两种GTF氨基催化区改良防龋质粒免疫小鼠的实验研究  

Immunization of Mice with Two Modified Anti-caries Plasmids Encoding Different Styles of S. sobrinus GTF Catalytic Region

作　　者：孙静华[1] 樊明文[1] 

机构地区：[1]武汉大学口腔生物医学工程教育部重点实验室,湖北武汉430079

出　　处：《口腔医学研究》2011年第2期89-92,96,共5页Journal of Oral Science Research

基　　金：国家自然科学基金资助项目(编号:30672325)

摘　　要：目的:通过比较编码表兄链球菌葡糖基转移酶氨基催化区CAT不同形式基因序列的两种改良防龋质粒免疫BALB/c小鼠后激发的特异性抗体水平差异,初步评价不同形式CAT在DNA防龋疫苗中的应用潜力。方法:利用分子克隆技术构建两种改良防龋质粒,在靶向防龋DNA疫苗pGJA-P/VAX中插入表兄链球菌OMZ176GTF-I CAT全基因序列构建pGJGAC/VAX;将基因公司合成的共计165bp的串联重复5次的GTF-I氨基催化区核心保守序列克隆到pGJA-P/VAX中构建pGJGA-5C/VAX。转染CHO细胞系,检测其表达。两种改良质粒经鼻腔滴注免疫BALB/c小鼠,检测血清和唾液中的特异性抗PAc,抗GLU和抗CAT抗体水平。结果:重组质粒的基因序列与预期相符。不同形式CAT基因序列的两种改良防龋质粒均可在真核细胞正确表达。pGJGAC/VAX和pGJGA-5C/VAX免疫动物后血清和唾液特异性抗PAc、抗GLU和抗CAT抗体均显著高于空载体pVAX1免疫组(P<0.05)。第10、12和14周,pGJGAC/VAX免疫组小鼠的血清特异性抗CAT抗体显著高于pGJGA-5C/VAX组(P<0.05)。两组间唾液特异性抗体未见显著性差异。pGJGA-5C/VAX在小鼠体内激发特异性抗体的速度较pGJGAC/VAX略快。结论:本实验构建的增加表兄链球菌GTF氨基催化区的两种改良防龋质粒,可在真核细胞中正确表达,两种质粒有效地诱导黏膜和系统体液免疫反应,pGJGAC/VAX和pGJGA-5C/VAX各有优势。Objective：To assess the potential of two different styles of S.sobrinus GTF catalytic region in an anti-caries DNA vaccine by evaluating the levels of the specific anti-PAc,anti-GLU,and anti-CAT antibodies in immunized mice.Methods：Two modified anti-caries DNA plasmids pGJGAC/VAX and pGJGA-5C/VAX by cloning different styles of the catalytic region of GTFs into the previous plasmid pGJA-P/VAX.The CHO cells were transfected and expression of fusion protein were detected using cell immunohistochemistry.Mice were immunized with pGJGAC/VAX,pGJGA-5C/VAX and pVAX1 via the intranasal route.Blood and saliva samples were collected every 2 weeks interval for assay of antibodies by ELISA.Results：The expressed protein of the two new recombinant plasmids could response to specific anti-PAc,anti-GLU and anti-CAT antibodies respectively.As for the antibody reactions,increase in the serum IgG and saliva IgA anti-PAc,anti-GLU and anti-CAT antibodies responses were seen in the pGJGAC/VAX group and the pGJGA-5C/VAX group after the second immunization（P〈0.05）.After 10,12 and 14 weeks,the anti-CAT IgG antibodies of pGJGAC/VAX group were higher than that of pGJGA-5C/VAX group（P〈0.05）.Conclusion：The two modified anti-caries plasmids encoding different styles of GTF CAT antigens could induce effective mucosal and systematic humoral responses.These two modified plasmids had different predominance.

关 键 词：龋齿 疫苗 DNA 表兄链球菌 葡糖基转移酶 

分 类 号：R780.1[医药卫生—口腔医学]