Accumulating evidence indicates that human herpesvirus 8 (HHV8), also known as Kaposi's sarcoma-associated herpesvirus (KSHV), is the etiological agent associated with the development of Kaposi's sarcoma (KS). In addition to KS, HHV8 is also associated with the development of primary effusion lymphoma (PEL) and some cases of multicentric Castleman's disease (MCD), both B-cell lymphoproliferative disorders (LPD) observed in humans with the acquired immunodeficiency syndrome (AIDS) also presenting with KS. Understanding how HHV8 is involved in these diverse disease manifestations is complicated with the lack of an accessible animal model that recapitulates these diseases. Recently, the investigators and others reported that rhesus macaques harbor a simian herpesvirus that is closely related to HHV8, referred to as rhesus rhadinovirus (RRV). Moreover, experimental inoculation of rhesus macaques with RRV strain 17577 indicates that in macaques previously inoculated with SIVmac239, RRV strain 17577 induces disease manifestations that possess features that resemble KS and MCD. The long-term objectives of this application are to further elucidate how SIV and RRV induce these disease manifestations and to develop this as an animal model to address HHV8-associated disease. To accomplish this, the following specific aims are proposed.
Specific Aim 1 : Role of immunosuppression in RRV strain 17677-associated disease. Rhesus macaques will be immunosuppressed by either SIVmac239 infection or by iatrogenic agents and inoculated intravenously with RRV strain 17577. Experimentally inoculated macaques will be monitored for viral load by semi-quantitative PCR, changes in leukocyte subsets in peripheral blood and lymph nodes, serological response to RRV, and for clinical signs of RRV-associated disease. Host response to virus infection will be analyzed for inflammatory cytokine expression and activity by RT-PCR, enzyme-linked immunosorbent assays (ELISA) and immunohistochemical staining on samples collected from peripheral blood, lymph node and retroperitoneal fibromatosis (RF) samples from RRV-, SIVmac239- and RRV/SIVmac239-infected samples. Differential expression of activity of inflammatory responses may indicate how RRV/SIVmac239 infection results in the development of RRV-associated disease.
Specific Aim 2 : Molecular genetic manipulation of the RRV genome. The investigators propose to initiate molecular genetic manipulation of the RRV genome to identify viral determinants of pathogenesis in SIVmac239-infected rhesus macaques. Viral orfs that they believe are important based upon in vitro analysis of infected tissue samples and in vitro cell culture systems will be targeted and nonsense mutations inserted to abrogate the expression of the viral gene product.
Specific Aim 3 : In vivo pathogenesis of mutant RRVs with defined nonsense mutations in targeted viral orfs. The results from these proposed studies should provide new insights into how RRV and SIV together induce RRV-associated disease, and serve as a means to elucidate the role of HHV8 in HHV8-associated disease.
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