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目的观察脂联素基因敲除小鼠主动脉组织病理学和组织化学的特性。方法SPF级6周龄雄性脂联素基因敲除小鼠纯合子(Adiponectin-/-)30只随机分为5组,第1、2、3组分别给予普通膳食喂养10、20、30周,第4组每2周经颈静脉注射空白腺病毒载体(即β-半乳糖甘酶腺病毒载体)并予普通膳食喂养30周,第5组每2周经颈静脉注射重组脂联素腺病毒载体并予普通膳食喂养30周;另随机选取SPF级6周龄雄性野生型小鼠(WT)6只为正常对照组,给予普通膳食喂养30周。采用酶法测定血糖浓度,放射免疫法测定血浆胰岛素和脂联素水平。分离小鼠胸主动脉置4%多聚甲醛固定,石蜡包埋,连续切片,行茜素红钙化染色。分离主动脉弓到髂骨分支的动脉,用比色法测定10%甲酸抽提的钙含量。超声破碎胸主动脉,用Bradford法测总蛋白含量,离心后取上清液采用对硝基苯酚法测定ALP活性。结果经普通膳食喂养的各组脂联素基因敲除小鼠与野生型小鼠在体重、血糖、血胰岛素水平方面无明显区别。与野生型小鼠、喂养10及20周的脂联素基因敲除小鼠相比,喂养30周的脂联素基因敲除小鼠出现了轻度的动脉钙化,其动脉的钙含量及ALP活性升高。通过对脂联素基因敲除小鼠进行外源性脂联素的补充,抑制了动脉钙化的出现及ALP活性升高。结论在普通膳食喂养30周后,脂联素基因敲除小鼠出现轻度的动脉钙化,其机制可能与动脉中升高的ALP活性有关;外源性脂联素的补充可抑制脂联素基因敲除小鼠动脉钙化的发生,提示脂联素为动脉钙化的保护因子。
Objective To observe the histopathological and histochemical features of aorta in mice with adiponectin knockout mice. Methods Thirty male Sprague - Dawley (Adiponectin - / -) mice of 6 weeks old were randomly divided into 5 groups. Groups 1, 2 and 3 were fed with normal diet for 10, 20 and 30 weeks, In group 4, blank adenovirus vector (ie, β-galactosidase adenovirus vector) was injected into the jugular vein every 2 weeks and fed for 30 weeks. Group 5 received intraperitoneal injection of recombinant adenovirus The animals were fed with common diet for 30 weeks. Six 6-week-old male wild-type mice (WT) of SPF grade were randomly selected as normal control group and fed with normal diet for 30 weeks. Serum glucose concentration was determined by enzymatic method and plasma insulin and adiponectin levels were determined by radioimmunoassay. Isolated mouse thoracic aorta 4% paraformaldehyde fixed, embedded in paraffin, serial sections, line alizarin red calcification. The aortic arch was dissected into the artery of the ilium branch and the calcium content extracted by 10% formic acid was determined by colorimetry. The thoracic aorta was sonicated. The content of total protein was measured by Bradford method. After centrifugation, the supernatant was taken and ALP activity was measured by p-nitrophenol method. Results Adiponectine knockout mice and wild type mice fed with ordinary diet had no significant differences in body weight, blood glucose and blood insulin levels. Adiponectine knockout mice that had been fed for 30 weeks had mild arterial calcification, calcium content in their arteries, and ALP, compared with wild type mice fed adiponectin knockout mice at 10 and 20 weeks Activity increased. Supplementation of exogenous adiponectin to adiponectin knockout mice inhibits the appearance of arterial calcification and increases ALP activity. CONCLUSIONS: Adiponectine knockout mice show mild arterial calcification after 30 weeks of general diet, and the mechanism may be related to the increased ALP activity in the arteries. Exogenous adiponectin supplementation can inhibit adiponectin The occurrence of arterial calcification in knockout mice suggests that adiponectin is a protective factor for arterial calcification.