Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 144-172).

Thesis (Ph. D.)--Case Western Reserve University, 2004. / [School of Medicine] Department of Pathology. Includes bibliographical references. Available online via OhioLINK's ETD Center.

MicroRNAs (miRNAs) are 20-22 nucleotide non-coding RNAs that can play important roles in developmental transitions by post-transcriptional regulation of mRNA translation and stability. We profiled miRNA expression in mouse thymocytes, mature T cells, and activated T cells, and identified distinctive patterns of miRNA expression during development, maturation, and activation of T cells. The miR-128 and miR-181 miRNA families are expressed at significantly higher levels in thymocytes. Examining the expression levels of these microRNAs in more detail, we observed that the expression pattern of these microRNA families distinguishes cells committed to lymphoid lineages from cells committed to myeloid lineages during normal mouse hematopoiesis. Extending this work to human malignancies, we determine that high miR-128 expression distinguishes lymphoid precursor derived malignancies from myeloid precursor derived malignancies. Little information is available regarding miRNA expression early after CD8 T cell activation. We demonstrate dynamic miRNA expression during early CD8 T cell activation, including the repression of miR-150, miR-181a, miR-26, miR-29 and miR-30, and the induction of miR-155, miR-31, miR-146, and the miR-17-92 cluster. We show that miR-31 is induced by calcium/Calcineurin signaling during acute CD8 T cell activation, and demonstrate elevated miR-31 expression in regulatory and memory T cell populations. We identify miR-31 targets in primary CD8 T cells and propose a model where miR-31 induction primes CD8 T cells for activation by promoting T cell survival, activation, and proliferation. Activation induced miRNA expression patterns are also found in some human malignancies. Chronic lymphocytic leukemia is typically thought to be a disease of resting lymphocytes. However, we demonstrate an activated B cell miRNA expression signature in CLL. Similarities in miRNA expression between activated B cells and CLL cells include high expression of miR-34a, miR-155, and miR-342-3p and low expression of miR-103, miR-181a and miR-181b. Additionally, we show that decreased levels of miR-29c and miR-223 in CLL are negative prognostic markers associated with shorter time to first therapy. These data indicate an activated B cell status for CLL cells and suggest that the expression level of individual miRNAs may predict clinical course in CLL.

T -2 toxin is an immunosuppressive mycotoxin that has been conjoined with several symptoms and diseases as early as the turn of the century, but whose mechanisms of action are still being investigated. Accordingly, this study was an attempt to determine the cytotoxic effects of T -2 toxin on normal human lymphocytes in vitro, with particular emphasis on mitochondrial viability, cellular and nuclear morphology as well as the localisation of the subcellular sites of toxin interaction. The cytotoxicity of T -2 toxin was assessed with the use of a methylthiazol tetrazolium (MTT) assay. This assay targeted the succinate dehydrogenase activity of the lymphocytic mitochondria, over a range of concentrations of T-2 toxin at various incubation times. The morphology of treated lymphocytes was analysed with the use of transmission electron microscopy and the localisation of the toxin was accomplished via immunocytochemistry. DNA fragmentation studies formed an integral part of the analyses. The cytotoxicity assay indicated that not only was cell viability inversely proportional to both the dose and exposure time, but that the eftects of the different doses were only evident at prolonged incubation times (12-24 hours). The electron microscopy studies showed that T-2 toxin (1,56 ug/ml) induced apoptosis (cell suicide) in normal human lymphocytes. This was determined by the observation of chromatin condensation and nuclear disintegration within the toxin treated lymphocytes. Apoptosis seemed to occur independently of mitochondrial damage at 6 hours of exposure to T-2 toxin. The presence of polyribosomes within the treated lymphocytes indicated that protein synthesis was not inhibited. Anti-T-2 toxin conjugated gold label was present in all areas of damage, particularly within the nuclei of the T-2 toxin treated lymphocytes. The DNA fragmentation results showed that T-2 toxin induced fragmentation in lymphocytes, the extent of which was directly proportional to the exposure time. It appears that the early signs of T-2 toxin induced apoptosis in normal human lymphocytes can be determined by damage to the nucleus. / Thesis (M.Med.)-University of Natal, Durban, 1998.

Recruitment of effector T cells to sites of infection is essential for an effective adaptive immune response. The inflammatory chemokine CCL5/RANTES activates its cognate receptor, CCR5, to initiate cellular functions including chemotaxis. This thesis describes the signaling events invoked by CCL5 and its ability to regulate the energy status of activated T cells. CCL5 treatment in ex vivo activated human T cells induced the activation of AMPK and downstream substrates ACC1, PFKFB2 and GSK-3. Evidence is provided that CCL5 treatment is able to induce glucose uptake in an mTOR-dependent manner. Using 2-deoxy-D-glucose, an inhibitor of glucose uptake, and Compound C, an inhibitor of AMPK, evidence is provided that demonstrate that CCL5-mediated chemotaxis is dependent on metabolic events, since these inhibitors perturb chemotaxis in a dose-dependent manner. Collectively, these studies suggest that CCL5 may also influence the metabolic status of activated T cells by simultaneously activating the AMPK and mTOR pathways.

Recruitment of effector T cells to sites of infection is essential for an effective adaptive immune response. The inflammatory chemokine CCL5/RANTES activates its cognate receptor, CCR5, to initiate cellular functions including chemotaxis. This thesis describes the signaling events invoked by CCL5 and its ability to regulate the energy status of activated T cells. CCL5 treatment in ex vivo activated human T cells induced the activation of AMPK and downstream substrates ACC1, PFKFB2 and GSK-3. Evidence is provided that CCL5 treatment is able to induce glucose uptake in an mTOR-dependent manner. Using 2-deoxy-D-glucose, an inhibitor of glucose uptake, and Compound C, an inhibitor of AMPK, evidence is provided that demonstrate that CCL5-mediated chemotaxis is dependent on metabolic events, since these inhibitors perturb chemotaxis in a dose-dependent manner. Collectively, these studies suggest that CCL5 may also influence the metabolic status of activated T cells by simultaneously activating the AMPK and mTOR pathways.

Diss. (sammanfattning) Göteborg : Univ. , 2007. / Härtill 3 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2002. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 6 uppsatser.