CAJ | 학술논문

结核分枝杆菌Rpf-DNA疫苗对Mtb感染小鼠免疫保护作用的研究
결핵분지간균Rpf-DNA역묘대Mtb감염소서면역보호작용적연구
Rpf-DNA vaccine for M. tuberculosis infected mice

万方数据

中国防痨杂志中國防癆雜誌 중국방로잡지
BULLETIN OF THE CHINESE ANTITUBERCULOSIS ASSOCIATION
2012年 8期 532-537 ,共6页

邢爱英%刘忠泉%刘洋%贾红彦%李自慧%郑晓静%曹廷明%杜凤娇%杜博平%古淑香%张宗德

형애영%류충천%류양%가홍언%리자혜%정효정%조정명%두봉교%두박평%고숙향%장종덕

分枝杆菌,结核%细菌蛋白质类%细胞因子类%疫苗,DNA 分枝桿菌,結覈%細菌蛋白質類%細胞因子類%疫苗,DNA
분지간균,결핵%세균단백질류%세포인자류%역묘,DNA

目的评价Mtb复苏因子DNA疫苗(resuscitation-promoting factor DNA,Rpf-DNA)对Mtb感染宿主的免疫保护作用.方法构建复苏因子D(Rpf D-DNA)和复苏因子E(Rpf E-DNA)质粒疫苗,180只BALB/c 小鼠(4周龄)分为6组,每组30只,分别用空质粒、Rpf D-DNA质粒、Rpf E-DNA质粒、BCG、生理盐水免疫,H37Rv标准株气溶胶感染.检测血清复苏因子(Rpf)抗体、IFN-γ; RpfD、E刺激淋巴细胞后进行淋巴细胞增殖、细胞杀伤实验；检测细胞上清IFN-γ、IL-12、IL-2;对肺组织进行病理学检测和细菌负荷(CFU)计数.结果 (1)疫苗免疫组血清Rpf D、Rpf E抗体水平；Rpf D组0.32±0.1,Rpf E组0.39±0.1,BCG组0.02±0.01,空质粒组0.01±0.0,生理盐水组0.09±0.04,空白对照组0.03±0.01,RpfD组与RpfE组抗体水平与BCG组、空质粒组、生理盐水组、空白对照组比较差异有显著统计学意义,F值分别为Rpf D:45.6,43.2,45.1,45.7；Rpf E:51.6,53.6,51.0,52.2;P值均＜0.01.血清IFN-γ:Rpf D组(43.9±24.8)pg/ml,Rpf E组(45.9±21.0)pg/ml,BCG组(21.0±11.0)pg/ml,空质粒组(7.9±4.9) pg/ml,生理盐水组(4.7±2.1) pg/ml,空白对照组(5.8±4.7)pg/ml,RpfD、RpfE质粒组与BCG组比较差异有统计学意义(F值分别为Rpf D:10.2；Rpf E:12.4;P值均＜0.05),与空质粒组、生理盐水组、空白对照组比较差异有显著统计学意义,F值分别为Rpf D:15.6,17.8,17.3;Rpf E:17.5,21.1,20.7;P值均＜0.01.(2)淋巴细胞增殖实验CCK-8:Rpf D组176.0±4.2,Rpf E组183.0±4.3,BCG组101.0±1.1,空质粒组100.0±6.8,生理盐水组104.0±8.4,空白对照组116.0±2.2,RpfD、RpfE质粒组与BCG组、空质粒组、生理盐水组、空白对照组比较差异有统计学意义,F值分别为Rpf D:9.5,10.1,10.0,9.0;Rpf E:11.2,12.9,11.7,10.3;P值均＜0.05.细胞杀伤实验:Rpf D组32.0±3.2,Rpf E组30.0±4.2,BCG组16.0±5.9,空质粒组3.3±1.5,生理盐水组6.7±0.5,空白对照组7.3±3.5,Rpf D、Rpf E质粒组与BCG组、空质粒组、生理盐水组、空白对照组比较差异有统计学意义,F值分别为Rpf D:8.8,14.5,13.7,11.9;Rpf E:8.1,12.5,11.6,11.1;P值均＜0.05.(3) Rpf D、Rpf E蛋白各自刺激淋巴细胞后上清细胞因子检测:IL-2:Rpf D组9.5±2.4,RpfE组9.2±1.2,BCG组2.4±2.1,空质粒组1.2±0.3,生理盐水组1.8±1.0,空白对照组1.5±0.7,Rpf D、Rpf E质粒组与其他各组比较差异有统计学意义,F值分别为Rpf D:12.5,14.6,13.5,13.9；Rpf E:12.0,13.6,13.1,13.2;P值均＜0.05.IFN-γ:Rpf D组22.2±5.7,Rpf E组28.7± 14.4,BCG组16.1±10.1,空质粒组9.8±1.6,生理盐水组13.2±2.1,空白对照组15.7±2.9,RpffD、RpfE质粒组与其他各组比较差异有统计学意义,F值分别为Rpf D:7.8,12.6,8.7,8.3;Rpf E:11.3,16.4,14.7,14.2;P值均＜0.05.结论实验表明复苏因子疫苗能诱导Mtb感染小鼠产生体液免疫和细胞免疫,有希望成为结核病候选疫苗之一.
목적 평개Mtb복소인자DNA역묘(resuscitation-promoting factor DNA,Rpf-DNA)대Mtb감염숙주적면역보호작용.방법 구건복소인자D(Rpf D-DNA)화복소인자E(Rpf E-DNA)질립역묘,180지BALB/c 소서(4주령)분위6조,매조30지,분별용공질립、Rpf D-DNA질립、Rpf E-DNA질립、BCG、생리염수면역,H37Rv표준주기용효감염.검측혈청복소인자(Rpf)항체、IFN-γ; RpfD、E자격림파세포후진행림파세포증식、세포살상실험；검측세포상청IFN-γ、IL-12、IL-2;대폐조직진행병이학검측화세균부하(CFU)계수.결과 (1)역묘면역조혈청Rpf D、Rpf E항체수평；Rpf D조0.32±0.1,Rpf E조0.39±0.1,BCG조0.02±0.01,공질립조0.01±0.0,생리염수조0.09±0.04,공백대조조0.03±0.01,RpfD조여RpfE조항체수평여BCG조、공질립조、생리염수조、공백대조조비교차이유현저통계학의의,F치분별위Rpf D:45.6,43.2,45.1,45.7；Rpf E:51.6,53.6,51.0,52.2;P치균＜0.01.혈청IFN-γ:Rpf D조(43.9±24.8)pg/ml,Rpf E조(45.9±21.0)pg/ml,BCG조(21.0±11.0)pg/ml,공질립조(7.9±4.9) pg/ml,생리염수조(4.7±2.1) pg/ml,공백대조조(5.8±4.7)pg/ml,RpfD、RpfE질립조여BCG조비교차이유통계학의의(F치분별위Rpf D:10.2；Rpf E:12.4;P치균＜0.05),여공질립조、생리염수조、공백대조조비교차이유현저통계학의의,F치분별위Rpf D:15.6,17.8,17.3;Rpf E:17.5,21.1,20.7;P치균＜0.01.(2)림파세포증식실험CCK-8:Rpf D조176.0±4.2,Rpf E조183.0±4.3,BCG조101.0±1.1,공질립조100.0±6.8,생리염수조104.0±8.4,공백대조조116.0±2.2,RpfD、RpfE질립조여BCG조、공질립조、생리염수조、공백대조조비교차이유통계학의의,F치분별위Rpf D:9.5,10.1,10.0,9.0;Rpf E:11.2,12.9,11.7,10.3;P치균＜0.05.세포살상실험:Rpf D조32.0±3.2,Rpf E조30.0±4.2,BCG조16.0±5.9,공질립조3.3±1.5,생리염수조6.7±0.5,공백대조조7.3±3.5,Rpf D、Rpf E질립조여BCG조、공질립조、생리염수조、공백대조조비교차이유통계학의의,F치분별위Rpf D:8.8,14.5,13.7,11.9;Rpf E:8.1,12.5,11.6,11.1;P치균＜0.05.(3) Rpf D、Rpf E단백각자자격림파세포후상청세포인자검측:IL-2:Rpf D조9.5±2.4,RpfE조9.2±1.2,BCG조2.4±2.1,공질립조1.2±0.3,생리염수조1.8±1.0,공백대조조1.5±0.7,Rpf D、Rpf E질립조여기타각조비교차이유통계학의의,F치분별위Rpf D:12.5,14.6,13.5,13.9；Rpf E:12.0,13.6,13.1,13.2;P치균＜0.05.IFN-γ:Rpf D조22.2±5.7,Rpf E조28.7± 14.4,BCG조16.1±10.1,공질립조9.8±1.6,생리염수조13.2±2.1,공백대조조15.7±2.9,RpffD、RpfE질립조여기타각조비교차이유통계학의의,F치분별위Rpf D:7.8,12.6,8.7,8.3;Rpf E:11.3,16.4,14.7,14.2;P치균＜0.05.결론 실험표명복소인자역묘능유도Mtb감염소서산생체액면역화세포면역,유희망성위결핵병후선역묘지일.