Cyclic adenosine monophosphate response in primary and subcultured bladder epithelial cells: inhibition by 12-O-tetradecanoylphorbol-13-acetate.
Cyclic adenosine monophosphate response in primary and subcultured bladder epithelial cells: inhibition by 12-O-tetradecanoylphorbol-13-acetate.
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原代和传代培养的膀胱上皮细胞中的环状单磷酸腺苷反应：12-O-十四烷酰佛波醇-13-乙酸酯的抑制。
DOI：
10.1016/0026-0495(93)90077-2
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Davis,BB
中科院分区：
文献类型：
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作者：
Thomas,DJ;Zenser,TV;Davis,BB
原代和首次传代的犬尿路上皮细胞(分别为DUC和DUC-P1)显示了环磷酸腺苷(CAMP)第二信使系统的活性催化亚基。在加入Forsklin后，cAMP水平显著增加，这引起cAMP水平随时间和剂量的增加而增加。细胞内cAMP水平的升高先于环核苷酸水平的中等水平的增加，并在被检查的最早时间(5分钟)观察到。Forsklin的最低有效浓度为1~10μ/L，细胞和培养上清液中cAMP水平升高达20~100倍。原代和传代细胞与0.1Mol/L，12-O-十四酰佛波醇-13-醋酸酯(TPA)预孵育60分钟，可降低Forsklin引起的cAMP水平升高的幅度。为了确定TPA在原代培养中的作用机制，使用了下列试剂：1μ摩尔/L星形孢子素和25μ摩尔/L鞘氨醇，35μ摩尔/L环己亚胺，3μ摩尔/L环己亚胺，3.0μ摩尔/L吲哚美辛，3μ/L，环核苷酸磷酸二酯酶抑制剂。星状孢子素和鞘氨醇是唯一能阻止TPA作用的药物。用0.2nmoL/L表皮生长因子、0.1nmo1/L-4μ-α(TPA的立体异构体)或1.0nmo1/L A23187评价TPA作用的特异性。与TPA相反，这些药物都没有减少Forsklin介导的cAMP增加。结果表明，在原代和传代细胞中，Forsklin对cAMP的反应性以及TPA对这种反应的调节。这种TPA效应似乎涉及蛋白激酶C，并且是TPA所特有的。因此，两个主要的细胞内信号通路(cAMP和磷脂酰肌醇[PI]周期)之间的串扰似乎存在于尿路上皮细胞中。
Primary and first-passage dog urothelial cells (DUC and DUC-P1, respectively) exhibited an active catalytic subunit for the cyclic adenosine monophosphate (cAMP) second messenger system. Dramatic increases in cAMP levels were observed following the addition of forskolin, which elicited a time- and dose-response-dependent increase in cAMP levels. Increases in intracellular cAMP levels preceded media increases in cyclic nucleotide levels and were observed at the earliest time examined (5 minutes). The lowest effective concentration of forskolin was between 1 and 10 μmol/L. cAMP level increases as large as 20- to 100-fold were observed in cells and media. Preincubation of primary and subcultured cells with 0.1 μmol/L 12-O-tetradecanoylphorbol-13-acetate (TPA) for 60 minutes reduced the magnitude of the forskolin-induced increase in cAMP levels. To determine the mechanism by which TPA elicits its effect in primary cultures, the following test agents were used: 1.0 μmol/L staurosporine and 25 μmol/L sphingosine, protein kinase C inhibitors; 35 μmol/L cycloheximide, a protein synthesis inhibitor; 3.0 μmol/L indomethacin, an inhibitor of prostaglandin synthesis; and 0.5 mmol/L RO-20-1724, a cyclic nucleotide phosphodiesterase inhibitor. Staurosporine and sphingosine were the only agents that prevented the effect of TPA. The specificity of the TPA effect was evaluated with the following test agents: 0.2 nmol/L epidermal growth factor (EGF), 0.1 μmol/L 4α-TPA (a stereoisomer of TPA), or 1.0 μmol/L A23187. In contrast to TPA, none of these agents reduced forskolin-mediated increases in cAMP. Results indicate forskolin cAMP responsiveness and regulation of this response by TPA in both primary and subcultured cells. This TPA effect appears to involve protein kinase C and is specific for TPA. Thus, cross-talk between two major intracellular signaling pathways (cAMP and phosphatidyl inosital [PI] cycle) appears to exist in urothelial cells.