轮状病毒感染乳鼠致病机制的实验研究

On pathogenesis of rotavirus diarrhea in suckling mouse

目的研究轮状病毒(rotavirus,RV)腹泻发病机制。方法生后7d乳鼠经口感染猴RV(SA-11)后不同时间点取小肠组织,观察形态学改变并用软件(Image pro plus5.1,IPP5.1)测定;免疫组化染色观察RV抗原分布;Phalloi-dine染色观察小肠绒毛丝状肌动蛋白分布;肠上皮细胞原位凋亡检测。结果乳鼠感染RV后小肠绒毛轻度充血水肿、上皮细胞广泛空泡状变性,顶部大量脂滴样结构,微绒毛排列紊乱或脱落,而细胞间连结未见明显结构改变;RV抗原分布主要在小肠绒毛的上部;肠组织丝状肌动蛋白含量有明显减少;小肠上皮细胞凋亡增加。结论RV主要感染小肠成熟绒毛上皮细胞;腹泻的发生与小肠绒毛上皮细胞骨架损害、微绒毛损害、细胞凋亡脱落和绒毛萎缩相关,但与肠上皮细胞间连接损害关系不大。

Objective To study the pathogenesis of rotavirus （RV） diarrhea. Methods Simian rotavirus （SA-11） was grown in cultured MA-104 cell. The viral titers of the culture supernatant were determined by plaque forming assay. KM mice aged 7 days were inoculated with the viral supernatant via feeding tube （ gavage）. Histological and ultramicrostructure changes of the small intestines were observed under light microscope and electron microscope. The values of crypt depth and villi height were measured with software （image pro plus 5.1, IPP5.1 ）. The distribution of the RV antigen in small intestine and the filamentous actin of the small intestine chorioepithelium were observed with immunohistochemical techniques. The apoptosis of the small intestine epithelium cells was observed with an in situ apoptosis detection kit. Results There were mild hyperemia, dropsy and extensive vacuolar degeneration of small intestine villi under light microscope. Plenty of lipid droplet-like structure at the top of the villi, microvilli malalignment or defluxion and enterocyte defluxion could be seen by electron microscope, but no obvious structure changes at the cell junctions were seen. The RV antigen mainly distributed at the top of the villi. The quantity of small intestine filamentous actin decreased and enterocyte apoptosis increased after RV infection. Conclusion RV mainly infects the mature villous epithelium. The presentation of RV diarrhea relates to the lesion of cytoskeleton, the microvilli lesion of the small intestine, enterocyte apoptosis and defluxion, villi atrophy, etc. , but may have no relationship with the structural changes of cell junctions in the small intestine epithelium.