Regulation of fructose-2,6-bisphosphate content in rat hepatocytes, perfused hearts, and perfused hindlimbs.
Regulation of fructose-2,6-bisphosphate content in rat hepatocytes, perfused hearts, and perfused hindlimbs.
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DOI：
10.1016/s0021-9258(18)34722-7
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发表时间：
1982-04
期刊：
影响因子：
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通讯作者：
L. Hue;P. Blackmore;H. Shikama;A. Robinson-Steiner;J. Exton
中科院分区：
文献类型：
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作者：
L. Hue;P. Blackmore;H. Shikama;A. Robinson-Steiner;J. Exton
已经开发出一种专门测量组织中 2, 6-二磷酸果糖的方法。消除提取物中存在的 6-磷酸己糖后，2, 6-二磷酸果糖被水解，并在与细菌 NADH 连接的荧光素酶的偶联酶测定中测量酸显示的 6-磷酸果糖。在所有研究的组织中都发现了果糖-2, 6-二磷酸；并且，按照浓度增加的顺序，附睾脂肪、肾、骨骼肌、肺、心脏、肝脏和脑含有0.1至2.3nmol/g组织。低于 0.1 nmol/g 的值无法准确测量，代表该方法的检测限。在存在 20 m2 丙氨酸、丙酮酸、甘油、L-山梨糖、乳酸、塔格糖、果糖和甘油醛的情况下，对来自喂食大鼠的分离肝细胞进行培养，导致果糖-2, 6-二磷酸含量按效果逐渐增强的顺序降低 2 至 10 倍。在20m2二羟基丙酮、半乳糖、甘露糖、木糖醇、阿拉伯糖或核糖存在下的孵育对2,6-二磷酸果糖的浓度影响很小或没有影响。当肝细胞在缺氧条件下或在线粒体解偶联剂羰基氰化物间氯苯腙存在下孵育时，2、6-二磷酸果糖含量分别降低了 40 和 608。在灌注的大鼠后肢肌肉中，胰岛素和肾上腺素分别导致 2, 6-二磷酸果糖增加 2 倍和 4 倍，并刺激糖酵解。电刺激引起肌肉收缩，乳酸产量增加 2.5 倍，但果糖 2, 6-二磷酸减少。它还消除了肾上腺素引起的这种代谢物的增加。在灌注的大鼠心脏中，肾上腺素引起蛋白激酶和磷酸化酶的激活，但不改变果糖-2, 6-二磷酸含量。
A method has been developed to specifically measure fructose-2, 6-bisphosphate in tissues. After elimination of the hexose-6-phosphate present in the extract, fructose-2, 6-bisphosphate is hydrolyzed and the acid-revealed fructose-6-phosphate is measured in a coupled enzymatic assay with bacterial NADH-linked luciferase. Fructose-2, 6-bisphosphate was found in all tissues studied; and, in order of increasing concentration, epididymal fat, kidney, skeletal muscle, lung, heart, liver, and brain contain 0.1 to 2.3 nmol/g of tissue. Values lower than 0.1 nmol/g cannot be accurately measured and represent the limit of detection of the method. Incubation of isolated hepatocytes from fed rats in the presence of 20 m~ alanine, pyruvate, glycerol, L-sorbose, lactate, tagatose, fructose, and glyceraldehyde caused, in order of increasing effect, a 2-to 10-fold decrease in fructose-2, 6-bisphosphate content. Incubation in the presence of 20 m~ dihydroxyacetone, galactose, mannose, xylitol, arabinose, or ribose had little or no effect on the concentration of fructose-2, 6-bisphosphate. When hepatocytes were incubated under anoxic conditions or in the presence of carbonyl cyanide m-chlorophenylhydrazone, a mitochondrial uncoupler, the fructose-2, 6-bisphosphate content was decreased by 40 and 608, respectively. In perfused rat hindlimb muscle, insulin and epinephrine, respectively, caused a 2-and 4-fold increase in fructose-2, 6-bisphosphate and a stimulation of glycolysis. Electrical stimulation caused muscular contraction and a 2.5-fold increase in lactate production, but decreased fructose-2, 6-bisphosphate. It also abolished the increase in this metabolite induced by epinephrine. In perfused rat hearts, epinephrine caused the activation of protein kinase and phosphorylase but did not alter the fructose-2, 6-bisphosphate content.