PKC{epsilon}-dependent and -independent effects of taurolithocholate on PI3K/PKB pathway and taurocholate uptake in HuH-NTCP cell line.
PKC{epsilon}-dependent and -independent effects of taurolithocholate on PI3K/PKB pathway and taurocholate uptake in HuH-NTCP cell line.
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牛磺石胆酸盐对 HuH-NTCP 细胞系中 PI3K/PKB 途径和牛磺胆酸盐摄取的 PKC{epsilon} 依赖性和非依赖性影响。
DOI：
10.1152/ajpgi.00177.2009
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发表时间：
2009
期刊：
影响因子：
--
通讯作者：
Anwer,MSawkat
中科院分区：
文献类型：
--
作者：
Schonhoff,ChristopherM;Yamazaki,Ai;Hohenester,Simon;Webster,CynthiaRL;Bouscarel,Bernard;Anwer,MSawkat
抑胆胆汁酸牛磺酸胆酸酯（TLC）抑制胆汁中有机阴离子的分泌和牛磺酸胆酸酯（TC）的肝脏摄取。TLC被认为可以通过大鼠肝细胞中磷酸肌苷-3激酶(PI3K)/ pkb依赖的新型蛋白激酶Cε (nPKCε)激活诱导Mrp2从管膜中恢复。本研究的目的是确定tlc诱导的TC摄取抑制是否也可能涉及稳定转染人Na+依赖性TC共转运多肽（NTCP）的HuH7细胞（HuH-NTCP细胞）中pi3k依赖性PKCε的激活。为了避免直接竞争摄取，用TLC对细胞进行预处理，清洗，然后用3h -TC孵育以测定TC的摄取。薄层色谱对TC的摄取有时间和剂量依赖性的抑制作用。TLC在不影响NTCP膜易位的情况下竞争性地抑制TC摄取。一种PI3K抑制剂未能逆转tlc诱导的TC摄取抑制和tlc抑制的PKB磷酸化。TLC确实激活了nPKCε，这可以通过增加膜易位和nPKCε- ser729磷酸化来证明。显性负npkc ε的过表达逆转了tlc诱导的PKB磷酸化抑制，但不影响TC摄取。最后，cAMP通过PI3K途径阻止了tlc诱导的TC摄取抑制，这种阻止是由于cAMP诱导的刺激和tlc诱导的TC摄取抑制的总和。综上所述，这些结果表明tlc诱导的PKB抑制，而不是TC摄取，是通过nPKCε介导的。nPKCε的激活和TLC对TC摄取的抑制不是通过PI3K/PKB途径介导的。
The cholestatic bile acid taurolithocholate (TLC) inhibits biliary secretion of organic anions and hepatic uptake of taurocholate (TC). TLC has been suggested to induce retrieval of Mrp2 from the canalicular membrane via the phosphoinositide-3-kinase (PI3K)/PKB-dependent activation of novel protein kinase Cε (nPKCε) in rat hepatocytes. The aim of the present study was to determine whether TLC-induced inhibition of TC uptake may also involve PI3K-dependent activation of PKCε in HuH7 cells stably transfected with human Na+-dependent TC-cotransporting polypeptide (NTCP) (HuH-NTCP cells). To avoid direct competition for uptake, cells were pretreated with TLC, washed, and then incubated with3H-TC to determine TC uptake. TLC produced time- and dose-dependent inhibition of TC uptake. TLC inhibited TC uptake competitively without affecting NTCP membrane translocation. A PI3K inhibitor failed to reverse TLC-induced TC uptake inhibition and TLC-inhibited PKB phosphorylation. TLC did activate nPKCε as evidenced by increased membrane translocation and nPKCε-Ser729phosphorylation. Overexpression of dominant negative-nPKCε reversed TLC-induced inhibition of PKB phosphorylation but not of TC uptake. Finally, cAMP prevented TLC-induced inhibition of TC uptake via the PI3K pathway, and the prevention is due to the sum of cAMP-induced stimulation and TLC-induced inhibition of TC uptake. Taken together, these results suggest that TLC-induced inhibition of PKB, but not of TC uptake, is mediated via nPKCε. Activation of nPKCε and inhibition of TC uptake by TLC are not mediated via the PI3K/PKB pathway.