Central tolerance to tumor-associated Ags is an immune-escape mechanism that significantly limits the TCR repertoires available for tumor eradication. The repertoires expanded in wild-type BALB/c and rat-HER-2/neu (rHER-2) transgenic BALB-neuT mice following DNA immunization against rHER-2 were compared by spectratyping the variable (V)β and the joining (J)β CDR 3. Following immunization, BALB/c mice raised a strong response. Every mouse used one or more CD8+ T cell rearrangements of the Vβ9-Jβ1.2 segments characterized by distinct length of the CDR3 and specific for 63-71 or 1206-1214 rHER-2 peptides. In addition, two CD4+ T cell rearrangements recurred in >50% of mice. Instead, BALB-neuT mice displayed a limited response to rHER-2. Their repertoire is smaller and uses different rearrangements confined to CD4+ T cells. Thus, central tolerance in BALB-neuT mice acts by silencing the BALB/c mice self-reactive repertoire and reducing the size of the CD8+ T cell component. CD8+ and CD4+ T cells from both wild-type and transgenic mice home to tumors. This definition of the T cell repertoires available is critical to the designing of immunological maneuvers able to elicit an effective immune reaction against HER-2-driven carcinogenesis.

PURPOSE For patients with early-stage breast cancer, predicting the risk of metastatic relapse is of crucial importance. Existing predictive models rely on agnostic survival analysis statistical tools (eg, Cox regression). Here we define and evaluate the predictive ability of a mechanistic model for time to distant metastatic relapse. METHODS The data we used for our model consisted of 642 patients with 21 clinicopathologic variables. A mechanistic model was developed on the basis of two intrinsic mechanisms of metastatic progression: growth (parameter α) and dissemination (parameter μ). Population statistical distributions of the parameters were inferred using mixed-effects modeling. A random survival forest analysis was used to select a minimal set of five covariates with the best predictive power. These were further considered to individually predict the model parameters by using a backward selection approach. Predictive performances were compared with classic Cox regression and machine learning algorithms. RESULTS The mechanistic model was able to accurately fit the data. Covariate analysis revealed statistically significant association of Ki67 expression with α ( P = .001) and EGFR expression with μ ( P = .009). The model achieved a c-index of 0.65 (95% CI, 0.60 to 0.71) in cross-validation and had predictive performance similar to that of random survival forest (95% CI, 0.66 to 0.69) and Cox regression (95% CI, 0.62 to 0.67) as well as machine learning classification algorithms. CONCLUSION By providing informative estimates of the invisible metastatic burden at the time of diagnosis and forward simulations of metastatic growth, the proposed model could be used as a personalized prediction tool for routine management of patients with breast cancer.

Ceramides are intramembrane diffusible mediators involved in transducing signals originated from a variety of cell surface receptors. Different adaptive and differentiative cellular responses, including apoptotic cell death, use ceramide-mediated pathways as an essential part of the program. Here, we show that human dendritic cells respond to CD40 ligand, as well as to tumor necrosis factor-α and IL-1β, with intracellular ceramide accumulation, as they are induced to differentiate. Dendritic cells down-modulate their capacity to take up soluble antigens in response to exogenously added or endogenously produced ceramides. This is followed by an impairment in presenting soluble antigens to specific T cell clones, while cell viability and the capacity to stimulate allogeneic responses or to present immunogenic peptides is fully preserved. Thus, ceramide-mediated pathways initiated by different cytokines can actively modulate professional antigen-presenting cell function and antigen-specific immune responses.

Introduction Type II collagen is a DR4/DR1 restricted target of self-reactive T cells that sustain rheumatoid arthritis. The aim of the present study was to analyze the T-cell receptor repertoire at the onset of and at different phases in rheumatoid arthritis.

In the present study, we use modified CDR3 beta-chain spectratyping (immunoscope) to dissect the effect of Mycobacterium tuberculosis (MT)-derived proteins on individual PLP139-151-specific cells in the SJL mouse strain. In this model, the immunoscope technique allows the characterization of a public TCR that involves rearrangement of Vbeta10 and Jbeta1.1 and a semi-private TCR characterized by rearrangement of Vbeta4 and Jbeta1.6. Both rearrangements are specific for PLP139-151 and sequences of the CDR3 region of the two beta-chains show a conserved motif for the public rearrangement and related but more variable sequences for the semi-private rearrangement. MT-derived proteins promote increase of IFN-gamma-secreting cells. However, we observe that the presence and amount of MT used during immunization have no effect on the frequency of usage, polarization and in vivo expansion of cells carrying the studied rearrangements. Rather, the strong Th1-promoting effect of adjuvant is possibly due to recruitment toward Th1 of a wider spectrum of TCR repertoires. Therefore, instead of having a comprehensive effect on the entire repertoire, MT modulates the immune response by affecting a subset of antigen-specific T cells whose polarization can be adapted to the environment. This step establishes the final balance between Th1 and Th2 and may be essential for the enhancement or protection of disease.

Dendritic cells (DCs) play a central role in the initiation and regulation of the immune response. The modalities by which DCs are committed to undergo apoptosis are poorly defined. Here it is shown that, unlike death receptor ligands, UVB radiation triggers apoptosis of human DCs very efficiently. UVB exposure is followed by the activation of caspases 8, 9, and 3, by the loss of mitochondrial transmembrane potential (⌬⌿m), and by cellular and nuclear fragmentation. Caspase inhibitors substantially prevented the occurrence of cellular and nuclear fragmentation but had no effect on UVB-induced ⌬⌿m dissipation. Importantly, mature DCs (MDCs) displayed relative resistance to UVB; UVB-induced caspase activation and apoptosis were substantially delayed compared to immature DCs (IDCs) . IntroductionMost tissues are equipped with interstitial dendritic cells (DCs) as an efficient alert system against foreign antigens for the exquisite ability of DCs in capturing macromolecules. 1,2 Tissue-associated DCs are commonly referred to as immature DCs (IDCs) because they lack some key surface accessory molecules and are, therefore, unable to trigger T-cell activation effectively. After antigen capture and processing, IDCs undergo extensive morphologic and biochemical changes and migrate to specialized lymphoid areas. Such mature DCs (MDCs) dismiss antigen capture functions, become competent antigen-presenting cells (APCs), and productively interact with T cells to initiate an antigen-specific immune response. 3 Two types of bone marrow-derived DCs colonize human skin from peripheral blood to serve as professional APCs of environmental antigens. These are Langerhans cells (LCs), which reside mostly in the basal and suprabasal layers of the epidermis, and dermal IDCs, which are confined within the dermis. [4][5][6] On antigen capture and maturation, mature LCs and MDCs relocate from the skin to draining lymph nodes in search of antigen-specific T cells.Ultraviolet B (280-320 nm) radiation is a major stress-inducing agent for most body surfaces. Exposure of the human skin to chronic and acute UVB irradiation is known to cause immunosuppression. 7 This results largely from local effects, including both massive depletion of LCs,8,9 likely due to apoptosis, 10,11 and functional impairment in the costimulatory abilities of the residual LCs. 12 UVB exposure may also suppress the T-cell stimulatory capacity of human DCs. 13 UV-induced intracellular mediators such as ceramide, moreover, may profoundly affect DC functions. 14 Little is known, however, about whether dermal human DCs trigger the apoptotic program under UVB exposure or how they activate a protective response to UVB-induced cellular stress.Here we show that high-dose UVB radiation may induce very efficient apoptosis of human DCs. This is associated with early mitochondrial changes and is mediated by multiple caspase activation, resulting in cytosolic and nuclear fragmentation. The upregulation of FLIP and bcl2, which occurs during DC maturation, may provide protection from ...

We examined the TCR repertoire used by naive SJL mice in their in vitro spontaneous response to proteolipid protein (PLP) 139–151 by Vβ-Jβ spectratyping and compared it to that used after immunization with the peptide. T cells from immunized mice use the public rearrangement Vβ10-Jβ1.1, but naive mice do not; in contrast, TCR CDR3-β rearrangements of Vβ18-Jβ1.2 and Vβ19-Jβ1.2 consistently are associated with the spontaneous response. T cells involved in spontaneous and induced responses can each recognize PLP139–151 presented in vivo, but its s.c. administration has different consequences for the two repertoires. Four days after immunization, T cells associated with spontaneous responsiveness appear in the draining lymph nodes but disappear by day 10 and never appear elsewhere. Simultaneously, Vβ10-Jβ1.1 T cells are likewise activated in the lymph nodes by day 4 and spread to the spleen by day 10. Eight- to 10-wk-old naive mice use a narrower repertoire of TCRs than do immunized age-matched mice. Induced Vβ10-Jβ1.1 T cells home to the CNS during experimental autoimmune encephalomyelitis, whereas we failed to detect Vβ18-Jβ1.2 and Vβ19-Jβ1.2 TCR rearrangements in the CNS. Thus, we observe that administration of PLP139–151 primes a T cell repertoire distinct from the one responsible for spontaneous responsiveness. This “immunized” repertoire substitutes for the naive one and becomes dominant at the time of disease onset.

To improve the current vaccine against tuberculosis, a recombinant strain of Mycobacterium bovis bacillus Calmette-Guérin (rBCG) expressing a Mycobacterium tuberculosis vaccine candidate antigen (MPT64) in strong association with the mycobacterial cell wall was developed. To deliver the candidate antigen on the surface, we fused the mpt64 gene to the sequence encoding the PE domain of the PE_PGRS33 protein of M. tuberculosis (to create strain H PE-⌬MPT64-BCG), which we have previously shown to transport proteins to the bacterial surface. In a series of protection experiments in the mouse model of tuberculosis, we showed that (i) immunization of mice with H PE-⌬MPT64-BCG provides levels of protection significantly higher than those afforded by the parental BCG strain, as assessed by bacterial colonization in lungs and spleens and by lung involvement