Exposure to Toll-Like Receptors Ligands Results in T Cell Activation and Disproportionate

EXPOSURE TO TOLL-LIKE RECEPTORS LIGANDS RESULTS IN T CELL ACTIVATION AND DISPROPORTIONATE CD4+ T CELL APOPTOSIS

Angel A. Luciano, Nicholas T. Funderburg, Wei Jang, Scott Sieg, and Michael Lederman

Center for AIDS Research, Case Western Reserve University and University Hospitals of Cleveland, Department of Pediatrics, Division of Neonatology; School of Medicine, Case Western Reserve University, Cleveland, OH.

Background: Activation of Toll-like receptors (TLRs) results in a cascade of events that among other outcomes, enhances the arming of adaptive immune defenses. Recent work has also implicated microbial TLR ligands in the pathogenesis of immune activation and cellular losses in chronic HIV infection.

Methods: We examined the effects of TLR ligand exposure on T cell activation in vitro. Peripheral blood mononuclear cells (PBMC) and purified T cell populations obtained from healthy controls were exposed to TLR ligands in vitro and examined for evidence of immune activation by flow cytometric analysis. PBMC from healthy donors were stimulated with individual TLR ligands (E coli peptidoglycan (PGN), Poly I:C, E. Coli lipopolysaccharide , Salmonella typhimurium flagellin, Bacillus Subtilis flagellin, imiquimod, single stranded Poly U (ssPolyU) and unmethylated DNA (CpG 2395) and activation markers (CD38, CD69, Ki 67) were monitored by flow cytometry at days 1 and 7. Cellular proliferation and apoptosis were measured by carboxy-fluorescein diacetate succinimidyl ester (CFSE) dye dilution and binding of Annexin
Results: Exposure to TLR ligands results in the differential and largely indirect activation of memory and effector CD4+ and CD8+ T cells but not naïve T cells. After exposure to each of 8 different ligands that activate TLRs 2, 3, 4, 5, 7, 8, and 9, induction of the ectoenzyme CD38 was seen on both CD4+ and CD8+ T lymphocytes. Also, CD8+ T cells gain expression of the C type lectin CD69 that has been associated with enhanced lymphocyte retention in secondary lymphoid organs (n=20). In contrast, CD4+ T cells uncommonly increase CD69 expression, but instead enter cell cycle (Ki67+) (n=20). Group medians for induction of CD69 and Ki67 were compared using the Wicoxon signed rank test. Despite activation and cell cycle entry, CD4+ T cells divide poorly and instead, disproportionately undergo activation-induced cell death.

Conclusions: Systemic exposure to microbial products may therefore increase immune activation, cellular sequestration and T cell turnover as may be seen in the setting of heightened microbial translocation through mucosal surfaces during sepsis, and in chronic infection with the human immunodeficiency virus.