Tallima Hatem, El Ridi Rashika
Zoology Department, Faculty of Science, Cairo University, Cairo 12613, Egypt.
Folia Parasitol (Praha). 2008 Sep;55(3):180-6. doi: 10.14411/fp.2008.025.
We have previously reported that Schistosoma mansoni larvae emerging from host lung at pH 7.5-7.8 and then fixed with diluted formaldehyde (HCHO) readily bind radiation-attenuated cercariae (RA) vaccine serum antibodies, as assessed by indirect membrane immunofluorescence (IF). Here we show that S. mansoni schistosomula emerging from lung pieces under 5% CO2 (pH < or = 7.3) readily bind RA vaccine serum antibodies, provided they have been incubated for 12 h at pH 7.5-7.8 in foetal calf serum-free RPMI medium, and fixed with diluted HCHO. Ex vivo larvae exposed during incubation to GW4869, a specific inhibitor of tegument-bound, neutral sphingomyelinase (nSMase) displayed significantly diminished binding of RA vaccine serum antibodies, thus suggesting that nSMase activity at pH > or = 7.5 leads to exposure of lung-stage larvae surface membrane antigens to specific antibody detection. More importantly, ex vivo larvae readily bound antibodies directed to dipeptidic multiple antigen peptide constructs, based on S. mansoni-specific sequences in S. mansoni glyceraldehyde 3-phosphate dehydrogenase (SG3PDH). Lung-stage schistosomula IF reactivity was diminished following antiserum absorption with recombinant SG3PDH. The data together indicate that intact ex vivo, as well as, 5-day-old in vitro-grown larvae express SG3PDH on their surface membrane. The findings are discussed in relation to the importance of surface membrane proteins as candidate vaccine antigens.
我们之前曾报道,曼氏血吸虫幼虫在pH 7.5 - 7.8时从宿主肺部逸出,然后用稀释甲醛(HCHO)固定,通过间接膜免疫荧光(IF)评估,其能轻易结合辐射减毒尾蚴(RA)疫苗血清抗体。在此我们表明,在5%二氧化碳(pH≤7.3)条件下从肺组织块中逸出的曼氏血吸虫童虫,只要在无胎牛血清的RPMI培养基中于pH 7.5 - 7.8孵育12小时,并用稀释的HCHO固定,就能轻易结合RA疫苗血清抗体。在孵育期间暴露于GW4869(一种与被膜结合的中性鞘磷脂酶(nSMase)的特异性抑制剂)的体外幼虫,其与RA疫苗血清抗体的结合显著减少,这表明在pH≥7.5时nSMase活性会导致肺期幼虫表面膜抗原暴露以进行特异性抗体检测。更重要的是,体外幼虫能轻易结合基于曼氏血吸虫甘油醛 - 3 - 磷酸脱氢酶(SG3PDH)中曼氏血吸虫特异性序列的二肽多抗原肽构建体的抗体。用重组SG3PDH吸收抗血清后,肺期童虫的IF反应性降低。这些数据共同表明,完整的体外幼虫以及体外培养5天的幼虫在其表面膜上表达SG3PDH。结合表面膜蛋白作为候选疫苗抗原的重要性对这些发现进行了讨论。