Oxidative stress and Mrp2 internalization.
Oxidative stress and Mrp2 internalization.
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DOI：
10.1016/j.freeradbiomed.2006.02.015
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发表时间：
2006-06
影响因子：
7.4
通讯作者：
Shuichi Sekine;Kousei Ito;T. Horie
中科院分区：
文献类型：
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作者：
Shuichi Sekine;Kousei Ito;T. Horie
肝脏的氧化应激有时伴有胆汁淤积。我们描述了在乙酸（EA）诱导的大鼠肝脏急性氧化应激下，多药耐药相关蛋白2/ atp结合盒转运蛋白家族2 （Mrp2/Abcc2）的内化，这是一种胆道转运蛋白，参与不依赖胆盐的胆汁流动。然而，信号通路和调控分子尚未被研究。在本研究中，我们利用分离的大鼠肝细胞偶联（IRCHs）研究了ea诱导Mrp2内化的机制。Mrp2指数，定义为IRCHs中Mrp2阳性管膜染色与细胞核数量的比值，通过EA处理显着降低。这种降低被非特异性蛋白激酶C （PKC）抑制剂Gö6850， Ca2+螯合剂EGTA消除，但不被蛋白激酶a （PKA）选择性抑制剂，Ca2+依赖性常规PKC （cPKC）抑制剂Gö6976或蛋白激酶G （PKG）抑制剂（1 μM）消除。此外，在EA处理后不久，观察到细胞内Ca2+水平和NO释放到培养基中。这两种增加以及Mrp2的内化都被EGTA完全阻断。综上所述，EA导致GSH、Ca2+升高、NO产生和nPKC激活的降低，最终导致Mrp2内化。
Oxidative stress in the liver is sometimes accompanied by cholestasis. We have described the internalization of multidrug resistance-associated protein 2/ATP-binding cassette transporter family 2 (Mrp2/Abcc2), a biliary transporter involved in bile-salt-independent bile flow, under ethacrynic acid (EA)-induced acute oxidative stress in rat liver. However, the signaling pathway and regulatory molecules have not been investigated. In the present study, we investigated the mechanism of EA-induced Mrp2 internalization using isolated rat hepatocyte couplets (IRCHs). The Mrp2 index, defined as the ratio of Mrp2-positive canalicular membrane staining in IRCHs per number of cell nuclei, was significantly reduced by treatment with EA. This reduction was abolished by a nonspecific protein kinase C (PKC) inhibitor Gö6850, a Ca2+chelator, EGTA, but not by a protein kinase A (PKA)-selective inhibitor, a Ca2+-dependent conventional PKC (cPKC) inhibitor Gö6976, or a protein kinase G (PKG) inhibitor (1 μM). Moreover, an increase in the intracellular Ca2+level and NO release into medium were observed shortly after the EA treatment. Both of these increases, as well as Mrp2 internalization, were completely blocked by EGTA. In conclusion, EA produced a reduction in GSH, Ca2+elevation, NO production, and nPKC activation in a sequential manner, finally leading to Mrp2 internalization.