2013 Annual Report
1a.Objectives (from AD-416):
Objective: Evaluate leukocyte patterns, gene expression profiles, and inflammatory mediators in adipose tissue under influence of dietary manipulation that leads to obesity.
1b.Approach (from AD-416):
The research approach to accomplish this objective employs an established animal model of diet-induced obesity, the C57BL/6J mouse strain, fed diets rich in milk fat or corn oil, compared with low-fat control diets matched for all nutrients except the level of milk fat or corn oil. At specified times of feeding, animals will be analyzed for metabolic changes induced by the diets (e.g., insulin sensitivity, RER), systemic inflammatory changes, and local inflammatory changes in intra-abdominal and subcutaneous adipose tissue depots. The inflammatory changes analyzed include gene (qPCR and/or expression arrays) and protein (ELISA and/or flow cytometry) expression of cytokines, chemokines and adhesion molecules, and leukocyte subsets phenotypically characterized (flow cytometry); and mechanistic studies will be carried out using targeted deletions of proteins potentially key to inflammatory cascades (e.g., CD11c knockouts, gamma delta T cell knockouts, TLR4 knockouts, etc., backcrossed to the C67BL/6J strain). The experimental approach analyzing anti-inflammatory factors (e.g, candidate genes such as IL-1ra or expression array analysis) influenced by changes in diet will utilize this animal model with focus on the early times following initiation of the high fat feeding and on times following reversal of the high-fat diet to a low-fat diet, a time we have found involves rapid resolution of inflammation that precede reductions in fat mass.
Feeding mice a high fat diet for several months causes inflammation in many tissues of the body, particularly fatty tissues (adipose tissues) in the abdomen, skeletal muscles, and liver. White blood cells migrate into these tissues and release proteins that change the normal functions of these tissues, thereby contributing to the development of type II diabetes. There are many different types of white blood cells, and our work has been to define the specific types that migrate into the tissues, and define the role these cells play in causing type II diabetes. In this study we analyzed one type of white blood cell called alpha beta T cells, and analyzed their influence in obesity-induced inflammation of skeletal muscle, a major organ that takes up glucose from the blood. In this study we wanted to evaluate the effect of alpha beta T cells on the body's response to insulin, and their role in causing inflammation of adipose tissue and skeletal muscle as the mice become obese. We found that both adipose tissue and skeletal muscle of obese mice have higher than normal counts of alpha beta T cells. We found that mice lacking alpha beta T cells (i.e., mice genetically engineered to be deficient in these cells) were protected against obesity-induced hyperglycemia (abnormally elevated glucose levels) and resistance to insulin (both measures of type II diabetes). We also found that these mice did not develop the inflammatory response in body tissues that is typical of normal obese mice. To confirm that the alpha beta T cells were responsible for the inflammation and type II diabetes in mice, we transfused alpha beta T cells into the mice that were lacking these cells, and found that they developed inflammation and type II diabetes very much like the normal obese mice. Several studies published in the recent scientific literature have shown that specific types of fats called fatty acids have the ability to modify inflammation, and the fatty acids may influence the white blood cells that are part of the excess tissue inflammation that occurs in obese individuals. In humans, a single high-fat meal increases inflammatory proteins and white blood cells in the blood. We found that short-term high fat feeding to mice increases two types of fatty acids called palmitic and oleic acid within adipose tissues. The oleic acid increase is highest in the fatty tissues next to the intestines. To determine if these fatty acids can influence the function of one type of white blood cell in adipose tissue, we isolated these cells (called macrophages) and studied them in tissue culture. If we exposed the macrophages to oleic acid they developed characteristics of macrophages that are anti-inflammatory (in other words, may prevent or reduce inflammation). If we exposed them to palmitic acid, they developed characteristics of proinflammatory macrophages (that can cause or promote inflammation). This indicated that these two fatty acids had opposite influences on macrophages. We then determined if these fatty acids had these effects on macrophages in the body when supplemented in the diet. Three-day supplementation of a normal high protein mouse diet with oleic acid induced an increase in anti-inflammatory characteristics of macrophages in the adipose tissues that surround the intestines, but supplementation with palmitic acid had no effect. Then for three days we fed mice a diet rich in milk fat (also high in both oleic acid and palmitic acid), and found increases in anti-inflammatory characteristics of macrophages in the fatty tissue next to the intestines. Thus, we have shown that short-term feeding of a high fat diet changes macrophages in the fat tissues next to the intestines, and that dietary oleic acid appears to be responsible for increases in the anti-inflammatory functions of these macrophages.