REGULATION OF RENIN AND PREECLAMPTIC HYPERTENSION

描述：(改编自《研究人员摘要》)许多组织分泌一种不活跃的肾素原形式。据信，产生活性肾素的43个氨基酸的前片段被移除，主要发生在肾脏。最近关于子痫前期转基因小鼠模型的数据支持生殖肾素在其发病机制中的作用，我们关于人类子痫前期蜕膜肾素表达增加的数据证实了这一作用。为了让肾素具有这样的功能，细胞对活性肾素的处理应该在局部进行。子宫肾素-血管紧张素系统的细胞和分子方面还没有得到充分的研究。在肾素基因的启动子中存在雌激素和孕激素反应元件，提示性激素在调节肾素原的表达中起作用。我们最近的数据表明，孕酮治疗子宫内膜间质细胞(体外蜕膜化)通过增加前肾素的表达和以激素特异性的方式增加前肾素到肾素的转化来增加肾素的活跃分泌，而人类妊娠的蜕膜细胞通过前肾素到肾素的过程。我们最近在HEK293细胞中的共转染实验和蜕膜细胞中组织蛋白酶B的抑制表明，组织蛋白酶B可能是一种前肾素转换酶。我们推测，蜕膜肾素的病理性升高可能直接引发子痫前期，性激素可能诱导原肾素及其原转换酶(PC)的表达，从而调节成熟肾素的分泌。因此，本研究的目的是：1)证明蜕膜基质细胞在体外分泌活性较高的成熟肾素，通过N末端测序测定；2)通过(I)用PC和组织蛋白酶B的探针进行Northern分析，以及(Ii)确定结构性和调节性肾素的分泌，以及(Iii)检测组织蛋白酶B抑制对原肾素加工的影响；(B)通过(I)用PC和组织蛋白酶B的探针进行Northern分析，确定负责原肾素激活的蜕膜细胞内切酶；(B)检测内切酶的表达调节；3)通过(A)与原肾素在小泡中的免疫显微镜共定位和(B)通过将原肾素和候选激活酶表达载体在标准细胞系中共转染并对成熟的活性肾素进行加工来验证功能，以确定候选激活酶是特异性的前肾素转换酶。这些研究将加深我们对孕酮在生殖肾素调节中的作用的理解，并可能为研究子痫前期的病理生理提供一个新的方向。

DESCRIPTION: (Adapted from the Investigator's Abstract) Many tissues secrete an inactive pro-form of renin. A 43 amino acid pro- segment removal to produce active renin is believed to occur, primarily in the kidney. Recent data on transgenic mouse model of preeclampsia support a role of reproductive renin in its pathogenesis and our data on increased decidual renin expression in human preeclampsia confirms this role. For renin to have such a functional role, cellular processing to active renin should occur locally. The cellular and molecular aspects of the uterine renin- angiotensin system have not been adequately investigated. Presence of estrogen and progesterone response elements in the promoter of renin gene suggests a role for the sex steroids in the regulation of prorenin expression. Our recent data suggest that progesterone treatment of endometrial stromal cells (in vitro decidualization) increases active renin secretion by increasing prorenin expression and by increasing conversion of prorenin to renin in a hormone- specific manner, and that decidual cells from human gestation process prorenin to renin. Our most recent co-transfection experiment in HEK293 cells and cathepsin B inhibition in decidual cells suggest that cathepsin B may function as a prorenin convertase. We hypothesize that pathologic increase in decidual renin may proximately initiate preeclampsia, and sex steroids may induce expression of prorenin and its proconvertase (PC), thereby regulating mature renin secretion. Therefore, the objectives are: 1) To demonstrate that decidual stromal cells in vitro secrete active mature renin, as assayed by activity and by N terminal sequencing; 2) To identify the decidual cell endoprotease responsible for prorenin activation by (i) northern analysis with probes for PCs and cathepsin B, and (ii) defining constitutive versus regulated renin secretion, and (iii) examining the effect of cathepsin B inhibition on prorenin processing; (b) To examine regulation of endoprotease expression; 3) To confirm that the candidate-activating enzyme is the specific prorenin convertase by (a) immuno-microscopic co-localization with prorenin in the vesicles and (b) functional verification by co-transfection of prorenin and candidate-activating enzyme expression vectors in a standard cell line and demonstration of processing to mature active renin. These studies will improve our understanding of the role of progesterone in the regulation of reproductive renin and may provide a novel direction for investigating the pathophysiology of preeclampsia.