REGULATION OF LACTOSYLCERAMIDE SYNTHESIS IN RENAL CELLS

我们发现在正常的人肾近端小管(PT)细胞中， 血浆低密度脂蛋白(LDL)具有浓度依赖性 乳糖基神经酰胺(LacCer)和酶的水平降低 参与LacCer的生物合成，即UDP-半乳糖：糖基神经酰胺 半乳糖基转移酶(GALT-2)。然而，进入的甲基低密度脂蛋白 细胞通过不依赖低密度脂蛋白受体的途径刺激该活性。 在正常甲状旁腺细胞中，GALT-2的表达约为对照组的三倍。相比之下，在 家族性高胆固醇血症患者和正常人的PT细胞 肾肿瘤PT细胞无低密度脂蛋白受体，低密度脂蛋白增加 GalT-2的活性大约是它的三倍。这导致了15倍的 FH-PT细胞中LacCer的细胞水平升高及其合成 在肿瘤PT细胞中发现了更复杂的神经鞘糖脂。我们已经净化了 并从人肾中部分鉴定了GalT-2。产生的抗体 特异性免疫沉淀法从培养物中提取GalT-2 正常的PT细胞。我们的总体目标是：1)追求分子 GALT-2的特性/表达及研究低密度脂蛋白的作用 M-低密度脂蛋白对甲状旁腺细胞mRNA水平和转化率的影响；2)测定 GALT-2在正常PT、FH-PT和肿瘤PT细胞中的细胞定位； 3)研究低密度脂蛋白及其修饰物的作用及其机制 低密度脂蛋白(M-LDL)对PT细胞GALT-2水平的影响。 假设。合成(生产过剩)和储藏 FH-PT细胞中的乳糖基神经酰胺是由于缺乏GALT-1调节所致 2个是低密度脂蛋白。有人建议进行研究，以阐明生化 低密度脂蛋白和M-低密度脂蛋白对GALT-2的作用机制及其调控 分子水平。这种研究提供了关于以下方面的基础知识 正常甲状腺细胞中LacCer合成是如何调节的 低密度脂蛋白介导的FH-PT和FH-PT中GALT-2调节缺失的病理生理机制 人肿瘤中的PT细胞。

We have found that in normal human kidney proximal tubular (PT) cells, plasma low density lipoproteins (LDL) exerts a concentration dependent decrease in the level of lactosylceramide (LacCer) and the enzyme involved in LacCer biosynthesis, i.e., UDP-galactose: gluco-sylceramide B1->4 galactosyltransferase (GalT-2). However, methyl-LDL which enters cells through a LDL receptor independent pathway stimulated the activity of GalT-2 approximately three fold in normal PT cells. In contrast, in PT cells from patients with familial hypercholesterolemia (FH), and human kidney tumor PT cells which do not have LDL receptors, LDL increased the activity of GalT-2 approximately three fold. This resulted in a 15 fold increase in the cellular level of LacCer in FH PT cells and the synthesis of more complex glycosphingolipids in tumor PT cells. We have purified and partially characterized GalT-2 from human kidney. Antibodies raised against this enzyme specifically immunoprecipitate GalT-2 from cultured normal PT cells. Our overall objects are: 1) to pursue molecular characterization/expression of GalT-2 and to study the effect of LDL, and M-LDL on the levels and the turnover of mRNA in PT cells; 2) to determine the cytolocalization of GalT-2 in normal PT, FH-PT and tumor PT cells; and 3) to study the effects and mechanisms of action of LDL and modified LDL (M-LDL) on the level of GalT-2 in PT cells. Hypothesis. The synthesis (overproduction) and storage of lactosylceramide in FH-PT cells is due to the lack of regulation of GalT- 2 by LDL. Studies are proposed that will elucidate biochemical mechanisms of action of LDL and M-LDL on GalT-2 and its regulation at the molecular level. such studies provide fundamental knowledge regarding how LacCer synthesis is regulated in normal PT cells and the pathophysiology of LDL mediated lack of regulation of GalT-2 in FH-PT and tumor PT cells in man.