Role of HIV-1 Nef protein carried by recombinant lentivirus in proliferation of primary effusion lymphoma cell line and vascular endothelial cells

Objective: To construct the recombinant lentivirus containing HIV-1 Nef gene and detect the protein expression in its target cell lines,such as 293T,BCBL-1 and EA.hy926,and explore the effect of Nef on cell proliferation.Methods:The fragment of Nef gene from expression vector pCI-neo-Nef was cloned into the lentivirus vector pHAGE-CMV-MCS-Izs-Green,then the recombinant vector pHAGE-Nef,packaging vector psPAX2 and envelope vector pMD2.G were cotransfected into the 293T cells.Culture media were harvested and filtered through a 0.45 μm filter to remove the cells.The viral titer was checked by observing the expression of green fluorescent protein(GFP).After infection of the recombinant lentivirus,the protein expression of Nef in 293T,BCBL-1 and EA.hy926 cells were detected by Western blot.The effect of Nef on cell proliferation was investigated by Cell Counting Kit-8 assay.Results:The recombinant lentivirus vector carrying HIV-1 Nef was constructed successfully.The viral titer was 1×107 TU/ml.293T,BCBL-1 and EA.hy926 cells could be efficiently infected by it.The expression of Nef in these cell lines could be detected by Western blot.In addition,Nef protein could inhibit the proliferation of BCBL-1 and EA.hy926 cells.Conclusion:Recombinant lentivirus with high titer and efficient infection of 293T,BCBL-1 and EA.hy926 cells,could be obtained quickly and simply by using the lentivirus vectors system,and Nef expression could be detected in 293T,BCBL-1 and EA.hy926 cells infected by lentivirus-Nef.Furthermore,our study suggested that Nef protein could inhibit the proliferation of BCBL-1 and EA.hy926 cells.