The Mechanism Research Of Periostin Regulates Migration, Invasion And Adhesion Of Human Endometrial Stromal Cells

Background & Objective:Endometriosis is a benign disease which affects almost 10% of women in their reproductive period. Currently, available therapies are surgical excision and various medical treatments which include gonadotropinreleasing hormone (GnRH) analogs, aromatase inhibitors, and progestins. However, these medical therapies are not suitable long term due to their numerous side effects, such as massive hemorrhage, perimenopausal stage symptoms and liver dysfunction. Moreover, the recurrence rate of endometriosis is still high following conservative surgery, unless patients accepted hysterectomy or reached menopause. Endometriosis is also associated with the occurrence of cancers and chronic diseases. Thus, it is necessary to continue seeking for an effective targeted therapy for endometriosis. At present, the etiology of endometriosis remains unclear, but the theory of reflux menses Sampson proposed in 1921 is still the leading theory. In recent years, Lang JH proposed the determinant of uterine eutopic endometrium for the supplementary of Sampson’s theory, including ectopic adhesion, invasion and proliferation of endometrial debris. Periostin (PN) belongs to the Fasciclin I family and is generally present at low level in most adult tissues, but plays a crucial role in oncology, injury or inflammation. High level of PN can promote cell migration, invasion, adhesion, and proliferation in many cancers. While our previous study confirmed that periostin (PN) was overexpressed in the eutopic and ectopic endometrial stroma of women with endometriosis by immunohistochemitry, the role of PN in the pathophysiology of endometriosis remains unknown. In this study, we aimed to investigate the effects of PN on endometrial stromal cells (ESCs) adhesion, migration, invasion and proliferation and to further study the mechanism under this process.Methods:1. The control endometria were obtained from 25 normal women. Eutopic and ectopic endometria were obtained from 32 women who underwent laparoscopy for ovarian endometriosis. Eutopic (EuSCs), ectopic (EcSCs) and normal ESCs (NSCs) were isolated and cultured. Then the identification and purity of ESCs were analyzed by cell immunofluorescence.2. The protein expression of PN, integrin-linked kinase 1 (ILK1) and p-Akt in NSCs, EuSCs and EcSCs were evaluated by western blot. And the level of PN mRNA was evaluated by qPCR.3. After receiving PN small interfering RNA (siRNA), the expression of PN, ILK1 and p-Akt in EcSCs were estimated again.4. We evaluated the abilities for migration, invasion, adhesion and proliferation in NSCs, EuSCs as well as EcSCs before and after receiving PN siRNA.Results:1. The identification and purity of ESCs:we successfully isolated and cultured ESCs. The purity of ESCs used for our study was (96.1 ± 2.2)%.2. The expression of PN, ILK1, and p-Akt in NSCs, EuSCs and EcSCs:the protein and mRNA levels of PN were upregulated in EuSCs (P< 0.05; P= 0.2261 in proliferative phase and P= 0.3385 in secretory phase) and EcSCs (P< 0.001; P< 0.001 in proliferative phase and P< 0.05 in secretory phase) compared with NSCs, while the mRNA level showed no significance between EuSCs and NSCs throughout the menstrual cycle. In EuSCs and EcSCs, the expressions of ILK1 and p-Akt were enhanced (P< 0.05).3. The influence of PN on the Expression of ILK1 and p-Akt in ESCs:the Expression of ILK 1 and p-Akt were downregulated after PN siRNA interference (P< 0.05)4. The influence of PN on the abilities for migration, invasion, adhesion and proliferation of ESCs:the abilities for migration and invasion of EuSCs and EcSCs were stronger than NSCs (P< 0.01; P< 0.001) and became lower after PN siRNA interference (P< 0.05; P< 0.01). EcSCs adhered to the fibronectin at the fastest speed (P< 0.05) and the ability was obviously inhibited after 60mins (P< 0.05). There were many fiber-like substances among EcSCs, which were dissolved after the loss of PN. However, ESCs proliferation showed no statistical significance before and after transfection of PN siRNA.Conclusion:1. NSCs, EuSCs and EcSCs were all derived from endometrial stroma.2. PN enhanced ESCs migration, invasion and adhesion due to the ILK1/Akt signal pathway. PN can also promote fibrogenes.3. PN may be a new clinical treatment target of endometriosis.