Regulatory T cells (Tregs

The clearance of dying cells is vital for re-establishing tolerance during inflammation and has potent immunoregulatory consequences. Because natural IgM plays a key role in the removal of apoptotic cells, we investigated whether the immune modulatory properties of apoptotic cells depended on its presence. Using an Ab-independent, Ag-induced model of inflammatory arthritis, we tested whether natural IgM is essential for the arthritis-suppressing properties of apoptotic cells. Whereas administration of apoptotic cells reduced joint inflammation and damage in normal mice accompanied by suppression of the Th17 response, no protection was afforded in secreted IgM-deficient (Sμ–) mice. The enhanced production of IL-10 by T cells from draining lymph nodes and splenic marginal zone B cells, driven by the infusion of apoptotic cells, was abrogated in the absence of natural IgM. Apoptotic cells were present shortly after administration in the splenic marginal zone, but their removal was substantially delayed in the absence of natural IgM. Incubation of apoptotic cells with natural IgM in vitro restored their arthritis-suppressing properties in Sμ– mice. Moreover, these IgM-coated apoptotic cells were cleared rapidly after injection from the spleens of Sμ– mice. Our results demonstrate that natural IgM is a critical factor in a chain of events triggered by the administration of apoptotic cells that promote IL-10–secreting B and T cells and restrain the development of inflammation.

IntroductionRetroviral T-cell receptor (TCR) gene transfer is an attractive strategy by which large numbers of antigen-specific T cells can be generated for adoptive transfer. [1][2][3][4] One of the advantages of this technique is that it can be used to circumvent possible impairment of autologous T-cell responses against tumor associated antigens as it bypasses central tolerance. 5 In addition, the introduced TCR specificity can be targeted against poorly immunogenic antigens and high affinity TCR can be selected for transfer. 6 This method has recently seen success in the first clinical trial of TCR gene therapy, where MART1 specific T cells, generated by retroviral gene transfer, were adoptively transferred into patients with metastatic melanoma. The engineered T cells engrafted in 15 of 17 patients, 2 of whom demonstrated long term tumor regression. 7 T cells engineered to express CEA-specific TCR also induced a decrease in CEA levels in 3 patients with metastatic colorectal cancer, and regression of tumor metastases in 1 patient, but were associated with a severe colitis in all 3 patients. 8 The most promising results were seen after adoptive transfer of TCR transduced T cells specific for the NY-ESO-1antigen which resulted in clinical responses in synovial cell sarcoma and in melanoma. 9 As the density of TCR on the surface of cells affects their functional avidity, inefficient TCR expression may impair the success of TCR gene therapy. [10][11][12][13][14] Several different strategies have been developed to increase the expression of introduced ␣ and ␤ chains by reducing the mispairing with endogenous TCR chains. The strategies include replacing the human TCR constant domain with murine sequences, the introduction of an additional disulphide bond into the constant regions, and the production of hybrid molecules consisting of the extracellular portion of TCR chains fused to the intracellular CD3 domain. [15][16][17][18] TCR ␣ and ␤ chains form a complex with 4 invariant CD3 chains: ␥,␦,⑀ and . This complex formation is required in order for the TCR to be expressed on the cell surface, and for signal transduction on antigen recognition. TCR introduced by retroviral gene transfer are likely to be in competition with endogenous TCR molecules for CD3 chains. We used a murine model system to explore whether the co-transfer of TCR genes together with the genes encoding the ␥,␦,⑀ and chains of the CD3 complex can augment TCR expression. Two different TCR, with specificity for Wilms' Tumor antigen 1 (WT1) and influenza nucleoprotein, were used to demonstrate that CD3 is rate limiting for the expression of introduced TCR in gene modified T cells. Co-transduction of CD3 and TCR genes resulted in up to 16-20 fold increase in TCR surface expression and tetramer binding compared with transduction of TCR genes alone. The increase in TCR expression was associated with increased T-cell avidity leading to improved recognition of low concentration of peptide antigen. In vivo TCRϩCD3 co-transduced T cells eradicate tumors fa...

Aging is associated with a dysfunctional endothelial phenotype as well as reduced angiogenic capabilities. Exercise exerts beneficial effects on the cardiovascular system, possibly by increasing/maintaining the number and/or function of circulating angiogenic cells (CACs), which are known to decline with age. However, the relationship between cardiorespiratory fitness (CRF) and age-related changes in the frequency of CACs, as well as the exercise-induced responsiveness of CACs in older individuals, has not yet been determined. One-hundred seven healthy male volunteers, aged 18-75 yr, participated in study 1. CRF was estimated using a submaximal cycling ergometer test. Circulating endothelial progenitor cells (EPCs), angiogenic T cells (T), and their chemokine (C-X-C motif) receptor 4 (CXCR4) cell surface receptor expression were enumerated by flow cytometry using peripheral blood samples obtained under resting conditions before the exercise test. In study 2, 17 healthy men (8 young men, 18-25 yr; 9 older men, 60-75 yr) were recruited, and these participants undertook a 30-min cycling exercise bout at 70% maximal O consumption, with CACs enumerated before and immediately after exercise. Age was inversely associated with both CD34 progenitor cells ( r = -0.140, P = 0.000) and T ( r = -0.176, P = 0.000) cells as well as CXCR4-expressing CACs (CD34: r = -0.167, P = 0.000; EPCs: r = -0.098, P = 0.001; T: r = -0.053, P = 0.015). However, after correcting for age, CRF had no relationship with either CAC subset. In addition, older individuals displayed attenuated exercise-induced increases in CD34 progenitor cells, T, CD4, T, and CD8CXCR4 T cells. Older men display lower CAC levels, which may contribute to increased risk of cardiovascular disease, and older adults display an impaired exercise-induced responsiveness of these cells. NEW & NOTEWORTHY Older adults display lower circulating progenitor cell and angiogenic T cell counts compared with younger individuals independently of cardiometabolic risk factors and cardiorespiratory fitness. Older adults also display impaired exercise-induced mobilization of these vasculogenic cells.

Adoptive transfer of antigen-specific T lymphocytes is an effective form of immunotherapy for persistent virus infections and cancer. A major limitation of adoptive therapy is the inability to isolate antigen-specific T lymphocytes reproducibly. The demonstration that cloned T-cell receptor (TCR) genes can be used to produce T lymphocyte populations of desired specificity offers new opportunities for antigen-specific T-cell therapy. TCR gene-modified lymphocytes display antigen-specific function in vitro, and were shown to protect against virus infection and tumor growth in animal models. A recent trial in humans demonstrated that TCR gene-modified T cells persisted in all and reduced melanoma burden in 2/15 patients. In future trials, it may be possible to use TCR gene transfer to equip helper and cytotoxic T cells with new antigen-specificity, allowing both T-cell subsets to cooperate in achieving improved clinical responses. Sequence modifications of TCR genes are being explored to enhance TCR surface expression, while minimizing the risk of pairing between introduced and endogenous TCR chains. Current T-cell transduction protocols that trigger T-cell differentiation need to be modified to generate "undifferentiated" T cells, which, upon adoptive transfer, display improved in vivo expansion and survival. Both, expression of only the introduced TCR chains and the production of naïve T cells may be possible in the future by TCR gene transfer into stem cells.

In this study, we generated human MHC Class I-restricted CD4+ T cells specific for Epstein-Barr virus (EBV) and cytomegalovirus (CMV), two herpesviridae associated with lymphoma, nasopharyngeal carcinoma and medulloblastoma, respectively. Retroviral transfer of virus-specific, HLA-A2-restricted TCR-coding genes generated CD4+ T cells that recognized HLA-A2/peptide multimers and produced cytokines when stimulated with MHC Class II-deficient cells presenting the relevant viral peptides in the context of HLA-A2. Peptide titration revealed that CD4+ T cells had a 10-fold lower avidity than CD8+ T cells expressing the same TCR. The impaired avidity of CD4+ T cells was corrected by simultaneously transferring TCR- and CD8-coding genes. The CD8 co-receptor did not alter the cytokine signature of CD4+ T cells, which remained distinct from that of CD8+ T cells. Using the xenogeneic NOD/SCID mouse model, we demonstrated that human CD4+ T cells expressing a specific TCR and CD8 can confer efficient protection against the growth of tumors expressing the EBV or CMV antigens recognized by the TCR. In summary, we describe a robust approach for generating therapeutic CD4+ T cells capable of providing MHC Class I-restricted immunity against MHC Class II-negative tumors in vivo.

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