Neuronal death and sphingolipid following occlusion of the middle cerebral artery

麻醉大鼠大脑中近端动脉干闭塞引起局灶性脑缺血。为了更好地确定局灶性脑缺血时神经酰胺、鞘磷脂、脑苷和神经节苷类在大鼠大脑皮层的代谢水平，我们测定了大脑中动脉闭塞引起局灶性脑缺血时神经酰胺、鞘磷脂、脑苷和神经节苷类的水平。鞘磷脂在缺血2小时开始下降，并持续下降96小时。相比之下，神经酰胺在缺血后6小时增加，在缺血后96小时增加到4.2倍，神经酰胺的脂肪酸组成完全为类似鞘磷脂的非羟基脂肪酸。羟基化脂肪酸连接的脑苷在缺血6小时时下降，而非羟基化脂肪酸连接的脑苷在缺血96小时内没有明显下降。神经节苷脂的水平没有可测量的变化。这些结果表明，神经酰胺在缺血早期通过鞘磷脂的破坏在大脑皮层产生。近年来，鞘磷脂降解产生的神经酰胺被认为是HL-60白血病细胞程序性细胞死亡、细胞生长和分化的第二信使。采用沙鼠短暂性前脑缺血再灌注模型，研究了沙鼠致死性和亚致死性缺血后海马鞘肌素水平的差异。采用薄层色谱法（TLC）从海马总脂中分离鞘磷脂，并采用气相液相色谱法（GLC）进行分析。致死性缺血后再循环中鞘磷脂水平的变化。30min时明显降低，60min时又恢复到对照水平。亚致死缺血后（缺血2分钟和获得性缺血耐受后缺血5分钟）的再循环未引起鞘磷脂水平的任何变化。

Focal cerebral ischemia was induced in anesthetized rats by occluding the stem of the proximal middle cerebral artery. To better define the sphingolipid metabolism during focal brain ischemia, levels of ceramide, sphingomyelin, cerebroside and gangliosides were determined in rat cerebral cortex during focal ischemia produced by occlusion of the middle cerebral artery. Sphingomyelin began to decrease at 2 hours of ischemia and continued to decrease for 96 hours. In contrast, ceramide increased at 6 hours and increased to 4.2-fold at 96 hours after ischemia, and the fatty acid composition of ceramide was solely nonhydroxylated fatty acid similar to sphingomyelin. Hydroxylated fatty acid-linked cerebroside decreased at 6 hours of ischemia, whereas any significant decrease of non-hydroxylated fatty acid-linked cerebroside didn't occur for 96 hours of ischemia. There were no measurable changes in the levels of gangliosides. These results suggested that ceramide was produced in the cerebral cortex by the breakdown of sphingomyelin during early ischemia.Recently ceramide generated by sphingomyelin degradation has been described as a second messenger involved in programd cell death, cell growth and differentiation in HL-60 leukemic cells. We studied on the differences in sphingomylin levels in gerbil hippocampus induced by lethal or sublethal ischemia using the transient forebrain ischemia-reperfusion gerbil model. Sphingomyelin was separated by thin layr chromatography (TLC) from hippocampal total lipids and analyzed by gas liquid chromatography (GLC). The sphingomyelin levels during recirculation after lethal ischemia (a 5-min.of ischemia) were significantly decreased at 30 min.and then returned to the control level at 60 min. The recirculation after a sublethal ischemia (2 min ischemia and 5 min.ischemia after acquired ischemic tolerance) did not induce any changes in sphingomyelin levels.