Nitric Oxide Regulation of Adrenal Steroidogenesis

描述(申请人提供)：血管内皮细胞释放a 生物活性介质(PGI2、一氧化氮(NO)和 作用于邻近的血管平滑肌细胞以影响血管 语气。在肾上腺皮质，随着肾上腺动脉变小并分成 毛细血管和血窦，平滑肌层丢失，腔隙 EC的一侧与类固醇生成细胞有密切的解剖接触。 因此，在肾上腺皮质的毛细血管中，这些介质可能会影响 邻近类固醇生成细胞的功能。我们把研究的重点放在 内皮源性NO在类固醇合成中的作用。我们发现，没有 供者抑制血管紧张素II(ALL)刺激的醛固酮释放。这个 NO的抑制作用不是通过环状GMP介导的，而是通过NO与 参与醛固酮生物合成的细胞色素P450酶。我们无法 检测ZG细胞一氧化氮合成、一氧化氮合酶活性及一氧化氮合酶蛋白的表达。 而肾上腺毛细血管内皮细胞则含有一氧化氮合酶并释放一氧化氮。我们将测试 假设在解剖学上与ZG细胞非常接近的ECs 肾上腺皮质，释放NO，调节或调节醛固酮的释放。这个 拟议的研究将调查外源、内源和 内皮源性NO对醛固酮释放的影响。这些研究将进行 体外培养牛ZG细胞和肾上腺毛细血管内皮细胞。另外，研究 将在灌流的大鼠肾上腺和麻醉的活体中进行 老鼠。这一假设将通过四个具体目标进行检验：(1)我们 将表征外源NO对培养细胞中醛固酮释放的影响 牛ZG细胞。一氧化氮供体，地他-农酸盐的作用将在 在氧气浓度不同的条件下，醛固酮会释放。 进一步的研究将确定NO抑制生物合成的步骤和 慢性一氧化氮处理对醛固酮生成和类固醇生成的影响 酵素。(2)由于ZG细胞没有一氧化氮合酶，我们将确定 转导ZG细胞增强一氧化氮合酶活性的内源性NO 携带一氧化氮合酶的腺病毒。(3)我们将描述 内皮源性NO对ZG细胞醛固酮释放的影响。利用共同孵化 ECS和ZG细胞，我们将确定激动剂对NO释放的影响 以及这种NO对ZG细胞释放醛固酮的作用。(4)我们会 确定内皮细胞一氧化氮在调节类固醇合成和肾上腺激素中的作用 麻醉状态下原位灌流肾上腺和活体肾上腺的血流 老鼠。

DESCRIPTION (provided by applicant): Vascular endothelial cells (ECs) release a number of biologically-active mediators (PGI2, nitric oxide (NO) and endothelin) that act on adjacent smooth muscle cells to influence vascular tone. In the adrenal cortex, as adrenal arteries get smaller and divide into capillaries and sinusoids, the smooth muscle layer is lost and the abluminal side of the EC comes in close anatomical contact with the steroidogenic cells. Thus, in the capillaries of the adrenal cortex, these mediators may influence the function of the adjacent steroidogenic cells. We have focused our studies on the role of endothelial-derived NO on steroidogenesis. We found that the NO donors inhibit angiotensin II (All)-stimulated aldosterone release. The inhibition by NO is not mediated by cyclic GMP but by NO binding to the cytochrome P450 enzymes involved in aldosterone biosynthesis. We were unable to detect NO synthesis, NO synthase (NOS) activity or NOS protein in ZG cells. However, adrenal capillary ECs contain NOS and release NO. We wilt test the hypothesis that ECs, which are in close anatomical proximity to ZG cells in the adrenal cortex, release NO and regulate or modulate aldosterone release. The proposed studies will investigate the effects of exogenous, endogenous and endothelial-derived NO on aldosterone release. These studies will be conducted in vitro in cultured bovine ZG cells and adrenal capillary ECs. Also, studies will be performed in the perfused rat adrenal gland and in vivo in anesthetized rats. The hypothesis will be tested by addressing four specific aims: (1) We will characterize the effect of exogenous NO on aldosterone release in cultured bovine ZG cells. The effect of the NO donor, deta-nonoate, will be tested on aldosterone release under conditions that vary the oxygen concentration. Additional studies will determine the biosynthetic step inhibited by NO and the effect of chronic NO treatment on aldosterone production and steroidogenic enzymes. (2) Since ZG cells do not have NOS, we will determine the effect of endogenous NO by conferring NOS activity on ZG cells by transducing the cells with an adenovirus containing NOS. (3) We will characterize the influence of endothelial-derived NO on ZG cell aldosterone release. Using co-incubation of ECs and ZG cells, we will determine the effect of agonists on the release of NO from ECs and the action of this NO on ZG cell aldosterone release. (4) We will determine the role of endothelial NO in regulating steroidogenesis and adrenal blood flow in the in situ perfused adrenal gland and in vivo in anesthetized rats.