusion:Overexpression of miR-202 inhibited the invading ability of ESCs by regulating the process of epithelial-mesenchymal transition(EMT).K-Ras is a functional target gene of miR-202.Overexpression of mir-202 inhibited the invasion and migration of ESCs by blocking the activation of K-ras / raf1 / MEK / ERK signaling pathway. |
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