Regulation of paracellular permeability in Caco-2 cell monolayers by protein kinase C.
Regulation of paracellular permeability in Caco-2 cell monolayers by protein kinase C.
复制标题
蛋白激酶 C 对 Caco-2 细胞单层细胞旁通透性的调节。
DOI：
10.1152/ajpgi.1993.265.5.g955
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Turk,J
中科院分区：
文献类型：
--
作者：
Stenson,WF;Easom,RA;Riehl,TE;Turk,J
Caco-2细胞是一种来源于人类结肠腺癌的肠上皮样细胞系。通过跨单层阻力和[~3H]甘露醇通过单层的渗透来评估这些细胞单层的细胞旁通透性。蛋白激酶C(PKC)激动剂佛波酯(50 NM)、卡巴胆碱(500微米)以及卡巴胆碱(50微米)和甘露醇二酯酶抑制剂单甘油酯(100微米)的联合应用增加了细胞旁通透性，表现为跨单层阻力减少和甘露醇渗透率增加。所有这些刺激对跨单层耐药的影响都被PKC抑制剂星形孢子素(3 NM)所抑制。氨基甲胆碱+单油酯的作用也可被M受体拮抗剂阿托品(0.1微米)所抑制。用每种刺激物处理单层都与PKC活性从胞浆转移到膜相关状态有关。佛波醇肉豆蔻酸醋酸酯或卡巴胆碱和单油脂联合刺激Caco-2细胞单层也与PKC的内源性底物Marcks蛋白的磷酸化有关。这些数据支持肠细胞旁通透性受肠细胞PKC活性调节的假说，并证明卡巴胆碱与M受体结合引起的细胞旁通透性增加是通过激活PKC介导的。
Caco-2 cells are an enterocyte-like cell line derived from a human colonic adenocarcinoma. Paracellular permeability was assessed in monolayers of these cells by transmonolayer resistance and by the permeation of [3H]mannitol across the monolayer. Paracellular permeability was increased by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (50 nM), carbachol (500 microM), and the combination of carbachol (50 microM) and monolein (100 microM), an inhibitor of diacylglycerol kinase, as manifested by a decrease in transmonolayer resistance and an increase in mannitol permeation. The effects of all of these stimuli on transmonolayer resistance were inhibited by staurosporine (3 nM), an inhibitor of PKC. The effects of carbachol plus monolein were also inhibited by atropine (0.1 microM), a muscarinic antagonist. Treatment of the monolayers with each of the stimuli was associated with translocation of PKC activity from cytosol to a membrane-associated state. Stimulation of Caco-2 cell monolayers with phorbol myristate acetate or with the combination of carbachol and monolein was also associated with phosphorylation of the MARCKS protein, an endogenous substrate of PKC. These data support the hypothesis that intestinal paracellular permeability is regulated by the activity of enterocyte PKC and demonstrate that the increase in paracellular permeability induced by binding of carbachol to the muscarinic receptor is mediated by activation of PKC.