T lymphocytes modulate early ischemia-reperfusion injury in the kidney; however, their role during repair is unknown. We studied the role of TCRbeta(+)CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs), known to blunt immune responses, in repair after ischemia-reperfusion injury to the kidney. Using a murine model of ischemic acute kidney injury we found that there was a significant trafficking of Tregs into the kidneys after 3 and 10 days. Post-ischemic kidneys had increased numbers of TCRbeta(+)CD4(+) and TCRbeta(+)CD8(+) T cells with enhanced pro-inflammatory cytokine production. Treg depletion starting 1 day after ischemic injury using anti-CD25 antibodies increased renal tubular damage, reduced tubular proliferation at both time points, enhanced infiltrating T lymphocyte cytokine production at 3 days and TNF-alpha generation by TCRbeta(+)CD4(+) T cells at 10 days. In separate mice, infusion of CD4(+)CD25(+) Tregs 1 day after initial injury reduced INF-gamma production by TCRbeta(+)CD4(+) T cells at 3 days, improved repair and reduced cytokine generation at 10 days. Treg manipulation had minimal effect on neutrophil and macrophage infiltration; Treg depletion worsened mortality and serum creatinine, while Treg infusion had a late beneficial effect on serum creatinine in bilateral ischemia. Our study demonstrates that Tregs infiltrate ischemic-reperfused kidneys during the healing process promoting repair, likely through modulation of pro-inflammatory cytokine production of other T cell subsets. Treg targeting could be a novel therapeutic approach to enhance recovery from ischemic acute kidney injury.

Healthy liver, intestine, lung, and skin harbor resident lymphocytes with conventional and unconventional phenotypes. Lymphocytes also have been detected in healthy mice kidneys; however, these cells have not been well studied and have been largely overlooked. To better characterize the intra-renal lymphocytes, we extensively perfused C57BL/6J mice with PBS and then isolated mononuclear cells for flow cytometry analysis. We observed T cells, B cells, and NK cells in normal mice kidneys after extensive perfusion. Approximately 50% of kidney T lymphocytes expressed intermediate levels of CD3 (CD3int T cells). Similar to liver and lung, a high percentage of unconventional CD3+CD4(-)CD8(-) double-negative T cells was observed in normal mice kidneys, from which 11% expressed B220 antigen. Unlike the spleen and blood, the classic CD4+ and CD8+ T lymphocytes in the kidney had a high proportion of activated CD69+ and effector/memory CD44- CD62L ligand phenotypes. Also, a small percentage of CD4+CD25+forkhead box p3+ and NKT cells was observed in perfused and exanguinated kidneys. In addition, a distinct TCR repertoire was found on intra-renal conventional and unconventional T cells compared with those from the spleen. Finally, after 24 h of renal ischemia reperfusion injury (IRI), increased production of cytokines IFN-gamma and TNF-alpha by CD4+ and CD8+ T cells, isolated from perfused kidneys, was observed. These data suggest that some of these cells harbored in the kidney could be implicated in the immune response of the IRI pathogenic process.

Kidney ischemia-reperfusion injury (IRI) is, in part, mediated by immune and inflammatory factors. Since microbial stimuli are known to alter immune and inflammatory responses, we hypothesized that differences in perinatal microbial status would modify renal injury following IRI. We performed bilateral renal IRI on 6-wk-old germ-free and control mice and studied the effects on kidney lymphocyte trafficking, cytokines, function, and structure. Compared with control mice, normal kidneys of germ-free mice exhibited more NKT cells and lower IL-4 levels. Postischemia, more CD8 T cells trafficked into postischemic kidneys of germ-free mice compared with control mice. Renal structural injury and functional decline following IRI were more severe in germ-free mice compared with control mice. When germ-free mice were conventionalized with the addition of bacteria to their diet, the extent of renal injury after IRI became equivalent to age-matched control mice, with similar numbers and phenotypes of T cells and NKT cells, as well as cytokine expression in both normal kidneys and postischemic kidneys of conventionalized germ-free mice and age-matched control mice. Thus microbial stimuli influence the phenotype of renal lymphocytes and the expression of cytokines of normal kidneys and also modulate the outcome of IRI.

There is no established modality to repair kidney damage resulting from ischemia-reperfusion injury (IRI). Early responses to IRI involve lymphocytes, but the role of B cells in tissue repair after IRI is unknown.

Green tea, a product of the dried leaves of Camellia sinensis, is the most widely consumed beverage in the world. The polyphenolic compounds from green tea (PGT) possess anti- inflammatory properties. We investigated whether PGT can afford protection against autoimmune arthritis, and also examined the immunological basis of this effect using the rat adjuvant arthritis (AA) model of human rheumatoid arthritis (RA). AA can be induced in the Lewis rat (RT.1l) by immunization with heat-killed Mycobacterium tuberculosis H37Ra (Mtb), and arthritic rats raise T cell response to the mycobacterial heat-shock protein 65 (Bhsp65). PGT (2-12 g/L, w/v) was fed to rats in drinking water for 1-3 wk followed by disease induction by Mtb injection. Thereafter, these rats were observed regularly and graded for signs of arthritis. Sub-groups of these rats were killed at defined time points, and their draining lymph node cells (LNC) were harvested and tested for T cell proliferative and cytokine responses. Furthermore, the sera collected from these rats were tested for anti-Bhsp65 antibodies. We observed that feeding 8 g/L PGT to Lewis rats for 9 d significantly reduced the severity of arthritis compared to the water-fed controls. Interestingly, PGT-fed rats had lower concentrations of the pro-inflammatory cytokine interleukin-17 (IL-17), but greater concentration of the immunoregulatory cytokine IL-10 than controls. PGT feeding also suppressed the anti-Bhsp65 antibody response. Thus, green tea induced changes in arthritis related immune response. We suggest further systematic exploration of dietary supplementation with PGT as an adjunct nutritional strategy for the management of RA.

T cells have been implicated in the early pathogenesis of ischemia reperfusion injury (IRI) of kidney, liver, lung, and brain. It is not known whether Ag-TCR engagement followed by Ag-specific T cell activation participates in IRI. T cell-deficient nu/nu mice are moderately resistant to renal IRI, which can be reversed upon reconstitution with syngeneic T cells. In this study, we found that nu/nu mice reconstituted with DO11.10 T cells, limited in their TCR repertoire, have significantly less kidney dysfunction and tubular injury after renal IRI compared with that in nu/nu mice reconstituted with wild-type T cells having a diverse TCR repertoire. CD4+ T cells infiltrating ischemic kidneys of nu/nu mice reconstituted with DO11.10 T cells exhibited lower IFN-γ production than that of wild-type controls. Frequency of regulatory T cells in kidneys of these mice was similar in both DO11.10 T cells and wild-type T cell recipient groups. DO11.10 mice immunized with OVA-CFA had significantly worse kidney function at 24 h after ischemia than those immunized with CFA alone. Thus, without T cell activation, diverse TCR repertoire was important for renal IRI in naive mice. However, once T cells were activated in an Ag-specific manner through TCR in DO11.10 mice, a restricted TCR repertoire no longer limited the extent of kidney injury. Thus, both TCR repertoire-dependent and -independent factors mediate T cell functions in kidney IRI.

Objective. Tolerization of T cells directed against a target autoantigen is a desired goal of experimental approaches for the treatment of autoimmune diseases, and novel and improved methods of tolerance induction are continuously being sought. Because most traditional methods of tolerance induction using soluble antigen are effective in the prevention of autoimmunity but fail to control established disease, this study was carried out to explore an innovative tolerogenic approach for the treatment of ongoing disease, using the rat adjuvantinduced arthritis (AIA) model of human rheumatoid arthritis.Methods. Lewis (RT.

Objective. Pretreatment of Lewis rats with soluble mycobacterial Hsp65 affords protection against subsequent adjuvant-induced arthritis (AIA). This study was aimed at unraveling the mechanisms underlying mycobacterial Hsp65-induced protection against arthritis, using contemporary parameters of immunity.Methods. Lewis rats were given 3 intraperitoneal injections of mycobacterial Hsp65 in solution prior to the initiation of AIA with heat-killed Mycobacterium tuberculosis. Thereafter, mycobacterial Hsp65-specific T cell proliferative, cytokine, and antibody responses were tested in tolerized rats. The roles of anergy and the indoleamine 2,3 dioxygenase (IDO)-tryptophan pathway in tolerance induction were assessed, and the frequency and suppressive function of CD4؉FoxP3؉ Treg cells were monitored. Also tested was the effect of mycobacterial Hsp65 tolerization on T cell responses to AIA-related mycobacterial Hsp70, mycobacterial Hsp10, and rat Hsp65.Results. The AIA-protective effect of mycobacterial Hsp65-induced tolerance was associated with a significantly reduced T cell proliferative response to mycobacterial Hsp65, which was reversed by interleukin-2 (IL-2), indicating anergy induction. The production of interferon-␥ (but not IL-4/IL-10) was increased, with concurrent down-regulation of IL-17 expression by mycobacterial Hsp65-primed T cells. Neither the frequency nor the suppressive activity of CD4؉FoxP3؉ T cells changed following tolerization, but the serum level of anti-mycobacterial Hsp65 antibodies was increased. However, no evidence was observed for a role of IDO or cross-tolerance to mycobacterial Hsp70, mycobacterial Hsp10, or rat Hsp65.Conclusion. Tolerization with soluble mycobacterial Hsp65 leads to suppression of IL-17, anergy induction, and enhanced production of anti-mycobacterial Hsp65 antibodies, which play a role in protection against AIA. These results are relevant to the development of effective immunotherapeutic approaches for autoimmune arthritis.