Overactivation of the androgen receptor exacerbates gravid uterine ferroptosis via interaction with and suppression of the NRF2 defense signaling pathway

Abstract

The mechanisms through which the androgen-dependent activation of the androgen receptor (AR) regulates gravid uterine ferroptosis remain unknown. We show that while co-exposure of pregnant rats to the androgen 5adihydrotestosterone (DHT) and insulin (INS) triggered uterine ferroptotic signaling cascades, additional treatment with the anti-androgen flutamide increased expression of the key ferroptosis-inhibitory proteins SLC7A11, GSH, and GPX4; reduced iron content; normalized levels of ferroptosisassociated Tfrc, Fpn1, and Ho1 mRNAs; reduced levels of proteins modified by 4-HNE (a marker of ferroptosis); and restored protein levels of NRF2, a key transcription factor regulating antioxidant defense signaling, in the gravid uterus. Furthermore, exposure to DHT alone increased uterine ferroptosis, and NRF2 abundance was negatively correlated with AR status. Coimmunoprecipitation and Western blot assays revealed that the AR physically interacted with endogenous NRF2, and this interaction was increased by DHT exposure in vivo. Our results suggest that AR overactivation and NRF2 suppression cooperate in the regulation of NRF2-targets in uterine ferroptosis.

Regulatory mechanisms of HMGB1 and its receptors in polycystic ovary syndrome‐driven gravid uterine inflammation

“…However, our findings are the first, to our knowledge, to demonstrate that in the uterus, the expression of HMGB1 protein and its interaction with AR is independent of hyperandrogenic stimulation. We and others have demonstrated that in vivo and in vitro exposure to DHT can increase uterine AR protein expression [48] and promote its interaction with other nuclear proteins such as melanoma antigen A11 [53] and nuclear factor erythroid 2‐related factor 2 [21]. While our study does not preclude the possibility that short‐term exposure to androgens may also regulate HMGB1 protein expression and interaction with AR in the uterus, the precise mechanism underlying the androgen‐independent crosstalk between HMGB1 and AR in the uterus in vivo appears to be more complex than we originally thought.…”

“…Band densitometry was performed using Image Laboratory (Version 5.0, Bio‐Rad), and the intensity of each protein band was normalized to the total protein in the individual sample. For quantification and to ensure standardization across blots, the expression of the target protein was normalized to the mean value for the control group on the blot, and then all the normalized values were statistically compared to assess the effect of the treatment groups as described previously [21,48]. When necessary, the membranes were stripped using Restore PLUS western blot stripping buffer (Thermo Fisher Scientific) for 15 min at room temperature, washed three times in TBST and then re‐probed [40,41,48].…”

“…All procedures were performed as described previously [21]. Briefly, uterine tissues were extracted with ice-cold lysis buffer.…”

“…We and others have reported in both in vivo and in vitro experimental settings that activation of the androgen-androgen receptor (AR) axis and development of insulin resistance increase pro-inflammatory responses and are associated with elevated inflammation in different cells, such as blood mononuclear, ovarian and endometrial cells in PCOS patients and PCOS-like rats [14][15][16][17][18]. In addition, animal studies have demonstrated that the overactivation of uterine AR in association with increased fetal loss may be in part due to elevated gravid uterine ferroptosis under the conditions of hyperandrogenism and insulin resistance [19][20][21]. Despite these important findings, how the PCOS-related manifestations impact specific intracellular molecules to regulate inflammatory responses in the uterus is still not fully understood.…”

Ferroptosis-Related Proteins Are Potential Diagnostic Molecular Markers for Patients with Preeclampsia

“…Multiple signaling pathways, including the mitogen-activated protein kinase (MAPK) signaling pathway, p53 signaling pathway, and the Hippo pathway, have been found to play important roles in ferroptosis [ 16 , 17 , 18 , 19 ]. In addition to the two classic ferroptosis-regulating genes, SLC7A11 and GPX4, a growing number of ferroptosis-related genes (FRGs) have been identified as contributors to some diseases [ 20 , 21 , 22 , 23 , 24 ]. As the research advanced, accumulating evidence indicates that ferroptosis is associated with a variety of human diseases, including carcinogenesis, degenerative diseases, cerebral hemorrhage, renal degeneration, and ischemia–reperfusion injury [ 25 , 26 , 27 , 28 , 29 , 30 ].…”