PI/PLC FUNCTION IN PREGNANT RAT MYOMETRIUM

最初资助期的研究表明， 磷脂酰肌醇信号通路(PSP)的激活结果 胞浆钙振荡(CCO)驱动的时相子宫肌层 宫缩。产生重复性胞浆钙瞬变 通过释放储存的细胞内钙，伴随着内流 细胞外钙离子。更多的研究表明 PSP信号转导蛋白的几种异构体 在大鼠子宫肌层组织中表达，包括五种亚型 磷脂酶C(PLC)。同步运动的生理学意义 这些PLC亚型在子宫肌层组织中的表达尚未见报道 被完全定义；然而，看起来不同的亚型 是通过不同的膜受体介导PSP激活所必需的 类(即G蛋白偶联与酪氨酸激酶偶联)。这个 在本修订版竞争中提出的研究的总体目标 继续应用是检验假设，即其中一个(或两个) 在子宫肌层中表达的PLC-γ亚型中的 对酪氨酸激活剂的反应产生时相收缩 激酶(TK)，包括凝血酶和血小板活化因子(PAF)。 具体目的是检验以下假设：1) 过钒酸盐(一种酪氨酸磷酸酶抑制剂)可增加PLC- 伽马磷酸化，肌醇磷酸(IP)生产， CCOs的产生和子宫肌层的阶段性收缩，2) 凝血酶引起的子宫肌层收缩是由TK介导的 激活，增加PLC-伽马磷酸化，IP产生，以及 CCOs的产生，3)PAF产生阶段性肌层收缩 TK激活介导的PLC-γ磷酸化增强，IP 生产和CCO的生成以及4)其中一个(或两个) PLC-伽马异构体是过钒酸盐、凝血酶和PAF所必需的 刺激激活PSP和胞内钙的产生 转瞬即逝。后一项研究将利用反义技术进行。 PLC-Gamma1和PLC-Gamma2与原发子宫的寡核苷酸 心肌细胞培养。这些拟议的研究将继续改善我们的 对子宫肌层潜在分子机制的理解 宫缩，并最终提高我们更有效地 治疗临床上重要的子宫收缩活动障碍， 尤其是早产，这在很大程度上导致了 美国的早产率过高。

Studies during the initial funding period have demonstrated that activation of the phosphatidylinositol signaling pathway (PSP) results in cytosolic calcium oscillation (CCO) driven phasic myometrial contractions. The repetitive cytosolic calcium transients are produced by the release of stored intracellular calcium, along with the influx of extracellular calcium. Additional studies have demonstrated that several isoforms of the signal transduction proteins involved in the PSP are expressed in rat myometrial tissue, including five isoforms for phospholipase C (PLC). The physiologic significance of the simultaneous expression of these multiple PLC isoforms in myometrial tissue is yet to be completely defined; however, it appears that different isoforms are necessary to mediate PSP activation via different membrane receptor classes (i.e. G-protein coupled v. tyrosine kinase coupled). The overall goal of the studies being proposed in this revised Competing Continuation Application is to test the hypothesis that one (or both) of the PLC-gamma isoforms expressed in myometrium are required for the generation of phasic contractions in response to activators of tyrosine kinase (TK), including thrombin and platelet activating factor (PAF). The specific aims are to test the following hypotheses: 1) that pervanadate (a tyrosine phosphatase inhibitor) produces increased PLC- gamma phosphorylation, inositol phosphate (IP) production, the generation of CCOs, and phasic myometrial contractions, 2) that myometrial contractions in response to thrombin are mediated by TK activation, increased PLC-gamma phosphorylation, IP production, and the generation of CCOs, 3) that PAF produces phasic myometrial contractions mediated by TK activation, increased PLC-gamma phosphorylation, IP production, and the generation of CCOs and 4) that one (or both) of the PLC-gamma isoforms are essential for the pervanadate, thrombin and PAF stimulated activation of the PSP and the generation of cytosolic calcium transients. The latter studies will be performed utilizing antisense oligonucleotides for PLC-gamma1 and PLC-gamma2 with primary uterine myocytes cultures. These proposed studies will continue to improve our understanding of the molecular mechanisms underlying myometrial contractions, and ultimately improve our ability to more effectively treat clinically important disturbances of uterine contractile activity, especially premature labor which contributes substantially to the excessive preterm delivery rate in the U.S.