啮齿类肝炎病毒感染大鼠模型的建立  

Establishment of rat model of rodent hepacivirus infection

作　　者：马鑫宇 官月婵 吴爽靖 谢明学[2] 毕研伟[2] 寸韡[2] MA Xin-yu;GUAN Yue-chan;WU Shuang-jing;XIE Ming-xue;BI Yan-wei;CUN Wei(Kunming Medical University,Kunming 650500,Yunnan Province,China;不详)

机构地区：[1]昆明医科大学,云南昆明650500 [2]中国医学科学院,北京协和医学院医学生物学研究所,云南昆明650000

出　　处：《中国生物制品学杂志》2022年第6期668-671,677,共5页Chinese Journal of Biologicals

基　　金：国家自然科学基金面上项目(81971947);云南省科技重大专项(2019ZF001)。

摘　　要：目的建立啮齿类肝炎病毒(rodent hepacivirus,RHV)感染大鼠模型。方法将体外转录获得的RHV rn1亚型(RHV-rn1)基因组全长RNA经大鼠肝内和尾静脉两种注射途径给药,于不同时间点经尾静脉采血,分离血清,RTqPCR法检测RHV-rn1基因组拷贝数。将肝脏注射感染病毒大鼠的血清(P0代RHV-rn1)经尾静脉注射进行大鼠个体传代,共传3代,获得P1、P2、P3代RHV-rn1。RT-qPCR法检测接种P0代RHV-rn1后不同时间点大鼠血清、RHV-rn1在体内传至P3代时大鼠不同脏器/组织及各代病毒血清的RHV-rn1基因组拷贝数。用不同剂量(10^(1)、10^(2)、10^(3)、10^(4)、10^(5)copies/μL)的P3代RHV-rnl经尾静脉感染大鼠,检测血清中的RHV-rn1基因组拷贝数。结果肝内注射组大鼠于注射后2 d出现高滴度病毒血症,至注射后14 d RHV-rn1基因组拷贝数开始稳定,约10^(6)copies/μL,可持续至49 d;尾静脉注射组大鼠至28 d未检测到病毒血症。接种P0代RHV-rn13 d后,大鼠血清中RHV-rn1基因组拷贝数为10^(5)copies/μL,随后逐渐上升,至14 d时,维持约在10^(6)copies/μL。RHV-rn1在体内传至P3代时,大鼠肝脏中的RHV-rn1基因组拷贝数达10^(7)copies/mg,高于其他组织器官1000倍以上。P1、P2、P3代RHV-rn1的拷贝数分别为1.12×10^(7)、7.5×10^(6)、1.65×10^(7)copies/μL。RHV-rn1半数感染的拷贝数为10^(2).5 copies。结论体外转录获得的RHV RNA经肝脏注射可使大鼠出现高滴度病毒血症,形成慢性感染,获得的RHV-rn1具有嗜肝性,且感染性较强,本实验成功建立了RHV感染的大鼠动物模型。Objective To establish a rat model of rodent hepacivirus(RHV)infection.Methods Full-length RNA of RHV subtype rn1(RHV-rn1)obtained by in vitro transcription was injected into rats by intrahepatic and intravenous routes respectively,of which the serum samples were collected at various time points and determined for RHV-rn1 genome copy number by RT-qPCR.The virus in sera of rats infected with RHV-rn1 by intrahepatic injection was subcultured in venous blood for three passages to obtain RHV-rn1 of P1,P2 and P3.The rat sera at various time points after inoculation with RHV-rn1 of P0,as well as the RHV-rn1 genome copy numbers in various organs/tissues of rats in which RHV-rn1 was subcultured to P3,were determined by RT-qPCR.Meanwhile,rats were injected with RHV-rn1 of P3(10^(1),10^(2),10^(3),10^(4)and 10^(5)copies/μL)by intravenous route,and determined for the RHV-rn1 genome copy number in sera.Results High titer viremia was observed in rats 2 d after intrahepatic injection with RHV-rn1,which was stable 14 d after injection;the RHV-rn1 genome copy number was about 10^(6)copies/μL,which last to 49 d after injection.However,no viremia was observed in the rats within 28 d after intravenous injection.The RHV-rn1 genome copy number in rats 3 d after inoculation with RHV-rn1 of P0 was 10^(5)copies/μL,which increased gradually and was maintained at about 10^(6)copies/μL 14 d after injection.The RHV-rn1 genome copy number in liver reached 10^(7)copies/mg,which was more than 1000 times higher than those in other organs.The RHV-rn1 genome copy numbers of P1,P2 and P3 were 1.12×10^(7),7.5×10^(6)and 1.65×10^(7)copies/μL respectively,while the median infective genome copy number was 10^(2).5 copies.Conclusion The RHV RNA obtained by transcription in vitro induced high titer viremia and chronic infection in rats by direct intrahepatic injection,which was hepatophilic and highly infectious.A rat model of RHV infection was successfully established in this study.

关 键 词：啮齿类肝炎病毒 大鼠 RT-qPCR法 基因组拷贝数 

分 类 号：Q95[生物学—动物学] R332[医药卫生—人体生理学]