Enhanced phagocytosis through inhibition of de novo ceramide synthesis
Enhanced phagocytosis through inhibition of de novo ceramide synthesis
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DOI：
10.1074/jbc.m206199200
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发表时间：
2003-01-10
影响因子：
4.8
通讯作者：
Shayman, JA
中科院分区：
文献类型：
--
作者：
Hinkovska-Galcheva, V;Boxer, L;Shayman, JA
Fc γ受体是IgG与嗜中性粒细胞结合并诱导嗜中性粒细胞吞噬作用的重要介质。COS-1细胞为研究这些受体提供了潜在有用的模型，因为用FcgammaRIIA转染使这些细胞具有吞噬作用。在COS-1细胞中Fc γ RIIA介导的吞噬作用期间，内源性神经酰胺水平在20分钟时增加52%（p < 0.01）。磷脂酶D活性增加62%（p < 0.01）。相应地，吞噬指数增加了3.7倍，由20分钟。两个神经酰胺形成的抑制剂被用来评估减少神经酰胺生成的后果。L-环丝氨酸，一种阻断丝氨酸棕榈酰转移酶活性的抑制剂，降低了鞘氨醇和神经酰胺的水平。在这些条件下，在2 mM L-环丝氨酸的存在下，吞噬指数增加100%。由神经酰胺合酶对二氢鞘氨醇或鞘氨醇的N-酰化产生的神经酰胺的形成被真菌毒素伏马菌素B-1抑制。当用5-50 μ m伏马菌素B-1处理细胞时，神经酰胺的细胞水平以浓度依赖性方式降低，而同时吞噬指数增加52%。与此同时，随着神经酰胺水平的下降，FcgammaRIIA活性的三个间接测量值发生了变化。Syk磷酸化，磷脂酶D的活性，和促分裂原活化蛋白（MAP）激酶磷酸化增加30分钟。当Syk磷酸化被阻断与piceatannol和细胞同样的挑战，磷脂酰肌醇3-激酶激活被阻断，但没有任何变化，无论是神经酰胺积累或MAP激酶激活观察。因此，神经酰胺的形成和MAP激酶的激活不依赖于Syk激酶在该系统中的活性。这些结果表明，COS-1细胞提供了一个有用的模型，在吞噬作用的研究鞘脂信号的重演。由从头合成形成的神经酰胺可能是调节吞噬作用的重要机制。
Fcgamma receptors are important mediators of the binding of IgG to and induction of phagocytosis in neutrophils. COS-1 cells provide a potentially useful model for studying these receptors because transfection with the FcgammaRIIA renders these cells phagocytic. During FcgammaRIIA-mediated phagocytosis in COS-1 cells, endogenous ceramide levels increased 52% by 20 min (p < 0.01). Phospholipase D activity increased by 62% (p < 0.01). Correspondingly, the phagocytic index increased by 3.7 fold by 20 min. Two inhibitors of ceramide formation were used to assess the consequences of reduced ceramide generation. L-Cycloserine, an inhibitor that blocks serine palmitoyltransferase activity, lowered both sphingosine and ceramide levels. Under these conditions, the phagocytic index increased 100% in the presence of 2 mM L-cycloserine. The formation of ceramide resulting from the N-acylation of dihydrosphingosine or sphingosine by ceramide synthase is inhibited by the fungal toxin fumonisin B-1. When cells were treated with 5-50 mum fumonisin B-1, the cellular level of ceramide decreased in a concentration-dependent manner, while simultaneously the phagocytic index increased by 52%. Concomitantly, three indirect measures of FcgammaRIIA activity were altered with the fall in ceramide levels. Syk phosphorylation, phospholipase D activity, and mitogen-activated protein (MAP) kinase phosphorylation were increased at 30 min. When Syk phosphorylation was blocked with piceatannol and cells were similarly challenged, phosphatidylinositol 3-kinase activation was blocked, but no changes in either ceramide accumulation or MAP kinase activation were observed. Ceramide formation and MAP kinase activation are therefore not dependent on Syk kinase activity in this system. These results indicate that COS-1 cells provide a useful model for the recapitulation of sphingolipid signaling in the study of phagocytosis. Ceramide formed by de novo synthesis may represent an important mechanism in the regulation of phagocytosis.