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目的及方法为探求 HBV 抗原特异性细胞毒 T 细胞(HBV-CTL),根椐 HBcAg 的 HLA 分子结合关键序列合成抗原多肽2段,分别与转铁蛋白(Tf)、抗 CD3单克隆抗体(CD3mAb)和 IL-2联合体外诱导 HBV-CTL,应用~3H-TdR 释放法分别测定 HBV-CTL 对 HBV DNA 转染的 HepG 2.2.15细胞特异性伤活性及无 HBV DNA 转染的 HePG2细胞的非特异性杀伤率。结果 HBV-CTL 对 HBV DNA 转染的2.2.15细胞特异性杀伤率分别为67%(多肽Ⅰ+Tf+CD3mAb+IL-2)、55%(多肽Ⅱ+Tf+CD3mAb+IL-2),明显高于 LAK 细胞组20%(IL-2)及 CD3-AK 细胞组34%(IL-2+CD3mAb+Tf)(P值<0.05)。而HBV-CTLs 对无 HBV DNA 转染的 HepG2细胞非特异性杀伤率,与 LAK 细胞及 CD3-AK 细胞相比,无显著性差异(P值>0.05)。HBV-CTLs 培养14-21天后,扩增倍数比 LAK 细胞高10倍,细胞表型分析显示 HBV-CTLs 是以 CD8+为主的异质细胞群。结论 HBcAg 合成多肽联合 Tf CD3mAb 及 IL-2,体外诱导 HBV-CTL,既可显著提高其增殖活性及增殖倍数,又可明显增强 HBV-CTL 对 HBV DNA 转染的2.2.15细胞的特异性细胞毒杀伤活性.
Objectives and Methods To explore the HBV antigen-specific cytotoxic T-cells (HBV-CTL), two antigenic peptides were synthesized based on the HLA sequences of HBcAg and Tc, CD3 mAb ) And IL-2 were induced in vitro. HBV-CTL specific cytotoxicity against HBV DNA-transfected HepG 2.2.15 cells and HepG2 cells transfected with no HBV DNA were determined by ~ 3H-TdR release assay Heterosexual killing rate. Results The specific killing rates of HBV-CTL on 2.2.15 cells transfected with HBV DNA were 67% (polypeptide I + Tf + CD3 mAb + IL-2), 55% (peptide II + Tf + CD3 mAb + IL- 2) (IL-2 + CD3mAb + Tf) was significantly higher in 20% (IL-2) and CD3-AK cell groups than in LAK cell group (P <0.05). However, non-specific cytotoxicity of HBV-CTLs on HepG2 cells transfected with HBV DNA was not significantly different from that of LAK cells and CD3-AK cells (P> 0.05). After cultured for 14-21 days, the amplification of HBV-CTLs was 10-fold higher than that of LAK cells. The phenotypic analysis of HBV-CTLs showed that CD8 + -based heterogeneous cell populations were found in HBV-CTLs. Conclusion The combination of HBcAg with Tf CD3 mAb and IL-2 can induce the proliferation of HBV-CTL in vitro and significantly enhance the specific cytotoxicity of HBV-CTL to 2.2.15 cells transfected with HBV DNA Toxic activity.