少突胶质细胞LDL受体对慢性低灌注型脑白质损伤的转归影响及作用机制

Cerebral white matter is mainly composed of the myelin which is the specific structure constituted by oligodendrocytes. White matter lesions are most likely to be resulted from brain hypoxic ischemic injury. However, mechanisms underlying chronic cerebral hypoperfusion-induced white matter injury remain covered. Our preliminary experiments showed that the expression of LDL receptor (LDLR) was decreased in white matter injury model of mice and oligodendrocytes. Nevertheless, the cholesterol content in corpus callosum and oligodendrocytes was unchanged. Moreover, lower FA value in corpus callosum was observed in DTI imaging for 10-week-old LDLR knock-out mice. Mounting evidences have demonstrated that LDLR may be involved in white matter damage independent of its classical role in mediating cholesterol endocytosis. With the supplementation of LV-LDLR(NPXY) and LV-LDLR(NPXY→AAVA), we aim to investigate the effect of LDLR in white matter injury, the involvment of intracellular NPVY motif in white matter improvements, and the potential molecular signaling. The function of NPVY motif and underlying pathway in white matter lesions are further discussed in LDLR knock-out mice and oligodendrocytes. This investigation may unveil a novel role of LDLR in ameliorating white matter lesions through its NPVY motif as well as provide a possible approach for the management of white matter injury.

脑白质主要由少突胶质细胞形成的髓鞘构成。脑白质损伤易继发于颅脑慢性低灌注改变，而其具体发病机制仍不清楚。我们前期通过建立脑白质损伤体内外模型发现：模型小鼠脑组织LDL受体（LDLR）表达下降，模型少突胶质细胞LDLR表达也下调，而组织和细胞的胆固醇含量并无改变。此外，10周龄LDLR敲除小鼠DTI成像表现为胼胝体FA值降低。这些证据均提示LDLR可能参与到脑白质损伤的病理改变中，而独立于其介导胆固醇入胞的功能。为此，本课题拟通过给予模型小鼠和少突胶质细胞外源性野生型LV-LDLR(NPXY)和突变型LV-LDLR(NPXY→AAVA)慢病毒载体，采用分子生物学、神经化学、形态学等方法评估胼胝体白质及细胞成髓鞘化情况，并探讨具体的信号效应分子。最后从LDLR敲除小鼠和细胞水平做进一步验证。论证LDLR能够通过其胞内NPVY元件对脑白质损伤起到改善作用，为脑白质损伤的防治提供新的干预靶点。

脑白质主要由成熟少突胶质细胞形成的髓鞘构成，极易受到慢性缺血性损伤，而损伤的具体病理生理机制还不清楚。LDLR作为少突系细胞主要的功能蛋白，主要负责摄取胆固醇来维持成年后的髓鞘化水平，其参与慢性缺血性脑白质损伤的具体机制也不清楚。我们通过双侧颈总动脉狭窄模型构建小鼠慢性脑缺血模型，通过CoCl2共培养或糖氧剥夺模型构建体外急慢性缺氧模型。在缺血缺氧损伤中，少突胶质细胞LDLR表达水平显著降低，少突分化及存活均受到严重打击。然而细胞及组织胆固醇水平并未发生显著改变。在人血清及小鼠水平发现了miR-344e/410-3p能够靶向抑制LDLR的表达，明确了LDLR的上游机制分子。给予LDLR过表达慢病毒能够显著改善慢性缺血相关的脱髓鞘改变。Ldlr-/-小鼠胼胝体有髓鞘损伤表现，Ldlr-/-少突前体细胞分化及髓鞘化能力受限。将Ldlr-/-少突前体细胞移植进慢性脑缺血小鼠胼胝体发现其无法改善缺血相关髓鞘脱失。通过对比野生型及突变型LDLR的作用发现，LDLR的髓鞘保护作用依赖于其天冬酰胺-脯氨酸-缬氨酸-酪氨酸（NPVY）元件与胞质内Shc分子的交互及下游MEK/ERK通路的激活。总之，我们的研究结果表明，在慢性脑缺血中，异常的少突胶质细胞LDLR通过细胞内信号转导作用损害髓鞘形成。保护少突胶质细胞LDLR不受损害可能是治疗缺血性脱髓鞘的新的干预靶点。