Niculescu F, Rus H, Shin S, Lang T, Shin M L
University of Maryland, School of Medicine, Department of Pathology, Baltimore 21201.
J Immunol. 1993 Jan 1;150(1):214-24.
Formation of sublytic terminal complement complexes (TCC) on nucleated cells produces transient increase in [Ca2+]i and activates protein kinase C. The present study is to evaluate whether TCC can generate endogenous signal messengers other than Ca2+ that regulate cell activities by measuring mass-levels of sn-1,2-diacylglycerol (DAG) and ceramide. As targets, lymphoblastoid human B cell lines JY25 and its mutant JY5 were used. JY5, cells deficient in glycosylphosphatidylinositol-anchored proteins with higher lytic susceptibility to human complement, are four times more efficient in forming C5b-9. When cells sensitized with limited anti-class II IgG were exposed to human serum to generate sublytic TCC, a sustained increase in DAG and ceramide was observed with a maximum 3.6-fold DAG increase over basal level in JY25 and 2.8-fold in JY5, and 6.3-fold ceramide increase in JY25 and 2.8-fold in JY5. The effect of TCC was evaluated with C7-deficient human serum (C7D) +/- C7 and also with C5b6, C7, C8, and C9 proteins. The DAG and ceramide increase by C7D + C7 over C7D control were 1.6- and 1.8-fold, respectively, in JY25, and 2.3-, and two-fold in JY5. TCC activation also induced an increased hydrolysis of sphyingomyelin and phosphatidylcholine. In addition, DAG increase by TCC was primarily achieved by C5b-7 and preincubation of cells with pertussis toxininhibited DAG increase, suggesting an involvement of a pertussis toxin-sensitive GTP-binding protein. As important signal transduction molecules, DAG and ceramide generated in response to TCC assembly, could participate in cell activation during inflammation and repair.
有核细胞上形成的亚溶血型末端补体复合物（TCC）会使细胞内钙离子浓度（[Ca2+]i）短暂升高，并激活蛋白激酶C。本研究旨在通过测量sn-1,2-二酰基甘油（DAG）和神经酰胺的质量水平，评估TCC是否能产生除Ca2+之外的内源性信号信使来调节细胞活动。以人淋巴母细胞系JY25及其突变体JY5作为研究对象。JY5细胞缺乏糖基磷脂酰肌醇锚定蛋白，对人补体的溶解敏感性更高，形成C5b-9的效率是JY25细胞的四倍。当用有限量的抗II类IgG致敏的细胞暴露于人类血清以产生亚溶血型TCC时，观察到DAG和神经酰胺持续增加，JY25细胞中DAG相对于基础水平最大增加3.6倍，JY5细胞中增加2.8倍；JY25细胞中神经酰胺增加6.3倍，JY5细胞中增加2.8倍。用缺乏C7的人血清（C7D）+/- C7以及C5b6、C7、C8和C9蛋白评估TCC的作用。在JY25细胞中，C7D + C7相对于C7D对照组使DAG和神经酰胺分别增加1.6倍和1.8倍，在JY5细胞中分别增加2.3倍和2倍。TCC激活还诱导了鞘磷脂和磷脂酰胆碱水解增加。此外，TCC导致的DAG增加主要由C5b-7引起，用百日咳毒素预处理细胞可抑制DAG增加，这表明百日咳毒素敏感的GTP结合蛋白参与其中。作为重要的信号转导分子，响应TCC组装而产生的DAG和神经酰胺可能参与炎症和修复过程中的细胞激活。
