Browsing by Subject "androgen"
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Item Connexin 43 Contributes to Estrogen Protection against Oxidative Stress in Cortical Astrocytes(2019-05) Kubelka, Nicholas K.; Singh, Meharvan; Uht, Rosalie M.; Schreihofer, Derek A.; Yang, Shaohua; Planz, John V.Age-related brain disorders are associated with the decline in the ability of brain cells to cope with homeostatic challenge. Although all major brain cell types have the capacity to respond to homeostatic challenges, astrocytes are particularly well-equipped to counteract these challenges. Here, we focused on Connexin 43 (Cx43) as a protein that is not only highly expressed in astrocytes, but whose expression is critical to inter-cellular communication that in turn, can influence cell viability. Most studies to date have focused on the expression (i.e., abundance) of Cx43. However, a critical limitation of these studies is that they did not thoroughly examine functionality of the Cx43 channels. In particular, there is a paucity of data describing the differential contributions of Cx43-containing hemichannels versus Cx43-containing gap junctions to cellular functions. We hypothesized the astrocyte Cx43 hemichannel as a yet unreported target of androgens and estrogens based on three notions. First, our laboratory has determined that astrocytes are a relevant and important target of such gonadal steroid hormones as estrogens (e.g., 17[beta]-estradiol (abbreviated herein as estradiol or E2)) and androgens (such as DHT), through which these hormones promote healthy brain cell function. Second, oxidative stress is associated with an increase in Cx43 opening. Finally, the Cx43 gene promoter contains functional estrogen response element (ERE) half sites, and estradiol, as well as other estrogenic compounds, decrease Cx43 channel opening in peripheral (non-CNS) tissue. Based on these notions, we hypothesized that gonadal androgens and estrogens will inhibit Cx43 hemichannel opening in cortical astrocytes as well. My data revealed that while E2, dihydrotestosterone (DHT), and the estrogenic metabolite of DHT (3[beta]diol) all protect primary cortical astrocytes from the mixed metabolic/oxidative insult, iodoacetic acid (IAA), only DHT decreased astrocyte Cx43 mRNA expression. Consistent with their cytoprotective effects, however, all three steroids decrease astrocyte Cx43 hemichannel opening, and antagonized the increased opening of Cx43 hemichannels induced by IAA. In an effort to pursue the mechanism by which these steroids reduced Cx43 hemichannel opening, we evaluated the phosphorylation of Cx43 at two key residues, Ser 368 and Tyr 265. Phosphorylation at these residues is associated with channel closing, and as such, we predicted that the three hormones would increase the phosphorylation of Cx43 at one or both of these residues. Whereas Tyr265 phosphorylation was unaffected any of the three hormones, DHT significantly reduced the phosphorylation of Cx43 at Ser368. These observations may indicate that while all three steroids contribute to astrocyte protection through a mechanism that involves blocking astrocyte Cx43 hemichannel opening, DHT may induce molecular changes in the astrocytes that are distinct from those induced by estradiol or 3[beta]diol. The knowledge gained through the experiments conducted not only enhance our understanding of how Cx43 hemichannels and Cx43 gap junctions influence astrocyte function and viability but also define Cx43 hemichannels as relevant targets of gonadal steroid hormone induced regulation of cell viability. Such knowledge may facilitate the development of more precise therapeutics (i.e., selectively targeting Cx43 hemichannels without activity at Cx43 gap junctions in the same cells or tissue), the benefit of which would be to better treat age-associated neurodegenerative disorders as well as disorders of peripheral tissueItem INDUCTION OF APOPTOSIS VIA TESTOSTERONE IN OXIDATIVELY DAMAGED DOPAMINERGIC CELLS(2014-03) Simmons, Olivia C.; Holmes, Shaletha S.; Cunningham, RebeccaFollowing an episode of stroke, reduction of blood supply to brain cells can lead to conditions of oxidative stress (OS) in brain cells, or neurons, specifically in the dopaminergic neurons. The loss of dopaminergic neurons manifests itself as Parkinson’s disease (PD). Classically, men have a higher incidence of developing PD post-stroke than females. This suggests a role of testosterone (T) in the development of PD after induction of OS in neurons. We postulate that T in OS-induced states will activate the pathway of apoptosis, or programmed cell death, via an enzyme called caspase to induce death of dopaminergic brain cells, and thus symptoms of PD. To test our hypothesis, we first pre-treated dopaminergic cell lines with hydrogen peroxide (H2O2) to simulate stroke-induced OS. The cells were then treated with differing concentrations of T (0, 1, 10, 100 nM), representing the physiologic ranges of T in humans. Expressions of pro-caspase-3 and pro-caspase-9, the uncleaved precursors to caspase-3 & caspase-9, respectively, in the cells were quantified using Western Blot analysis. Statistical significance of our findings was reported using ANOVA and Fisher’s post hoc analysis with SAS software. Our experiments showed a trend of decreased expression of pro-caspase-9, and a significant decrease in pro-caspase-3 expression in the H+T treatment conditions as compared to the control conditions. These results point to the apoptotic cell death pathway via caspase-3 and caspase-9 as the mechanism by which increased T levels lead to PD in stroke patients. Purpose (a): Following ischemic stroke, reduction of blood supply to brain cells can lead to conditions of oxidative stress (OS) in neurons, specifically in the dopaminergic neurons of the substantia nigra (SN). The loss of dopaminergic neurons manifests itself as Parkinson’s disease (PD). Classically, men have a higher incidence of developing PD post-stroke than females. This suggests a role of testosterone (T) in the development of PD after induction of OS in neurons. We postulate that T in OS-induced states will activate the caspase pathway of apoptosis to induce dopaminergic cell death, and thus symptoms of PD. Methods (b): To test our hypothesis, we first pre-treated N27 dopaminergic cell lines with hydrogen peroxide (H2O2) to simulate stroke-induced OS. The cells were then treated with differing concentrations of T (0, 1, 10, 100 nM), representing the physiologic ranges of T in humans. Expressions of pro-caspase-3 and pro-caspase-9, the uncleaved precursors to caspase-3 & caspase-9, respectively, in the cells were quantified using Western Blot analysis. Statistical significance of our findings was reported using ANOVA and Fisher’s post hoc analysis with SAS software and p <0.05 as significant. Results (c): Our experiments showed a trend of decreased expression of pro-caspase-9, and a significant decrease in pro-caspase-3 expression in the H+T treatment conditions as compared to the control conditions. Conclusions (d): These results point to the apoptotic pathway via caspase-3 and caspase-9 as the mechanism by which increased T levels lead to PD in stroke patients.Item The role of lipid rafts in androgen's neurotoxic effects(2021-08) Fadeyibi, Oluwadarasimi M.; Cunningham, Rebecca L.; Cunningham, J. Thomas; Sumien, Nathalie; Yan, Liang-Jun; Park, InWooSex differences have been observed in multiple oxidative stress (OS) associated neurodegenerative diseases. Androgens, the major male sex hormone, through a membrane androgen receptor (mAR) localized to lipid rafts in the plasma membrane exacerbates OS. The goal of this study is to determine if interfering with mAR localization to cholesterol-rich lipid rafts decreases androgen induced neurotoxicity under OS environments. We hypothesize that cholesterol-rich caveolar lipid rafts are necessary for androgens to induce OS generation in neurons via the mAR localized within the plasma membrane. Nystatin was used to deplete cholesterol-rich caveolar lipid rafts in N27 cells prior to testosterone exposure in oxidative stressed N27 cells. Cell viability and protein analysis of OS, apoptosis, and mAR localization were conducted. Our results show that the loss of lipid rafts blocked androgen-induced OS in cells by decreasing the localization of mAR to caveolar lipid rafts. These results are consistent with our clinical findings that showed a relationship between hyperlipidemia and androgens on dementia in men but not women.