Specific loss of adipocyte CD248 improves metabolic health via reduced white adipose tissue hypoxia, fibrosis and inflammation

Background: A positive energy balance promotes white adipose tissue (WAT) expansion which is characterized by activation of a repertoire of events including hypoxia, inflammation and extracellular matrix remodelling. The transmembrane glycoprotein CD248 has been implicated in all these processes in different malignant and inflammatory diseases but its potential impact in WAT and metabolic disease has not been explored.Methods: The role of CD248 in adipocyte function and glucose metabolism was evaluated by omits analyses in human WAT, gene knockdowns in human in vitro differentiated adipocytes and by adipocyte-specific and inducible Cd248 gene knockout studies in mice.Findings: CD248 is upregulated in white but not brown adipose tissue of obese and insulin-resistant individuals. Gene ontology analyses showed that CD248 expression associated positively with pro-inflammatory/pro-fibrotic pathways. By combining data from several human cohorts with gene knockdown experiments in human adipocytes, our results indicate that CD248 acts as a microenvironmental sensor which mediates part of the adipose tissue response to hypoxia and is specifically perturbed in white adipocytes in the obese state. Adipocytespecific and inducible Cd248 knockouts in mice, both before and after diet-induced obesity and insulin resistance/glucose intolerance, resulted in increased microvascular density as well as attenuated hypoxia, inflammation and fibrosis without affecting fat cell volume. This was accompanied by significant improvements in insulin sensitivity and glucose tolerance.Interpretation: CD248 exerts detrimental effects on WAT phenotype and systemic glucose homeostasis which may be reversed by suppression of adipocyte CD248. Therefore, CD248 may constitute a target to treat obesity-associated co-morbidities. (C) 2019 The Authors. Published by Elsevier B.V.</p

Nutritional regulation of genome-wide association obesity genes in a tissue-dependent manner

Background: Genome-wide association studies (GWAS) have recently identified several new genetic variants associated with obesity. The majority of the variants are within introns or between genes, suggesting they affect gene expression, although it is not clear which of the nearby genes they affect. Understanding the regulation of these genes will be key to determining the role of these variants in the development of obesity and will provide support for a role of these genes in the development of obesity. Methods We examined the expression of 19 GWAS obesity genes in the brain and specifically the hypothalamus, adipose tissue and liver of mice by real-time quantitative PCR. To determine whether these genes are nutritionally regulated, as may be expected for genes affecting obesity, we compared tissues from fasting and non-fasting animals and tissues from mice consuming a high fat high sucrose diet in comparison to standard rodent chow. Results We found complex, tissue-dependent patterns of nutritional regulation of most of these genes. For example, Bat2 expression was increased ~10-fold in the brain of fed mice but was lower or unchanged in the hypothalamus and adipose tissue. Kctd15 expression was upregulated in the hypothalamus, brain and adipose tissue of fed mice and downregulated by high fat feeding in liver, adipose tissue and the hypothalamus but not the remainder of the brain. Sh2b1 expression in the brain and Faim2 expression in adipose tissue were specifically increased >20-fold in fed mice. Tmem18 expression in adipose tissue but not the brain was reduced 80% by high fat feeding. Few changes in the expression of these genes were observed in liver. Conclusions These data show nutritional regulation of nearly all these GWAS obesity genes, particularly in the brain and adipose tissue, and provide support for their role in the development of obesity. The complex patterns of nutritional and tissue-dependent regulation also highlight the difficulty that may be encountered in determining how the GWAS genetic variants affect gene expression and consequent obesity risk in humans where access to tissues is constrained.Cellular and Physiological Sciences, Department ofMedicine, Faculty ofReviewedFacult

Nutritional regulation of genome-wide association obesity genes in a tissue-dependent manner

Abstract Background Genome-wide association studies (GWAS) have recently identified several new genetic variants associated with obesity. The majority of the variants are within introns or between genes, suggesting they affect gene expression, although it is not clear which of the nearby genes they affect. Understanding the regulation of these genes will be key to determining the role of these variants in the development of obesity and will provide support for a role of these genes in the development of obesity. Methods We examined the expression of 19 GWAS obesity genes in the brain and specifically the hypothalamus, adipose tissue and liver of mice by real-time quantitative PCR. To determine whether these genes are nutritionally regulated, as may be expected for genes affecting obesity, we compared tissues from fasting and non-fasting animals and tissues from mice consuming a high fat high sucrose diet in comparison to standard rodent chow. Results We found complex, tissue-dependent patterns of nutritional regulation of most of these genes. For example, Bat2 expression was increased ~10-fold in the brain of fed mice but was lower or unchanged in the hypothalamus and adipose tissue. Kctd15 expression was upregulated in the hypothalamus, brain and adipose tissue of fed mice and downregulated by high fat feeding in liver, adipose tissue and the hypothalamus but not the remainder of the brain. Sh2b1 expression in the brain and Faim2 expression in adipose tissue were specifically increased >20-fold in fed mice. Tmem18 expression in adipose tissue but not the brain was reduced 80% by high fat feeding. Few changes in the expression of these genes were observed in liver. Conclusions These data show nutritional regulation of nearly all these GWAS obesity genes, particularly in the brain and adipose tissue, and provide support for their role in the development of obesity. The complex patterns of nutritional and tissue-dependent regulation also highlight the difficulty that may be encountered in determining how the GWAS genetic variants affect gene expression and consequent obesity risk in humans where access to tissues is constrained.</p

Altered pancreatic growth and insulin secretion in WSB/EiJ mice.

These data suggest that insulin secretion in WSB mice is blunted specifically in vivo, either due to a reduced insulin requirement and/or due to factors that are absent or destroyed in vitro. These studies also highlight the role of post-natal growth in determining adult β-cell mass. Mice are important animal models for the study of metabolic physiology and the genetics of complex traits. Wild-derived inbred mouse strains, such as WSB/EiJ (WSB), are unrelated to the commonly studied mouse strains and are valuable tools to identify novel genes that modify disease risk. We have previously shown that in contrast to C57BL/6J (B6) mice, WSB mice fed a high fat diet do not develop hyperinsulinemia or insulin resistance, and had nearly undetectable insulin secretion in response to an intraperitoneal glucose challenge. As hyperinsulinemia may drive obesity and insulin resistance, we examined whether defects in β-cell mass or function could contribute to the low insulin levels in WSB mice. In young WSB mice, β-cell mass was similar to B6 mice. However, we found that adult WSB mice had reduced β-cell mass due to reduced pancreatic weights. Pancreatic sizes were similar between the strains when normalized to body weight, suggesting their pancreatic size is appropriate to their body size in adults, but overall post-natal pancreatic growth was reduced in WSB mice compared to B6 mice. Islet architecture was normal in WSB mice. WSB mice had markedly increased insulin secretion from isolated islets in vitro. These data suggest that insulin secretion in WSB mice is blunted specifically in vivo, either due to a reduced insulin requirement and/or due to factors that are absent or destroyed in vitro. These studies suggest that WSB mice may provide novel insight into mechanisms regulating insulin secretion and also highlight the role of post-natal growth in determining adult β-cell mass

Diabetes genes identified by genome-wide association studies are regulated in mice by nutritional factors in metabolically relevant tissues and by glucose concentrations in islets

Background: Genome-wide association studies (GWAS) have recently identified many new genetic variants associated with the development of type 2 diabetes. Many of these variants are in introns of known genes or between known genes, suggesting they affect the expression of these genes. The regulation of gene expression is often tissue and context dependent, for example occurring in response to dietary changes, hormone levels, or many other factors. Thus, to understand how these new genetic variants associated with diabetes risk may act, it is necessary to understand the regulation of their cognate genes. Results We identified fourteen type 2 diabetes-associated genes discovered by the first waves of GWAS for which there was little prior evidence of their potential role in diabetes (Adam30, Adamts9, Camk1d, Cdc123, Cdkal1, Cdkn2a, Cdkn2b, Ext2, Hhex, Ide, Jazf1, Lgr5, Thada and Tspan8). We examined their expression in metabolically relevant tissues including liver, adipose tissue, brain, and hypothalamus obtained from mice under fasted, non-fasted and high fat diet-fed conditions. In addition, we examined their expression in pancreatic islets from these mice cultured in low and high glucose. We found that the expression of Jazf1 was reduced by high fat feeding in liver, with similar tendencies in adipose tissue and the hypothalamus. Adamts9 expression was decreased in the hypothalamus of high fat fed mice. In contrast, the expression of Camk1d, Ext2, Jazf1 and Lgr5 were increased in the brain of non-fasted animals compared to fasted mice. Most notably, the expression levels of most of the genes were decreased in islets cultured in high glucose. Conclusions These data provide insight into the metabolic regulation of these new type 2 diabetes genes that will be important for determining how the GWAS variants affect gene expression and ultimately the development of type 2 diabetes.Cellular and Physiological Sciences, Department ofMedicine, Faculty ofSurgery, Department ofReviewedFacult

Whole pancreas insulin content.

<p>These studies were performed in young mice between 6 and 7 weeks of age. A. Total pancreatic insulin content. B. Pancreas weight. C. Pancreatic insulin content per gram of pancreas. (n = 5–6 WSB per diet, n = 10 B6 per diet) *p<0.05, ***p<0.001 vs. B6 mice consuming the same diet. ++p<0.01 vs. chow-fed mice of the same strain.</p

Islet vascularization at 4 weeks of age.

<p>A. Islet vascularization of chow-fed C57BL/6J (top) and WSB/EiJ (bottom) mice. Vasculature (CD31) is stained in green, while insulin is stained red to demarcate islet areas. B. Percent of islet area stained for CD31. (n = 5 per strain).</p

Islet structure and size, β-cell mass, and α-cell mass at twenty weeks of age.

<p>A. Representative islets from B6 (left) and WSB (right) mice fed the chow diet. Insulin is stained red, glucagon green, and cell nuclei are marked by DAPI staining (blue). No alteration to islet structures was found in islets from the high fat-fed mice. B. The average cross-sectional areas of all islets found in each mouse were determined then averaged for each strain (average number of islets per mouse examined: B6 chow- 167, B6 HFD 187, WSB chow 129, WSB HFD 171). C. Distribution of islet sizes in B6 (black; chow-fed solid, high fat diet (HFD)-fed striped) and WSB mice (white; chow-fed solid, high fat diet (HFD)-fed striped). In the upper group, although there was substantial overlap in the sizes, B6 mice tended to have larger maximal islet sizes (both diets). D. Percent of pancreatic tissue area stained for insulin. E. Pancreatic weight of mice used in these experiments. F. Pancreatic weights normalized to total body weight in chow and high fat-fed WSB and B6 mice. G. β-cell mass, calculated as the % staining area multiplied by the pancreatic weight. H. Percent of pancreatic area stained for glucagon. I. α-cell mass calculated as for β-cell mass. (n = 5 mice per strain per diet). *p<0.05, **p<0.01, ***p<0.001 vs. B6 mice consuming the same diet. +p<0.05, +++p<0.001 vs. chow-fed of the same strain.</p

Islet structure and size, β-cell mass, and α-cell mass in young WSB mice.

<p>Studies were performed on mice at 4 weeks of age. A. Representative islets from B6 (left) and WSB (right) mice. Insulin (red), glucagon (green) and DNA (blue) are stained. B. Mean islet sizes (average number of islets per mouse examined: B6 chow- 166, B6 HFD 130, WSB chow 108, WSB HFD 115). C. Islet size distribution of B6 (black; chow solid, high fat diet (HFD) striped) and WSB (white; chow solid, high fat diet white striped) mice. The range of islet sizes observed was similar in all groups. D. Percent of pancreatic area stained for insulin. E. Pancreas weight for stained tissues. F. β-cell mass. G. Percent of pancreatic area stained for glucagon. H. α-cell mass. (n = 5 mice per strain per diet). **p<0.01 vs. B6 mice consuming the same diet.</p