Activity of the Bile Salt Export Pump (ABCB11) Is Critically Dependent on Canalicular Membrane Cholesterol Content
Activity of the Bile Salt Export Pump (ABCB11) Is Critically Dependent on Canalicular Membrane Cholesterol Content
复制标题
DOI：
10.1074/jbc.m808667200
复制
发表时间：
2009-04-10
影响因子：
4.8
通讯作者：
Elferink, Ronald P. J. Oude
中科院分区：
文献类型：
--
作者：
Paulusma, Coen C.;de Waart, D. Rudi;Elferink, Ronald P. J. Oude
ATP8B1突变导致严重的遗传性肝病。该疾病的特征是胆汁盐排泄受损（胆汁淤积），但ATP8B1功能受损导致胆汁淤积的机制尚不清楚。ATP8B1是一种4型p型atp酶，是磷脂酰丝氨酸的翻转酶。atp8b1缺陷小鼠显示胆从肝细胞管状（顶端）膜提取胆固醇的显著增加。在这里，我们研究了一种假设，即从管膜中提取不成比例的胆固醇会损害胆盐转运体ABCB11的活性，从而导致胆汁淤积。通过单次肝灌注，我们发现Atp8b1缺乏不仅abcb11介导的转运，而且abcc2介导的转运至少减少了4倍。我们发现，胆固醇抑制型atp8b1缺陷小鼠的小管膜具有显著降低的胆固醇/磷脂比率，即0.75 +/- 0.24，而野生型为2.03 +/- 0.71。用甲基- β -环糊精体外去除小鼠肝质膜上的胆固醇，证明了膜上胆固醇含量与atp依赖的牛磺胆酸运输之间的近似线性关系。abcc2介导的运输活性在膜胆固醇消耗的30%时不受影响，但在膜胆固醇消耗的70%时下降到可以忽略不计的水平。这些影响是可逆的，因为肝膜的胆固醇充盈完全恢复了Abcb11-和abcc2介导的转运。我们的数据表明，膜胆固醇含量是ABCB11/ABCC2转运活性的关键决定因素，为ATP8B1疾病的病因提供了解释，并提出了一种保护管膜免受管腔胆盐过载的新机制。
Mutations in ATP8B1 cause severe inherited liver disease. The disease is characterized by impaired biliary bile salt excretion (cholestasis), but the mechanism whereby impaired ATP8B1 function results in cholestasis is poorly understood. ATP8B1 is a type 4 P-type ATPase and is a flippase for phosphatidylserine. Atp8b1-deficient mice display a dramatic increase in the biliary extraction of cholesterol from the canalicular ( apical) membrane of the hepatocyte. Here we studied the hypothesis that disproportionate cholesterol extraction from the canalicular membrane impairs the activity of the bile salt transporter, ABCB11, and as a consequence causes cholestasis. Using single pass liver perfusions, we show that not only ABCB11-mediated transport but also Abcc2-mediated transport were reduced at least 4-fold in Atp8b1 deficiency. Weshow that canalicular membranes of cholestatic Atp8b1-deficient mice have a dramatically reduced cholesterol to phospholipid ratio, i.e. 0.75 +/- 0.24 versus 2.03 +/- 0.71 for wild type. In vitro depletion of cholesterol from mouse liver plasma membranes using methyl-beta-cyclodextrin demonstrated a near linear relation between cholesterol content of the membranes and ATP-dependent taurocholate transport. Abcc2-mediated transport activity was not affected up to 30% of membrane cholesterol depletion but declined to negligible levels at 70% of membrane cholesterol depletion. These effects were reversible as cholesterol repletion of the liver membranes completely restored Abcb11- and Abcc2-mediated transport. Our data demonstrate that membrane cholesterol content is a critical determinant of ABCB11/ABCC2 transport activity, provide an explanation for the etiology of ATP8B1 disease, and suggest a novel mechanism protecting the canalicular membrane against luminal bile salt overload.