乙肝核酸疫苗对荷瘤小鼠免疫保护作用的实验研究

目的 观察乙肝病毒(hepatitis B virus, HBV)核酸疫苗pCMV-S2S免疫小鼠后的特异性细胞毒性T淋巴细胞(CTL)应答, 及对表达HBV preS2S抗原的SP2/0-S2S肿瘤细胞攻击的保护作用.方法 将BALB/c小鼠随机分为3组, 于0、4及8周分别接种pCMV-S2S(实验组)、乙肝表面抗原蛋白疫苗(HBsAg,对照组1)、空载质粒( pCMV,对照组2 )各3次.末次免疫后4周,每组各取3只小鼠,以乳酸脱氢酶(LDH)释放法检测特异性CTL活性.各组其它小鼠均用于肿瘤细胞攻击实验:分别于一侧胁部皮下种植SP2/0-S2S细胞,同时于另一侧种植不表达HBV preS2S抗原的SP2/0-CMV 细胞作阴性对照.种植肿瘤细胞(荷瘤)后观察成瘤时间、肿瘤大小和荷瘤后小鼠的生存时间及生存率.结果 pCMV-S2S组CTL活性为(34.21±1.38)%,高于HBsAg组(19.64±1.50)%]和pCMV组(3.45±1.89)%](P＜0.05).荷瘤后10 d,pCMV-S2S组小鼠SP2/0-S2S细胞的成瘤率为58.3%,低于SP2/0-CMV细胞的成瘤率(91.7%)(P＜0.05);两对照组SP2/0-S2S细胞与SP2/0-CMV细胞的的成瘤率无明显差异.荷瘤后pCMV-S2S组初次出现死亡小鼠的时间为24 d,较两对照组均延迟了13 d,平均生存时间为(31±1) d,较两对照组延长(P＜0.05);4周生存率(75%)高于两对照组(P＜0.05), 两对照组之间生存时间及生存率差异无统计学意义.结论 HBV核酸疫苗pCMV-S2S能在小鼠体内诱生较强的特异性CTL活性,对荷瘤小鼠具有特异性免疫保护作用,可望用于HBV感染的免疫保护.

Immunoprotection Role of a Hepatitis B Virus DNA Vaccine Responsible to BALB/c mice

OBJECTIVE: To investigate the specific immune response and the immunoprotection effects of cytotoxic T-lymphocyte (CTL), which is induced by hepatitis B virus (HBV) DNA vaccine pCMV-S2S, on BALB/c mouse with the attack of SP2/0-S2S cells, a BALB/c mouse myeloma cell line stably expressing the HBV preS2S antigen. METHODS: BALB/c mice were divided into 3 groups: experimental group (intramuscularly injected with pCMV-S2S), control group 1 (HBsAg vaccine immunizing) or control group 2 (plasmid pCMV immunizing), which were immunized thrice at week 0.4 and 8 respectively. Specific CTL activities of 3 mice of each group were measured by lactate dehydrogenase (LDH) release assays, 4w after the last boost. The other mice of each group were subcutaneously inoculated with SP2/0-S2S cells into one lateral of flanks. SP2/0-CMV cell without expressing preS2S was subcutaneously inoculated into another lateral of mouse flanks. The time of visible tumor formation and the size of tumor were recorded and observed. The life span and survival rate of mice after loaded with the tumor cells were analyzed by Log-Rank statistics and Kaplan-Meier Survival curve respectively. RESULTS: The specific CTL lysis value of pCMV-S2S group mice was (34.21 +/- 1.38)%, higher than that of HBsAg group (19.64 +/- 1.50)%] or pCMV group (3.45 +/- 1.89)%] (P < 0.05). The visible SP2/0-S2S tumor formation rate in pCMV-S2S group mice was 58.3% lower than that in the pCMV group (91.7%), 10 d after inoculated with the tumor cells (P < 0.05). The tumor formation rates of the two control groups were no obvious difference. The date of the first mouse dying in pCMV-S2S group was 24d, delayed for 13d, compared with that in the two control groups. The average life span of pCMV-S2S group mice after loaded with the tumor cells was (31 +/- 1) d, longer than that of the two control groups (P < 0.05). The 4w survival rate of the pCMV-S2S group mice was obviously increased (75%), compared with the two control groups (P < 0.05). The average life span and the 4w survival rate of the two control group mice were no obvious difference. CONCLUSION: The hepatitis B virus DNA pCMV-S2S vaccination may elicit substantial HBV preS2S-specific CTL response in vivo, which gives the mouse the immunoprotection against the attack of SP2/0-S2S cells. The results indicate that pCMV-S2S may be used to the immunoprotection from HBV infection.