OBJECTIVE-We have previously reported a highly diabetogenic CD8 T-cell clone, G9C8, in the nonobese diabetic (NOD) mouse, specific to low-avidity insulin peptide B15-23, and cells responsive to this antigen are among the earliest islet infiltrates. We aimed to study the selection, activation, and development of the diabetogenic capacity of these insulin-reactive T-cells.
RESEARCH DESIGN AND METHODS-We generated a T-cell receptor (TCR) transgenic mouse expressing the cloned TCR V␣18/V6 receptor of the G9C8 insulin-reactive CD8 T-cell clone. The mice were crossed to TCRC␣ Ϫ/Ϫ mice so that the majority of the T-cells expressed the clonotypic TCR, and the phenotype and function of the cells was investigated.RESULTS-There was good selection of CD8 T-cells with a predominance of CD8 single-positive thymocytes, in spite of thymic insulin expression. Peripheral lymph node T-cells had a naïve phenotype (CD44 lo , CD62L hi ) and proliferated to insulin B15-23 peptide and to insulin. These cells produced interferon-␥ and tumor necrosis factor-␣ in response to insulin peptide and were cytotoxic to insulin peptide-coated targets. In vivo, the TCR transgenic mice developed insulitis but not spontaneous diabetes. However, the mice developed diabetes on immunization, and the activated transgenic T-cells were able to transfer diabetes to immunodeficient NOD.scid mice.CONCLUSIONS-Autoimmune CD8 T-cells responding to a low-affinity insulin B-chain peptide escape from thymic negative selection and require activation in vivo to cause diabetes.
Objective
Apoptotic destruction of insulin-producing pancreatic β-cells is involved in the aetiology of both type 1 and type 2 diabetes. AMP-activated protein kinase (AMPK) is a sensor of cellular energy charge whose sustained activation has recently been implicated in pancreatic β-cell apoptosis and in islet cell death post-transplantation. Here, we examine the importance of β-cell AMPK in cytokine-induced apoptosis and in the cytotoxic action of CD8+ T cells.
Research Design and Methods
Clonal MIN6 β-cells or CD1 mouse pancreatic islets were infected with recombinant adenoviruses encoding enhanced green fluorescent protein (eGFP/Null), constitutively-active AMPK (AMPK CA), or dominant-negative AMPK (AMPK DN) and exposed or not to tumour necrosis factor-α (TNF-α) interleukin-1β (IL-1β) and interferon-γ (IFN-γ). Apoptosis was detected by monitoring the cleavage of caspase-3 and DNA fragmentation. The cytotoxic effect of CD8+ purified T cells was examined against pancreatic islets from NOD mice infected with either Null or the AMPK DN-expressing adenoviruses.
Results
Exposure to cytokines, or expression of AMPK CA, induced apoptosis in clonal MIN6 β-cells and CD1 mouse pancreatic islets. By contrast, over-expression of AMPK DN protected against the proapoptotic effect of these agents, in part by preventing decreases in cellular ATP, and lowered the cytotoxic effect of CD8+ T cells towards NOD mouse islets.
Conclusions
Inhibition of AMPK activity enhances islet survival in the face of assault by either cytokines or T cells. AMPK may therefore represent an interesting therapeutic target to suppress immune mediated β-cell destruction, and may increase the efficacy of islet allografts in Type 1 diabetes.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.