Proliferation of Ag-specific T cells is central to the development of protective immunity. The concomitant stimulation of the TCR and CD28 programs resting T cells to IL-2-driven clonal expansion. We report that a prolonged occupancy of the TCR and CD28 bypasses the need for autocrine IL-2 secretion and sustains IL-2-independent lymphocyte proliferation. In contrast, a short engagement of the TCR and CD28 only drives the expansion of cells capable of IL-2 production. TCR/CD28- and IL-2-driven proliferation revealed a different requirement for PI3K and for the mammalian target of rapamycin (mTOR). Thus, both PI3K and mTOR activities were needed for T cells to proliferate to TCR/CD28-initiated stimuli and for optimal cyclin E expression. In contrast, either PI3K or mTOR were sufficient for IL-2-driven cell proliferation as they independently mediated cyclin E induction. Interestingly, rapamycin delayed cell cycle entry of IL-2-sufficient T cells, but did not prevent their expansion. Together, our findings indicate that the TCR, CD28, and IL-2 independently control T cell proliferation via distinct signaling pathways involving PI3K and mTOR. These data suggest that Ag persistence and the availability of costimulatory signals and of autocrine and paracrine growth factors individually shape T lymphocyte expansion in vivo.

Muscle injury induces a classical inflammatory response in which cells of the innate immune system rapidly invade the tissue. Macrophages are prominently involved in this response and required for proper healing, as they are known to be important for clearing cellular debris and supporting satellite cell differentiation. Here, we sought to assess the role of the adaptive immune system in muscle regeneration after acute damage. We show that T lymphocytes are transiently recruited into the muscle after damage and appear to exert a pro-myogenic effect on muscle repair. We observed a decrease in the cross-sectional area of regenerating myofibers after injury in Rag2-/- γ-chain-/- mice, as compared to WT controls, suggesting that T cell recruitment promotes muscle regeneration. Skeletal muscle infiltrating T lymphocytes were enriched in CD4+CD25+FOXP3+ cells. Direct exposure of muscle satellite cells to in vitro induced Treg cells effectively enhanced their expansion, and concurrently inhibited their myogenic differentiation. In vivo, the recruitment of Tregs to acutely injured muscle was limited to the time period of satellite expansion, with possibly important implications for situations in which inflammatory conditions persist, such as muscular dystrophies and inflammatory myopathies. We conclude that the adaptive immune system, in particular T regulatory cells, is critically involved in effective skeletal muscle regeneration. Thus, in addition to their well-established role as regulators of the immune/inflammatory response, T regulatory cells also regulate the activity of skeletal muscle precursor cells, and are instrumental for the proper regeneration of this tissue.

Genetic programs promoting cell cycle progression, DNA repair, and survival are coordinately induced in developing T cells and require rapid turnover of effector molecules. As the COP9 signalosome (CSN) has been placed at the crossroads of these programs in lower organisms, we addressed its role by conditionally deleting CSN5/JAB1 , its catalytic subunit, in developing thymocytes. CSN5/JAB1 del/del thymocytes show defective S phase progression and massive apoptosis at the double-negative (DN) 4 -double-positive (DP) transition stage, which is paralleled by altered turnover of selected CSN-controlled substrates, including p53, I B-␣ , and ␤ -catenin. Combined dysregulation of the p53 and NF-B pathways affects thymocyte survival by altering the mRNA and protein levels of selected Bcl-2 family members. Genetic complementation analysis performed on p53 ؊ / ؊ , Bcl-xL/Bcl-2A1, or T cell receptor transgenic backgrounds indicates that CSN5/JAB1 acts at distinct developmental stages to coordinate proliferation, survival, and positive selection of thymocytes by controlling the induction of defi ned genetic programs acting downstream of CSN-regulated transcription factors. CSN5/JAB1-defi cient mice ( CSN5/JAB1 del/del ). The lack of CSN5/JAB1 mRNA and protein in DN and DP thymocytes was demonstrated by semiquantitative RT-PCR and Western blot analysis (Fig. S1 B). In addition, genomic DNA prepared from sorted DN and DP/single-positive (SP) cells was analyzed by PCR using primers detecting WT, fl oxed, or deleted alleles (Fig. S1 C). Most T cells in CSN5/JAB1 del/del mice showed a complete CSN5/JAB1 deletion, as indicated by the presence of the deleted allele in thymocytes and peripheral T cells. At the DN stage, CSN5/JAB1 del/del thymocytes yielded exclusively the PCR product corresponding to deleted allele. In peripheral T cells, however, we detected considerable levels of the CSN5/ JAB1 fl ox allele, suggesting that the residual mature T cells had escaped Cre-mediated deletion (Fig. S1 D). In support of this interpretation, peripheral T cells, although severely reduced in number, displayed a predominant memory-eff ector phenotype (CD44 high CD62L Ϫ ) in CSN5/JAB1-defi cient animals, suggesting homeostatic expansion of a small pool of phenotypically normal, mature T lymphocytes (Fig. S2).Thymic morphology, thymocyte counts, and subset distribution were comparatively assessed 5 -6 wk after birth in CSN5/JAB1 fl ox/fl ox mice expressing the LckCre transgene (herein defi ned as CSN5/JAB1 del/del ) and in littermates lacking LckCre . Thymi from CSN5/JAB1 del/del animals showed a severely hypoplastic medulla ( Fig. 1 A ), a marked decrease in the proportion of DP and SP thymocytes ( Fig. 1 B ), and an 80 -90% reduction in thymocyte number ( Fig. 1 C ), whereas the various subsets of DN thymocytes, as determined by the expression of the CD44 and CD25 stage-specifi c markers, were not signifi cantly altered compared with the CSN5/JAB1 fl ox/fl ox littermate controls, both percentage-wise ( Fig. 1 B ) and in absolute numbers ( F...

Nonmyeloablative hematopoietic cell transplantation can cure patients with hematologic malignancies but has reported limited success against solid tumors. This is possibly because of profound peripheral tolerance mechanisms and/or suboptimal tumor recognition by effector T lymphocytes. We report that in mice developing spontaneous prostate cancer, nonmyeloablative minor histocompatibility mismatched hematopoietic stem cell transplantation, and donor lymphocyte infusion of unmanipulated lymphocytes combined with posttransplant tumor-specific vaccination circumvents tumor-specific tolerance, allowing acute tumor rejection and the establishment of protective immunosurveillance. Although donor-derived tumor-specific T cells readily differentiated into effector cells and infiltrated the tumor soon after infusion, they were alone insufficient for tumor eradication, which instead required the concomitance of minor histocompatibiltiy antigen-specific CD8 + T-cell responses. The establishment of protective immunosurveillance was best induced by posttransplant tumor-specific vaccination. Hence, these results provide the proof of principle that tumor-specific T-cell responses have to be harnessed together with minor histocompatibility responses and sustained by posttransplant tumor-specific vaccination to improve the efficacy of allotransplantion for the cure of solid tumors.

Prep1 is a homeodomain transcription factor that acts by dimerizing withi/i fetal liver cells transplanted showed the same defects as the Prep1 i/i mice did. Among PBC family members, Pbx2 and very low levels of Pbx3 were observed in the thymi of wild-type mice. In Prep1 i/i mice, the level of Pbx2 protein was profoundly decreased, while for Pbx3 no definitive conclusion could be reached. Therefore, the deficient postnatal Tlymphocytic potential of the Prep1 hematopoietic progenitors depends on the combined, not compensated, absence of Prep1 and at least Pbx2.

Lysosomal ␤-galactosylceramidase deficiency results in demyelination and inflammation in the nervous system causing the neurological Krabbe disease. In the Twitcher mouse model of this disease, we found that neurological symptoms parallel progressive and severe lymphopenia. Although lymphopoiesis is normal before disease onset, primary and secondary lymphoid organs progressively degenerate afterward. This occurs despite preserved erythropoiesis and leads to severe peripheral lymphopenia caused by reduced numbers of T cell precursors and mature lymphocytes. Hematopoietic cell replacement experiments support the existence of an epigenetic factor in mutant mice reconcilable with a progressive loss of autonomic axons that hampers thymic functionality. We propose that degeneration of autonomic nerves leads to the irreversible thymic atrophy and loss of immune-competence. Our study describes a new aspect of Krabbe disease, placing patients at risk of immune-related pathologies, and identifies a novel target for therapeutic interventions.

We have previously shown that chymotrypsin-cleaved soluble uPAR (D2D388-274) elicits migration of monocytic cells through interaction with FPRL-1, a G protein-coupled receptor that is homologous to the fMLP receptor. Here, we report that D2D388-274 also modulates the ability of monocytes to migrate in response to other chemokines. Pretreatment of monocytes with increasing amounts of D2D388-274 prevents cell migration in response to MCP-1, RANTES and fMLP. We demonstrate that D2D388-274 does not inhibit MCP-1 receptor binding, elicit CCR2 internalization and prevent MCP-1-induced intracellular Ca 2+ increase. Thus, CCR2 receptor desensitization cannot account for D2D388-274-mediated inhibition of MCP-1-induced cell migration. Rather, we show that pretreatment of monocytes with D2D388-274 dramatically decreases chemokine-induced integrin-dependent rapid cell adhesion by interacting with FPRL-1. Together, our results indicate that chemokine-dependent cell migration can be regulated not only by homologous and heterologous receptor desensitization, but also by inhibition of integrindependent cell adhesion, an important step in cell transmigration.

Resistance and tolerance mechanisms participate to the interplay between host and pathogens. IL-17-mediated response has been shown to be crucial for host resistance to respiratory infections, whereas its role in host tolerance during chronic airway colonization is still unclear. Here, we investigated whether IL-17-mediated response modulates mechanisms of host tolerance during airways chronic infection by P. aeruginosa. First, we found that IL-17A levels were sustained in mice at both early and advanced stages of P. aeruginosa chronic infection and confirmed these observations in human respiratory samples from cystic fibrosis patients infected by P. aeruginosa. Using IL-17a−/− or IL-17ra−/− mice, we found that the deficiency of IL-17A/IL-17RA axis was associated with: i) increased incidence of chronic infection and bacterial burden, indicating its role in the host resistance to P. aeruginosa; ii) reduced cytokine levels (KC), tissue innate immune cells and markers of tissue damage (pro-MMP-9, elastin degradation, TGF-β1), proving alteration of host tolerance. Blockade of IL-17A activity by a monoclonal antibody, started when chronic infection is established, did not alter host resistance but increased tolerance. In conclusion, this study identifies IL-17-mediated response as a negative regulator of host tolerance during P. aeruginosa chronic airway infection.