目的 建立小鼠巨细胞病毒(MCMV)经鼻腔急性感染同种异型皮肤移植BALB/c小鼠间质性肺炎模型.方法 ① 25只供体C57BL/6雌鼠与100只受体BALB/c雌鼠背部皮肤移植后,每只小鼠腹腔注射环孢素A(CsA,12 mg/kg),连续14 d.② 移植受体小鼠随机分为5组,其中4组每只鼻腔接种30 μl不同剂量MCMV悬液(102、103、104、105 PFU/只),另一组为正常对照组,小鼠鼻腔接种原代小鼠胚胎成纤维细胞(MEF)悬液30 μl/只.在接种后第5、9、14、21天处死小鼠,无菌取肺组织分别做HE染色、透射电镜、PCR、RT-PCR检测MCMV Smith毒株的即刻早期(IE)和晚期基因糖蛋白B(gB)基因转录产物,并进行原位杂交和免疫组织化学实验.结果 104、105 PFU实验组小鼠肺组织有局灶性的病理损害,并发现感染后第14天肺组织病理改变最明显;透射电镜检测到疱疹样病毒颗粒;PCR、RT-PCR检测实验组MCMV IE和gB基因及其转录产物均为阳性;两组原位杂交和免疫组织化学检测均可见肺间质上皮细胞中存在病毒核酸和蛋白;并在感染后的21 d内,105 PFU组小鼠相比对照组有明显的临床表现.结论 成功建立同种异型皮肤移植后MCMV急性感染所致的小鼠间质性肺炎模型.%Objective To establish a mouse model of interstitial pneumonitis with acute murine cytomegalovirus ( MCMV ) after allogeneic skin transplantation. Methods ① 25 of C57BL/6 donor mice and 100 0f BALB/c recipient mice were underwent back skin graft. Cyclosporine A ( CsA,12 mg/kg ) was subsequently given for 14 days.(② All recipient animals were randomly divided into five groups ( n =20 ), four experimental groups were infected intranasally with MCMV infected mouse embryonic fibroblasts ( MEF ) at different concentrations ( 102 ,103,104 and 105 PFU ) in a volume of 30 μl, and the control group was inoculated with 30 μl of MEF cell suspension.After infection days 5,9,14 and 21, mice were sacrificed and lung samples were collected for HE staining, transmission electron microscope( TEM ), PCR/RT-PCR. in situ hybridization, and immunohistochemistry. Results Focal pathological abnormality in the lung was observed in the mice with high dose of virus ( 104 and 105 PFU ).Herpes virus particles were found by TEM in the epithelial of the lung tissue. MCMV DNA was detected by PCR for immediate early( IE ) genes and late genes glycoprotein B( gB ) , and it was localized in lung cells by in situ hybridization. Viral mRNA for IE and gB were also present suggesting lytic infection in the lung tissue. MCMV protein was revealed in the interstitial lung epithelial cells by immunohistochemistry. Clinical manifestations were only observed in the group with 105 PFU in 21 days after infection. Conclusion The study demonstrates that a mouse model of interstitial pneumonitis is successfully established after allogeneic mouse skin transplantation.
展开▼
