Kimura M, Ogihara M
Biochemical Pharmacology Group, Faculty of Pharmaceutical Sciences, Josai University, Keyakidai Sakado, Saitama, Japan.
Eur J Pharmacol. 1997 May 26;327(1):87-95. doi: 10.1016/s0014-2999(97)89682-3.
We investigated whether or not insulin and cAMP-elevating agents induce the proliferation of adult rat hepatocytes during the early and late phases of primary culture. Adult rat hepatocytes synthesized a significant amount of DNA when cultured in the presence of 10(-7) M insulin for 3 h. Under these conditions, the number of nuclei increased within 4 h. Hepatocyte DNA synthesis and proliferation were not essentially affected by the initial plating densities. Other cAMP-elevating agents, such as glucagon, forskolin and dibutyryl cAMP, as well as beta-adrenoceptor agonists (i.e., metaproterenol and isoproterenol) alone had no effect on either hepatocyte DNA synthesis or proliferation in primary culture. In contrast, these agents potentiated both processes at concentrations as low as 10(-7) M when cultured in combination with 10(-7) M insulin. The stimulatory effects of beta-adrenoceptor agonists and other cAMP-elevating agents were significantly blocked by the cAMP-dependent protein kinase inhibitor, H-89 (N-[2-(p-(bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride; 10(-7) M). The mitogenic effect of insulin upon hepatocytes was almost completely suppressed by genistein (5 x 10(-6) M), wortmannin (10(-7) M) and by rapamycin (10 ng/ml). These results show that insulin rapidly induced the proliferation of adult rat hepatocytes in primary culture. The mitogenic effects of insulin were potentiated by beta-adrenoceptor agonists and cAMP-elevating agents. The effects of beta-adrenoceptor agonists and cAMP-elevating agents may be mediated through cAMP-dependent protein kinase. In addition, the activation of receptor tyrosine kinase, phosphoinositide 3-kinase and p70 ribosomal protein S6 kinase may be involved in the insulin signal transduction pathway.
我们研究了胰岛素和cAMP升高剂在原代培养的早期和晚期是否能诱导成年大鼠肝细胞增殖。成年大鼠肝细胞在含有10(-7)M胰岛素的培养基中培养3小时后，合成了大量DNA。在此条件下，4小时内核数量增加。肝细胞DNA合成和增殖基本上不受初始接种密度的影响。其他cAMP升高剂，如胰高血糖素、福斯可林和二丁酰cAMP，以及β-肾上腺素能受体激动剂（即间羟异丙肾上腺素和异丙肾上腺素）单独对原代培养中的肝细胞DNA合成或增殖均无影响。相反，当与10(-7)M胰岛素联合培养时，这些试剂在低至10(-7)M的浓度下能增强这两个过程。β-肾上腺素能受体激动剂和其他cAMP升高剂的刺激作用被cAMP依赖性蛋白激酶抑制剂H-89（N-[2-(对-(溴肉桂酰胺基)乙基]-5-异喹啉磺酰胺二盐酸盐；10(-7)M）显著阻断。胰岛素对肝细胞的促有丝分裂作用几乎完全被染料木黄酮（5×10(-6)M）、渥曼青霉素（10(-7)M）和雷帕霉素（10ng/ml）抑制。这些结果表明，胰岛素能在原代培养中迅速诱导成年大鼠肝细胞增殖。β-肾上腺素能受体激动剂和cAMP升高剂能增强胰岛素的促有丝分裂作用。β-肾上腺素能受体激动剂和cAMP升高剂的作用可能通过cAMP依赖性蛋白激酶介导。此外，受体酪氨酸激酶、磷酸肌醇3-激酶和p70核糖体蛋白S6激酶的激活可能参与胰岛素信号转导途径。
