胰岛素和表皮生长因子均可刺激分离的脂肪细胞中的脂肪生成及乙酰辅酶A羧化酶活性。乙酰辅酶A羧化酶测定中匀浆程序对于避免人为误差的重要性。
Both insulin and epidermal growth factor stimulate lipogenesis and acetyl-CoA carboxylase activity in isolated adipocytes. Importance of homogenization procedure in avoiding artefacts in acetyl-CoA carboxylase assay.
作者信息
Haystead T A, Hardie D G
出版信息
Biochem J. 1986 Mar 1;234(2):279-84. doi: 10.1042/bj2340279.
Abstract
Epidermal growth factor (EGF) stimulates lipogenesis by 3-4-fold in isolated adipocytes, with a half-maximal effect at 10 nM-EGF. In the same batches of cells insulin stimulated lipogenesis by 15-fold. Freezing and prolonged homogenization of adipocytes results in release of large quantities of pyruvate carboxylase from broken mitochondria, and sufficient pyruvate can be carried through into assays for this enzyme to cause significant interference with assays of acetyl-CoA carboxylase in crude adipocyte extracts. This may account for the high amount of citrate-independent acetyl-CoA carboxylase activity reported to be present in adipocyte extracts in some previous publications. This problem may be eliminated by homogenizing very briefly without freezing. By using the modified homogenization procedure, EGF treatment of adipocytes was shown to produce an effect on acetyl-CoA carboxylase activity almost identical with that of insulin. Both messengers increase Vmax. without significant effect on the Ka for the allosteric activator, citrate.
摘要
表皮生长因子（EGF）可使分离的脂肪细胞中的脂肪生成增加3至4倍，在10 nM - EGF时达到最大效应的一半。在同一批细胞中，胰岛素可使脂肪生成增加15倍。脂肪细胞的冷冻和长时间匀浆会导致大量丙酮酸羧化酶从破碎的线粒体中释放出来，并且足够的丙酮酸会带入该酶的检测中，从而对粗制脂肪细胞提取物中乙酰辅酶A羧化酶的检测产生显著干扰。这可能解释了一些先前出版物中报道的脂肪细胞提取物中存在的大量不依赖柠檬酸的乙酰辅酶A羧化酶活性。通过在不冷冻的情况下非常短暂地匀浆可以消除这个问题。通过使用改良的匀浆程序，研究表明EGF处理脂肪细胞对乙酰辅酶A羧化酶活性产生的影响与胰岛素几乎相同。两种信使分子均增加最大反应速度（Vmax），而对变构激活剂柠檬酸的米氏常数（Ka）没有显著影响。
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Both insulin and epidermal growth factor stimulate lipogenesis and acetyl-CoA carboxylase activity in isolated adipocytes. Importance of homogenization procedure in avoiding artefacts in acetyl-CoA carboxylase assay.胰岛素和表皮生长因子均可刺激分离的脂肪细胞中的脂肪生成及乙酰辅酶A羧化酶活性。乙酰辅酶A羧化酶测定中匀浆程序对于避免人为误差的重要性。
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