【作者】 郝敏；

【导师】 贾雪梅；

【作者基本信息】 安徽医科大学 ， 人体解剖与组织胚胎学， 2009， 硕士

【摘要】 目的:①观察高脂饮食大鼠肾脏组织光镜下形态学变化;②观察高脂饮食大鼠肾脏组织中细胞凋亡的情况;③观察高脂饮食大鼠Bcl-2和Bax表达的变化;④通过形态计量学分析高脂饮食对大鼠肾脏组织的影响。方法:SD雄性大鼠20只,随机分为高脂饮食组10只、对照组10只。对照组在室温及稳定湿度条件下,全价颗粒饲料喂养;高脂饮食组给予高脂饮食(饲料成分为胆固醇2%、猪油10%、基础饲料88%)。于6W后测量两组血清学指标。取肾脏组织分别进行以下处理:①HE染色;②脱氧核糖核酸末端转移酶介导的缺口末端标记(Teminal deoxynucleotidyl transferase-mediated dUTP nick end labeling ,TUNEL)法测定肾组织中凋亡细胞;③免疫组织化学染色;④利用计算机图像分析软件分别计算Bcl-2平均光密度(mean optical density,MOD)、Bax MOD、肾小体平均直径、肾小体平均周长、近曲小管平均直径和远曲小管平均直径。结果:①高脂饮食组血清甘油三酯(triglyceride,TG)明显高于对照组(0.74±0.09 vs 0.49±0.05);高脂饮食组血清总胆固醇(total cholesterol,TC)明显高于对照组(1.19±0.18 vs 0.92±0.19);而高脂饮食组高密度脂蛋白胆固醇(high density lipoprotein cholesterol,HDL-C)水平较之对照组明显下降(0.71±0.12 vs 1.03±0.16),差异均具有显著性(P<0.05);②HE染色切片中,对照组肾脏组织结构正常,高脂饮食组与对照组相比较,肉眼光镜下肾脏组织结构无明显病理变化。③凋亡细胞检测结果显示,对照组和高脂饮食组均能观察到细胞核着色为棕黄色的凋亡细胞,对照组肾组织内凋亡细胞较少,高脂饮食组凋亡细胞明显增多,肾小管的凋亡细胞明显多于肾小球,肾间质也有少量凋亡细胞分布。对照组凋亡细胞数为3.57±1.38/高倍镜视野,高脂饮食组凋亡细胞数为15.45±5.07/高倍镜视野,差异具有显著性(P﹤0.05)。④免疫组织化学染色结果提示,Bcl-2和Bax在对照组和高脂饮食组肾组织中均有表达,阳性反应物定位于细胞浆,呈棕黄色颗粒。Bcl-2主要定位于肾小管上皮细胞,在肾小球也有少量表达。Bax主要定位于肾小管上皮细胞,在肾小球也有少量表达。高脂饮食组大鼠肾脏组织中Bcl-2 MOD值为0.2450±0.0216, Bax MOD值为0.2763±0.0912,与对照组Bcl-2 MOD(0.2941±0.0204)和Bax(0.1506±0.3334)MOD值比较,差异均具有显著性(P﹤0.05)。⑤肾脏组织形态计量学分析结果表明,高脂饮食组肾小体平均直径明显小于对照组(229.5126±37.3912 vs 262.4389±38.5034);高脂饮食组肾小体平均周长明显小于对照组(830.0191±136.0398 vs 974.8371±128.1884);高脂饮食组近曲小管管腔平均直径明显小于对照组(95.1354±16.1624 vs 102.2602±16.0461);高脂饮食组远曲小管管腔平均直径明显小于对照组(78.6235±15.6967 vs 89.8674±21.7171),差异均具有显著性(P <0.05)。结论:①高脂饮食大鼠肾脏组织中凋亡细胞数量增多;②高脂饮食大鼠肾脏组织中Bcl-2表达减弱,Bax表达增强;③高脂饮食大鼠肾脏组织肾小体平均直径、肾小体平均周长、近曲小管平均直径、远曲小管平均直径均减小;④细胞凋亡可能是高脂饮食引起肾脏损害的机制之一,且Bcl-2、Bax的表达及Bcl-2/Bax影响着肾脏组织细胞凋亡的发生。更多还原

【Abstract】 Objectives①To observe the morphologic change of the kidney tissue of the high-fat-diet rats under light microscope;②To observe the cell apoptosis in the kidney tissue of the high-fat-diet rats;③To observe the change of expression of Bcl-2 and Bax in kidney tissue of the high-fat-diet rats;④To analyze the influence of high-fat diet on kidney tissue of rats by morphometrics.Methods 20 SD male mice were divided into two groups: ten to high-fat-diet group and ten to the controlled group. The controlled group were fed on complete feed in the room temperature and steady humidity, while the high-fat-diet group were given high-fat food (which was made up of 2% cholesterol, 10% lard and 88% basic feeding stuff). After 6 weeks their blood serum was tested, in which the kidney were taken to :①Do Hematoxylin-Eosin(HE) staining;②Measure the apoptosis cells in the kidney tissue by teminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method;③Do immunohistochemical staining;④Calculate the Bcl-2 mesn optical density(MOD), Bax MOD, average diameter of renal corpuscle, average perimeter of renal corpuscle,average diameter of proximal convoluted tubules, average diameter of distal convoluted tubules by computer picture analysis software.Results①The blood serum triglyceride (TG), of high-fat-diet group was evidently higher than the controlled group((0.74±0.09 vs 0.49±0.05); The blood serum total cholesterol (TC) of high-fat-diet group was evidently higher than the controlled group(1.19±0.18 vs 0.92±0.19); while the high density lipoprotein cholesterol(HDL-C) of high-fat diet group was evidently low compared with the controlled group and the difference was significant(P<0.05);②HE staining showed that the structure of kidney tissue was normal. Compared with it, the structure of kidney tissue of the high-fat-diet group had no evident pathological under the light microscope.③The result of the apoptosis test by TUNEL method suggested that there were apoptosis cells with brown nucleolus in both groups. Yet the controlled group had a few and a small amount of apoptosis cells could be seen in the glomerular,and more could be found in the tubular in the high-fat-diet group.Besides, there were also some in renal interstitium.The number of apoptosis cells of controlled group was 3.57±1.38/HP,while that of the high-fat-diet group was 15.45±5.07/HP. The difference was significant(P﹤0.05).④The result of immunohistochemisty staining showed that Bcl-2 and Bax were expressed in both groups, positive reactants were located in cytoplasm and were brown grains. Bax was located mainly in the renal tubular epithelial cells,and a small amount could also be found in glomerular. And Bcl-2 was mainly located in the renal tubular epithelial cells,and a small amount could also be found in glomerular.The number of Bcl-2MOD in the kidney tissue of high-fat-diet group was 0.2450±0.0216, and Bax MOD was 0.2763±0.0912.Compared to Bcl-2 MOD(0.2941±0.0204)and Bax(0.1506±0.3334)MOD of the controlled group, the difference was significant.(P﹤0.05)⑤The morphometric analysis of the kidney tissue suggested that the average diameter of renal corpuscle of high-fat-diet was evidently smaller than that of the controlled group ( 229.5126±37.3912 vs 262.4389±38.5034),the average perimeter of renal corpuscle of the high-fat-diet was evidently smaller than that of the controlled group ( 830.0191±136.0398 vs 974.8371±128.1884),the average diameter of proximal convoluted tubules of the high-fat-diet group was evidently smaller than that of the controlled group, (95.1354±16.1624 vs 102.2602±16.0461),and the average diameter of distal convoluted tubules of high-fat-diet group was evidently smaller than the controlled group. (78.6235±15.6967 vs 89.8674±21.7171). The difference was distinct (P<0.05).Conclusion①The amount of apoptosis in the kidney tissue of the a high-fat-diet rats increased;②The expression of Bcl-2 in the kidney tissue of high-fat-diet rats was weakened while the expression of Bax was reinforced.③The average diameter and the average perimeter of renal corpuscle of kidney tissue decreased.The average diameter of proximal convoluted tubules and distal convoluted tubules of the high-fat-diet rats decreased;④Apoptosis was probably one of the mechanism of toxic injury to the kidney induced by lipid.What’s more, the expression of Bcl-2, Bax and Bcl-2/Bax influenced the happening of apoptosis.更多还原