Regulation of glycogen metabolism in primary cultures of rat hepatocytes. Restoration of acute effects of insulin and glucose in cells from diabetic rats.
Regulation of glycogen metabolism in primary cultures of rat hepatocytes. Restoration of acute effects of insulin and glucose in cells from diabetic rats.
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发表时间：
1986-01
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通讯作者：
T. B. Miller;A. Garnache;J. Cruz;R. McPherson;C. Wolleben
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作者：
T. B. Miller;A. Garnache;J. Cruz;R. McPherson;C. Wolleben
严重胰岛素缺乏大鼠肝脏中糖原沉积和糖原合成调节的缺陷可以在体内胰岛素给药后数小时内逆转。使用原代培养的肝细胞分离正常和糖尿病大鼠在无血清的化学成分确定的培养基中，本研究解决了胰岛素的慢性作用，以促进胰岛素和葡萄糖的直接影响的短期调控的酶控制糖原代谢。原代培养物在胰岛素、三碘甲腺原氨酸和皮质醇存在下维持1-3天。在第1天，在四氧嘧啶糖尿病培养物中，10（-7）M胰岛素在15分钟或1小时内不能急性激活糖原合成酶，而在正常肝细胞培养物中胰岛素急性激活合成酶。到第3天，在从四氧嘧啶糖尿病大鼠分离的肝细胞中，胰岛素在15分钟内使合酶I的百分比增加约30%，正常细胞也是如此。胰岛素对合成酶激活的急性效应与磷酸化酶α的变化无关。而糖原合成酶磷酸酶活性不能被证明是急性胰岛素的影响，在糖尿病细胞的总活性恢复到正常对照值在3天的培养期。30 mM葡萄糖激活正常大鼠培养肝细胞中糖原合酶的急性效应在培养1天后在从四氧嘧啶或自发性糖尿病（BB/W）大鼠分离的肝细胞中缺失。培养3天后，在相同条件下，葡萄糖在10分钟内使糖原合成酶活性增加50%。这些研究清楚地表明，胰岛素以慢性方式与甲状腺激素和类固醇协同作用，以促进胰岛素和葡萄糖对肝糖原合成的急性调节。
Defects in the deposition of glycogen and the regulation of glycogen synthesis in the livers of severely insulin-deficient rats can be reversed, in vivo, within hours of insulin administration. Using primary cultures of hepatocytes isolated from normal and diabetic rats in a serum-free chemically defined medium, the present study addresses the chronic action of insulin to facilitate the direct effects of insulin and glucose on the short term regulation of the enzymes controlling glycogen metabolism. Primary cultures were maintained in the presence of insulin, triiodothyronine, and cortisol for 1-3 days. On day 1 in alloxan diabetic cultures, 10(-7) M insulin did not acutely activate glycogen synthase over a period of 15 min or 1 h, whereas insulin acutely activated synthase in cultures of normal hepatocytes. By day 3 in hepatocytes isolated from alloxan diabetic rats, insulin effected an approximate 30% increase in per cent synthase I within 15 min as was also the case for normal cells. The acute effect of insulin on synthase activation was independent of changes in phosphorylase alpha. Whereas glycogen synthase phosphatase activity could not be shown to be acutely affected by insulin, the total activity in diabetic cells was restored to normal control values over the 3-day culture period. The acute effect of 30 mM glucose to activate glycogen synthase in cultured hepatocytes from normal rats after 1 day of culture was missing in hepatocytes isolated from either alloxan or spontaneously diabetic (BB/W) rats. After 3 days in culture, glucose produced a 50% increase in glycogen synthase activity during a 10-min period under the same conditions. These studies clearly demonstrate that insulin acts in a chronic manner in concert with thyroid hormones and steroids to facilitate acute regulation of hepatic glycogen synthesis by both insulin and glucose.