中国寄生虫学与寄生虫病杂志

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细菌脂多糖诱导小鼠B细胞对日本血吸虫发育的影响

汤宏斌1,程腊英1,熊涛2,董惠芬2 *,蒋明森2   

  1. 1 武汉大学动物实验中心,武汉430071;2 武汉大学基础医学院人体寄生虫学教研室,武汉430071
  • 出版日期:2015-10-30 发布日期:2016-01-06

The Effect of Lipopolysaccharide-Induced Mouse B Cell Activation on Schistosoma japonicum Development

TANG Hong-bin1,CHENG La-ying1,XIONG Tao2,DONG Hui-fen2 *,JIANG Ming-sen2   

  1. 1 Center for Animal Experiment of Wuhan University/Animal Biosafety Level-3 Laboratory,Wuhan 430071,China; 2 Department of Medical Parasitology,School of Basic Medical Science,Wuhan University,Wuhan 430071,China
  • Online:2015-10-30 Published:2016-01-06

摘要:

目的  研究细菌脂多糖诱导BALB/c小鼠B淋巴细胞对日本血吸虫发育的影响。  方法  取T细胞缺陷的BALB/c nude小鼠18只,以及T/B细胞联合缺陷的BALB/c SCID小鼠23只,每鼠感染日本血吸虫尾蚴(30±1)条。用细菌脂多糖(LPS)腹腔注射诱导(100 μg/ml,0.2 ml/只,2周1次,共2~3次)nude小鼠(NL组,9只)和SCID小鼠(SL组,12只)。同时设置对照组,即以PBS处理nude小鼠(N组,9只)和SCID小鼠(S组,11只)。感染后28 d、36 d,分别解剖N、NL、S和SL各组小鼠(n=4/5,4/5,5/6,6/6),肝门静脉灌注法收集血吸虫成虫,计算成虫检获率,测量体长,记录雌雄合抱数。取肝组织经5% KOH溶液消化后,计数虫卵,计算每克肝组织虫卵数。摘除眼球取血,检测外周血中转化生长因子(TGF-β)、γ干扰素(IFN-γ)和白介素10(IL-10)水平。取脾组织制备脾细胞悬液,检测脾淋巴细胞中调节性B细胞(regulatory B cells,Bregs)所占比例。  结果  感染后28 d,NL、N组雄虫体长分别为(7.66±2.85)mm和(5.28±1.64)mm(P<0.01),雌虫体长分别为(9.64±1.99)mm和(7.49±1.63)mm(P<0.01)。感染后36 d,NL组每克肝组织虫卵数(1 088±297)明显高于N组(715±404)(P<0.05),SL组的(217±33)明显低于S组(573±160)(P<0.01)。感染后28 d、36 d,N组脾淋巴细胞中CD1dhiCD5+CD19+Bregs所占比例分别为(12.73±0.96)%和(37.15±3.04)%(P<0.05),与NL组比较差异无统计学意义(P>0.05)。感染后28 d,N、NL组血清中TGF-β水平分别为(101.75±46.72)pg/ml和(260.90±45.34)pg/ml,IFN-γ水平分别为(7.91±1.62)pg/ml和(14.11±3.72)pg/ml,两组比较差异均有统计学意义(P<0.01)。感染后36 d,S、N组血清中IL-10分别为(41.85±3.14)pg/ml、(66.25±4.16)pg/ml,SL、NL组分别为(44.48±3.87)pg/ml、(72.22±17.76)pg/ml,两种小鼠比较差异具有统计学意义(P<0.01),但LPS处理组与对照组比较差异无统计学意义(P>0.05)。  结论  LPS可诱导日本血吸虫感染BALB/c小鼠B细胞释放TGF-β等细胞因子,促进日本血吸虫雌雄成虫的早期发育。

关键词: 日本血吸虫;B淋巴细胞;调节性B细胞;白介素10;生长趋化因子&beta, ;干扰素&gamma

Abstract:

Objective  To investigate the effect of lipopolysaccharide(LPS)-induced B cell activation on the development of Schistosoma japonicum.  Methods  Eighteen BALB/c nude mice deficient in T cells and 23 BALB/c SCID mice deficient in T and B cells were used in this study. Each was infected with 30±1 S. japonicum cercariae. The nude (n=9, NL group) and SCID(n=12, SL group) mice then received 2-3(every two weeks) intraperitoneal injections with LPS (100 μg/mL, 0.2 mL for each mouse). The remaining nude(n=9, N group) and SCID(n=11, S group) mice received PBS injection as control. The mice were sacrificed on days 28 and 36 after infection(n=4/5, 4/5, 5/6, 6/6 for N, NL, S and SL groups, respectively), and adult worms were collected by hepatic portal vein perfusion. The collecting rate of the adult worms was calculated, the body-length measured, and pairs of worms recorded. The liver tissue was collected and digested with 5% KOH, and the number of eggs per gram of liver tissue was calculated. The levels of TGF-β, IFN-γ and IL-10 in peripheral blood were evaluated. Spleen cell suspension was prepared for detecting the proportion of regulatory B cells(Bregs) in splenic lymphocytes.  Results  On day 28 after infection, the body-lengths of male worms in NL and N groups were (7.65±2.85) mm and (5.28±1.64) mm (P<0.01), and those of female worms were (9.64±1.99) mm and (7.49±1.63) mm (P<0.01), respectively. On day 36 after infection, the number of eggs per gram of liver tissue was significantly higher in the NL group than in the N group(1 088±297 vs 715±404, P<0.05), and significantly lower in the SL group than in the S group(217±33 vs 573±160, P<0.01). The proportions of CD1dhiCD5+CD19+ Bregs in N group on days 28 and 36 after infection were(12.73±0.96)% and(37.15±3.04)% (P<0.05), respectively, with no significant difference with that of NL group. The serum levels of TGF-β and IFN-γ on day 28 after infection were significantly different between N and NL groups(TGF-β, 101.75±46.72 vs 260.90±45.34 pg/mL; IFN-γ, 7.91±1.62 vs 14.11±3.72 pg/mL, both P<0.01). Similarly, significant difference was found for the plasma level of IL-10 on day 36 after infection between the S and N groups (41.85±3.14 vs 66.25±4.16 pg/mL, P<0.01), and between the SL and NL groups (44.48±3.87 vs 72.22±17.76 pg/mL, P<0.01), but not between the LPS groups and the control groups.  Conclusion  LPS can induce the release of cytokines (e.g. TGF-β) from B cells of mice infected with S. japonium, to facilitate the early development of adult female and male worms.

Key words: Schistosoma japonicum, B lymphocyte, Breg, IL-10, TGF-β, INF-γ