cGAMP与幽门螺杆菌蛋白抗原肌肉注射诱导BALB/c小鼠免疫应答初步分析

Preliminary analysis of immune response in BALB/c mice immunized with cyclic guanosine monophosphate-adenosine monophosphate and protein antigens from Helicobacter pylori by intramuscular injection

目的初步分析肌肉注射免疫环鸟-腺二核苷酸(cyclic guanosine monophosphate-adenosine monophosphate,cGAMP)与幽门螺杆菌(Helicobacter pylori,H.pylori)蛋白抗原在BLAB/c小鼠中诱导的免疫应答。方法将BALB/c小鼠随机分成4组,cGAMP组、抗原组(H.pylori蛋白抗原)、cGAMP+抗原组及对照组(未免疫),肌肉注射免疫小鼠。间接ELISA法测定免疫后小鼠血清中抗原特异性IgG抗体、胃组织及粪便上清液中IgA抗体;CCK-8法检测小鼠脾淋巴细胞体外增殖水平;酶联免疫斑点法(ELISpot)分析脾淋巴细胞分泌抗原特异性细胞因子水平。结果经肌肉注射免疫,cGAMP+抗原组可诱导小鼠产生显著高于对照组的血清IgG抗体应答(P<0.01);可诱导胃肠道黏膜产生显著高于单独抗原组的特异性IgA抗体应答(P<0.05);体外孵育H.pylori蛋白抗原和脾淋巴细胞,促进了淋巴细胞的增殖,cGAMP+抗原组显著高于单独抗原组(P<0.01);与单独抗原组及对照组比较,cGAMP+抗原组免疫小鼠诱导的脾淋巴细胞中抗原特异性IFNγ分泌细胞数量最多(P<0.01)。结论cGAMP通过肌肉注射途径可诱导BLAB/c小鼠对H.pylori蛋白抗原的体液免疫应答及脾脏中抗原特异性淋巴细胞免疫反应,其具有作为肌肉途径免疫H.pylori蛋白疫苗佐剂的潜质。

Objective To analyze the immune response against Helicobacter pylori(H.pylori)in BALB/c mice immunized with cyclic guanosine monophosphate-adenosine monophosphate(cGAMP)and protein antigens by intramuscular injection.Methods BALB/c mice divided randomly into 4 groups were immunized with the protein antigens of H.pylori,cGAMP,antigens+cGAMP,and saline(as the control group)respectively.Then,antigen-specific IgG antibody in serum and IgA in gastric tissue and faeces were detected by ELISA.The proliferation levels of murine spleen cells were measured in vitro with the CCK-8 assay,while the levels of antigen-specific cytokines secreted from murine spleen were quantified with the enzyme-linked immunospot assay(ELISpot).Results Intramuscular administration of protein antigens from H.pylori together with cGAMP induced specific IgG levels which were significantly higher than those in comparing to the control group(P<0.01),while the antigen-specific IgA response was higher than that in the antigen group(P<0.05).The proliferation level of lymphocytes from mice injected with both antigens and cGAMP were significantly higher than that in antigen group(each P<0.01).The levels of splenic IFNγ-producing lymphocytes in response to H.pylori antigens and cGAMP immunization were significantly higher than those in the corresponding groups without using cGAMP(each P<0.01).Conclusion By using cGAMP as an adjuvant,H.pylori antigens could elicit significantly stronger humoral and cellular immune responses in mice than those induced by antigens alone.The cGAMP might be used for the development of H.pylori vaccine for intramuscular injection as demonstrated by its stronger adjuvant activity.