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目的观察旋毛虫幼虫对胃上皮细胞的侵入及侵入前后虫体与细胞蛋白的变化,筛选幼虫侵入相关蛋白。方法将旋毛虫感染性幼虫接种至胃上皮细胞(SGC-7901)单层,37℃5%CO2条件下培养不同时间后在倒置显微镜下观察幼虫侵入情况;培养18h后分别提取虫体与细胞蛋白,进行SDS-PAGE与Western blot分析。结果旋毛虫幼虫培养6h时已侵入细胞单层,18h在幼虫头端与尾端可见鞘的形成。SDS-PAGE结果表明,幼虫与细胞共培养后比仅在培养基中培养的幼虫增加了3条蛋白带,减少了4条蛋白带;细胞与幼虫共培养后细胞蛋白增加了3条蛋白带,减少了2条蛋白带。Western blot分析显示,幼虫与细胞共培养后虫体蛋白多了3条被旋毛虫感染鼠血清识别的反应带(58、21、18ku),少了3条反应带(98、86、31ku);细胞与幼虫共培养后细胞蛋白多了6条被旋毛虫感染鼠血清识别的反应带(96、62、40、34、29、22ku),少了2条反应带(98、15ku)。结论旋毛虫幼虫可侵入体外培养的胃上皮细胞单层;幼虫与细胞共培养后增加的被感染鼠血清识别的虫体与细胞蛋白可能是幼虫分泌的侵入相关蛋白。
Objective To observe the invasion of gastric epithelial cells by Trichinella spiralis larvae and the changes of the parasite and cell proteins before and after invasion, and screen the larva for invasion of related proteins. Methods The larvae invaded into the monolayer of gastric epithelial cells (SGC-7901) and inoculated with 5% CO2 at 37 ℃ for different time. The infected larvae were observed under an inverted microscope. After culturing for 18 hours, , Analyzed by SDS-PAGE and Western blot. Results Trichinella spiralis larvae had invaded into the cell monolayer when cultured for 6 hours, and the formation of sheath was observed at the head and tail of larvae at 18 hours. The results of SDS-PAGE showed that after larvae co-cultured with larvae, larvae increased by 3 bands and decreased by 4 bands compared with larvae cultured in medium alone. After co-cultured with larvae, the cell proteins increased by 3 bands, Reduced two protein bands. Western blot analysis showed that after larvae co-cultured with the cells, there were three more reaction bands (58,21,18ku) recognized by serum of Trichinella infected mice and three fewer bands (98,86,31ku); After co-culture of cells with larvae, there were six more cellular proteins (96, 62, 40, 34, 29, 22 ku) recognized by serum of Trichinella infected mice and two fewer reaction bands (98 and 15 ku). Conclusions Trichinella spiralis larvae can invade the monolayer of gastric epithelial cells cultured in vitro. The parasites and cellular proteins recognized by the sera of infected mice co-cultured with larvae may be the invasion-related proteins secreted by larvae.