Generation of vFLIP Transgenic Mice: A Model to Study KSHV-Associated Lymphomagenesis

Abstract

Primary effusion lymphoma (PEL) is a distinct subtype of aggressive non-Hodgkin's lymphoma (NHL), specifically associated with infection by Kaposi's sarcoma-associated herpesvirus (KSHV) and occurring more frequently in HIV-infected individuals. Several in vitro observations suggest that vFLIP, a viral protein expressed during latency, is an important viral oncogene. It is essential for the survival of KSHV-infected PEL cells, mainly by constitutively activating the NF-kB pathway. In order to assess the role of vFLIP in the pathogenesis of PEL, we knocked a cDNA encoding vFLIP, preceded by a loxP-flanked neoR-Stop cassette and followed by Frt-flanked IRES-eGFP sequences, into the ubiquitously expressed ROSA26 locus. A specifically restricted expression of the transgene in CD19+ B-cells has been achieved by crossing the ROSA26. vFLIP knock-in mice with mice expressing cre recombinase under the control of the CD19 promoter. These mice have also been crossed with transgenic mice expressing KSHV LANA, which is considered to also be a viral oncogene, to assess a potential synergistic effect of these two KSHV latent proteins in the lymphomagenic process of PEL. vFLIP expression in the CD19+ B-cells results in splenomegaly, with an increase in both T and B-cells, and with a relative increase of the T versus B-cell ratio. Although primary follicles were enlarged, the expression of vFLIP in the CD19+ B-cells results in lack of germinal center formation in the spleen, lymph nodes and intestine, and in partially impaired class-switching recombination. vFLIP transgenic mice had an increased number of plasmablast-like cells expressing lambda light chain, reminiscent of a phenomenon seen in KSHV-associated multicentric Castleman's disease (MCD). These results indicate that by constitutively activating the NF-κB pathway in pre-germinal center B-cells expressing CD19, the normal B-cell differentiation is impaired, and provide clues about possible aberrant differentiation in PEL and MCD.