PI KINASES, CANALICULAR TRANSPORTERS AND CHOLESTASIS

细胞内胆汁淤积和调节ATP依赖的胆管转运体SPGP(牛磺胆酸盐)、mrp2(非胆汁酸有机阴离子)、mdr1(有机阳离子)、mdr3(磷脂酰胆碱转位酶)和新描述的mrp3(甘胆酸盐)的机制尚不清楚。需要一个完整的微管网络和囊泡运输，起始于跨高尔基网络，结束于胆小管膜。两者都与PI3-激酶活性有关，并被PI3-激酶抑制剂Wortmannin和LY294002阻断。完整细胞中PI3-激酶的激活加速了胆汁酸的分泌。我们认为，PI-3-激酶脂类产物对依赖于ATP的小管转运蛋白的囊泡运输及其在小管膜上的活性是必不可少的。我们还假设，在某些形式的胆汁淤积中，这些磷脂的生成是有缺陷的。利用生化、分子和共聚焦荧光显微镜技术以及新的试剂和实验模型，包括基因工程P85-/-PI3K小鼠，将确定小管转运蛋白的特异性运输以及PI3K1及其脂质产生者在胆汁分泌中的作用。这些研究应该能阐明胆汁淤积的细胞内机制。此外，激活特定的PI3K功能可能选择性地增强胆小管的转运，在胆汁淤积症中具有治疗潜力。

Mechanisms responsible for intracellular cholestasis and regulation of intracellular trafficking of the ATP-dependent canalicular transporters spgp (taurocholate), mrp2 (nonbile acid organic anions), mdr1 (organic cations), mdr3 (phosphatidylcholine translocase) and newly described mrp3 (glycocholate) are poorly understood. An intact microtubular network and vesicular trafficking originating in the trans-Golgi network and concluding in the bile canalicular membrane are required. Both are associated with PI 3-kinase activity and are blocked by Wortmannin and LY294002, which are PI 3-kinase inhibitors. Activation of PI 3- kinase in intact cells accelerates bile acid secretion. We propose that PI 3-kinase lipid products are essential for vesicular trafficking of the ATP-dependent canalicular transporters as well as for their activity in the canalicular membrane. We also postulate that generation of these phospholipids is defective in certain forms of cholestasis. Using biochemical, molecular and confocal fluorescence microscopic techniques and novel reagents and experimental models, including genetically engineered p85-/- PI 3- kinase mice, the specific trafficking of canalicular transporters and the role of PI 3-kinase and its lipid producers in bile secretion will be determined. These studies should elucidate intracellular mechanisms of cholestasis. Furthermore, activation of specific PI 3-kinase functions may selectively enhance canalicular transport and have therapeutic potential in cholestasis.