Analysis of expression and function of vacuolar-type H^+-ATPase at bladder

在包括人类在内的哺乳动物中，存在于膀胱表面细胞的管腔细胞质膜上，分泌质子使膀胱内的尿液酸化。为了研究它们的表达过程，我们建立了小鼠膀胱表面细胞再生的实验模型。在该模型中，用胰蛋白酶处理表皮细胞后3小时，用V-ATP酶蛋白脂基因的cDNA通过北方印迹杂交测量的V-ATP酶mRNA的表达水平显著增加。另一方面，使用三种不同的细胞级分，表层细胞丰富级分，中间和基底细胞丰富级分，和全细胞的蛋白质印迹，揭示了几乎相同的水平的V-ATP酶蛋白质的表达。这些结果表明，V-ATP酶存在于内膜系统的迁移到管腔细胞质膜的膀胱表浅细胞从中间到表浅细胞分化过程中。我们还研究了V-ATP酶在建立粘膜屏障和血-尿屏障中的作用。

Vacuolar-type H^+-ATPases were present in luminal cytoplasmic membrane of bladder superficial cells in mammals including human ; which secrete protons to acidify the urine in the bladder. In order to study their expression process, we have developed an experimental model for regeneration of superficial cells of mouse bladder. In this model, the expression level of V-ATPase mRNA,which was measured by Northern blot hybridization using cDNA of V-ATPase proteolipid gene, increased significantly 3 hours after exfoliation of superficial cells by trypsin treatment. On the other hand, western blotting using three different cells fractions, superficial cells rich fraction, intermediate and basal cells rich fraction, and whole cells, revealed almost same levels of V-ATPase protein expression. These results suggested that V-ATPases present in endomembrane systems moved into luminal cytoplasmic membrane of bladder superficial cells during cell differentiation from intermediate to superficial cells. We also examined the role of V-ATPase in establishing mucosal barrier against infection as well as blood-urine barrier.