Phillip G. Kopf

ID 52920 Poster Board 424 Kratom is a tropical evergreen tree indigenous to Southeast Asia whose leaves and decoctions have long been used in traditional folk medicine for the relief of pain and enhancement of vitality. In recent years kratom use has increased in the United States, where an estimated 10-15 million people use the herb for the self-management of pain and opioid withdrawal. Kratom leaves contain over 40 active alkaloids. Previous studies in our laboratory demonstrated that mitragynine was the only kratom alkaloid that enhanced cortisol secretion in a human adrenocortical cell line. To further investigate the molecular mechanism by which mitragynine enhances cortisol secretion, we examined the effect of mitragynine on the expression of the steroidogenic enzymes and cofactors involved in cortisol production. HAC15 cells were exposed to vehicle or mitragynine (10 μM). After 72 h, mRNA expression of the following steroidogenic enzymes, cofactors, and receptors was examined: CYP11A1, HSD3B2, CYP21A2, CYP11B1, StAR, FDXR, FDX1, and MC2R. While mRNA expression was upregulated for all of these steroidogenic factors, CYP11B1 was induced to the greatest extent (fold change: vehicle, 1.0±0.2; 10 μM mitragynine, 18.4±1.5; n=6). Analysis of protein levels of these steroidogenic factors is underway. These data indicate that mitragynine enhances cortisol secretion in vitro via upregulation of the cortisol synthesis enzymatic pathway. This mechanism may contribute to anti-inflammatory and analgesic actions of kratom in vivo. Additional studies are currently being done to determine what receptor is mediating this stimulation of the cortisol pathway.

Polybrominated diphenyl ethers (PBDEs) have been previously shown to alter various endocrine biosynthetic pathways. Growing epidemiological evidence suggests that PBDEs alter cardiovascular function. The goal of this study was to examine the effects of BDE-47 on adrenal corticosteroid pathways that play vital roles in cardiovascular homeostasis and pathophysiology. The effect of BDE-47 on aldosterone and cortisol secretion was characterized in a human adrenocortical cell line. HAC15 cells were exposed to various concentrations of BDE-47 (1 nM to 100 μM). Cell viability, corticosteroid secretion, gene expression of enzymes involved in corticosteroid synthesis, and metabolic activity was examined. Additionally, Sprague Dawley male rats were orally exposed to BDE-47 (10 or 100 µg/kg), 5 days per week for 16 weeks. Organ weights and plasma corticosteroid levels were measured. In HAC15 cells, basal and stimulated aldosterone and cortisol secretion was significantly increased by BDE-47. Gene expression of several enzymes involved in corticosteroid synthesis and mitochondrial metabolism also increased. In Sprague Dawley rats, adrenal but not heart, kidney, or liver weights, were significantly increased in BDE-47 treatment groups. Plasma corticosterone levels were significantly increased in the 100 µg BDE-47/kg treatment group. No change in plasma aldosterone levels were observed with BDE-47 exposure. These data indicate that BDE-47 disrupts the regulation of corticosteroid secretion and provides further evidence that PBDEs are potential endocrine disruptors. Future studies will determine the underlying molecular mechanism of altered corticosteroid production and examine whether these alterations result in underlying cardiovascular disease in our rodent model of 16-week BDE-47 exposure.

The health benefits of the natural polyphenol trans‐resveratrol may play an important role in preventing a variety of diseases. Resveratrol has been shown to reduce blood pressure and improve metabolic diseases such as type 2 diabetes mellitus and obesity. Our previous studies examined the role of K⁺ channels in the vasorelaxation responses to trans‐resveratrol in the rat tail artery. During these studies, we uncovered a novel transient contraction prior to the sustained relaxation effect of trans‐resveratrol. Thus, the purpose of this study was to determine the role of the endothelium in these vascular contraction and relaxation responses to trans‐resveratrol. We additionally sought to determine if the cis‐isomer of resveratrol exerts any of the same vascular effects as the trans‐isomer. The vascular responses to trans‐resveratrol were examined in rat tail arteries with intact or denuded endothelium over a 2‐hr period. Additionally, the vascular responses to trans‐ and cis‐resveratrol were compared in rat tail arteries with intact endothelium. Both the transient contractile response and the persistent relaxation response to trans‐resveratrol were similar in the arterial rings with intact or denuded endothelium. There was a significant correlation between the initial contraction‐enhancing action of trans‐resveratrol and the magnitude of the sustained relaxation for vessels with both intact and denuded endothelium. Moreover, we demonstrated that cis‐resveratrol produced a significantly greater relaxation response as compared to trans‐resveratrol without the initial contractile response. These data demonstrate the role of the vascular smooth muscle in the vascular responses to resveratrol and the potential clinical benefits of the cis‐isomer of resveratrol as compared to the trans‐isomer.

Hyperaldosteronism is associated with hypertension, cardiac hypertrophy, and congestive heart failure. Steroidogenic factors facilitate aldosterone secretion by increasing adrenal blood flow. Angiotensin II (Ang II) decreases adrenal vascular tone through release of zona glomerulosa (ZG) cell-derived vasodilatory eicosanoids. However, ZG cell-mediated relaxations of bovine adrenal cortical arteries to Ang II are not altered by AT1 or AT2 receptor antagonists. Since traditional Ang II receptors do not mediate these vasorelaxations to Ang II, we investigated the role of Ang II metabolites. Ang III was identified by LC/MS as the primary ZG cell metabolite of Ang II. Ang III stimulated ZG cell-mediated relaxations of adrenal arteries with greater potency than Ang II. Furthermore, ZG cell-mediated relaxations of adrenal arteries by Ang II are attenuated by aminopeptidase inhibition, while Ang III-stimulated relaxations persisted. Ang IV had little effect as compared to Ang II. Moreover, ZG cell-mediated relaxations of adrenal arteries by Ang II were attenuated by an Ang III antagonist, but not by an Ang (1-7) antagonist. In contrast, Ang II and Ang III were equipotent in stimulating aldosterone secretion from ZG cells and were unaffected by aminopeptidase inhibition. Additionally, aspartyl- and leucyl-aminopeptidases, that convert Ang II to Ang III, are the primary peptidase expressed in ZG cells. This was confirmed by enzyme activity. These data indicate that intra-adrenal metabolism of Ang II to Ang III is required for ZG cell-mediated relaxations of adrenal arteries but not aldosterone secretion. These studies define an important new role of Ang III in the adrenal gland.