Stumpo D J, Kletzien R F
J Cell Physiol. 1981 Apr;107(1):11-9. doi: 10.1002/jcp.1041070103.
Primary cultures of parenchymal cells isolated from adult rat liver by a collagenase perfusion procedure and maintained as a monolayer in a serum-free culture medium were used to study gluconeogenesis and the role that the glucocorticoids play in the control of this pathway. These cells carried out gluconeogenesis from three-carbon precursors (alanine and lactate) in response to glucagon and dexamethasone added alone or in combination. Maximum glucose production was observed with cells pretreated for several hours with dexamethasone and glucagon prior to addition of substrate and glucagon (8- to 12-fold increase over basal glucose production). Half-maximum stimulation of gluconeogenesis was seen with 3.6 X 10(-10) M glucagon and 3.6 X 10(-8) M dexamethasone. Maximum stimulation was observed with 10(-7) M glucagon and 10(-6) M dexamethasone. The length of time of dexamethasone pretreatment was found to be important in demonstrating the effect of glucocorticoids on glucagon-stimulated gluconeogenesis. Treatment of cells with dexamethasone for 2 hours did not result in an increase in glucose production over identical experimental conditions in the absence of dexamethasone, whereas pretreatment for 5 hours (1.2-fold increase) or 15 hours (1.7-fold increase) did result in an increase in glucose production. The results establish that the adult rat liver parenchymal cells in primary culture are a valid model system to study hepatic gluconeogenesis. In addition, we have established directly that the glucocorticoids amplify the glucagon stimulation of gluconeogenesis.
通过胶原酶灌注法从成年大鼠肝脏分离出实质细胞，并在无血清培养基中维持单层培养，利用这些原代培养细胞研究糖异生作用以及糖皮质激素在该途径调控中所起的作用。这些细胞在单独添加或联合添加胰高血糖素和地塞米松的情况下，能够利用三碳前体物质（丙氨酸和乳酸）进行糖异生。在添加底物和胰高血糖素之前，先用胰高血糖素和地塞米松预处理细胞数小时，观察到最大葡萄糖生成量（比基础葡萄糖生成量增加8至12倍）。当胰高血糖素浓度为3.6×10⁻¹⁰ M、地塞米松浓度为3.6×10⁻⁸ M时，糖异生作用受到半最大刺激。当胰高血糖素浓度为10⁻⁷ M、地塞米松浓度为10⁻⁶ M时，观察到最大刺激效果。发现地塞米松预处理的时间长度对于证明糖皮质激素对胰高血糖素刺激的糖异生作用的影响很重要。在相同实验条件下，用2小时地塞米松处理细胞，与未用地塞米松处理相比，葡萄糖生成量并未增加，而预处理5小时（增加1.2倍）或15小时（增加1.7倍）确实导致葡萄糖生成量增加。结果表明，原代培养的成年大鼠肝脏实质细胞是研究肝脏糖异生作用的有效模型系统。此外，我们还直接证实了糖皮质激素可增强胰高血糖素对糖异生作用的刺激。
