Hypothesis. The central hypothesis of this proposal is that CD36, ABCD2, and mitoNEET mediate the protective effects of stearic acid on gestational diabetes and on the acquired diabetes of the progeny. Rationale. The goal of this proposal is to test the novel hypothesis that stearic acid ameliorates gestational diabetes mellitus (GDM) and subsequent diabetes in the progeny of GDM mothers through a triad of CD36, ABCD2, and mitoNEET proteins. We will use the CSTBLKS/J-Lep/*'* mouse that develops GDM around day 16 of gestation. We will examine the mechanisms of stearic acid on CD36, ABCD2, and mitoNEET expression and function in both GDM and on long-term metabolic and cardiovascular outcomes in the progeny of the db/+ dams. Finally, we will use individual genetic knockout models for each protein to determine the extent to which each protein mediates the action of stearic acid in both GDM and diabetes of the progeny. The uniqueness and potential clinical relevance of this approach is the mechanistic analysis of a possible means for treatment of both GDM and pursuant diabetes mellitus acquired by the offspring. We propose in vivo studies that can be further dissected by in vitro studies with isolated tissues and cells, which will enable us to thoroughly explore the protective mechanism of action of stearic acid. 1. To determine the impact of stearic acid on the development and extent of GDM. Db/+ dams will be used to examine whether stearic acid affects the development of GDM by measuring oral glucose tolerance, vascular responsiveness, and adipocyte function. The expression and subcellular localization of CD36, ABCD2, and mitoNEET in adipose will be quantified and functionality of the proteins assessed through measurements of fatty acid biochemistry (p-oxidation, fatty acid composition, and angiotensin II secretion). Similarly, we will also examine the effect of stearic acid treatment fed exclusively to the db/+ dams on the extent of diabetes in the offspring. Finally, we will thoroughly examine the mechanism of action of stearic acid on fatty acid biochemistry in insulin resistant 3T3-L1 adipocytes.
This aim i s important because we will mechanistically evaluate a potential palliative intervention for both GDM and the accompanying diabetes of the offspring. 2. To determine the duration which stearic acid improves the diabetic phenotype in db/db mice. Db/db progeny will be treated with stearic acid and the duration and extent of the protective effect investigated. The effect of withdrawal of stearic acid on the lasting benefit of the intervention will also be examined. We will measure blood pressure, vascular responsiveness, weight and glucose. We will examine the mechanisms of stearic acid action by determining the expression and subcellular location of CD36, ABCD2, and mitoNEET in adipose and assessing function of each protein through measurements of adipose fatty acid biochemistry.
This aim i s important because it will provide data on the cardiovascular benefit and sustainability of stearic acid intervention. 3. To determine whether the beneficial effects of stearic acid are mediated by CD36, ABCD2, and/or mitoNEET. We will cross mice null for CD36, ABCD2, or mitoNEET with db/+ mice and assess the efficacious effects of stearic acid in ameliorating GDM and the ensuing diabetic phenotype of the double crossed db/db progeny lacking each individual gene. We will measure OGTT, vascular responsiveness, and adipose fatty acid biochemistry in db/+ dams. Furthermore, we will assess the effect of individual gene knockout on the ability of stearic acid to affect the development of diabetes in the db/db mice. This will be accomplished through measurements of weight, glucose, insulin, blood pressure, and vascular responsiveness. Mechanistically, the effects of gene ablation will be dissected through comprehensive analyses of isolated primary adipose biochemistry.
This aim i s critical because each double crossed mouse line will provide the definitive mechanistic data regarding the role of each protein in mediating the beneficial effects of stearic acid. ? ? ?
