Modulation de la réponse lymphocytaire T par IL4I1 - RESUME : L’enzyme Interleukin-four Induced Gene 1 (IL4I1), qui dégrade la phénylalanine, est exprimée par des cellules présentatrices d’antigène (CPA) en réponse à des stimuli pro-inflammatoires. Elle affecte la prolifération et les fonctions lymphocytaires T et pourrait donc participer au rétrocontrôle des réponses immunitaires. Cependant, les mécanismes d’action d’IL4I1 restent encore mal connus. Mon projet de thèse a comporté deux axes. Dans un premier axe, j’ai participé à la caractérisation du rôle d’IL4I1 dans la différenciation des lymphocytes T CD4+ naïfs en lymphocytes T régulateurs et T auxiliaires effecteurs. Dans mon deuxième axe, qui représente la majeure partie de mon travail, j’ai analysé l’action d’IL4I1 sur les évènements précoces de l’activation T. J’ai observé qu’IL4I1 inhibe la phosphorylation de ZAP70 dès les premières minutes d’activation. Ce défaut se propage aux trois voies de signalisation principales : la voie calcique, la voie des MAP kinases et la voie NFκB. Elle retentit secondairement sur la capacité à former des synapses avec les CPA, ainsi que sur l’expression de différents marqueurs membranaires d’activation. L’activité enzymatique d'IL4I1 n’est pas responsable du défaut d’activation observé. L’étude des synapses CPA-T a montré une polarisation des vésicules de sécrétion contenant IL4I1 en direction de la cellule T ainsi qu’occasionnellement leur présence au contact du lymphocyte T. Des expériences complémentaires confirment la fixation d’IL4I1 sur les lymphocytes T. Ces données suggèrent un nouveau mécanisme d’action d’IL4I1 dépendant de sa liaison à un récepteur membranaire de la cellule T. - Mots clefs : Interleukin-4 induced gene 1 (IL4I1), enzyme immunosuppressive, signalisation du lymphocyte T / Modulation of the T cell response by IL4I1 - SUMMARY: The enzyme Interleukin-four Induced Gene 1 (IL41I), which degrades phenylalanine, is expressed by antigen presenting cells (APC) in response to pro-inflammatory stimuli. IL4I1 modifies the proliferation and function of T lymphocytes, and may participate in the negative feedback of the immune response. Its mechanism of action remains poorly understood.My thesis project included two tasks. In the first task, I participated in the characterisation of the role of IL4I1 in naive CD4+ T cell differentiation into regulatory and helper T lymphocytes. In the second task, and the main part of my thesis, I have studied the effect of IL4I1 on early T cell activation. I observed that ZAP70 phosphorylation was rapidly decreased after TCR stimulation. This alteration was transmitted to the three main downstream signalling pathways: calcium fluxes, the MAP kinase pathway, and the NFκB pathway. The enzymatic activity of IL4I1 was not responsible for the observed decreased activation. Analysis of the APC-T cell synapse showed the polarised secretion of IL4I1 toward the T cell. Labelling of IL4I1 was sometimes detected directly on T lymphocytes. Complementary experiments indicate that IL4I1 binds to T lymphocytes. These data suggest a new mechanism of action of IL4I1 dependent on its ability to bind a membrane receptor on T lymphocytes. - Key-words: Interleukin-4 induced gene 1 (IL4I1), Immunosuppressive Enzyme, T lymphocyte signalling.

This research is part of an effort to develop non-mammalian surrogate immunoessays with the earth worm Lumbricus terrestris to assess immunotoxic potential of xenobiotics to mammals. The objective was to determine if earthworm immunoessays, namely E- and S- rosette formation and phagocytosis, are sensitive to a known mammalian immunotoxin, the PCB Arclor 1254. Results are presented in terms of PCB exposure and tissue concentrations during uptake/depuration.

Call number: LD2668 .T4 1978 L38 / Master of Science

published_or_final_version / Surgery / Doctoral / Doctor of Philosophy

Patients that have undergone organ transplantation are life-long treated with immunosuppressant drugs and these have to be monitored regularly to get the desired effect of suppressing the immune system. To monitor the drug concentration normally a venous blood sample is collected at a clinic but the use of dried blood spots (DBS) as a matrix for drug monitoring for immunosuppressant drugs will make home sampling possible for this patient group. The aim of this study was to develop and validate a bioanalytical method for quantifying cyclosporine A and tacrolimus in dried blood spots. The method consist of punching out a 5 mm disc from a blood spot , followed by extracting the spot in a 96-well hydrophobic filter plate with 150 µL extraction solution containing internal standard (ascomycin and cyclosporine A d12) in a methanol water solution (80:20v/v%). The extract is then centrifuged through the filter plate down in a 96-deep well plate and injected on the LC-MS/MS, with an analysis time of 2.5min. The method will be validated in accordance with the guidelines set by the European Medicines Agency with additions specific to DBS. The method is not fully validated but will be in due time. The validated parameters show a robust and fast analysing method that has the prospects of being used for analysing DBS samples for patients and in the future can possibly be used by patients in home environment.

by Wong Dart-man. / Bibliography: leaves 107-114 / Thesis (M.Phil.)--Chinese University of Hong Kong, 1982

Castanospermine (CS) is an alkaloid isolated from the nut of the Australian chestnut tree, Castanospermum australae. It has been shown to have potent anti-inflammatory effects, as evidenced by its ability to inhibit the clinical manifestations of passively induced autoimmune encephalomyelitis and passive adjuvant arthritis in rats and the efferent phase of contact hypersensitivity in mice. The purpose of this study was to determine if CS has immunosuppressive, as well as anti-inflammatory, properties. Contact hypersensitivity in mice to picryl chloride was chosen as the in vitro model of cell mediated immune reactivity. Mice were sensitised with picryl chloride and treated with CS, at doses of 150 and 300 mg/kg/day, twice daily for seven days, beginning at the time of sensitisation. Passive transfer of spleen cells from treated animals transferred significantly less contact hypersensitivity (P<0.05) to naive mice than did cells from control mice. This suggests that CS inhibited the generation of picryl chloride reactive effector cells. This inhibition was not due to a depletion of the T cell phenotype responsible for contact hypersensitivity, CD4+, as FACS analysis showed no alteration in CD4+/CD8+ ratio in CS treated mice. In vitro studies, using antigen-specific cell lines, showed that CS inhibits antigen specific T-cell proliferation in a dose dependent fashion. Studies on the kinetics of this inhibition revealed that CS inhibits an early step, before 24 hours of culture have elapsed, in the T-cell proliferative response. Experiments were designed to examine if this early event was antigen processing by the accessory cells in the culture, or an early event in T-cell replication itself. CS not only failed to inhibit the antigen processing step, but when processing was carried out in the presence of CS the subsequent T-cell proliferation was enhanced. The results also indicated that CS, when added with preprocessed antigen, was inhibiting T-cell proliferation in a dose dependent manner. Subsequent studies examining the role of several key molecules in T-cell proliferation showed that CS did not effect the expression of the receptors for IL-2 or transferrin, nor did it alter the expression of the adhesion molecules LFA-1 or ICAM-1. The precise molecular mechanism by which CS inhibits contact hypersensitivity in vivo and T-cell proliferation in vitro still remains to be determined.

C57B1/6 mice bearing Lewis lung carcinoma CLLC) are suppressed in their ability to generate cytotoxic T lymphocytes against the LLC tumor associated antigens. Since LLC have previously been shown to secrete the immunosuppressive factor PGE2, indomethacin, a prostaglandine synthetase inhibitor, was administered to LLC-bearing mice in their drinking water to prevent the immunosuppression typical of tumor bearers. The indomethacin treated tumor-bearers displayed an increase in their cytotoxic responses to the LLC tumor associated antigens. Thus the immunosuppression found in mice bearing LLC could be due to prostaglandin synthesis. Production of PGE2 by tumor cells may be a mechanism by which the tumors escape immune mediated destruction.Ball State UniversityMuncie, IN 47306

Thesis (Ph. D.)--University of Hong Kong, 2009. / Includes bibliographical references (leaves 188-245). Also available online.

Thesis (Ph. D.)--Oregon State University, 2010. / Printout. Includes bibliographical references (leaves 132-156). Also available on the World Wide Web.