重组结核疫苗AEC/BC02诱导豚鼠的Ⅰ型超敏反应

Type I hypersensitivity induced by recombinant tuberculosis vaccine AEC / BC02 in guinea pigs

目的评价重组结核疫苗AEC/BC02诱导豚鼠Ⅰ型超敏反应炎性介质的释放。方法将18只雄性豚鼠分成3组:AEC/BC02疫苗组、阴性对照组和阳性对照组,每组6只,分别经后腿肌肉注射AEC/BC02疫苗、生理盐水和卵清蛋白(ovalbumin,OVA),共7针,间隔10 d。于末次免疫后14 d,各组豚鼠经心脏采血,收集抗凝血,加入不同刺激物;豚鼠采血后,吸入二氧化碳致死,分离豚鼠气管,置于含不同刺激物的KBS缓冲液中;剪开分离气管后的豚鼠腹腔,分别灌入含不同刺激物的KBS缓冲液;AEC/BC02疫苗组用重组蛋白Ag85b、EC及BCG-CpG-DNA(简称CpG)刺激,阴性对照组未刺激,阳性对照组用OVA蛋白刺激。按组胺检测试剂盒和半胱氨酰白三烯检测试剂盒说明书,采用ELISA法检测外周血、气管和腹腔特异性刺激后释放的组胺和半胱氨酰白三烯含量。结果 AEC/BC02疫苗组外周血释放的组胺含量(41.0±23.4)ng/ml与阳性对照组(46.4±28.2)ng/ml比较,差异无统计学意义(P>0.05);但气管和腹腔肥大细胞释放的组胺含量[气管:(27.4±10.8)ng/ml和腹腔:(9.7±8.1)ng/ml]均明显低于阳性对照组[气管:(71.4±21.1)ng/ml,腹腔:(23.5±12.4)ng/ml)],释放的半胱氨酰白三烯含量[气管:(236.4±63.5)pg/ml,腹腔:(173.4±31.3)pg/ml]也均明显低于阳性对照组[气管:(458.0±208.2)pg/ml,腹腔:(259.2±30.4)pg/ml],差异均有统计学意义(P<0.05)。结论肌肉注射途径下,AEC/BC02疫苗给药7针不会诱导豚鼠严重的Ⅰ型超敏反应。

Objective To evaluate the release of inflammatory mediators of type I hypersensitivity induced by recombinant tuberculosis vaccine AEC/BC02 in guinea pigs. Methods Eighteen male guinea pigs were divided into three groups, 6 for each. The guinea pigs in test group were injected i.m. with AEC/BC02 vaccine in behind limbs, while those in negative control group with physiological saline, and those in positive control group with ovalbumin （OVA）, for 7 times each at an interval of 10 d. Heart blood samples were collected from various groups 14 d after the last injection and added with various stimulators. The guinea pigs were killed by inhalation with carbon dioxide, of which the tracheas were isolated and put into the KBS buffer containing various stimulators. The abdominal cavities of guinea pigs after removal of tracheas were poured with KBS buffer containing various stimulators. Recombinant Ag85b, EC and BCG-CpG-DNA were used as stimulators in test group, while OVA in positive control group, and no stimulators in negative control group. The histamine and cysteinyl leukotriene contents in peripheral blood, trachea and abdominal cavity were determined by ELISA using the corresponding kits. Results The histamine content released from peripheral blood in AEC/BC02 group was （41.0 ± 23. 4） ng/ml, which showed no significant difference with that in positive control group （46. 4 ± 28.2） ng/ml （P 〉 0. 05）, while those from trachea [（27.4 ± 10. 8） ng/ml] and abdominal cavity [（9.7 ± 8. 1 ） ng/mll were significantly lower than those in positivecontrol group [trachea： （71.4 ± 21. 1） ng/ml; abdominal cavity： （23. 5 ± 12. 4） ng/mlJ （P 〈 0. 05）. The cysteinyl leukotriene contents released from trachea [（236. 4 ± 63.5） pg/ml] and abdominal cavity [（173.4 ± 31.3） pg/ml] in test group were also significantly lower than those in positive control group [trachea： （458. 0 ± 208. 2） pg/ml; abdominal cavity： （259.2 ± 30. 4） pg/ml ] （P 〈 0. 05 ）. Conclusion The intramuscular injection with AEC/BC02 vaccine for 7 times induced no severe type I hypersensitivity in guinea pigs.