Understanding how the antigenically foreign fetus escapes attack by maternal lymphoid cells has long been a goal of reproductive immunologists. The present thesis work analyzes the expression and regulation of maternal immunity to self Ia antigen, as well as to embryonic antigens common to fetal and tumor cells but absent from normal adult somatic tissues. Initial studies revealed that the in vitro proliferative capacity of thymus and bone marrow-derived T cells is enhanced during syngeneic murine pregnancy. Pregnancy was also found to promote the manifestation of autoreactive splenic T cells capable of mediating syngeneic graft-versus-host reactions. In addition, primigravid isopregnant mice harbour splenic T cells which are specifically sensitized to syngeneic fetal antigens. Both alpha-fetoprotein and a population of B-like pregnancy-associated splenic suppressor cells were shown to be potent regulators of autoimmune lymphoproliferative activity. Monoclonal antibodies (clone PSBA-1) were generated which, in the presence and absence of complement, abrogate the in vitro inhibitory activity of such suppressor cells. Moreover, a significant proportion of gravid females which were treated with PSBA-1 antibodies resorbed their embryos. Thus, a B-like suppressor cell appears to play a crucial immunoregulatory role during murine pregnancy.

A chamber technique was developed allowing recovery of cytokines and T-cells from human DTH reactions. These cells were cloned, and were enriched, relative to the blood, for T-cells reactive to the antigen eliciting the DTH. Antigen-specific CD4+ clones, had two phenotypes; an IL-2/IFN-g producing group not providing help for Ig synthesis, and a group providing help but not producing IL-2 or IFN-g. Cytotoxicity and TNF production were found in both groups. TNF and IFN-g were present in the chambers over DTH reactions. Anergic patients demonstrated deficient mononuclear cell delivery to DTH sites which was restored to normal by co-injection of MLC supernatants with antigen. T-cell clones from such sites were similar to those from normal DTH sites. Additional experiments established that MLC supernatants contained 2 distinct factors, one restoring DTH in anergic patients, and another, which protected anergic rats from lethal peritonitis.

Trypanosoma musculi is a protozoan parasite which produces a characteristic, self-limiting murine infection of approximately three weeks duration; the infection comprises a growth phase, a plateau phase and an elimination phase. Following clearance of parasitaemia, a mouse is cured and immune to reinfection. The present studies examine the immune mechanisms which operate during the elimination phase. / Passive transfer of plasma from an immune mouse to an infected recipient brings about rapid and complete clearance of parasitaemia in C57BL/6 mice. This curative activity is labile to heat treatment for 30 minutes at 56$ sp circ$C. A protein A- derived immunoglobulin fraction of immune plasma (IP) shares these properties. Further purification shows that the curative activity resides primarily in the IgG2a subclass, and that this antibody is intrinsically heat-labile. Complement component C3 (but not the lytic C5-C9 sequence) is necessary for antibody-mediated cure of infection. Cellular elements (macrophages) are also essential for elimination of parasitaemia to occur. The ultimate T. musculi effector mechanism thus requires the interaction of both humoral and cellular components.

Graft versus host disease (GVHD) is a major complication of bone marrow transplantation and may also occur naturally through the transplacental passage of maternal lymphocytes to the fetus. GVHD is associated with a broad spectrum of immunological disorders as well as distinct pathological changes. Yet, the exact nature of the effector cells involved remains to be elucidated. Studies on natural killer cell activity in bone marrow transplant patients have led to the speculation that natural killer cells may be major effectors of GVHD-induced immunopathogenesis. / To determine the role played by natural killer cells in GVHD, a murine model of neonatal GVHD was established. Natural killer cell function was evaluated in lymphoid target organs at various times post induction and was found to be significantly augmented by seven days post-injection. The phenotype of the effector cell was determined by antibody depletion studies to be asialo GM1 positive, partially Thy-1 positive and glass wool non-adherent. Immunostaining indicated an increased frequency of asialo GM-1 positive cells in the spleens of GVHD-induced mice. These effectors were further determined to be predominantly of donor origin. / The in vivo role of these asialo GM-1 positive cells in the actual pathogenesis of GVHD was also investigated. Asialo GM1 positive cells were depleted from either the donor inoculum prior to injection or from the recipient in vivo post-injection. Only the depletion of asialo GM1 positive cells from the host in vivo was effective in reducing GVHD-associated morbidity. / The possible role of placental suppressive factors in the protection of the fetus was also examined using this model. Extracts derived from placental tissue were found to be sufficiently potent to suppress in vivo the development of acute GVHD. / Our findings indicate that donor-derived asialo GM-1 positive killer cells play a major role in the development of acute GVHD. In addition, factors associated with the placenta may provide a sufficient functional barrier to prevent the development of GVHD in the fetus as the result of the transplacental passage of maternal lymphocytes.

Direct lympho-stromal interactions constitutes an intrinsic part of T-cell differentiation. However the precise role of cellular complexes is far from clear. The objective of the present work was to gain an insight into this question. An in vitro model was previously developed in the laboratory, which detects the adherence of thymocytes to a thymic stromal cell line, E-5. Using this model, we describe here a novel lympho-stromal interaction involving L3T4$ sp+$/Ly-2$ sp+$ "cortical type" thymocytes and medullary epithelial cell. These adherent thymocytes appear at day 16 of murine gestation and their number vary during ontogenesis and leukemogenesis. After contact with epithelial cells, thymocytes become totally refractory to form further complexes, are activated and undergo some phenotypic modifications. All these results strongly suggest that this novel lympho-stromal interaction takes place in the transition of "cortical type" thymocytes into "medullary type" thymocytes.

Since the discovery of Dendritic cells (DCs) in 1975, their role as potent antigen presenting cells has been a field of intense interest and research. Candidate DC vaccines take advantage of the role of DCs in one of two ways. The first technique relies on the in vivo isolation of DCs which are then subsequently employed directly as adjuvants. In the second method, hematopoeitic growth factors are added to stimulate the expansion and proliferation of these cells in vivo. Due to the dearth of DCs, research has focused on the addition of growth factors such as FLT3-L (fms-like tyrosine kinase 3-ligand) and GM-CSF (granulocyte/macrophage colony-stimulating factor) which have proven invaluable for the expansion of this cell type. The goal of our study has been to deliver these growth factors intramuscularly using a plasmid vector. This work has shown that DCs are efficiently generated in vivo following a single intramuscular co-injection of cDNA encoding GM-CSF and FLT3-L.

Recurrent pregnancy loss is associated with anti-phospholipid antibodies (aPL), in particular in women with anti-phospholipid syndrome. aPL are a large family of autoantibodies, directed against phospholipids and phospholipid-binding proteins (e.g., prothrombin and beta2-glycoprotein I). Although the association between aPL and recurrent pregnancy loss is well documented, little is known about which aPL subset is responsible. To address this, we characterized a panel of six murine monoclonal aPL, which were subsequently administered to pregnant mice via passive transfer. While both anti-beta2GPI antibodies evaluated caused significant increase in fetal resorption, the effects of anti-PT antibodies varied from no effect to increased pregnancy loss. Similarly, neither lupus anticoagulant activity nor murine cross-reactivity predetermined aPL pathogenicity. We conclude that there is no obvious common reactivity between the aPL capable of increasing pregnancy loss, suggesting that a combination of reactivities, rather than one aPL specificity in particular, may lead to aPL-mediated pregnancy loss.

Many liver diseases such as Alcoholic Liver Disease and Hepatitis C are associated with the production of autoantibodies. One of these autoantibodies is Rheumatoid Factor (RF), which binds to IgG and can aid in host defense. Since little is known about the role of RF in liver disease, we characterized the production of this autoantibody in three murine models of liver damage using alcohol, anti-Fas antibodies and carbon tetrachloride. RF was induced in response to chronic alcohol consumption as well as intraperitoneal injection of anti-Fas antibodies. RF was not produced in response to treatment with carbon tetrachloride for four weeks. Concurrent treatment with an E. coli glycolipoprotein which induces RF protected against liver damage as measured by a decrease in liver enzymes. Importantly, RF induced by this glycolipoprotein did not contribute to damage in the liver, suggesting that in the context of liver damage RF is not necessarily pathological.

Superantigens trigger the polyclonal activation of human T cells. We exploited this property to gain insight into the mechanisms governing CD4 T cell expansion and to study the functional heterogeneity of naive and memory CD4 T cell subsets. We show that the amount of TCR ligand affects the evolution of a T cell response in two ways: by shaping the diversity of the T cell population recruited in the proliferative pool and by affecting the progression of these precursors into cell cycle. These two processes characterize a hierarchy of recruitment of cells that strongly correlates with the efficiency of TCR engagement but not with the relative precursor frequency in the non-immune repertoire. Remarkably, once established, the distribution of T cell clones within a selected repertoire is maintained by the characteristic of T cells to expand at a rate that is independent of quantitative differences in ligand exposure. Moreover, at optimal ligand concentrations that lead to the simultaneous expansion of all responsive T cell clones, we observed a marked clone-specific heterogeneity in the capacity to secrete cytokines. The functional heterogeneity of different CD4+ T cell subpopulations was also studied using a superantigen model. Based on the expression of CD45RA and CCR7, four distinct subsets of CD4+ T lymphocytes can be identified: naive (CD45RA+ CCR7+), central memory (TCM, CD45RA- CCR7+), effector memory (TEM, CD45RA- CCR7-) and a previously uncharacterized subset (CD45RA+ CCR7-). In CD8 T cells, this subset has been shown to comprise "terminally differentiated" effector memory cells. The four subsets show different functional sensitivities for the superantigens, the TEM population being the most sensitive and the naive cells being the least sensitive, as measured by the upregulation of activation markers. We show that the CD4+ CD45RA+CCR7- subpopulation, which is rarely detectable in healthy individuals, is enriched in T cells having str

Different immunotherapeutic strategies have been attempted against cancer, ranging from the use of cytokines like IL-2, to the use of dendritic cells, like Provenge, or the combinations of gene-modified, cytokine producing cellular vaccines, like GVAX. Immunotherapy has progressed slowly because either the induced immune response was not effective or because the heterogeneity of cancer can limit the effectiveness of inducing an immune response against a single antigen. Previous work in our lab has found that fusing GMCSF with γ-chain family cytokines produced novel immune modulatory effects and that cells treated with these fusion cytokines could be effective in treating diseases in mice. We generated a fusion of GMCSF and IL-21 (GIFT-21) with the aim of stimulating distinct, but complementary, elements of the innate and adaptive immune system. GIFT-21's aberrant interactions with its cognate receptors on macrophages induced an unanticipated pro-inflammatory response, resulting in B16 melanoma cancer rejection in C57Bl/6 mice and a significant survival advantage in NOD SCID mice. We further explored this phenomenon by treating mice with dendritic cells (DC) derived by treating monocytes with GIFT-21. B16 and D2F2/neu breast cancer growth was inhibited only in mice treated with antigen naïve GIFT-21 DCs. This effect was lost in CD8-/- and CCR2-/- mice and when mice were treated with β2 microglobulin deficient GIFT-21 DCs, and we confirmed that GIFT-21 DCs migrated to and sampled from the tumors to present tumor antigens to CCL2 recruited CD8+ T cells via MHCI.We also generated a separate fusion of GMCSF and IFN-γ (GIFY) with the aim of inducing a pro-inflammatory response by DCs and improving upon their antigen presenting properties. We confirmed that GIFY had superior cytostatic properties to IFN-γ and that it could induce DCs to present antigens to CD8+ T cells akin to the combination of GMCSF and IFN-γ. However GIFY was ineffective at preventing tumor growth when injected as a purified recombinant in mice bearing B16 melanoma tumors and when used as an adjuvant for DCs in the context of a tumor vaccine against EG-7 lymphoma.We conclude that GIFT-21 and its associated cellular products may serve as a novel therapeutic platform for the treatment of cancer and that GIFY may be appropriate in other settings. / Différentes stratégies immunothérapeutiques ont été considérées contre le cancer, allant de l'utilisation de cytokines comme l'IL-2, aux cellules dendritiques, tel que Provenge, ou à la combinaison de cellules modifiées génétiquement pour exprimer des cytokines produisant des vaccins cellulaires, tel que GVAX. Ces méthodes immunothérapeutiques ont progressé lentement puisque d'une part, la réponse immunitaire n'était pas efficace, et d'une autre part, l'hétérogénéité du cancer peut limiter l'efficacité à produire une réponse immunitaire contre un antigène particulier. Des recherches antérieures au sein de notre laboratoire ont révélé que la fusion de GMCSF avec la famille des cytokines de la chaîne gamma du récepteur de l'IL-2 ont produit des effets immunomodulateurs sans précédent. De plus, les cellules traitées avec ces cytokines fusionnées pourraient être efficaces dans le traitement de maladies chez les souris.Nous avons généré la fusion de GMCSF et IL-21 (GIFT-21) dans le but de stimuler des éléments distincts, mais complémentaires, du système immunitaire inné et adaptif. Les interactions aberrantes de GIFT-21 avec ses récepteurs des macrophages ont produit une réponse pro-inflammatoire inattendue, résultant en un rejet de la lignée cellulaire B16 des souris C57B1/6 et une survie significative des souris NOD SCID. Nous avons exploré davantage ce phénomène en injectant des souris avec des cellules dendritiques (DC) dérivées en traitant des monocytes avec GIFT-21. La croissance des cancers B16 et D2F2/neu a seulement été inhibée dans les souris injectées avec des cellules dendritiques traitées avec GIFT-21 sans pré-incubation avec un antigène. Par contre, cet effet n'a pas été observé dans les souris CD8-/- et CCR2-/-, ni lorsque nous avons injecté des souris avec des cellules dendritiques GIFT-21 déficientes en microglobuline β2. Nous avons confirmé que les cellules dendritiques GIFT-21 ont migré vers les tumeurs et les ont échantillonées afin de présenter des antigènes à travers MHCI à des cellules T CD8+ recrutées par CCL2. Nous avons également généré une fusion de GMCSF et IFN-γ (GIFY) dans le but d'induire une réponse pro-inflammatoire chez les cellules dendritiques et d'ainsi améliorer leur habileté a présenter des antigènes. Nous avons confirmé que GIFY possède des propriétés cytostatiques supérieures à IFN-γ et qu'il pourrait induire les cellules dendritiques à présenter des antigènes à travers des cellules T CD8+ similaires à la combinaison de GMCSF et IFN-γ. Par contre, GIFY s'est montré inefficace à prévenir la croissance des tumeurs lorsqu'injecté en tant que recombinante purifiée dans des souris avec des tumeurs B16 et lorsqu'utilisé en tant qu'adjuvant sous forme de vaccin pour des cellules dendritiques contre un lymphome EG-7. Nous pouvons conclure que GIFT-21 et ses produits cellulaires pourront servir de plate-forme thérapeutique sans précédent pour le traitement du cancer et que GIFY pourrait être approprié dans d'autres contextes.