Background: Cyclosporine is an immunosuppressive agent used to treat immune-mediated and inflammatory diseases in dogs. We have developed a pharmacodynamic (PD) assay that measures interleukin-2 (IL-2) produced by activated T cells to measure the immunosuppressive effects of cyclosporine. Hypothesis/objectives: Our retrospective study extracted data from samples submitted to our laboratory to obtain descriptive statistics, to determine whether assay results predicted treatment effectiveness, and to determine whether cyclosporine formulation or breed affected PD responses. Animals: 1,110 samples were analyzed over 4 years. Methods: Extracted data was analyzed to determine whether there was a relationship between assay results and clinical control, and whether either formulation or breed affected results. Results: We found no relationship between assay results and control of signs, and found that breed did not affect results. At comparable doses, proprietary modified cyclosporine was more immunosuppressive than proprietary non-modified cyclosporine, and both proprietary and generic modified formulations had similar efficacy.

One of the major focuses for our lab involves examining the immunosuppressive properties of various agents and receptors in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). This dissertation encompasses an investigation of cannabidiol in the EAE model, the endocannabinoid CB1 receptor in the EAE model, staphylococcal superantigens (SAgs) as immunosuppressive agents, and various aspects of the EAE model. The first chapter covers the existing literature pertinent to these studies, the second and third chapters cover the material, methods, and results from the studies, and the fourth chapter is a discussion of how those results fit into the existing body of literature. A fifth chapter has also been included which covers two additional studies designed to develop alternative EAE models for our lab; however, both studies turned out differently than expected. One of the most interesting developments from this final chapter was the discovery of an age dependent difference in the memory T cell response of older mice, which allows for more robust disease to be induced when cells from 6 month old mice are used in the passive EAE (P-EAE) model as opposed to mice 10 weeks of age.

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Staphylococcus aureus causes chronic mastitis in bovines that is difficult to treat with current therapeutics. The goal of this research is to provide information about and improve innate immune responses to infection. Infection can result in host cell apoptosis or programmed cell death. Many pathogens can inhibit apoptosis; thereby acquiring a replicative niche, a reprieve from immune responses, and an escape from treatments. We hypothesize that S. aureus inhibits apoptosis in dendritic cells (DC). To investigate our hypothesis, DC were infected with live S. aureus (LSA), γ-irradiated S. aureus (ISA), or Streptococcus agalactiae (Strep ag.) for 2 hours. Stimulations of DC included ultraviolet light (UV) and lipoteichoic acid (LTA). Results indicate that γ-irradiated S. aureus can inhibit UV-induced apoptosis by upregulating LTA. This research provides information about S. aureus infections, but further research is needed to improve responses to this type of infection. One way to improve innate immune responses to infection is by supplementing bovines with OmniGen-AF®, a probiotic that restores neutrophil function during immunosuppression. To determine the mechanism by which OmniGen-AF® functions, wildtype, MyD88 KO, and TLR4 KO mice were fed either normal chow or supplemented with OmniGen-AF® for two weeks. Mice were immunosuppressed with dexamethasone and challenged with LTA. LTA overcame immunosuppression in a TLR4-depenent manner regardless of supplementation with OmniGen-AF®. Overall this research supplies knowledge about S. aureus inhibition of apoptosis in DC and S. aureus LTA activation of PMN regardless of immunosuppression or supplementation with OmniGen-AF®. / Master of Science

En transplantation, la morbidité attribuable aux évènements de sur-immunosuppression dépasse celle des rejets et aucun outil clinique ne diagnostique cette condition. Précédemment, nous avons découvert et validé longitudinalement l'association entre la production du facteur de nécrose tumorale des monocytes intermédiaires stimulés par des peptides du virus de l'Epstein-Barr et la sur-immunosuppression. L'hypothèse générale de mes recherches est qu'il est possible de développer un test diagnostiquant la sur-immunosuppression. Spécifiquement, j'ai étudié une cohorte transversale prospective unicentrique pour valider extrinsèquement les valeurs diagnostiques du test et étudier les sous-populations de lymphocytes B à des fins d'optimisation. Cette cohorte de 71 greffés rénaux expose des valeurs diagnostiques semblables à la cohorte exploratoire, soit des sensibilité et spécificité respectives de 83 et 68 % pour le seuil diagnostique prédéfini. Cette association s'est montrée indépendante des caractéristiques cliniques, de la fonction rénale et du régime immunosuppresseur. Ensuite, nous avons développé et validé une seconde étape diagnostique ayant une haute spécificité, basée sur un seuil de ≤ de 19 % de lymphocytes Bm5 et early Bm5 sanguins, pour confirmer le diagnostic de sur-immunosuppression. Au final, ce test en deux étapes classe les deux tiers des patients dans la catégorie sur-immunosupprimés ou contrôles, avec des valeurs prédictives positive et négative de 91 et 89 % respectivement. L'analyse multivariée ajustée montre que le risque de sur-immunosuppression chez les patients classés comme positifs est plus de huit fois supérieur à celui des contrôles. En conclusion, mon projet de maîtrise a permis l'optimisation et la validation d'un biomarqueur diagnostiquant la sur-immunosuppression en greffe rénale. Une cohorte de validation multicentrique est en cours. / In the transplantation field, the morbidity attributable to over-immunosuppression events exceeds that of rejections and no clinical tool diagnoses this condition. Previously, we discovered and validated longitudinally the association between the production of tumor necrosis factor of intermediate monocytes stimulated by peptides of the Epstein-Barr virus and over-immunosuppression.The general hypothesis of my research is that it is possible to develop a test diagnosing overimmunosuppression. Specifically, I studied a prospective unicentric cross-sectional cohort to extrinsically validate the diagnostic values of the test and to study the subpopulations of B lymphocytes for optimization purposes.This cohort of 71 renal transplant recipients exhibits diagnostic values similar to the exploratory cohort, which are a sensitivity and specificity of 83 and 68 % respectively for the predefined diagnostic threshold. This association has been shown to be independent of clinical features, renal function and immunosuppressive regimen. Then, we developed and validated a second diagnostic step with high specificity, based on a threshold of ≤ 19 % of Bm5 and early Bm5 blood lymphocytes, to confirm the diagnosis of overimmunosuppression. In the end, this two-step test classifies two thirds of patients in the overimmunosuppressed or controls category, with positive and negative predictive values of 91 and 89 %, respectively. The adjusted multivariate analysis shows that the risk of overimmunosuppression in patients classified as positive is more than eight times greater than that of controls.In conclusion, my master's project allowed the optimization and validation of a biomarker diagnosing over-immunosuppression in renal transplant patients. A multicenter validation cohort is underway.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第25166号 / 医博第5052号 / 新制||医||1071(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 藤田 恭之, 教授 上野 英樹, 教授 河本 宏 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Yes / T-lymphocytes (T-cell) therapy offers a treatment for cancers. Developing this technology in the future provides the opportunity to revolutionise treatment and to make cancer a chronic condition. T-cells in themselves are a type of lymphocytes (itself a type of white blood cell) that play a central role in cell mediated immunity. They can be distinguished from other lymphocytes, such as B-cells and natural killer cells (NK cells), by the presence of a T-cell receptor (TCR) on the cell surface. T-cells have the capacity to destroy diseased cells, but tumours present a considerable challenge that reduces their impact. As cancer cells are frequently ‘invisible’ to the immune system, and they create an environment that suppresses T-cell activity., genetic engineering of T-cells can be used therapeutically to overcome these challenges. T-cells can be taken from the blood of cancer patients and then modified to recognise and destroy cancer-specific antigens.