| Objective Epstein–Barrvirus(EBV)is closely associated with many human diseases,including a variety of deadly human malignant tumours such as Burkitt lymphoma,Hodgkin’s disease and nasopharyngeal carcinoma.However,due to the lack of ideal animal models,the biological characteristics of EBV,particularly its function in tumourigenesis,have not been determined.Chinese tree shrews(Tupaia belangeri chinensis)are small animals that live in tropical and subtropical regions and closely related to primates,have been used to establish a variety of animal models and have some irreplaceable advantages for studying human diseases.Here,we used tree shrew as a model for EBV infection by intravenous injection,establishing a foundation for studying the infection and pathogenic mechanism of EBV and for the development of vaccines and medicines.Methods Thirteen healthy and adult(1–1.5 years of age)male or female tree shrews were involved.Ten tree shrews(Ts1-Ts10)were inoculated with EBV by intravenous injection of 500μl of the viral solution and as negative controls,the other three tree shrews were inoculated with fresh RPMI 1640 medium containing 10% FBS and 1×antibiotics using the same procedure.Blood was collected at regular intervals from the femoral artery or vein of tree shrews to isolate PBMCs and serum.They were euthanized at times ranging from 3 to 21 weeks after inoculation with the virus,and their tissues were assayed for pathological changes.The q PCR assay was used to detect EBV copy number in PBMCs.EBV-related gene expression in PBMCs was detected by the RT-PCR assay.EBV antibodies in serum of tree shrews were determined by ELISA.Pathology changes of tissues were examined by HE staining and EBERs were detected by EBER-ISH.The western blot assay was used to analyse the EBV related protein expression of LMP1,EBNA1 and EBNA2 in spleens and livers of tree shrews.Results Eight of 10 tree shrews showedthe evidence of EBV infected.Three of the 13 tree shrews(Ts1,2,8)died prior to the end of the experiment,but there was no evidence that the deaths resulted from EBV infection.Another 2 animals(Ts3,Ts4)displayed severe weakness and were euthanized at week 12 and week 5.The remaining 8 animals appeared to be normal and displayed no weakness after EBV inoculation.1.The results of q PCR showed that of the 8 EBV-infected tree shrews,EBV copy number increased intermittently and appeared to fluctuate in 3 tree shrews.However,in the other 5 tree shrews,EBV copy number increased to the highest value at week 2 or 4 and followed by a decline.2.The results of RT-PCR showed that the expression of EBV-related genes was detected in varying degrees in 8 EBV-infected tree shrews.3.The results of ELISA showed that the level of VCA Ig G increased to varying degrees in all tree shrews in which the EBV copy number increased.However,the level of EA Ig G was not elevated.4.HE staining showed that compare with the negative controls,splenic corpuscle hyperplasia was detected in the spleens of four(Ts2,3,7,10)of the 8 infected tree shrews.In the liver,extensive inflammatory cell infiltration was detected in only 1(Ts2)of the 8 infected tree shrews.Extensive lymphocyte proliferation was observed in 3 animals(Ts3,7,10)with mesenteric lymph node enlargement.5.EBER-ISH staining showed that EBER-positive cells were detected in the spleens of 3 tree shrews(Ts2,7,10)and mesenteric lymph nodes of 3 tree shrews(Ts3,7,10).6.The results of western blot showed that LMP1 was detected in the spleens of all 8 EBV-infected tree shrews,EBNA1 expression was detected in the spleens of 4 tree shrews(Ts2,3,9,10),but no EBNA2 expression was detected in spleens and no EBV-related protein expression was detected in livers of any animals.Conclusion These findings suggestthat EBV can infect tree shrews via intravenous injection.The presented model offers some advantages for exploring the pathophysiology of EBV infection in humans. |
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