摘要
目的 研究MF59佐剂[一种由角鲨烯、山梨糖醇三油酸酯（Span85）、吐温80和柠檬酸缓冲液组成的水包油乳剂]对热灭活BCG（hBCG）免疫小鼠诱导免疫应答的影响,验证自制MF59增强hBCG诱导细胞免疫反应的作用.方法 BALB/c （SPF级）小鼠分成A～G组,每组8只,于第0、2、4周皮下注射0.2 ml MF59、低剂量hBCG（0.025 mg/ml）、中剂量hBCG（0.25 mg/ml）、高剂量hBCG（2.5 mg/ml）、MF59＋低剂量hBCG、MF59＋中剂量hBCG、MF59＋高剂量hBCG.末次免疫2周,解剖小鼠.取小鼠脾细胞和腹腔巨噬细胞,并在体外经BCG PPD刺激培养,ELISA检测培养上清γ干扰素（IFN-γ）、白细胞介素（IL）-1β、Ib-2、IL-4、IL-12含量,酶联免疫斑点法（ELISPOT）检测脾细胞分泌IFN-γ、IL-2和IL-4的斑点形成细胞数（SFCs）.所有试验组的平均值比较采用一元方差分析、每两组平均值比较采用最小显著差异法,以P＜0.05为差异有统计学意义.结果 MF59作为高剂量hBCG的佐剂（G组）免疫小鼠,其脾细胞体外经BCG PPD刺激培养,分泌IFN γ的SFCs[（151.3±66.6）个]、IL-2的SFCs[（247.8±58.0）个]和IL-4的SFCs[（65.8±24.6）个]显著高于其他组[其他组IFN-γ、IL-2和IL-4的SFCs的最高平均值分别为（30.4±13.0）个、（37.1±10.8）个和（16.4±9.7个）]（分泌IFN-γ的SFCs比较,t=3.2,P=0.007;分泌IL-2的SFCs比较,t=3.6,P=0.003;分泌IL-4的SFCs比较,t=3.0,P=0.01）.MF59与不同剂量hBCG联合免疫小鼠,其腹腔巨噬细胞体外经BCG PPD刺激培养,培养上清IL-1β水平分别为（663.3±177.5） pg/ml、（813.8±193.4）pg/ml和（742.3±316.1）pg/ml,均显著高于A、B和C组[分别为（104.7±65.8）pg/ml、（82.9±54.8） pg/ml和（66.5±53.5） pg/ml]（E、F和G与A组比较：t=2.9,P=0.01;t=3.5,P=0.004;t=2.5,P=0.031.E、F和G与B组比较：t=3.1,P=0.007;t=3.6,P=0.003;t=2.6,P=0.024.E、F和G与C组比较：t=3.0,P=0.01;t=3.5,P=0.004;t=2.5,P=0.031）.A、C和G组小鼠的腹腔巨噬细胞体外经
Objective To study the effects of MF59 （an oil-in-water emulsion composed of small droplets of squalene surrounded by a monolayer of nonionic detergents （Span85 and Tween 80）） as adjuvant on immune responses to the heat-killed Bacillus Calmette-Guérin （hBCG） to prove MF59 enhancing the cellular immune responses to hBCG.Methods BALB/c mice were divided into seven groups,which were immunized with MF59 （group A）,low doses of hBCG （0.025 mg/ml） （group B）,middle doses of hBCG （0.25 mg/m1） （group C）,high doses of hBCG （2.5 mg/m1） （group D）,MF59＋ low doses of hBCG （group E）,MF59＋ middle doses of hBCG （group F） and MF59＋ high doses of hBCG （group G） for three times at intervals of two weeks,respectively.The mice were sacrificed two weeks after the last immunization.The splenic lymphocytes and peritoneal macrophages were isolated and cultured with BCG-PPD.Sandwich ELISA and ELISPOT assay were used to detect BCG-PPD-specific cytokines in culture supernatants and the number of IFN-γ,IL-2,IL-4 producing spot forming cells （SFCs）,respectively.One-way analysis of variance with least-significant-difference （LSD） comparisons was used to test differences among the levels of cytokines and the number of SFCs.P values of ＜ 0.05 were considered statistically significant.Results The number of IFN-γ,IL-2,IL-4 producing SFCs in group G （151.3±66.6,247.8±58.0,and 65.8± 24.6） was higher than that in other groups （the biggest means of IFN-γ,IL-2,IL-4 producing SFCs were （30.4± 13.0）,（37.1±10.8）,and（16.4±9.7）,respectively） （t=3.2,P =0.007 for IFN-γ producing SFCs;t =3.6,P =0.003 for IL-2 producing SFCs;t=3.0,P =0.01 for IL-4 producing SFCs）.The levels of IL-1β in the supernatants from BCG-PPD-stimulated peritoneal macrophages were higher in groups E （663.3 ± 177.5 pg/ml）,F （813.3 ± 193.4 pg/ml）,and G （742.3±316.1 pg/ml） than that in groups A （104.7±65.8 pg/ml）,B （82.9±54.8 pg/ml） and C （66.5±53.5pg/ml） （gro
出处
《中国防痨杂志》
CAS
2014年第6期440-446,共7页
Chinese Journal of Antituberculosis
