【摘要】 目的以空肠弯曲菌(campylobacterjejuni,CJ)菌体全抗原为对照,以基因重组的该菌融合黏附蛋白(TrxA/PEB1)作为CJ疫苗免疫大鼠,并对其神经安全性进行实验性评价。方法分别以纯化的TrxA/PEB1和CJPen19菌体全抗原经全身免疫增强法免疫大鼠,初次免疫后第2、4、5周采用ELISA法动态观察抗体滴度,并分别于第3、5周分离坐骨神经原纤维及行甲苯胺蓝染色光镜观察。采用ELISA法检测第5周抗血清与GM1抗原的结合反应。神经外膜下注射抗TrxA/PEB1和抗CJ全菌抗原的特异性抗血清,观察其神经病理学改变。结果(1)经两种抗原免疫后,大鼠均产生高滴度特异性IgG,且各自与两种抗原发生交叉反应;(2)以CJPen19全抗原免疫后抗血清可与GM1抗原结合,而TrxA/PEB1免疫者则否;(3)CJPen19免疫组有60%大鼠坐骨神经出现以轴索变性为主的免疫性损伤,而TrxA/PEB1融合蛋白免疫者则未发生;(4)CJPen19免疫血清神经外膜下注射后全部大鼠均发生轴索变性为主的病变,且75%为显著病变,而TrxA/PEB1免疫血清外膜下注射后却未引起明显病变。结论CJPen19全菌抗原和TrxA/PEB1纯化蛋白作为免疫原均可激发大鼠有效体液免疫应答,但与CJ全菌抗原相比,TrxA/PEB1蛋白所产生的全身免疫反应未引起周围神经损伤,其抗血清不含针对GM1抗原的抗体,对外周神经无免疫损伤更多还原

【Abstract】 Objective To explore the nerve safety of the vaccine of recombinant campylobacter jejuni (CJ) periplasmic solute-binding protein 1 (PEB1) produced by genetic engineering process. Methods Wistar rats were immunized by systemic immunization with whole cell antigen of CJ Pen19 and its fusion protein (TrxA/PEB1) respectively. The titers of anti-CJ and anti-TrxA/PEB1 in sera were detected by ELISA at the week of 2nd, 4th and 5th respectively. Animals were sacrificed and their sciatic nerves were taken on the day of 21st and 35th after immunization. Histological study, including teasing fibers and cross-section for toluidine blue stain, was finished under light microscope and electron microscope. The titers of specific IgG anti- CJ GM1 and anti-TrxA/PEB1 IgG antibody was measured by ELISA. Histological study for sciatic nerve was also performed on the 7th day after injection perineurially with immunized serum. Results (1) High titers of specific IgG antibody were detected after immunization with CJ Pen19 whole antigen and purified TrxA/PEB1 systemically. Cross-reaction was demonstrated between the two immunity reactions. (2) OD values of cross-reaction between the specific anti-CJ IgG antibody and antigen GM1 was 4 times more than that of control group, but not increase for antibody of specific anti-TrxA/PEB1 IgG. (3) There are 60% incidence of fibrils undergoing axon degeneration in the sciatic nerves of rats after systemic immunization, but no pathologic change was seen in the TrxA/PEB1 immunized group. (4) Axon degeneration in the fibers of sciatic nerves was appeared in all rats after perineurial injection with CJ Pen19 immunized serum, and presented severe pathological damage in 75% of the sciatic nerves. However there were no obvious pathologic change in the group of perineurial injection with TrxA/PEB1 immunized serum. Conclusions Significant and effective humoral immune response could be produced by immunization with both whole antigen of CJ Pen19 and purified TrxA/PEB1 protein in rats. However in contrast to the whole antigen of CJ Pen19, no any pathological damage could be found on peripheral nerves after systemic immunization with the purified recombinant PEB1 protein of CJ. There was no anti-GM1 antibody in the anti-TrxA/PEB1 serum, which could be a reason for the immunization with TrxA/PEB1 as less possible to damage peripheral nerve. It was indicated that even the cellular immunity evoked by TrxA/PEB1 was not promoting the damage to nerve. It indicated the safety to use TrxA/PEB1 as vaccine, particularly escaped from immunizing damage on peripheral nerve.更多还原