The Role of Feline Glycoprotein A Repetitions Predominant (GARP) in Regulatory T Cell Mediated Immune Dysfunction During FIV Lentiviral Infection.

摘要

Feline Immunodeficiency Virus (FIV) is an important pathogen for companion animal medicine which is homologous in structure and pathogenesis to the human lentivirus, HIV, and therefore serves as an appropriate model for the study of HIV-AIDS in a naturally infected host. Our lab has described the contributions of T regulatory (Treg) cells to the disease progression and immunologic dysfunction associated with FIV. We previously reported that membrane-associated TGFbeta (mTGFbeta) on Treg cells mediates suppressor function against CD4+ T helper (Th) cells, and that FIV infection of cats results in activation of these mTGFbeta+CD4 +CD25+ Treg cells in vivo. This dissertation focuses on the role of a novel protein, Glycoprotein A Repetitions Predominant (GARP), during FIV infection and provides the first description of GARP in the feline system, demonstrating that it serves as a membrane anchor for TGFbeta on the surface of feline Treg cells. We examine co-expression of TGFbeta and GARP on CD4+CD25+ Treg cells from chronically FIV-infected cats and demonstrate that GARP+ Treg cells are activated during infection and function as highly effective suppressors of CD4+ Th cells. Having demonstrated that GARP+ Treg cells are activated during infection, we next investigated the mechanism of activation. To determine if FIV infection was sufficient to directly activate GARP+ Treg cells, purified CD4+CD25+ cells from FIV negative cats were infected in vitro and phenotypic and functional changes analyzed. Between 3 and 6 days post in vitro infection, GARP and TGFbeta expression were increased on CD4+CD25 + cells. These CD4+CD25+GARP + cells also expressed the Treg specific transcription factor, FoxP3, demonstrating that FIV infection alone induces an activated regulatory cell phenotype. To determine if these cells were functionally activated, 6 day in vitro FIV-infected CD4+CD25+ cells were co-cultured with ConA activated CD4+CD25- Th cells for 24 hours and IL2 levels I the culture supernatant measured. In vitro infection-activated CD4+CD25+ cells were capable of suppressing IL2 production from activated Th cells. From these data, we hypothesize that the in vivo activation of GARP:TGFbeta expressing Treg cells by direct FIV infection contributes to the CD4+ T cell anergy via ligation of TGFbeta-RII on the surface of CD4+CD25- Th cells. Progression through the chronic stage of infection is characterized by a generalized CD4+ T cell depletion but despite this, the percentage of Treg cells in the CD4+ population remains constant. Having previously demonstrated that soluble TGFbeta is sufficient to convert activated CD4+CD25- Th cells into suppressor cells, we asked whether the expanded CD4+CD25+GARP +mTGFbeta+ population identified during FIV infection could engage in TGFbeta-mediated signaling with TGFbeta-RII+CD4 +CD25- Th and convert these cells into regulatory cells, thereby maintaining Treg numbers by recruitment of new Treg cells from the Th pool during FIV infection. Here, we demonstrate that GARP+ Treg cells activated during FIV-infection in vivo are capable of converting activated Th cells into phenotypic and functional Treg cells in vitro. This process is mediated by mTGFbeta on the Treg cells and TGFbeta-RII on the target Th cells as the addition of TGFbeta/ TGFbeta-RII blocking antibodies prior to co-culture abrogated conversion. Taken together, these data suggest that FIV infection is characterized by the expansion and activation of infected GARP+ Treg cells and continued immune dysfunction results from conversion of FIV-activated Th cells into regulatory cells. Death of FIV-infected T cells coupled to the conversion of Th cells leads to a continual cycle of Th depletion consistent with CD4 decline culminating in AIDS pathogenesis. In addition to providing a mechanistic perspective for disease progression, this discovery suggests a novel point of therapeutic intervention. Further research aimed at providing more detailed longitudinal data for conversion dynamics over the course of infection and the specificities of the TGFbeta-mediated conversion signaling pathways will provide important information for future Treg-based immunotherapy development.