TLR4-Associated IRF-7 and NFκB Signaling Act as a Molecular Link Between Androgen and Metformin Activities and Cytokine Synthesis in the PCOS Endometrium

Context: Low-grade chronic inflammation is commonly seen in polycystic ovary syndrome (PCOS) patients with elevated levels of inflammatory cytokines in the endometrium. Objective: This work aimed to increase the limited understanding of the mechanisms underlying cytokine synthesis and increased endometrial inflammation in PCOS patients. Methods: Endometrial biopsy samples were collected from non-PCOS (n = 17) and PCOS (n = 22) patients either during the proliferative phase of the menstrual cycle or with hyperplasia. Endometrial explants were prepared from PCOS patients and underwent pharmacological manipulation in vitro. The expression and localization of toll-like receptor 2 (TLR2)/4, key elements of innate immune signal transduction and nuclear factor kappa B (NF kappa B) signaling pathways, and multiple cytokines were comprehensively evaluated by Western blotting, immunohistochemistry, and immunofluorescence in endometrial tissues. Results: We demonstrated the distribution of protein expression and localization associated with the significantly increased androgen receptor, TLR2, and TLR4-mediated activation of interferon regulatory factor-7 (IRF-7) and NF kappa B signaling, cytokine production, and endometrial inflammation in PCOS patients compared to non-PCOS patients with and without endometrial hyperplasia. In vitro experiments showed that 5-dihydrotestosterone (DHT) enhanced androgen receptor, TLR4, IRF-7, and p-NF kappa B p65 protein expression along with increased interferon alpha (IFN alpha) and IFN gamma abundance. The effects of DHT on IRF-7, p-NF kappa B p65, and IFN abundance were abolished by flutamide, an antiandrogen. Although 17 beta-estradiol (E2) decreased p-IRF-7 expression with little effect on TLR-mediated IRF7 and NF kappa B signaling or on cytokine protein levels, exposure to metformin alone or in combination with E2 suppressed interleukin-1 receptor-associated kinase 4 (IRAK4), p-IRF-7, IRF-7, I kappa B kinase alpha (IKK alpha), p-NF kappa B p65, IFN gamma, and tumor necrosis factor alpha protein expression. Conclusion: Cytokine synthesis and increased endometrial inflammation in PCOS patients are coupled to androgen-induced TLR4/IRF-7/NF kappa B signaling, which is inhibited by metformin treatment.

Automated summary

Cytokine synthesis and increased endometrial inflammation in PCOS patients are associated with androgen-induced TLR4/IRF-7/NFκB signaling, which can be inhibited by metformin treatment.