【作者】 杨益宁；

【导师】 孙子林；

【作者基本信息】 东南大学 ， 内科学， 2006， 硕士

【摘要】 目的:观察糖基化终产物(advanced glycation end products,AGEs)对人肾系膜细胞(human renal mesangial cell,HRMC)合成和分泌Ⅳ型胶原及层粘连蛋白(laminin,LN)的影响以及药物盐酸氨基胍(aminoguanidine,AG)的干预作用;观察AGEs对HRMC基质金属蛋白酶-2(matrix metallo proteinase-2,MMP-2)表达的影响以及AG的干预效应。方法:1、运用体外制备的糖基化修饰的牛血清白蛋白(AGE-bovine serum albumin,AGE-BSA)干预HRMC,以及氨基胍单独或与AGE-BSA共同干预HRMC;2、放射免疫分析法检测HRMC上清液中Ⅳ型胶原、LN含量;3、逆转录聚合酶链式反应检测MMP-2 mRNA表达,蛋白免疫印迹检测MMP-2蛋白表达。结果:1、AGE-BSA呈浓度、时间依赖性增加HRMC上清中Ⅳ型胶原和LN的含量(P<0.05);2、盐酸氨基胍可减低HRMC上清中由AGE-BSA升高的Ⅳ型胶原、LN含量(P<0.05);3、HRMC的MMP-2 mRNA和蛋白表达水平随着AGE-BSA浓度增加和干预时间延长而降低(P<0.05);4、盐酸氨基胍与AGE-BSA共同干预HRMC,可部分恢复MMP-2 mRNA和蛋白的表达(P<0.05)。结论:1、AGE-BSA诱导HRMC合成和分泌Ⅳ型胶原、LN增加,提示AGEs可能引起胞外基质不恰当积聚;2、AGE-BSA降低HRMC的MMP-2表达,提示AGEs可能通过MMP-2参与糖尿病肾病的发生发展;3、盐酸氨基胍可以下调由AGE-BSA诱导的Ⅳ型胶原、LN合成分泌增加,可抑制由AGE-BSA诱导的HRMC MMP-2表达减低,提示盐酸氨基胍可能通过干预AGE-BSA与细胞的相互作用,降低AGE-BSA的生物学效应,影响胞外基质和MMP-2水平,从而发挥肾保护效应。更多还原

【Abstract】 Objectives:To investigate the effects of advanced glycation end products (AGEs) on the expression of matrix metalloproteinase-2 (MMP-2),the content of typeⅣcollagen and laminin in supernatants of cultured human renal mesangial cells (HRMC). To explore the influence of aminoguanidine (AG) on the MMP-2 expression, the effects of AG on the level of typeⅣcollagen and laminin in supernatants of cultured HRMC.Methods:1.HRMC were incubated with different concentrations of AGE-bovine serum albumin (AGE-BSA) for different time, and exposed to different concentrations of aminoguanidine for 24 hours with or without AGE-BSA(200 mg·l-1). 2.The contents of typeⅣcollagen and laminin in the media were detected by radioimmunoassay. 3.The mRNA and protein expression of MMP-2 in HRMC were measured by RT-PCR and Western blotting respectively.Results:1.AGE-BSA increased the level of typeⅣcollagen and laminin time- and dose- dependently in supernatants of cultured HRMC(P<0.05). 2.Aminoguanidine can reduce AGE-BSA-induced HRMC synthesizes and sectetes typeⅣcollagen and laminin. 3.AGE-BSA decreased the expression of MMP-2 in the level of mRNA and protein time- and dose- dependently (P<0.05). 4.The declined expressions of MMP-2 mRNA and protein in AGE-BSA treated HRMC were increased by exposure to aminoguanidine (P<0.05).Conclusions:1.AGE-BSA increases the content of typeⅣcollagen and laminin in cultured HRMC. These findings suggest that AGEs may induce the extracellular matrix imbalance. 2.AGE-BSA down-regulates the expression of MMP-2 mRNA and protein in HRMC. The data indicates that MMP-2 might mediate the role of AGEs in the development of diabetic nephropathy. 3.AG inhibits the upregulation of typeⅣcollagen and laminin in HRMC by AGE-BSA. The depressed expressions of MMP-2 mRNA and protein in HRMC incubated with AGE-BSA are increased by exposure to AG. These results suggest aminoguanidine might play their role of reno-protection via supression of typeⅣcollagen and laminin and upregulation of MMP-2.更多还原