汉滩病毒感染与整合素β3表达的相关性

目的　探讨整合素 β3表达与汉滩病毒 (HTNV)感染的关系。 方法　将人整合素 β3、αv及αⅡb真核表达载体分别及共转染至CHO细胞 ,采用间接免疫荧光测定 (IFA)、流式细胞计数(FCM)检测外源基因的表达 ;然后用HTNVA9株感染稳定转染的CHO细胞 ,应用IFA和FCM检测病毒抗原。结果　整合素 β3在共转染组细胞中高效表达 ,且目的蛋白主要定位于细胞膜上 ;β3单转染组目的蛋白虽有一定量的细胞膜表达 ,但表达量低于共转染组 (P <0 0 5 ) ;αv和αⅡb单转染组目的蛋白的表达量明显低于共转染组 (P <0 0 1) ,且定位发生变化。病毒感染率共转染细胞较高 ,CHO/αvβ3和CHO/αⅡbβ3细胞分别为 6 0 1%和 5 5 9% ;未转染 β3的细胞感染率较低 ,CHO细胞仅为 1% ;而单转染 β3细胞介于两者之间 ( 38 7% )。结论　汉滩病毒感染率与整合素 β3表达水平密切相关 ,整合素 β3可以促进HTNV的入胞作用

The relationship between cellular infection by Hantaan virus and expression of beta3 integrin

OBJECTIVE: To investigate the relationship between cellular entry of Hantaan virus (HTNV) and expression of beta3 integrin in beta3-integrin-deficient and HTNV-insusceptible China hamster ovary (CHO) cells. METHODS: Eukaryotic expression vector encoding human integrin beta3 and eukaryotic expression vector harboring human integrin alphav or alphaIIb subunit cDNA were transfected into HTNV non-permissive CHO cells individually or collectively. Screening for stable transfectant clones was performed using G418 selective (culture medium. The exogenous gene expression was analyzed qualitatively and quantitatively by immunofluorescence assay (IFA) and flow cytometry (FCM). Various modified CHO cells and untransfected CHO cells were infected using HTNV A9. At various time points after infection, HTNV antigens in infected cells were detected qualitatively and quantitatively by IFA, FCM. RESULTS: Highly-effective surface expression of beta3 integrin was measured in CHO/alphavbeta3 and CHO/alphaIIbbeta3, while weaker surface expression was detected in CHO/beta3 (P < 0.05). Expression of alphav or alphaIIb integrin in the individually transfected group was significantly lower than in the cotransfected group (P < 0.01) and the sites of localization changed. In contrast, effective surface expression was not seen when pcDNA3 was transfected alone. The infection rate of CHO/alphavbeta3 (60.1%) and CHO/alphaIIbbeta3 (55. 9%) cells were significantly higher than that of CHO/beta3 (38.7%) cells, while the infection rate of CHO/beta3 was significantly higher than that of CHO/alphav, CHO/pcDNA3 and CHO cells respectively. There was a close relationship between the positive percentage of HTNV A9-infected cells and expression of beta3 integrin. CONCLUSION: These results indicated that cellular entry of HTNV was related to the expression of beta3 integrin.