Hinman Jason D, Duce James A, Siman Robert A, Hollander William, Abraham Carmela R
Department of Medicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
J Neurochem. 2004 Apr;89(2):430-41. doi: 10.1046/j.1471-4159.2004.02348.x.
Ultrastructural disruption of myelin sheaths and a loss of myelin with age are well-documented phenomena in both the human and rhesus monkey. Age-dependent activation of calpain-1 (EC 3.4.22.52) has been suggested as a plausible mechanism for increased proteolysis in the white matter of the rhesus monkey. The present study documents activation of calpain-1 throughout brain white matter in aged animals, evidenced by immunodetection of the activated enzyme as well as a calpain-derived spectrin fragment in both tissue section and Triton X-100-soluble homogenate of subcortical white matter from the frontal, temporal, and parietal lobes. Separation of myelin fractions from brain stem tissue into intact and floating myelin confirmed previous reports of an age-related increase in activated calpain-1 in the floating fraction. Measurements of calpain-1 activity using a fluorescent substrate revealed an age-related increase in calpain-1 proteolytic activity in the floating myelin fraction consistent with immunodetection of the activated enzyme in this fraction. Double-immunofluorescence demonstrated co-localization of activated calpain-1 with human leukocyte antigen-DR (HLA-DR), a marker for activated microglia, suggesting that these cells represent the major source of the increase in activated calpain-1 in the aging brain. These data solidify the role of calpain-1 in myelin protein metabolism and further implicate activated microglia in the pathology of the aging brain.
髓鞘的超微结构破坏以及随着年龄增长髓鞘的丧失在人类和恒河猴中都是有充分记录的现象。钙蛋白酶-1（EC 3.4.22.52）的年龄依赖性激活被认为是恒河猴白质中蛋白水解增加的一种可能机制。本研究记录了老年动物全脑白质中钙蛋白酶-1的激活，这通过在额叶、颞叶和顶叶皮质下白质的组织切片以及Triton X-100可溶性匀浆中对活化酶以及钙蛋白酶衍生的血影蛋白片段进行免疫检测得以证实。将脑干组织的髓鞘部分分离为完整髓鞘和漂浮髓鞘，证实了先前关于漂浮部分中活化钙蛋白酶-1随年龄增加的报道。使用荧光底物测量钙蛋白酶-1活性显示，漂浮髓鞘部分中钙蛋白酶-1的蛋白水解活性随年龄增加，这与该部分中活化酶的免疫检测结果一致。双重免疫荧光显示活化的钙蛋白酶-1与人白细胞抗原-DR（HLA-DR）共定位，HLA-DR是活化小胶质细胞的标志物，表明这些细胞是衰老大脑中活化钙蛋白酶-1增加的主要来源。这些数据巩固了钙蛋白酶-1在髓鞘蛋白代谢中的作用，并进一步表明活化的小胶质细胞与衰老大脑的病理学有关。
