Tsp10多肽免疫血清在体外对旋毛虫侵入肠上皮细胞及幼虫发育的影响

Effect of Tsp10 polypeptide immune sera on in vitro invasion of intestinal epithelial cells by Trichinella spiralis and larval development

目的观察Tsp10多肽免疫血清在体外对旋毛虫侵入肠上皮细胞及幼虫发育的影响。方法将不同免疫血清与含有旋毛虫肠道感染性幼虫的半固体培养基混合后，接种至正常小鼠肠上皮细胞（intestinal epithelial cells，IEC）单层上，37℃5％CO2培养2h、36h时分别观察幼虫侵入率、细胞单层受损面积及发育至Ⅱ～Ⅳ期幼虫的百分比；免疫荧光试验检测受损肠上皮细胞中的旋毛虫抗原。结果幼虫在半固体培养基培养2h时，Tsp10多肽免疫血清组的幼虫侵入率与细胞单层受损面积（46．3％，24．3mm^2）均明显低于T7噬菌体免疫血清组（94．1％，65．3mm^2）及正常小鼠血清组（92．5％，65．7mmz）（P〈0．01）；培养36h时，Tsp10多肽免疫血清组Ⅱ～Ⅳ期幼虫的百分比（16％）亦明显低于T7噬菌体免疫血清组（37％）及正常小鼠血清组（38％）（P〈0．01）。Tsp10多肽及ES抗原免疫血清的免疫荧光试验结果显示，受损的IEC呈亮绿色荧光，而应用T7噬菌体免疫血清及正常小鼠血清时则为阴性。结论Tsp10多肽免疫血清可阻止部分旋毛虫幼虫对肠上皮细胞的侵入与发育，Tsp10可能是旋毛虫的主要侵入蛋白之一。

To observe the effect of sera from mice immunized with Tsp10 polypeptide on the in vitro invasion of intestinal epithelial ceils by Trichinella spiralis and larval development, the semisolid culture medium containing the T. spiralis intestine infective larvae were mixed with different immune sera, then inoculated on normal mouse intestinal epithelial cells （IECs） monolayer and incubated at 37 ℃ under 5% CO2 for 2 hours and 36 hours. The larval invasion rate, damage degree to cell monolayer, and the percentage of L2-L4 larvae were respectively observed. Immunofluorescent test （IFT） was used to detect T. spiralis antigens in the damaged IEC. When the larvae were cultured in semisolid medium for 2 hours, the larval invasion rate （46.3% ） and damage degree to cell monolayer （24.3 mm^2） of Tsp10 polypeptide immune serum group were obviously lower than 94.1% and 65.3 mm2of T7 phage immune serum group and 92.5% and 65.7 mm^2 of normal mouse serum groups （P 〈0.01）. When the larvae were cultured with Tsp10 polypeptide immune sera for 36 hours, the percentage of II-IV stage larvae （16%） was significantly lower than that of those cultured in T7 phage immune sera （37%） and in normal mouse sera （38%） （P〈0.01）. The results of IFT showed that the damaged IECs were positively stained by Tsp10 polypeptide and ES antigen immune sera, respectively, but not by T7 phage immune serum and normal mouse serum. The Tsp10 polypeptide immune sera could prevent partially the invasion of T. spiralis larvae into IEC and larval development.