All ETDs from UAB

Advisory Committee Chair

Sasanka Ramanadham

Advisory Committee Members

John J Shacka

Joanne E Murphy-Ullrich

Selvarangan Ponnazhagan

Gene P Siegal

Document Type

Dissertation

Date of Award

2014

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Type 1 diabetes is a metabolic disease characterized by hyperglycemia, an absolute deficiency in insulin, and the autoimmune-mediated destruction of pancreatic islet ß-cells. Since exogenous insulin does not effectively duplicate native ß-cell function and the need persists for life, strategies to prevent ß-cell death or promote ß-cell survival are expected to provide beneficial outcomes. Based on reports that Akt1 promotes ß-cell survival and proliferation, administration of viral vectors modified to effectively deliver constitutively active (CA)-Akt1 to ß-cells would be expected to be beneficial for the ß-cell. In freshly isolated islets, CA-Akt1 promoted ex vivo islet cell survival and ß-cell survival. Furthermore, in mice treated with streptozotocin to induce ß-cell death, administration of Ad5RGDpK7.RIP-CA-Akt1 conferred improved glucose tolerance and ß-cell viability versus vectors with reporter gene alone or without increased infectivity motifs. Taken together, CA-Akt1 is effective in promoting ß-cell survival and proliferation and Ad5RGDpK7.RIP-CA-Akt1 could be used in combination with other ß-cell centered regimens providing increased beneficial outcomes. The result of failure in self-tolerance and autoreactive immune cell infiltration of the islet is destruction of ß-cells. An increasing body of evidence implicates Ca2+-independent phospholipase A2ß (iPLA2ß) induction during diabetes and its participation in ß-cell apoptosis; therefore, we assessed the effects of iPLA2ß inhibition on type 1 diabetes development. The iPLA2ß hydrolyzes glycerophospholipids generating a free fatty acid and a lysophospholipid, both capable of being further metabolized into bioactive inflammatory products. Inhibition of iPLA2ß with FKGK18, a fluoroketone-based inhibitor specific for iPLA2ß, administered to NOD mice conferred significantly reduced diabetes incidence and insulitis, improved glucose homeostasis, higher circulating insulin, and ß-cell preservation. FKGK18 administration also inhibited production of TNFα from CD4+ T-cells and antibodies from B-cells, suggesting modulation of immune cell responses by iPLA2ß-derived lipid products. Further, adoptive transfer of diabetes by CD4+ T-cells to immunodeficient and diabetes-resistant NOD.scid mice was mitigated by FKGK18 pretreatment and TNFα production from CD4+ T-cells was reduced by inhibitors of cyclooxygenase and 12-lipooxygenase, which metabolize arachidonic acid to generate bioactive inflammatory eicosanoids. Adoptive transfer of diabetes was not prevented, however, when NOD.scid mice were administered FKGK18-pretreated T-cells or FKGK18 administered concurrently with T-cell transfer. Taken together, these observations suggest that iPLA2ß-derived lipid signals modulate immune cell responses and suggest that early inhibition of iPLA2ß may be beneficial in ameliorating autoimmune-mediated destruction of ß-cells and subsequent prevention of T1D development.

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