氢氧化铝对人乳头瘤病毒16型L2E6E7融合蛋白免疫增强作用的初步研究

Adjuvant effects of aluminum hydroxide on a recombinant human papillomavirus 16L2E6E7 fusion protein

目的 初步评价Al(OH)3对重组HPV16的L2E6E7融合蛋白(简称HPV16L2E6E7)的免疫佐剂作用.方法 将108只雌性C57BL/6小鼠随机分成Al(OH)3组(85.7 μg/只,肌肉注射)、HPV16L2E6E7组(120.0μg/只,肌肉注射)、A1 (OH)3+HPV16L2E6E7组[每只 Al(OH)3 85.7μg+HPV16L2E6E7120.0μg,肌肉注射],每组36只,免疫程序为0、3、7d.ELISPOT法检测IFN-γ,间接ELISA法检测IgG抗体水平.另将40只皮下接种TC-1肿瘤细胞(1×104/只)的雌性C57BL/6小鼠随机分为PBS组(100.0 μL)、Al (OH)3组(85.7μg/只,肌肉注射)、HPV16L2E6E7组(120.0μg,/只,肌肉注射)、Al(OH)3+ HPV16L2E6E7组[每只Al(OH)385.7μg+HPV16L2E6E7 120.0μg,肌肉注射],每组10只.免疫程序为0、3、7d.观察肿瘤形成时间、体积变化及成瘤率,观察期为53 d.结果 免疫后第10、14、21、28、35和42d,HPV16L2E6E7组与Al(OH)3+HPV16L2E6E7组IgG抗体滴度都高于Al(OH)3对照组,差异具有统计学意义(P均＜0.01),免疫后21、28、35、42 d Al(OH)3+ HPV16L2E6E7组特异性IgG抗体显著高于HPV16L2E6E7组(P均＜0.01);免疫后14、21、28d Al(OH)3+ HPV16L2E6E7组E7特异性IFN-γ水平显著高于HPV16L2E6E7组(P均＜0.01);小鼠抑瘤实验中,HPV16L2E6E7组抑瘤率为60％,Al(OH)3+ HPV16L2E6E7抑瘤率为40％,两组无明显差异(P＞0.05),但与对照组比较都有显著性差异(P＜ 0.01,P＜0.05).结论 初步评价Al(OH)3能与HPV16L2E6E7较好吸附,可诱导C57BL/6雌性小鼠产生更强的特异性体液和细胞免疫应答,具有免疫增强作用.

Objective To evaluate the adjuvant effects of Al(OH)3 on a recombinant human papillomavirus 16L2E6E7 fusion protein (HPV16L2E6E7).Methods There were 108 female C57BL/6 mice randomly divided into Al(OH)3 group [85.7 μg for each mouse,intramuscular injection (im)],HPV16L2E6E7 group (120.0 μg for each mouse,im) and Al(OH)3 + HPV16L2E6E7 group (85.7μg Al(OH)3 and 120.0 μg HPV16L2E6E7 for each mouse,im),36 mice in each group,and the injection were given on 0,3,7 d.ELISA and ELISPOT were used to detect IgG and IFN-γ levels.Forty female C57BL/6 mice subcutaneously inoculated with 1 × 104 TC-1 tumor cells were randomly divided into PBS group (100.0 μL,im),Al(OH)3 group (85.7 μg for each mouse,im),HPV16L2E6E7 group (120.0μg for each mouse,im),Al(OH)3 + HPV16L2E6E7 (85.7 μg Al(OH)3 and 120.0μg HPV16L2E6E7 for each mouse,im),10 mice in each group,the injection were given on 0,3,7 d.The tumor formation time,tumor volume and antitumor rates were recorded throughout 53-day observation.Results The IgG titers of HPV16L2E6E7 and Al (OH)3 + HPV16L2E6E7 group were significantly higher than control group (P all ＜ 0.01) on 10,14,21,28,35 and 42 d.The specific IgG antibody titers of Al(OH)3 + HPV16L2E6E7 group were significantly higher than HPV16L2E6E7 group (P all ＜0.01) on 21,28,35,42 d.The IFN-γ levels of Al(OH)3 + HPV16L2E6E7 group were significantly higher than HPV16L2E6E7 group (P all ＜ 0.01) on 14,21,28 d.In TC-1 cell tumor model of female C57BL/6 mice,the antitumor rate of HPV16L2E6E7 was 60％ and it decreased to 40％ in Al(OH)3 + HPV16L2E6E7 group (P ＞ 0.05).Compared to the control group,there were no significant differences (P ＜ 0.01,P ＜ 0.05).Conclusions HPV16L2E6E7 fusion protein accompanied by Al(OH)3 adjuvant can induce stronger antigen-specific humoral and cellular immune responses which demonstrates that it can improve the immunogenicity of HPV16L2E6E7.