Kamada T, McMillan D E, Otsuji S
Department of Laboratory Medicine, Faculty of Medicine, Kagoshima University, Japan.
Diabetes Res Clin Pract. 1992 May;16(2):85-90. doi: 10.1016/0168-8227(92)90077-5.
We studied metabolic pool size of polyphosphoinositides and phosphatidate of erythrocyte membranes from normal and diabetic subjects using 32P for 20-h incubation, a sufficiently long period to reach isotopic equilibrium between monoesterphosphate bond and gamma-phosphate of ATP. Phosphatidylinositol 4-monophosphate (PtdIns4P), phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) and phosphatidate were the phospholipids labelled. Metabolic pools of individual phospholipids were estimated, based on their proportionate and absolute radioactivity. A significant decline in radioactivity of phosphatidate and PtdIns(4,5)P2 was seen in erythrocytes from the diabetic subjects, indicating suppression of the metabolically labile pool of these two phospholipids. There was no significant change in PtdIns4P radioactivity between the groups. The direct effect of insulin on phosphorylation of polyphosphoinositides and phosphatidate was also evaluated by a short incubation period of erythrocyte membranes with [gamma-32P]-ATP. Added insulin increased the incorporation of 32P into phosphatidate in a dose-dependent manner that reached a steady state at 2 nM. We conclude that the metabolically labile pool size of phosphatidate is decreased and that of polyphosphoinositides is altered in erythrocyte membranes from diabetic patients.
我们使用³²P进行20小时孵育，研究了正常受试者和糖尿病患者红细胞膜中多磷酸肌醇和磷脂酸的代谢池大小，这一足够长的时间可使单酯磷酸键与ATP的γ-磷酸之间达到同位素平衡。被标记的磷脂有磷脂酰肌醇4-单磷酸（PtdIns4P）、磷脂酰肌醇4,5-二磷酸（PtdIns(4,5)P2）和磷脂酸。根据各磷脂的相对放射性和绝对放射性估计其代谢池大小。糖尿病患者红细胞中磷脂酸和PtdIns(4,5)P2的放射性显著下降，表明这两种磷脂的代谢不稳定池受到抑制。两组之间PtdIns4P放射性无显著变化。还通过用[γ-³²P]-ATP对红细胞膜进行短时间孵育来评估胰岛素对多磷酸肌醇和磷脂酸磷酸化的直接作用。添加的胰岛素以剂量依赖方式增加³²P掺入磷脂酸，在2 nM时达到稳定状态。我们得出结论，糖尿病患者红细胞膜中磷脂酸的代谢不稳定池大小减小，多磷酸肌醇的代谢不稳定池大小发生改变。
