突变型单核细胞趋化蛋白-1构建及其对单核细胞趋化蛋白-1的干预作用  

Construction of mutated monocyte chemoattractant protein 1 and its intervention with monocyte chemoattractant protein 1

作　　者：孙改河[1] 沈燕[1] 张愔[1] 王喜英[1] 金雨琦[1] 苏天水[1] 

机构地区：[1]郑州大学第一附属医院检验科,郑州450052

出　　处：《郑州大学学报（医学版）》2007年第3期458-460,共3页Journal of Zhengzhou University(Medical Sciences)

基　　金：河南省科技攻关基金资助项目0496060902;河南省高校青年骨干教师资助项目资助

摘　　要：目的构建表达N端2-8位氨基酸缺失的突变型单核细胞趋化蛋白-1(mMCP-1),并观察其对单核细胞趋化蛋白-1(MCP-1)的干预作用。方法Megaprimer-PCR构建mMCP-1-pVAX1(pVMm)。以脂质体将pVMm转染真核细胞系C2C12,ELISA分析mMCP-1的表达情况。采用趋化实验分析mMCP-1对病毒性心肌炎小鼠外周血单个核细胞(PMNCs)的趋化活性及其对MCP-1的干预作用。结果①酶切和测序显示野生型MCP-1真核表达载体(pVM)和pVMm构建成功。pVMm在真核细胞C2C12中有效表达。②mMCP-1不具有趋化病毒性心肌炎小鼠PMNCs的活性。与单纯使用MCP-1相比,以不同浓度的mMCP-1和MCP-1同时行趋化实验,被趋化的细胞明显减少(P<0.01),25μg/LmMCP-1可阻断10μg/LMCP-1对病毒性心肌炎小鼠PMNC的趋化活性。结论小鼠MCP-1N端2-8位氨基酸决定其趋化活性。mMCP-1可用于干预MCP-1的作用。Aim： To construct and express N-terminus deleted monocyte chemoattractant protein-1 （mutated MCP-1, mMCP-1 ） and observe its biological activity. Methods ：The plasmid pVMm carrying murine mMCP-1 coding gene was construtted by Megaprimer-PCR. Lipofectamine was used to transfect pVMm into C2C12 and the expression of mMCP-1 was detected by ELISA. Chemoattractant activity of mMCP-1 to peripheral blood mononuclear cells（PMNCs） of mice with viral myocarditis and the intervention of mMCP-1 with the role of MCP-1 were analyzed by chemotaxis assay. Results： pVM carrying murine wild-type MCP-1 coding gene and pVMm were successfully constructed according to enzyme digestion and sequencing, mMCP-1 was expressed by C2C12 after transfected with pVMm. mMCP-1 was not able to chemoattract the PMNCs of mice with viral myocarditis. Compared with MCP-1 alone, the chemoattractant cells were reduced by different concentrations of mMCP-1 and 10 μg/L MCP-1 （ P 〈 0.01 ）. 25 μg/L mMCP-1 could block the chemoattractant activity of 10 p.g/L MCP-1 to PMNCs of mice with viral myocarditis. Conclusion： N-terminus 2-8 amino acid of murine MCP-1 is the region with chemoattract activity, mMCP-1 could interfere with the role of MCP-1.

关 键 词：单核细胞趋化蛋白-1 趋化因子 趋化活性 干预作用 

分 类 号：R446.61[医药卫生—诊断学]