LDLR基因敲除高胆固醇血症大鼠的脑白质改变

Changes of white matter in LDLR knockout hypercholesterolemic rats

目的探讨低密度脂蛋白受体（low density lipoprotein receptor, LDLR）纯合子突变高胆固醇血症大鼠脑白质组织病理学改变与野生型大鼠的差异。方法选取30只LDLR^-/-大鼠和28只野生型大鼠，分别在15、18和26周龄时采用酶联免疫法测定血浆胆固醇水平，通过透射电子显微镜观察脑胼胝体区域轴索结构，通过蛋白质印迹法对胼胝体区髓鞘碱性蛋白（myelin basic protein, MBP）进行定量分析。在26周龄时通过快蓝染色法染髓鞘，免疫荧光染色法进一步检测脑白质区MBP表达水平并观察神经胶质细胞形态学改变。结果与野生型大鼠相比，LDLR^-/-大鼠血浆胆固醇浓度明显升高，26周时可高达3.3倍。电镜结果显示，LDLR^-/-大鼠自15周出现轴索损伤，且随时间进展逐渐加重。26周时，LDLR^-/-大鼠蛋白质印迹分析显示胼胝体区MBP表达水平显著降低，快蓝染色显示胼胝体区神经纤维出现疏松、弥散性空泡形成及有髓神经纤维缺失。此外，还发现LDLR^-/-大鼠少突胶质细胞数量明显减少，星形胶质细胞及小胶质细胞大量激活。结论LDLR^-/-大鼠会出现自发性高胆固醇血症，在成年早期脑白质区即存在轴索损伤、髓鞘脱失、少突胶质细胞减少以及星形胶质细胞和小胶质细胞异常激活。

Objective To investigate the difference between histopathological changes of brain white matter in low-density lipoprotein receptor （LDLR） homozygous mutation rats with hypercholesterolemia and wild-type rats. Me,hods Thirty LDLR^-/- rats and 28 wild-type rats were selected. Plasma cholesterol levels were measured by enzyme-linked immunosorbent assay at 15, 18 and 26 weeks old respectively. The axonal structure of the corpus callosum area was observed by transmission electron microscopy. The myelin basic protein （MBP） of the corpus callosum area was quantitatively analyzed by Western blotting. In addition, at 26 weeks old, the myelin sheaths were stained by fast blue staining. The expression level of MBP in white matter was further detected by immunofluorescence staining, and the morphological changes of glial cells were observed. Results Compared with the wild-type rats, the plasma cholesterol concentration in LDLR^-/- rats increased significantly, and it could be as high as 3.3 times at 26 weeks. The results of electron microscopy showed that the LDLR^-/- rats had axonal injury at 15 weeks and aggravated gradually over time. At 26 weeks, Western blot analysis of the LDLR^-/- rats showed that the MBP expression level of the corpus callosum area decreased significantly. Fast blue staining showed loosening of nerve fibers, diffuse vacuole formation, and myelinated nerve fiber loss in the corpus callosum area. In addition, it was also found that the number of oligodendrocytes in LDLR^-/- rats was significantly reduced, and large numbers of astrocytes and microglia were activated. Conclusions LDLR^-/- rats will have spontaneous hypercholesterolemia.Axonal injury, demyelination, decreased oligodendrocytes, as well as the abnormal activation of astrocytes and microglia are present in the early adult brain white matter area.