No description available.

The immune system must be able to mount a response against pathogens and transformed cells while remaining tolerant to healthy host tissue. A key process for ensuring this self-tolerance is the negative selection of self-reactive thymocytes. Expression of Programmed Death-1 (PD-1), a co-inhibitory member of the CD28 family associated with dampened peripheral immune responses,was found to be upregulated in 20-40% of thymocytes undergoing negative selection in the HYcd4model of thymic development. Although analysis of gene and protein expression directly ex vivo indicates that PD-1- and PD-1+ thymocytes are equally apoptotic, PD-1+ thymocytes appear to be protected from apoptosis in an in vitro stimulation assay. Analysis of HYcd4PD-1-/- mice indicates that thymocytes receive a higher intensity signal in the absence of PD-1. Future work utilizing HYcd4PD-1-/- mice will increase our understanding of the role of PD-1 in thymic negative selection. / Immunology

No description available.

No description available.

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

Rationale: Eosinophils are “end cell” leucocytes, associated with allergy, asthma and helminthiasis. At sites of inflammation, eosinophils may modulate immune response through expression of the extra-hepatic tryptophan-catabolising enzyme, Indoleamine 2, 3-dioxygenase (IDO). Kynurenines, products of tryptophan cleavage, induce apoptosis of T-cells, including thymocytes. Eosinophils naturally home to the thymi in mammals. Thymus is a primary lymphoid organ, where T-cells develop and undergo selection. My hypothesis is that eosinophils homing to the thymi participate in T-cell development through their expression of IDO. Methods: Immunohistochemistry revealed eosinophils in thymic tissue. Immunocytochemistry and flow cytometry were used to locate IDO protein expression in the thymus particularly in thymic eosinophils. RT-PCR and real-time PCR determined the presence of IDO mRNA in the thymus. Results: thymic eosinophils express IDO and infiltrate compartments associated with negative selection. The highest IDO transcription correlated with the influx of eosinophils and prevalence of immature thymocytes. / Experimental Medicine

PNP is a crucial enzyme in purine metabolism, and its inherited defects result in severe T-lineage immune deficiency in humans. I hypothesized that PNP deficiency disrupts the development of late CD4-CD8- double negative (DN) thymocytes and induces mitochondrial-mediated apoptosis of CD4+CD8+ double positive (DP) thymocytes. By using PNP-deficient (PNP-/-) mice as well as an OP9-DL1 co-culture system simulating PNP-deficient conditions, I demonstrated that PNP deficiency interferes with the maturation of DN thymocytes at the transition from DN3 to DN4 stage. Although PNP deficiency does not affect the generation or proliferation of DP thymocytes, PNP-/- DP thymocytes were observed to undergo apoptosis at a higher rate. My results suggest that apoptosis is induced through a mitochondrial mediated pathway. Additionally, re-introduction of PNP into PNP-/- thymocytes protected the cells from the toxic effects of deoxyguanosine by preventing the formation of deoxyguanosine triphosphate, indicating that the toxic metabolite in PNP deficiency is deoxyguanosine.

PNP is a crucial enzyme in purine metabolism, and its inherited defects result in severe T-lineage immune deficiency in humans. I hypothesized that PNP deficiency disrupts the development of late CD4-CD8- double negative (DN) thymocytes and induces mitochondrial-mediated apoptosis of CD4+CD8+ double positive (DP) thymocytes. By using PNP-deficient (PNP-/-) mice as well as an OP9-DL1 co-culture system simulating PNP-deficient conditions, I demonstrated that PNP deficiency interferes with the maturation of DN thymocytes at the transition from DN3 to DN4 stage. Although PNP deficiency does not affect the generation or proliferation of DP thymocytes, PNP-/- DP thymocytes were observed to undergo apoptosis at a higher rate. My results suggest that apoptosis is induced through a mitochondrial mediated pathway. Additionally, re-introduction of PNP into PNP-/- thymocytes protected the cells from the toxic effects of deoxyguanosine by preventing the formation of deoxyguanosine triphosphate, indicating that the toxic metabolite in PNP deficiency is deoxyguanosine.

CD45 is a protein tyrosine phosphatase that is expressed on all nucleated hematopoietic cells. The major substrates of CD45 in thymocytes and T cells are the Src family kinases Lck and Fyn. The role of CD45 in thymocyte development and T cell activation via its regulation of Src family kinases in T cell receptor signaling has been studied extensively. However, the role of CD45 in processes that affect thymocyte development prior to the expression of the T cell receptor has not been explored. The overall hypothesis of this study was that CD45 is a regulator of spreading, migration, proliferation, and differentiation of early thymocytes during development in the thymus and the absence of CD45 would alter the outcome of thymocyte development. The first aim was to determine how CD45 regulates CD44-mediated signaling leading to cell spreading. The interaction between CD44 and Lck was first examined. CD44 associated with Lck in a zinc-dependent and a zinc-independent manner. Mutation analysis localized the zinc-dependent interaction to the membrane proximal region of CD44, but did not involve individual cysteine residues on CD44. CD44 and Lck co-localized in microclusters upon CD44-mediated cell spreading. CD45 co-localized with Lck and CD44 in microclusters and with F-actin in ring structures. The recruitment of CD45 to microclusters may be a mechanism of how CD45 negatively regulates CD44-mediated spreading. The second specific aim was to determine the role of CD45 in migration, proliferation, and progression and differentiation of early thymocytes. CD45 negatively regulated CXCL12-mediated migration, and positively regulated the proliferation and progression of CD117- DN1 thymocytes. Absence of CD45 led to an altered composition of thymic subsets. The CD45-/- thymus contained decreased numbers of ETPs and an aberrant CD117- DN1 population that lacked CD24, TCRbeta, and CCR7 expression. There were also increased thymic NK and gamma/delta T cells, but decreased NKT cells. In addition, a novel intermediate between DN1 and DN2 that required Notch for progression was identified. Overall, this study identified new roles for CD45 in early thymocytes and provided a better picture of how the development of T cells, a central component of the immune system, is regulated.

CD45 is a protein tyrosine phosphatase that is expressed on all nucleated hematopoietic cells. The major substrates of CD45 in thymocytes and T cells are the Src family kinases Lck and Fyn. The role of CD45 in thymocyte development and T cell activation via its regulation of Src family kinases in T cell receptor signaling has been studied extensively. However, the role of CD45 in processes that affect thymocyte development prior to the expression of the T cell receptor has not been explored. The overall hypothesis of this study was that CD45 is a regulator of spreading, migration, proliferation, and differentiation of early thymocytes during development in the thymus and the absence of CD45 would alter the outcome of thymocyte development. The first aim was to determine how CD45 regulates CD44-mediated signaling leading to cell spreading. The interaction between CD44 and Lck was first examined. CD44 associated with Lck in a zinc-dependent and a zinc-independent manner. Mutation analysis localized the zinc-dependent interaction to the membrane proximal region of CD44, but did not involve individual cysteine residues on CD44. CD44 and Lck co-localized in microclusters upon CD44-mediated cell spreading. CD45 co-localized with Lck and CD44 in microclusters and with F-actin in ring structures. The recruitment of CD45 to microclusters may be a mechanism of how CD45 negatively regulates CD44-mediated spreading. The second specific aim was to determine the role of CD45 in migration, proliferation, and progression and differentiation of early thymocytes. CD45 negatively regulated CXCL12-mediated migration, and positively regulated the proliferation and progression of CD117- DN1 thymocytes. Absence of CD45 led to an altered composition of thymic subsets. The CD45-/- thymus contained decreased numbers of ETPs and an aberrant CD117- DN1 population that lacked CD24, TCRbeta, and CCR7 expression. There were also increased thymic NK and gamma/delta T cells, but decreased NKT cells. In addition, a novel intermediate between DN1 and DN2 that required Notch for progression was identified. Overall, this study identified new roles for CD45 in early thymocytes and provided a better picture of how the development of T cells, a central component of the immune system, is regulated.