Increased L-CPT-1 activity and altered gene expression in pancreatic islets of malnourished adult rats: a possible relationship between elevated free fatty acid levels and impaired insulin secretion

Intrauterine growth restriction is associated with chronically elevated levels of serum fatty acids and reduced glucose-stimulated insulin secretion. Lipid metabolism in pancreatic beta cells is critical for the regulation of insulin secretion, and the chronic exposure to fatty acids results in higher palmitate oxidation rates and an altered insulin response to glucose. Using a rat model of isocaloric protein restriction, we examined whether pre- and postnatal protein malnutrition influences the properties of pancreatic islet carnitine palmitoyltransferase-1 (liver isoform, L-CPT-1), a rate-limiting enzyme that regulates fatty acid oxidation in mitochondria. The activity of L-CPT-1 in pancreatic islets increased in the low protein (LP), although the L-CPT-1 mRNA levels were unaffected by malnutrition. The susceptibility of enzyme to inhibition by malonyl-CoA was unaltered and the content of malonyl-CoA was reduced in LP cells. Because the mitochondrial oxidation of fatty acids is related to the altered expression of a number of genes encoding proteins involved in insulin secretion, the levels of expression of insulin and GLUT-2 mRNA were assessed. A reduced expression of both genes was observed in malnourished rats. These results provide further evidence that increased L-CPT-1 activity and changes in gene expression in pancreatic islets may be involved in the reduced insulin secretion seen in malnourished rats.     

Adaptive responses by mouse fetus to a maternal HLE diet by downregulating SREBP1: a microarray- and bio-analytic-based study

Maternal diet has long been recognized as a significant factor affecting offspring development and health, but the target genes affected by a maternal high-lipid diet are currently unknown. In this study, the gene expression profile of neonatal mouse liver was analyzed using gene chips to identify genes with significant up- or downregulated expression levels due to maternal high-fat diet during gestation. Real-time PCR and Western blotting were used to measure key genes selected using microarray. Serum lipid, glucose, and insulin levels in adult offspring from dams fed with chow or a high-lipid diet were measured using commercial kits. Results indicate that the expression of genes involved in cholesterol and fatty acid synthesis were significantly inhibited, while the expression of genes involved in glycolysis were significantly decreased by maternal high-lipid diet during gestation. SREBP1 might be the key gene regulating genes involved in fatty acid, glucose, and cholesterol metabolism in response to a maternal high-fat diet.     

Gene Expression Profile Analysis of Type 2 Diabetic Mouse Liver

Liver plays a key role in glucose metabolism and homeostasis, and impaired hepatic glucose metabolism contributes to the development of type 2 diabetes. However, the precise gene expression profile of diabetic liver and its association with diabetes and related diseases are yet to be further elucidated. In this study, we detected the gene expression profile by high-throughput sequencing in 9-week-old normal and type 2 diabetic db/db mouse liver. Totally 12132 genes were detected, and 2627 genes were significantly changed in diabetic mouse liver. Biological process analysis showed that the upregulated genes in diabetic mouse liver were mainly enriched in metabolic processes. Surprisingly, the downregulated genes in diabetic mouse liver were mainly enriched in immune-related processes, although all the altered genes were still mainly enriched in metabolic processes. Similarly, KEGG pathway analysis showed that metabolic pathways were the major pathways altered in diabetic mouse liver, and downregulated genes were enriched in immune and cancer pathways. Analysis of the key enzyme genes in fatty acid and glucose metabolism showed that some key enzyme genes were significantly increased and none of the detected key enzyme genes were decreased. In addition, FunDo analysis showed that liver cancer and hepatitis were most likely to be associated with diabetes. Taken together, this study provides the digital gene expression profile of diabetic mouse liver, and demonstrates the main diabetes-associated hepatic biological processes, pathways, key enzyme genes in fatty acid and glucose metabolism and potential hepatic diseases.     

Liver and muscle metabolic changes induced by dietary energy content and genetic selection in rainbow trout (Oncorhynchus mykiss)

We combined genetic selection and dietary treatment to produce a model to study metabolic pathways involved in genetic and nutritional control of fat deposition in fish muscle. Two experimental lines of rainbow trout, selected for a lean (L) or fat (F) muscle, were fed with diets containing either 10 or 23% lipids from the first feeding, up to 6 mo. At the end of the feeding trial, trout were distinguished by very different muscle fat content (from 4.2 to 10% wet weight), and line x diet interactions were observed for parameters related to fat storage. We analyzed the activity and gene expression of key enzymes involved in lipid metabolism (fatty acid synthase, hydroxyacyl-CoA dehydrogenase, carnitine palmitoyltransferase 1 isoforms, and peroxisome proliferator-activated receptor alpha) and glycolysis (hexokinase 1 and pyruvate kinase) as well as energy production (isocitrate dehydrogenase, citrate synthase, and cytochrome oxidase) in the liver and the white muscle of rainbow trout. The lipid-rich diet repressed the activity of the lipogenic enzymes and stimulated enzymes involved in fatty acid oxidation and glycolysis in liver but had little effect on muscle enzymes assessed in this study. Regarding the selection effect, enzyme activity and expression suggest that compared with the L line, the F line presented reduced hepatic fatty acid oxidation as well as reduced mitochondrial oxidative capacities and enhanced glucose utilization in both liver and muscle. Very few line x diet interactions were found, suggesting that the two factors (i.e., dietary energy content and selection) used in this study to modify muscle lipid content exerted some additive but mostly independent effects on these metabolic actors.     

Gene expression in human NAFLD

Despite the high prevalence of nonalcoholic fatty liver disease (NAFLD), little is known of its pathogenesis based on study of human liver samples. By the use of Affymetrix GeneChips (17,601 genes), we investigated gene expression in the human liver of subjects with extreme steatosis due to NAFLD without histological signs of inflammation (liver fat 66.0 +/- 6.8%) and in subjects with low liver fat content (6.4 +/- 2.7%). The data were analyzed by using sequence-based reannotation of Affymetrix probes and a robust model-based normalization method. We identified genes involved in hepatic glucose and lipid metabolism, insulin signaling, inflammation, coagulation, and cell adhesion to be significantly associated with liver fat content. In addition, genes involved in ceramide signaling (MAP2K4) and metabolism (UGCG) were found to be positively associated with liver fat content. Genes involved in lipid metabolism (PLIN, ACADM), fatty acid transport (FABP4, CD36), amino acid catabolism (BCAT1), and inflammation (CCL2) were validated by real-time PCR and were found to be upregulated in subjects with high liver fat content. The data show that multiple changes in gene expression characterize simple steatosis.     

Liver X Receptor Agonists Augment Human Islet Function through Activation of Anaplerotic Pathways and Glycerolipid/Free Fatty Acid Cycling

Recent studies in rodent models suggest that liver X receptors (LXRs) may play an important role in the maintenance of glucose homeostasis and islet function. To date, however, no studies have comprehensively examined the role of LXRs in human islet biology. Human islets were isolated from non-diabetic donors and incubated in the presence or absence of two synthetic LXR agonists, TO-901317 and GW3965, under conditions of low and high glucose. LXR agonist treatment enhanced both basal and stimulated insulin secretion, which corresponded to an increase in the expression of genes involved in anaplerosis and reverse cholesterol transport. Furthermore, enzyme activity of pyruvate carboxylase, a key regulator of pyruvate cycling and anaplerotic flux, was also increased. Whereas LXR agonist treatment up-regulated known downstream targets involved in lipogenesis, we observed no increase in the accumulation of intra-islet triglyceride at the dose of agonist used in our study. Moreover, LXR activation increased expression of the genes encoding hormone-sensitive lipase and adipose triglyceride lipase, two enzymes involved in lipolysis and glycerolipid/free fatty acid cycling. Chronically, insulin gene expression was increased after treatment with TO-901317, and this was accompanied by increased Pdx-1 nuclear protein levels and enhanced Pdx-1 binding to the insulin promoter. In conclusion, our data suggest that LXR agonists have a direct effect on the islet to augment insulin secretion and expression, actions that should be considered either as therapeutic or unintended side effects, as these agents are developed for clinical use.     

Activation of nuclear receptor NR5A2 increases Glut4 expression and glucose metabolism in muscle cells

NR5A2 is a nuclear receptor which regulates the expression of genes involved in cholesterol metabolism, pluripotency maintenance and cell differentiation. It has been recently shown that DLPC, a NR5A2 ligand, prevents liver steatosis and improves insulin sensitivity in mouse models of insulin resistance, an effect that has been associated with changes in glucose and fatty acids metabolism in liver. Because skeletal muscle is a major tissue in clearing glucose from blood, we studied the effect of the activation of NR5A2 on muscle metabolism by using cultures of C2C12, a mouse-derived cell line widely used as a model of skeletal muscle. Treatment of C2C12 with DLPC resulted in increased levels of expression of GLUT4 and also of several genes related to glycolysis and glycogen metabolism. These changes were accompanied by an increased glucose uptake. In addition, the activation of NR5A2 produced a reduction in the oxidation of fatty acids, an effect which disappeared in low-glucose conditions. Our results suggest that NR5A2, mostly by enhancing glucose uptake, switches muscle cells into a state of glucose preference. The increased use of glucose by muscle might constitute another mechanism by which NR5A2 improves blood glucose levels and restores insulin sensitivity.     

Immunological impact of Wharton’s Jelly mesenchymal stromal cells and natural killer cell co-culture

In palmipeds, overfeeding leads to hepatic steatosis, also called “foie gras” which is the result of many metabolic mechanisms. In order to understand these mechanisms, we decided to measure the expression of genes implicated in lipid metabolism during 12 hours (h) following the last meal of the overfeeding period. We have shown that there is a precocious expression (within 2 h) of fatty acid synthase and acyl CoA synthetase long-chain 1 in liver and muscle of mule ducks in addition with a later peak. Furthermore, di-acyl glycerol acyl transferase presents the highest induction of expression in liver and it is overexpressed quite a long time, positioning this enzyme as a key factor in hepatic steatosis. These observations are quite similar in muscle. Lipoprotein secretion is upregulated later in postprandial period, with an upregulation of apolipoprotein and microsomal triglycerides transfer protein beginning at 5 h in liver or muscle. Regarding hepatic re-uptake of lipid, lesser variations are observed, suggesting that fatty acid binding protein and very low-density lipoprotein receptor (VLDLR) are already at their maximum expression specifically in these tissues. In muscle, VLDLR and LDLR upregulation is observed 5 h after the meal, associated with an overexpression in the adipose tissue of lipase maturation factor 1 involved in the maturation of lipoprotein lipase. These findings will allow us to better understand the kinetic treatment in lipid metabolism after a meal in overfed ducks. This first report on kinetic expression will allow researcher to better target their sampling time knowing the optimal point of expression of each gene.