捻转血矛线虫与胃癌细胞共培养系统建立

Establishment of a nematode-cell co-culture system of Haemonchus contortus and gastric cancer cells

为研究宿主感染捻转血矛线虫的早期免疫应答,本试验使用细胞小室建立了捻转血矛线虫-胃癌细胞气液界面共培养系统。捻转血矛线虫在共培养系统中以弦运动和圆运动的方式生活。细胞远红外荧光染色结果显示,线虫以细胞蛋白为食维持基本营养,进食的方式可能为叮咬胃癌细胞后吸食。脱鞘幼虫在体外培养系统中生长发育速度缓慢,共培养0,4,7 d时的虫体长度分别为(981.5±24.3),(1113.9±24.4),(1157.0±47.6)μm,其中7 d时与宿主感染4 d体内的虫体大小相似。捻转血矛线虫和细胞在共培养系统中互相胁迫,随着共培养时间的增加,线虫呈现活力下降和蜷缩状的生理状态,同时贴壁细胞逐渐从胶原板脱落并进入坏死状态。相较于共培养0 d时,共培养4 d时胃癌细胞(MGC)坏死的比例由12.91%增加到53.25%,胃腺癌细胞(BGC)坏死的比例由2.63%增加到24.81%。线虫还能诱导细胞表达大量的IL-33,MGC与线虫共培养4,7 d时IL-33的表达量分别上升至共培养0 d时的2.520,4.180倍;BGC与线虫共培养4,7 d时IL-33的表达量分别上升至共培养0 d时的1.732,5.150倍,这与宿主早期感染时IL-33的高表达一致。本试验为阐明捻转血矛线虫早期感染时宿主的免疫应答分子机制提供了新方法。

To study the host early immune response to Haemonchus contortus,an air-liquid nematode-cell co-culture system was built using trans-well inserts.Microscopic examination revealed two locomotive behaviours of H.contortus,either a sinusoidal motion or a circular burrowing one.The larvae ingested cells based on the Far-Red labeled cells and the filling of intestinal tract with labeled proteins from cells.The length of larvae at 0,4 and 7 days post co-culture(dpc)were(981.5±24.3),(1113.9±24.4)and(1157.0±47.6)μm respectively,which at 7 dpc was similar to that in the host at 4 dpi.Reduced vitality of larvae was observed after co-culture,and larvae presented a state of curled up.Cells were dislodged from the collagen pad by larvae at the same time,and necrotic.The necrosis rate of MGC at 4 dpc increased from 12.91%to 53.25%compared to 0 dpc,and that of BGC increased from 2.63%to 24.81%.IL-33 gene expression in cells was significantly increased after co-culture,consistent with a high expression in the host.The expression level of IL-33 in MGC at 4 and 7 dpc were 2.520-fold and 4.180-fold higher than that at 0 dpc.The relative expression of IL-33 in BGC at 4 and 7 dpc raised to 1.732 and 5.150 compares to that at 0 dpc as well.Our results provide a new method to elucidate the molecular mechanism of the host’s immune response in the early infection of H.contortus.