Inflammation following tissue damage promotes lymphocyte recruitment, tissue remodeling, and wound healing while maintaining self tolerance. Endogenous signals associated with tissue damage and cell ...death have been proposed to initiate and instruct immune responses following injury. In this study, we have examined the effects of elevated levels of a candidate endogenous danger signal, heat shock cognate protein 70 (hsc70), on stimulation of inflammation and autoimmunity following cell damage. We find that damage to pancreatic beta cells expressing additional cytosolic hsc70 leads to an increased incidence of diabetes in a transgenic mouse model. Steady-state levels of activated APC and T cell populations in the draining lymph node were enhanced, which further increased following streptozotocin-induced beta cell death. In addition, proinflammatory serum cytokines, and lymphocyte recruitment were increased in hsc70 transgenic mice. Islet Ag-specific T cells underwent a greater extent of proliferation in the lymph nodes of mice expressing hsc70 following beta cell damage, suggesting elevated Ag presentation following release of Ag in the presence of hsc70. These findings suggest that an elevated content of hsc70 in cells undergoing necrotic or apoptotic cell death can increase the extent of sterile inflammation and increase the susceptibility to autoimmunity.
T‐cell death is a fundamental process that is intricately regulated at multiple phases during T‐cell differentiation, tolerance induction and the decline of the immune response. Caspase 3 is a ...crucial molecule regulating both mitochondrial and death receptor apoptotic pathways and therefore we were interested in examining the role of caspase 3 in T cells. Using P14 and H‐Y CD8⁺ TCR‐transgenic models, our analysis has shown that caspase 3 is not required for thymic negative selection. In addition, caspase 3 does not play a prominent role in the contraction phase following acute viral infection, nor clonal deletion of CD8⁺ T cells under tolerizing conditions. Surprisingly, our studies demonstrate that caspase 3 was not required for the induction of CD8⁺ T‐cell anergy in vivo, contrary to published reports using CD4⁺ T cells. Therefore, these results demonstrate that caspase 3 is not essential in CD8⁺ T cells for multiple forms of thymic or peripheral tolerance, nor the contraction phase after an acute anti‐viral response.
Abstract
Protein kinase B (PKBα/Akt1) a PI3K-dependent serine-threonine kinase, promotes T cell viability in response to many stimuli and regulates homeostasis and autoimmune disease in vivo. To ...dissect the mechanisms by which PKB inhibits apoptosis, we have examined the pathways downstream of PKB that promote survival after cytokine withdrawal vs Fas-mediated death. Our studies show that PKB-mediated survival after cytokine withdrawal is independent of protein synthesis and the induction of NF-κB. In contrast, PKB requires de novo gene transcription by NF-κB to block apoptosis triggered by the Fas death receptor. Using gene-deficient and transgenic mouse models, we establish that NF-κB1, and not c-Rel, is the critical signaling molecule downstream of the PI3K-PTEN-PKB signaling axis that regulates lymphocyte homeostasis.
Elucidating the signaling events that promote T-cell tolerance versus activation provides important insights for manipulating immunity in vivo. Previous studies have suggested that the absence of ...PKCθ results in the induction of anergy and that the balance between the induction of the transcription factors NFAT, AP1 and NF-κB plays a key role in determining whether T-cell anergy or activation is induced. Here, we examine whether Bcl-10 and specific family members of NF-κB act downstream of PKCθ to alter CD8⁺ T-cell activation and/or anergy. We showed that T cells from mice deficient in c-Rel but not NF-κB1 (p50) have increased susceptibility to the induction of anergy, similar to T cells from PKCθ-deficient mice. Surprisingly T cells from Bcl-10-deficient mice showed a strikingly different phenotype to the PKCθ-deficient T cells, with a severe block in TCR-mediated activation. Furthermore, we have also shown that survival signals downstream of NF-κB, are uncoupled from signals that mediate T-cell anergy. These results suggest that c-Rel plays a critical role downstream of PKCθ in controlling CD8⁺ T-cell anergy induction.
Abstract
Regulatory innate lymphoid cells (ILCregs) are an emerging family of immunoregulatory cells that suppress innate and adaptive immunity in the context of diseases, including cancer. However, ...there is currently a lack of lineage-defining surface markers or transcription factors (TFs) that distinguishes ILCregs from other subsets of ILCs. Moreover, the functional role of ILCregs within the tumour microenvironment (TME) has not been well established. In this study, we uncovered a novel population of CD103-expressing CD56 +ILCregs within the TME that correlated with poor recurrent-free survival (RFS) in patients with epithelial ovarian carcinoma (EOC). CD103 +ILCregs expressed distinct surface markers and TFs that distinguishes them from canonical CD56 +natural killer (NK) cells. Moreover, intratumoural CD103 +ILCregs expressed markers (i.e., CD39, CD69, CD101, TIGIT) and transcriptomic profiles that are shared with Tregs, suggesting that they may have immunoregulatory properties. We found that ascites-derived TGF-β drives NK cell conversion into CD103 +ILCs that can suppress autologous CD4 +T cells in vitro. Furthermore, intratumoural CD103 +ILCregs suppressed autologous CD8 +T cells in vitro. Finally, we found that the abundance of CD103 +ILCregs in the TME were associated with reduced levels of both activated CD4 +T cells and CD8 +T cells. Overall, we have identified CD103 +ILCregs as distinct immunoregulatory cells that suppresses T cells within the TME of patients with EOC. Further untangling of the complex networks of immune regulation, including ILCregs, may give rise to novel therapeutic targets to enhance anti-tumour immunity and improve clinical response rates to immunotherapies.
This work was supported by a CIHR foundation award (CIHR FDN #143220), a TRANSCAN CIHR grant (CIHR – TRN #184713), a TRI grant from the Ontario Institute for Cancer Research (TRI OICR GCS #108796), and the Terry Fox Research Institute (TFRI-iTNT #1060). This project was also generously funded in memory of Mr. JIM JoTak under the Canadian Cancer Society (CCSRI #706152).
Background and Aims: Accurate endoscopic detection of premalignant lesions and early cancers in the colon is essential for cure, since prognosis is closely related to lesion size and stage. Although ...it has great clinical potential, autofluorescence endoscopy has limited tumor-to-normal tissue image contrast for detecting small preneoplastic lesions. We have developed a molecularly specific, near-infrared fluorescent monoclonal antibody (CC49) bioconjugate which targets tumor-associated glycoprotein 72 (TAG72), as a contrast agent to improve fluorescence-based endoscopy of colon cancer. Methods: The fluorescent anti-TAG72 conjugate was evaluated in vitro and in vivo in athymic nude mice bearing human colon adenocarcinoma (LS174T) subcutaneous tumors. Autofluorescence, a fluorescent but irrelevant antibody and the free fluorescent dye served as controls. Fluorescent agents were injected intravenously, and in vivo whole body fluorescence imaging was performed at various time points to determine pharmacokinetics, followed by ex vivo tissue analysis by confocal fluorescence microscopy and histology. Results: Fluorescence microscopy and histology confirmed specific LS174T cell membrane targeting of labeled CC49 in vitro and ex vivo. In vivo fluorescence imaging demonstrated significant tumor-to-normal tissue contrast enhancement with labeled-CC49 at three hours post injection, with maximum contrast after 48 h. Accumulation of tumor fluorescence demonstrated that modification of CC49 antibodies did not alter their specific tumor-localizing properties, and was antibody-dependent since controls did not produce detectable tumor fluorescence. Conclusions: These results show proof-of-principle that our near-infrared fluorescent-antibody probe targeting a tumor-associated mucin detects colonic tumors at the molecular level in real time, and offer a basis for future improvement of image contrast during clinical fluorescence endoscopy.
PKCθ Signals Activation versus Tolerance In Vivo Berg-Brown, Nancy N.; Gronski, Matthew A.; Jones, Russell G. ...
The Journal of experimental medicine,
03/2004, Letnik:
199, Številka:
6
Journal Article
Recenzirano
Odprti dostop
Understanding the pathways that signal T cell tolerance versus activation is key to regulating immunity. Previous studies have linked CD28 and protein kinase C-θ (PKCθ) as a potential signaling ...pathway that influences T cell activation. Therefore, we have compared the responses of T cells deficient for CD28 and PKCθ in vivo and in vitro. Here, we demonstrate that the absence of PKCθ leads to the induction of T cell anergy, with a phenotype that is comparable to the absence of CD28. Further experiments examined whether PKCθ triggered other CD28-dependent responses. Our data show that CD4 T cell–B cell cooperation is dependent on CD28 but not PKCθ, whereas CD28 costimulatory signals that augment proliferation can be uncoupled from signals that regulate anergy. Therefore, PKCθ relays a defined subset of CD28 signals during T cell activation and is critical for the induction of activation versus tolerance in vivo.
Differentiation of CD8 single-positive (SP) T-cells is predicated by the ability of lymphocyte progenitors to integrate multiple signaling cues provided by the thymic microenvironment. In the thymus ...and the OP9-DL1 system for T-cell development, Notch signals are required for progenitors to commit to the T-cell lineage, and necessary for their progression to the CD4
+
CD8
+
(DP) stage of T-cell development. However, it remains unclear whether Notch is a prerequisite for the differentiation of DP cells to CD8 SP stage of development. Here, we demonstrate that Notch receptor-ligand interactions allow for efficient differentiation and selection of conventional CD8 T-cells from bone marrow (BM)-derived hematopoietic stem cells (HSCs). However, BM-HSCs isolated from
Itk
−/−
Rlk
−/−
mice gave rise to T-cells with decreased IFNγ production, while gained the ability to produce IL-17. We further reveal that positive- and negative-selection
in vitro
are constrained by peptide-MHC (pMHC) class I expressed on the OP9 cells. Finally, using an MHC class I-restricted T-cell receptor (TCR) transgenic model, we show that the commitment of DP precursors to the CD8 T-cell lineage is dependent on Notch signaling. Our findings further establish the requirement for Notch receptor-ligand interactions throughout T-cell differentiation, including the final step of CD8 SP selection.
Self-reactive T cells in the body are controlled by mechanisms of peripheral tolerance that limit their activation and induction of immune pathology. Our understanding of these mechanisms has been ...advanced by the use of tissue-specific promoters to express neo-self-antigens. Here, we present findings using the RIP-gp (rat insulin promoter-glycoprotein) transgenic mouse, which expresses the lymphocytic choriomeningitis virus glycoprotein (LCMV-gp) specifically in the pancreatic beta islet cells. T cells responsive to this antigen remain ignorant of the LCMV-gp expressed by the islets, and breaking tolerance is dependent upon the maturation status of antigen-presenting cells, the avidity of the T-cell receptor ligation, and the level of major histocompatibility complex expression in the pancreas. Furthermore, decreased activity of Casitas B-lineage lymphoma b, a negative regulator of T-cell receptor signaling, can allow recognition and destruction of the pancreatic islets. This review discusses the roles of these factors in the context of anti-tissue responses, both in the setting of autoimmunity and in anti-tumor immunity.
The development of nonviral, peptide-based constructs able to elicit protective in vivo CTL responses represents a major challenge in the design of future vaccines. We report the design of branched ...peptide delivery vehicles, termed loligomers, that facilitate the import, processing, and presentation of CTL epitopes onto nascent MHC class I molecules. These complexes are then effectively displayed on the surface of APCs. The intracellular delivery of CTL epitopes by loligomers prolonged the expression of Ag-MHC class I complexes on the surface of APCs in comparison with free CTL epitope alone. Furthermore, the injection of CTL epitope-containing loligomers into mice led to the generation of in vivo CTL responses and the induction of autoimmune disease in an animal model. Synthetic epitope-carrying, peptide-based delivery vehicles may represent useful components to be included in the formulation of future vaccines.
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