Kubitz R, D'urso D, Keppler D, Häussinger D
Medizinische Universitätsklinik, Heinrich Heine Universität, Düsseldorf, Germany.
Gastroenterology. 1997 Nov;113(5):1438-42. doi: 10.1053/gast.1997.v113.pm9352844.
BACKGROUND & AIMS: Circumstantial evidence suggests a regulation of biliary secretion by transporter insertion and retrieval into and from the canalicular membrane. This study was undertaken to provide direct evidence for such a process.
Osmosensitivity of the subcellular localization of the mrp2 gene-encoded conjugate export pump (MRP2) was studied by immunofluorescence and confocal laser scanning microscopy of isolated hepatocyte aggregates and in perfused rat liver.
MRP2 was localized largely in membranes of the pseudocanaliculi formed by isolated hepatocyte aggregates during hypo-osmotic exposure, whereas after hyperosmotic exposure MRP2 was also detectable in intracellular vesicles. In perfused liver, the EAG15 antibody specific for rat MRP2 and the ZO-1 antibody specific for tight junctions produced immunostaining of the canalicular membrane. However, the relative amount of MRP2 increased significantly in the pericanalicular region with increasing perfusate osmolarity, as shown by confocal microscopy of intracellular vesicles containing MRP2 (but not ZO-1) and by computed densitometry. The osmodependent distribution of MRP2 between the canalicular membrane and intracellular, pericanalicular vesicles occurred within 30 minutes and was fully reversible.
The findings provide direct evidence for an osmosensitive dynamic insertion and retrieval of the canalicular MRP2 transporter into and out of the canalicular membrane.
间接证据表明，通过转运体插入胆小管膜及从该膜上回收可调节胆汁分泌。本研究旨在为这一过程提供直接证据。
通过对分离的肝细胞聚集体以及灌注大鼠肝脏进行免疫荧光和共聚焦激光扫描显微镜检查，研究mrp2基因编码的结合物输出泵（MRP2）亚细胞定位的渗透压敏感性。
在低渗暴露期间，MRP2主要定位于分离的肝细胞聚集体形成的假胆小管膜中，而在高渗暴露后，在细胞内囊泡中也可检测到MRP2。在灌注肝脏中，对大鼠MRP2特异的EAG15抗体和对紧密连接特异的ZO-1抗体可使胆小管膜产生免疫染色。然而，如通过对含有MRP2（而非ZO-1）的细胞内囊泡进行共聚焦显微镜检查和计算机密度测定所示，随着灌注液渗透压升高，MRP2在胆小管周围区域的相对含量显著增加。MRP2在胆小管膜与细胞内胆小管周围囊泡之间的渗透压依赖性分布在30分钟内发生，且完全可逆。
这些发现为胆小管MRP2转运体对渗透压敏感的动态插入和从胆小管膜上回收提供了直接证据。
