干酪乳杆菌细胞壁成分诱导小鼠自身免疫性心肌炎肾上腺髓质素的表达

Experimental Study of Adrenomedullin in Autoimmune Myocarditis Induced by Immunization of Mice with Lactobacillus Casei Cell Wall Element

目的探计肾上腺髓质素(AM)mRNA和蛋白在干酪乳杆菌细胞壁成分(LCWE)诱导小鼠自身免疫性心肌炎动物模型心肌组织中的表达水平和意义。方法4～6周Balb/c雄性小鼠45只,随机分为实验组30只,对照组15只。实验组于d0、3、5、10经腹部皮下注射LCWE,对照组以磷酸盐缓冲溶液(PBS)代替LCWE。于实验d14、21、28分别留取血清和心肌标本。采用RT-PCR及免疫组织化学方法检测小鼠心肌组织AMmRNA及蛋白表达水平,并检查心肌组织病理变化。ELISA法检测小鼠血清抗肌凝蛋白抗体。结果实验组小鼠心肌炎性细胞浸润及心肌细胞变性坏死较明显,对照组心肌组织正常。实验组血清存在高浓度抗肌凝蛋白抗体。免疫组织化学显示,实验组AM表达水平显著高于对照组(P<0.05),与病变积分呈显著正相关(r=0.81P<0.01)。RT—PCR结果显示,实验组小鼠心肌AMmRNA表达水平高于对照组(P<0.05),与病变积分呈显著正相关(r=0.71P<0.01)。结论AM参与LCWE诱导小鼠自身免疫性心肌炎,心肌AM表达水平可作为判断心肌炎性损伤程度的客观指标。

Objective To study adrenomedullin（AM） mRNA and protein expression level in myocardium of autoimmune myocarditis animal models induced by immunization of mice with lactobacillus casei cell wall element（ LCWE）. Methods Forty-five Balb/c male mice were randomly divided into experimental group （ n = 30） and control group （ n = 15） ,which were intraperitoneally injected with LCWE and phosphate buffered solution（PBS） at day 0,3,5 and 10, respectively. Sera and myocardium ssrnples were gained 14,21 and 28 days after the first immunization. AM expression levels were determined by semiquantitative reverse transcriptase-polymerase chain reaction（RT- PCR） and immunchistochemistry, and mycardial histopathological lesions were observed. The anti-myosin antibodies in different stages were examined by an ELISA. Results There were myocardial necrosis or inflammatory infiltration in the experimental group, but myocardial lesions were not found in the control group. Anti-myosin antibodies were detected in sera of experimental mice, but not in control group. Immunchistochemistry findings demonstrated that AM expression level was higher in the experimental group than in the control group（ P 〈 0.05）. The dynamic changes of AM protein expression showed a significantly positive correlation with the changes of myocardial histopathologic scores（ r = 0.81 P〈 0.01 ）. RT - PCR findings demonstrated that AM rnRNA expression level was higher in the myocardium of experimental group than in the control group（ P 〈 0.05）. The dynamic changes of AM rnRNA expression showed a significantly positive correlation with the changes of myocardial histopathologic ,scores（ r = 0.71 P 〈 0.01 ）. Conclusions AM is involved in autoimmune myocarditis mice models induced by LCWE. The expression level of AM can be regarded as a objective indicator for myocardial damage degree.