UCLA

A hallmark of age-related immunosenescence is the accumulation of late-differentiated memory CD8 T cells in peripheral blood with features of replicative senescence, such as inability to proliferate, absence of CD28 expression, shortened telomeres, loss of telomerase activity, enhanced activation, and increased secretion of inflammatory cytokines. Oligoclonal expansions of these late-differentiated memory CD8 T cells in peripheral blood are correlated with increased risk of morbidity and mortality in elderly humans. The studies described in this thesis focus on furthering our understanding of CD8 T cell replicative senescence in humans and its relationship to aging and HIV disease, with special emphasis on immune features of gut-associated lymphoid tissue (GALT), which contains 40-65% of lymphocytes (as opposed to peripheral blood, which houses only 2%) and is an area of high antigenic exposure.

We first review the data for CD8 T cell replicative senescence in peripheral blood. This includes a comprehensive discussion of experiments defining the phenotype and function of senescent cells in vitro. We then present evidence that CD8 T cell replicative senescence occurs in vivo, and discuss how senescent cells contribute to, and are affected by, dysregulated immunity in aging and HIV disease, ultimately contributing towards "inflammaging" and an immune risk phenotype (IRP) predictive of morbidity and mortality. We next move the focus of our studies to the GALT and compare blood and gut (specifically rectosigmoid colon) CD8 T cells, focusing on age-related phenotypic and functional alterations previously linked to senescence in peripheral blood. Overall, our results support the hypothesis that gut CD8 T cells have profiles suggestive of greater "aging" than those in peripheral blood. These results include a significant increase in RA-CD8+ (memory), CD28-CD8+ (differentiated), RA-CD28-CD8+ (differentiated memory), DR+CD38+CD8+ (activated), CD8 &alpha/&alpha, RO+PD-1+CD8+, and a (non-significant) decrease in baseline telomerase activity. However, other criteria from this study (lower levels of CD57 expression, lack of inverted CD4:CD8 ratio, increase in KI-67 (proliferation) expression, and proliferative potential that is similar to blood) suggest that gut CD8 T cells may not be more senescent than peripheral blood. Our data suggest that additional work (such as looking at CD4 cells, other regions of the gut, and CD8 T cell gut senescence in aging and HIV disease) is needed to further elucidate this relationship. We then describe the development and optimization of a novel protocol to test our hypothesis of compartment-specific differences of in vitro proliferative dynamics of CD8 T cells. The protocol involves a 5 day culture of mononuclear leukocyte populations from blood and gut, respectively, labeled with CFSE and BrdU and stimulated with anti-CD2/3/28-linked microbeads. The specific experimental variables we have optimized include: mode of T cell stimulation, CFSE concentration, inclusion of second proliferation marker BrdU, culture duration, initial cell concentration and inclusion of autologous irradiated feeder cells. Finally, we transition back to peripheral blood in order to examine a link between peripheral blood T cell senescence and a mild presentation of Kaposi's sarcoma in treated HIV-infected subjects with long-term suppression of HIV replication. This study indicates these subjects exhibit some features of immunosenescence relative to treated controls without sarcoma (increased frequency of CD57+ and CD28- CD4 and CD8 T cells, decreased proportion of naïve CD27+CD28+CD45RA+ CD4 and CD8 cells, increased expression of CCR5 in CD4 and CD8 T cells), indicating immunologic perturbations associated with immune senescence might be casually associated with development of this disease. Together, results from this research further our understanding of CD8 T cell senescence, especially in the rectosigmoid colon region of the GALT, and pave the way for further research elucidating the relationship of CD8 T cell immunosenescence and pathologies such as aging, HIV disease and Kaposi's sarcoma.