D-半乳糖诱导大鼠脑损伤的糖基化机制

目的D-半乳糖（D-galactose）诱导大鼠体内不同糖基化水平，研究其脑损伤发生的机理。方法采用不同剂量D-半乳糖150、75、37．5mg／（kg·d）]分别腹腔注射（ip）处理大鼠8周，诱导糖基化状态和脑损伤。采用硫代巴比妥酸（TBA）比色法测定糖化血红蛋白，硝基四氮唑蓝（NBT）比色法测定血清果糖胺；按文献方法分别测定血红细胞醛糖还原酶活性和晚期糖基化终末产物（AGEs）含量及脑组织中AGEs含量，羟胺法和比色法分别测定SOD和GSH-Px活性，硫代巴比妥酸法测定MDA含量；以Fura-2／AM作为钙荧光指示剂，双波长荧光分光光度法检测脑海马神经细胞胞质Ca^2＋]i的变化；透射电镜观察脑海马神经细胞线粒体的变化。结果D-半乳糖处理8周后，大鼠血红细胞醛糖还原酶活性升高，糖化产物形成增多；脑组织中AGEs及脑细胞胞质Ca^2＋]i含量明显升高，SOD及GSH-Px活性下降，MDA含量升高（P〈0．01，P〈0．05），海马神经细胞线粒体出现病理性改变。结论D-半乳糖通过诱导体内蛋白糖基化和脑组织AGEs大量生成，降低抗氧化能力及胞质Ca^2＋]i超负荷等，导致脑细胞损伤。

Glycation Mechanism of Brain Damage Induced by D-Galactose in Rats

Objective To prepare different levels of glycation in rats induced by D-galactose and to understand the role of advanced glycation end-products （AGEs） in the mechanism of brain damage. Methods Rats were administrated intraperitoneally with D-galactose in a dose of 150 mg/kg, 75 mg/kg and 37.5 mg/kg, respectively, for 8 weeks to induce the glycation condition and brain damage. After treatment, the levels of HbAIC and fructosamine in serum were detected with thiobarbituric acid （TBA） and nitrotetrazolium blue chloride （NBT） photoreduction method, respectively; According to the literature method, the activity of aldose reductase and content of AGEs in plasma and in brain tissues were measured, and the activity of SOD and GSH-Px, the contents of MDA in brain tissues were measured by hydroxylamine method, chemical colorimetry and TBA method. Isolated hippocampal cells were loaded with Ca^2＋-sensitive fluorescent indicator Fura-2/AM, and the changes of Ca^2＋]i in these cells were detected by double beam fluorospectrophotometry, and the morphologic changes of mitochondria in hippocampal cells were observed by transmission electron microscopy. Results After treatment of D-galactose for 8 weeks, the activity of serum aldose reductase and the content of AGEs were increased significantly, and the levels of AGEs and MDA in brain tissues and calcium ion in hippocampal cells were also enhanced significantly （ P 〈 0.01 or P 〈 0.05）. However, the activities of SOD and GSH-Px decreased （ P 〈 0.01 or P 〈 0.05）. Additionally, pathological changes of mitochondria in hippocampal cells were observed. Conclusion D-galactose induces protein glycation in vivo and AGEs formation in brain tissues, which reduce the antioxidative ability and result in overload of calcium ion in hippocampal cells, leading to damage to brain cells.