新生小鼠轮状病毒全身扩散的实验研究

Experimental Study of Rotavirus Infection in Whole Body of Newborn Mouse

目的探讨新生小鼠发生轮状病毒(RV)全身扩散后对RV易感器官及靶器官造成的损害。方法收集RV流行高峰期腹泻患儿粪便,提取RV,经抗原ELISA检测和聚丙烯酰胺凝胶电泳(PAGE)证实为RV长型,作为实验用病毒株。选用健康新生昆明小鼠,经灌胃与腹腔注射二种途径接种RV,第3天后处死小鼠。地高辛标记引物探针,取各脏器标本进行PCR扩增后进行原位杂交,检测RV基因,光镜与电镜下观察病理变化,并与健康对照组小鼠比较。结果腹腔注射组新生小鼠受累的器官明显多于灌胃组,且组织病理改变也重于灌胃组。光镜下:RV灌胃组小肠绒毛、胃固有层、心肌细胞有病理改变;RV腹腔注射组小肠绒毛、胃固有层、心、肝、肾有病理改变。电镜下:RV腹腔注射组可见小肠绒毛、肝、毛细胆管、肾细胞器及细胞结构改变。肠绒毛变短,核膜结构破坏,细胞内出现大量吸收小泡,粗面内质网扩张;肝细胞线粒体肿胀、凝集病变尤为明显,细胞核固缩、崩解,嵴膜模糊不清,粗面内质网扩张,肝细胞中存在大量脂滴,细胞之间淋巴细胞和浆细胞浸润显著;毛细胆管明显扩张,微绒毛脱落;肾近曲小管线粒体轻度肿胀。原位杂交:RV灌胃组小肠上皮细胞呈阳性;RV腹腔注射组肠、肾近曲小管上皮细胞呈阳性。原位PCR:RV灌胃组小肠绒毛、肠腺细胞、肾近曲小管和集合管上皮细胞呈阳性;RV腹腔注射组肠、肾、肝、心脏、胰呈阳性。其余各组小鼠各组织器官均为阴性。结论RV可向新生小鼠全身扩散,RV具有一定的泛嗜性,可造成多脏器结构改变。

Objective To explore the hurt of susceptibility organs and critical orgens followed by rotavirns （RV） infection of whole body in newborn mouse. Methods RV strain was derived from the stool samples of patients with RV diarrhea and was proved to be long type by methods of ELISA and polyaerylamide gel eleetrophoresis （PAGE）. RV was inoculated by the pathways of taking orally and injected to abdominal cavities, respectively. The pathological changes of the newborn mouse model infected with human natural RV by light microscope and electron microscope. The gene probe was marked by digoxin. The direct prove of RV infection in these organs was got by the detection of in situ PCR. Results Pathological changes were found in the small intestinal villus,lamina propria of the stomach and the heart cells of the mice taken RV orally. The mice with intraabdominal RV injection showed pathological changes of the cells in the small intestinal villus, liver and kidneys observed by electron microscope. Shortened small intestinal villus, nuclear membrane disorganization, massive vacuolization, mitochondrial swelling and rough endoplasmic retieulum dilation were observed in the cells of small intestinal. In the liver of the mice, marked mitochondrial swelling and agglutination, cell nucleus pyknosis or collapse, presence of numerous lipid droplets and vacuoles were found in the liver cells, with lymphocyte and plasmacyte infiltration. Obvious dilatation and shedding of the microvillus were found in eholangioles. The mitochondria of the proximal convoluted renal tubule showed mild swelling, but the cells in the heart and lung did not display obvious changes. Conclusion RV can damage lots of extra intestinal organs of the newborn mice if RV diffuses to the whole body of the mice.