RESUMO
In women of childbearing age, extensive decidualization, shedding and remodeling of the endometrium during the menstrual cycle are fundamental for successful pregnancy. The role of prostaglandins (PGs) in menstruation has long been proposed in humans, and the rate-limiting enzyme cyclooxygenase was shown to play a key role in endometrial breakdown and shedding in a mouse menstrual-like model in our previous study. However, the specific types of PGs involved and their respective roles remain unclear. Therefore, our objective was to investigate the mechanism through which PGs regulate endometrial disintegration. In this study, the microscopy was observed by HE; the protein levels of prostaglandins E1 (PGE1), prostaglandins E2 (PGE2), prostaglandin F2α (PGF2α) and Prostaglandin I2 (PGI2) were detected by ELISA; the mRNA level of Pfgfr2, Vascular Endothelial Growth Factor(Vegf), Angiostatin and Hypoxia inducible factor-1α (Hif1α) were examined by real-time PCR; PTGFR Receptor (PTGFR), VEGF, Angiostatin and HIF-1α protein levels were investigated by western blotting; the locations of protein were observed by Immunohistochemistry; HIF-1α binding PTGFR promoter was detected by Chromatin Immunoprecipitation (ChIP) and real-time PCR. We found that the concentrations of PGE1, PGE2, and PGF2α all increased significantly during this process. Furthermore, Ptgfr mRNA increased soon after Progesterone (P4) withdrawal, and PTGFR protein levels increased significantly during abundant endometrial breakdown and shedding processes. PTGFR inhibitors AL8810 significantly suppressed endometrial breakdown and shedding, promoted Angiostatin expression, and reduced VEGF-A expressions and vascular permeability. And HIF-1α and PTGFR were mainly located in the luminal/gland epithelium, vascular endothelium, and pre-decidual zone. Interestingly, HIF-1α directly bound to Ptgfr promoter. Moreover, a HIF-1α inhibitor 2-methoxyestradiol (2ME) significantly reduced PTGFR expression and suppressed endometrial breakdown which was in accord with PTGFR inhibitor's effect. Similar changes occurred in human stromal cells relevant to menstruation in vitro. Our study provides evidence that PGF2α/PTGFR plays a vital role in endometrial breakdown via vascular changes that are regulated by HIF-1α during menstruation.
RESUMO
Aims: Hyperhomocysteinemia (HHcy) has been considered as a risk factor for cardiovascular disease, Alzheimer's disease, nonalcoholic fatty liver, and many other pathological conditions. Vitamin B6, Vitamin B12, and folate have been used to treat HHcy in clinics. However, at present, clinical therapies of HHcy display unsatisfactory effects. Here, we would like to explore a new mechanism involved in homocysteine (Hcy) metabolic disorders and a novel target for HHcy treatment. The key enzymes involved in Hcy metabolism deserve more insightful investigation. Methylenetetrahydrofolate reductase (MTHFR) is a key enzyme regulating the intracellular Hcy metabolism. Until now, the effect of post-translational modification on the bioactivity of MTHFR still remains unclear. This study aimed at exploring the relationship between MTHFR S-sulfhydration and its bioactivity, and at identifying the contribution of an elevated Hcy level on MTHFR bioactivity. Results: By both in vivo and in vitro studies, we observed the following results: (i) The bioactivity of MTHFR was positively associated with its S-sulfhydration level; (ii) MTHFR was modified at Cys32, Cys130, Cys131, Cys193, and Cys306 by S-sulfhydration under physiological conditions; (iii) Hydrogen sulfide (H2S) deficiency caused the decrease of MTHFR S-sulfhydration level and bioactivity in HHcy, which resulted in further aggravation of HHcy; and (iv) H2S donors reversed the decreased bioactivity of MTHFR in HHcy, thus reducing the excessive Hcy level. Innovation and Conclusion: Our study suggested that H2S could improve MTHFR bioactivity by S-sulfhydration, which might provide a candidate therapeutic strategy for HHcy. Antioxid. Redox Signal. 36, 1-14.
