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Browsing by Author "Handa, Robert, advisor"

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    Dihydrotestosterone attenuates endotoxin, cytokine, and hypoxia-induced vascular inflammation
    (Colorado State University. Libraries, 2011) Osterlund, Kristen Leanne, author; Handa, Robert, advisor; Gonzales, Rayna, committee member; Amberg, Gregory, committee member; Garrity, Deborah, committee member; Tobet, Stuart, committee member
    Vascular inflammation plays a key role in the etiology of cardiovascular disease, particularly stoke. Vascular inflammation is under the control of several transcription factors, including nuclear factor kappa B and hypoxia inducible factor-1 alpha (HIF-1α). Activation of these transcription factors can lead to the production of inflammatory mediators such as cyclooxygenase-2 (COX-2). COX-2 plays a role in vascular inflammation, cerebral ischemia-induced injury, and has been implicated as a source of reactive oxygen species (ROS). Inflammatory mediators, such as endotoxin or cellular breakdown products released following injury, are known to signal through the Toll-like receptor 4 (TLR4). TLR4 activation leads to NFκB activation and subsequent production of COX-2. Like COX-2, TLR4 has also been implicated in injury-induced oxidative stress and cerebral ischemia damage. Previous studies have demonstrated that gonadal steroid hormones can also modulate vascular inflammation. Both protective and detrimental effects of androgens on the cardiovascular system have been reported. Since the potent androgen receptor (AR) agonist dihydrotestosterone (DHT) can be converted to 3β-diol, an estrogen receptor (ER) β-selective agonist, I hypothesized that ERβ may mediate some of the protective effects of androgens, while the AR may mediate some of the detrimental effects. The overall goal of this dissertation was to determine the mechanisms by which androgens can influence the vascular inflammatory response under both physiological and pathophysiological conditions. The hypothesis to be tested was that DHT influences vascular inflammation under both physiological and pathophysiological conditions. In my first set of experiments, using Western blot, I found that DHT increases expression of the vascular inflammatory mediator COX-2 under physiological conditions in human coronary artery vascular smooth muscle (VSM) cells and human brain VSM cells. This effect of DHT was attenuated in the presence of the AR antagonist bicalutamide. This data indicates that the pro-inflammatory effect of DHT under normal physiological conditions is AR mediated. In my second set of experiments, I examined the effects of DHT on vascular inflammation under a variety of pathophysiological conditions. Surprisingly, I found that DHT decreased cytokine-induced COX-2 expression and oxidative stress, endotoxin-induced COX-2 and TLR4 expression in human VSM cells. Furthermore, DHT also decreased hypoxia induced HIF-1α and COX-2 expression in human brain VSM cells and rat pial arteries. Finally, I found that DHT decreased hypoxia with glucose deprivation (HGD)-induced HIF-1α, COX-2 and TLR4 expression in human brain VSM cells. DHT`s anti-inflammatory effects during cytokine or HGD-induced inflammation in human brain VSM cells were not blocked by the AR antagonist bicalutamide, indicating that they were not AR mediated. These results led me to my second hypothesis, that DHT's anti-inflammatory effects are ERβ-mediated. In my third set of experiments, I found that the DHT metabolite/ERβ selective agonist 3β-diol also decreased cytokine-induced COX-2 expression in human brain VSM cells. Furthermore, DHT's ability to reduce cytokine-induced COX-2 expression in human brain VSM cells was inhibited by the non-selective estrogen receptor antagonist ICI 182,780 and the selective ERβ antagonist PHTPP. The mRNAs for steroid metabolizing enzymes in the pathway necessary to convert DHT to 3β-diol were detected in human brain VSM cells, as were AR and ERβ mRNAs. Therefore, DHT appears to be protective against cerebrovascular inflammation via conversion to 3β-diol and subsequent activation of ERβ in human brain VSM cells. The results of these studies indicate that: 1) DHT increases COX-2 expression under unstimulated/physiological conditions via an AR-dependent mechanism. 2) DHT decreases cytokine-, endotoxin,-hypoxia, and HGD-induced COX-2 expression via an AR-independent mechanism. 3) DHT decreases cytokine-induced reactive oxygen species. 4) DHT decreases hypoxia-induced HIF-1α expression. 5) DHT decreases HIF-1α and TLR4 expression during HGD via an AR-independent mechanism. 6) DHT's effect to attenuate cytokine-induced COX-2 expression is ERβ-mediated.
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    Estrogen receptors alpha and beta: Opposing roles in hypothalamic-pituitary-adrenal axis function and stress-related behaviors
    (Colorado State University. Libraries, 2008) Weiser, Michael James, author; Handa, Robert, advisor
    Estradiol has reported effects on mood ranging from anxiogenic to anxiolytic and depressant to anti-depressant. These opposing actions of estradiol may be explained by the existence of two distinct estrogen receptor (ER) systems, ER alpha (ERα) and ER beta (ERβ). Furthermore, there exists a sex difference in stress-related psychiatric disorders such as anxiety and depression, for which women are more susceptible than men. Common to the pathology of these disorders is a dysfunctional hypothalamic-pituitary-adrenal (HPA) axis where glucocorticoid negative feedback is impaired leading to chronically high levels of circulating glucocorticoids. The HPA axis is the main neuroendocrine axis that governs physiological responses to stressors. In rodents, basal and stress-induced activity of the HPA axis is higher in females than in males. This suggests that, if transferable to humans, the sex difference observed in HPA axis function in animal models may help explain the female predisposition for certain psychiatric disorders. The studies described in this dissertation were aimed at characterizing the distinct roles for ERα and ERβ in HPA axis activity and stress-related behaviors. The studies in Chapter 3 examine the effect of estradiol signaling through ERα or ERβ on glucocorticoid negative feedback of the HPA axis. Results indicate that estradiol impairs glucocorticoid-dependent negative feedback by activating ERα specifically at the level of the paraventricular nucleus (PVN). The studies in Chapter 4 examine the effect of estradiol signaling through ERα or ERβ on anxiety-like and depressive-like behaviors. Results indicate that selective activation of ERα is anxiogenic and depressant, whereas selective activation of ERβ is anxiolytic and antidepressant. Finally, the studies in Chapter 5 examine the effect of estradiol signaling through ERβ on behavior and HPA axis activity induced by glucocorticoid receptor (GR) activation in the central nucleus of the amygdala (CeA). Results indicate that delivery of a GR agonist to the CeA is anxiogenic and augments the HPA axis response to a stressor, and peripheral administration of an ERβ agonist blocks this effect. Collectively, these studies point to an antagonistic relationship between estradiol signaling through ERα and ERβ with respect to HPA axis activity and stress-related behaviors.
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    Sex-dependent function and regulation of the hypothalamic pituitary adrenal axis
    (Colorado State University. Libraries, 2019) Heck, Ashley L., author; Handa, Robert, advisor; Bouma, Gerrit, committee member; Hentges, Shane, committee member; Florant, Gregory, committee member
    Physiological responses to stressors are largely governed by a neuroendocrine axis, the hypothalamic pituitary adrenal (HPA) axis. Whereas HPA activation is necessary for body wide adaptation to a stressor via the production of glucocorticoids, its excessive or inappropriate activation can increase risk for a number of diseases. Importantly, many of these stress-related diseases exhibit a strong sex bias in prevalence, which may be related to sex differences in the activity of the HPA axis. Thus, the studies described in this dissertation examine sex differences in the regulation and function of the HPA axis in rodents to further unravel the sex-dependent vulnerability often characteristic of stress-related diseases in humans. In Chapters 2 and 3, sex differences in glucocorticoid negative feedback at the level of corticotropin releasing hormone (Crh) in the hypothalamic paraventricular nucleus (PVN), an important factor limiting HPA axis activation, are explored. Results of Chapter 2 indicate that male C57BL/6 mice exhibit a more rapid response of PVN Crh expression to the removal of glucocorticoid negative feedback due to androgen actions, likely via upstream regulatory neurons. Results of Chapter 3, alternatively, show more robust glucocorticoid receptor (GR) mediated negative feedback on PVN Crh in females, but only on a day of their reproductive cycle when circulating estrogen levels are low. Thus, a complex interplay among androgen/ estrogen actions and glucocorticoid regulatory mechanisms appears to drive sex-dependent PVN Crh expression to potentially influence sex-biased HPA activity and stress-related disease risk. In Chapters 4 and 5, the response of the HPA axis to chronic stress, a factor which is more etiologically relevant for human disease risk, is examined. The results of Chapter 4 demonstrate that female C57BL/6 mice exhibit time-of-day dependent changes in the basal and acute stress-induced activity of the HPA axis following chronic variable stress (CVS). Male mice, conversely, appear mostly resistant to the effects of CVS on HPA function until socially isolated (Chapter 5). These findings establish an essential foundation for the use of the C57BL/6 mouse, a strain typically more resistant to the effects of CVS, in future studies of sex-dependent HPA axis regulation following chronic stress.