Vesicular exocytosis contributes to volume-sensitive ATP release in biliary cells
Vesicular exocytosis contributes to volume-sensitive ATP release in biliary cells
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DOI：
10.1152/ajpgi.00355.2003
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发表时间：
2004-04-01
影响因子：
4.5
通讯作者：
Fitz, JG
中科院分区：
文献类型：
--
作者：
Gatof, D;Kilic, G;Fitz, JG
细胞外ATP是一种有效的自分泌/旁分泌信号，通过激活嘌呤能受体调节广泛的肝脏功能。在胆管上皮中，细胞体积的增加通过磷酸肌醇3-激酶（PI 3-激酶）依赖性机制刺激ATP释放。由于PI 3-激酶也调节囊泡胞吐，这些研究的目的是确定体积刺激的囊泡胞吐是否有助于细胞ATP释放。在人胆管癌细胞系中，通过使用质膜标记物FM 1 -43测量胞吐作用，而通过使用腺苷酸酶-ATP测定评估ATP释放。在基础条件下，胆管细胞表现出组成性胞吐率为1.6%/min，和低水平的细胞外ATP检测到在48.2任意光单位。低渗暴露诱导的胆管细胞体积增加导致胞吐率增加10倍，ATP释放增加35倍。囊泡胞吐和ATP释放均与细胞体积成正比，并且均表现出相似的调节特性，包括：1）依赖于完整的PI 3-激酶，2）通过抑制PKC而减弱，以及3）在低渗暴露前通过激活PKC而增强。这些发现表明，胆管细胞体积的增加刺激ATP释放和囊泡胞吐通过类似的调节模式。细胞体积、PKC和PI 3-激酶之间的功能相互作用调节胞吐作用，从而调节胆管细胞中ATP释放和嘌呤能信号传导。据推测，蛋白激酶C是参与招聘的体积敏感的囊泡池容易释放的状态。
Extracellular ATP is a potent autocrine/paracrine signal that regulates a broad range of liver functions through activation of purinergic receptors. In biliary epithelium, increases in cell volume stimulate ATP release through a phosphoinositide 3-kinase (PI3-kinase)-dependent mechanism. Because PI3-kinase also regulates vesicular exocytosis, the purpose of these studies was to determine whether volume-stimulated vesicular exocytosis contributes to cellular ATP release. In a human cholangiocarcinoma cell line, exocytosis was measured by using the plasma membrane marker FM1-43, whereas ATP release was assessed by using a luciferase-luciferin assay. Under basal conditions, cholangiocytes exhibited constitutive exocytosis at a rate of 1.6%/min, and low levels of extracellular ATP were detected at 48.2 arbitrary light units. Increases in cholangiocyte cell volume induced by hypotonic exposure resulted in a 10-fold increase in the rate of exocytosis and a robust 35-fold increase in ATP release. Both vesicular exocytosis and ATP release were proportional to cell volume, and both exhibited similar regulatory properties including: 1) dependence on intact PI3-kinase, 2) attenuation by inhibition of PKC, and 3) potentiation by activation of PKC before hypotonic exposure. These findings demonstrate that increases in cholangiocyte cell volume stimulate ATP release and vesicular exocytosis through similar regulatory paradigms. Functional interactions among cell volume, PKC, and PI3-kinase modulate exocytosis, thereby regulating ATP release and purinergic signaling in cholangiocytes. It is hypothesized that PKC is involved in the recruitment of a volume-sensitive vesicular pool to a readily releasable state.