Preliminary Study Of KSHV Viral Interferon Regulatory Factor 1 Induces Angiogenesis And Tumorigenesis

Background:Kaposi's sarcoma-associated herpesvirus(KSHV)infection is associated with Kaposi's sarcoma(KS)closely,and KSHV has been proposed to be the etiological agent of KS.KS starts as a proliferation of endothelial-type cells,with an early onset inflammatory and abnormal leaky blood vessel expansion.Viral interferon regulartory factor 1(vIRF1)is an oncogene encoded by KSHV ORF K9.It can induce NIH 3T3 cells to tansform.NIH 3T3 cells which was overexpressed vIRF1,formed tumors after injection into nude mice.Recently,the research of vIRF1 was focused on the IFN transcriptional regulation,inhibiting the transcriptional activation of p53 and preventing p53-mediated apoptosis.However,the effect of vIRF1 on angiogenesis was not mentioned.Objective:To explore whether KSHV vIRF1 could modulate angiogenesis and observe its effect on tumorigenesis.And preliminary study the mechanism of it.Methods:First,HUVEC cells were infected by recombinant lentivirus containing KSHV K9 gene or its control(Mock),respectively.Got cells named HUVEC/vIRF1 or HUVEC/Mock expressing green fluorescent protein(GFP)by flow cytometry assay(FCM)screening.Next,explored the possible roles of KSHV vIRFl in HUVEC cells proliferation,colony formation,migration,invasion,VEGF expression and capillary tube formation ability by cell counting kit-8(CCK-8),colony formation assay,transwell assay,Western blot and capillary tube formation analysis.In vivo,explored the effect on angiogenesis and tumorigenesis of vIRFl by the chick embryo chorioallantoic membrane(CAM)and nude mice models.Then detected the expression of tumorigenesis and angiogenesis related proteins in HUVEC/vIRF1 and HUVEC/Mock.And verified some of them by Western blot.Results:According to CCK-8 assay and colony formation assay,vIRF1 protein could promote the cell proliferation and colony formation.Transwell assay showed that vIRF1 promoted migration and invasion.The expression of VEGF in HUVEC/vIRFl was more than control.Capillary tube formation assay indicated that vIRF1 could induce angiogenesis in vitro.And vIRFl could also enhance the angiogenesis and tumorigenesis on the CAM and the nude mice models in vivo.vIRF1 could up-regulated the expression of EZR protein and down-regulated the expression of A2M,CAV1,TGM2 and PTPN1 by iTRAQ assay and Western blot.Conclusions:KSHV vIRF1 protein could promote cell proliferation,migration,and invasion.It could induce angiogenesis and tumorigenesis on CAM and nude mice.vIRFl may promote angiogenesis and tumor formation by up-regulating the expression of VEGF,EZR protein,etc,and down-regulating the expression of A2M,CAV1,TGM2 and PTPN1,etc.