Daniel Lindén

Opposing Effects of Adiponectin Receptors 1 and 2 on Energy Metabolism

The adipocyte-derived hormone adiponectin regulates glucose and lipid metabolism and influences the risk for developing obesity, type 2 diabetes, and cardiovascular disease. Adiponectin binds to two different seven-transmembrane domain receptors termed AdipoR1 and AdipoR2. To study the physiological importance of these receptors, AdipoR1 gene knockout mice (AdipoR1−/−) and AdipoR2 gene knockout mice (AdipoR2−/−) were generated. AdipoR1−/− mice showed increased adiposity associated with decreased glucose tolerance, spontaneous locomotor activity, and energy expenditure. However, AdipoR2−/− mice were lean and resistant to high-fat diet–induced obesity associated with improved glucose tolerance and higher spontaneous locomotor activity and energy expenditure and reduced plasma cholesterol levels. Thus, AdipoR1 and AdipoR2 are clearly involved in energy metabolism but have opposing effects.

Adipocyte Differentiation-Related Protein Promotes Fatty Acid Storage in Cytosolic Triglycerides and Inhibits Secretion of Very Low–Density Lipoproteins

Objective—We investigated the role of adipocyte differentiation-related protein (ADRP) in triglyceride turnover and in the secretion of very low–density lipoprotein (VLDL) from McA-RH7777 cells and primary rat hepatocytes. Methods and Results—An increase in the expression of ADRP increased triglyceride accumulation in cytosolic lipid droplets and prevented the incorporation of fatty acids into secretable triglycerides, thereby reducing the secretion of triglycerides as well as of apolipoprotein B-100 (apoB-100) and apoB-48 VLDL. The ability of ADRP to block the secretion of apoB-100 VLDL1 decreased with increasing quantities of fatty acids in the medium, indicating a saturable process and emphasizing the importance of sequestering of fatty acids for the effect of ADRP on VLDL secretion. Knockdown (small interfering RNA) of ADRP decreased the pool of cytosolic lipid droplets but increased only the secretion of apoB-48 VLDL1. Additionally, there was an increased flow of fatty acids into &bgr;-oxidation. Conclusions—ADRP is essential for the accumulation of triglycerides in cytosolic lipid droplets. An increase in ADRP prevents the formation of VLDL by diverting fatty acids from the VLDL assembly pathway into cytosolic triglycerides, whereas a decrease of the protein increases the sorting of fatty acids to &bgr;-oxidation and promotes the secretion of apoB-48 VLDL1.

Liver-directed overexpression of mitochondrial glycerol-3-phosphate acyltransferase results in hepatic steatosis, increased triacylglycerol secretion and reduced fatty acid oxidation

Glycerol‐3‐phosphate acyltransferase (GPAT) catalyzes the first committed step in triacylglycerol (TAG) and phospholipid biosynthesis. GPAT activity has been identified in both ER and mitochondrial subcellular fractions. The ER activity dominates in most tissues except in liver, where the mitochondrial isoform (mtGPAT) can constitute up to 50% of the total activity. To study the in vivo effects of hepatic mtGPAT overexpression, mice were transduced with adenoviruses expressing either murine mtGPAT or a catalytically inactive variant of the enzyme. Overexpressing mtGPAT resulted in massive 12‐and 7‐fold accumulation of liver TAG and diacylglycerol, respectively but had no effect on phospholipid or cholesterol ester content. Histological analysis showed extensive lipid accumulation in hepatocytes. Furthermore, mtGPAT transduction markedly increased adipocyte differentiation‐related protein and stearoyl‐CoA desaturase‐1 (SCD‐1) in the liver. In line with increased SCD‐1 expression, 18:1 and 16:1 in the hepatic TAG fraction increased. In addition, mtGPAT overexpression decreased ex vivo fatty acid oxidation, increased liver TAG secretion rate 2‐fold, and increased plasma TAG and cholesterol levels. These results support the hypothesis that increased hepatic mtGPAT activity associated with obesity and insulin resistance contributes to increased TAG biosynthesis and inhibition of fatty acid oxidation, responses that would promote hepatic steatosis and dyslipidemia.—Lindén, D., William‐Olsson, L., Ahnmark, A., Ekroos, K., Hallberg, C., Sjögren, H. P., Becker, B., Svensson, L., Clapham, J. C., Oscarsson, J., Schreyer, S. Liver‐directed overexpression of mitochondrial glycerol‐3‐phosphate acyltransferase results in hepatic steatosis, increased triacylglycerol secretion and reduced fatty acid oxidation. FASEB J. 20, 434–443 (2006)

Hepatic PGC-1β Overexpression Induces Combined Hyperlipidemia and Modulates the Response to PPARα Activation

Objective—Previous studies have indicated that the hyperlipidemia and gene expression changes induced by a short-term high-fat diet (HFD) are mediated through the peroxisome proliferator-activated receptor &ggr; coactivator (PGC)-1&bgr;, and that in vitro both PGC-1&bgr; and PGC −1&agr; increase PPAR&agr;-mediated transcriptional activities. Here, we examined the in vivo effects of these two coactivators in potentiating the lipid lowering properties of the PPAR&agr; agonist Wy14,643 (Wy). Methods and Results—C57BL/6 mice were fed chow or HFD and transduced with adenoviruses encoding PGC-1&agr; or PGC-1&bgr;. On chow, hepatic PGC-1&bgr; overexpression caused severe combined hyperlipidemia including elevated plasma apolipoprotein B levels. Hepatic triglyceride secretion, DGAT1, and FAT/CD36 expression were increased whereas PPAR&agr; and hepatic lipase mRNA levels were reduced. PGC-1&bgr; overexpression blunted Wy-mediated changes in expression levels of PPAR&agr; and downstream genes. Furthermore, PGC-1&bgr; did not potentiate Wy-stimulated fatty acid oxidation in primary hepatocytes. PGC-1&bgr; and PGC-1&agr; overexpression did not alter SREBP-1c, SREBP-1c target gene expression, nor hepatic triglyceride content. On HFD, PGC-1&bgr; overexpression decreased hepatic SREBP-1c, yet increased FAS and ACC&agr; mRNA and plasma triglyceride levels. Conclusions—Hepatic PGC-1&bgr; overexpression caused combined hyperlipidemia independent of SREBP-1c activation. Hepatic PGC-1&bgr; overexpression reduced the potentially beneficial effects of PPAR&agr; activation on gene expression. Thus, inhibition of hepatic PGC-1&bgr; may provide a therapy for treating combined hyperlipidemia.

Ageing Fxr Deficient Mice Develop Increased Energy Expenditure, Improved Glucose Control and Liver Damage Resembling NASH

Nuclear receptor subfamily 1, group H, member 4 (Nr1h4, FXR) is a bile acid activated nuclear receptor mainly expressed in the liver, intestine, kidney and adrenal glands. Upon activation, the primary function is to suppress cholesterol 7 alpha-hydroxylase (Cyp7a1), the rate-limiting enzyme in the classic or neutral bile acid synthesis pathway. In the present study, a novel Fxr deficient mouse line was created and studied with respect to metabolism and liver function in ageing mice fed chow diet. The Fxr deficient mice were similar to wild type mice in terms of body weight, body composition, energy intake and expenditure as well as behaviours at a young age. However, from 15 weeks of age and onwards, the Fxr deficient mice had almost no body weight increase up to 39 weeks of age mainly because of lower body fat mass. The lower body weight gain was associated with increased energy expenditure that was not compensated by increased food intake. Fasting levels of glucose and insulin were lower and glucose tolerance was improved in old and lean Fxr deficient mice. However, the Fxr deficient mice displayed significantly increased liver weight, steatosis, hepatocyte ballooning degeneration and lobular inflammation together with elevated plasma levels of ALT, bilirubin and bile acids, findings compatible with non-alcoholic steatohepatitis (NASH) and cholestasis. In conclusion, ageing Fxr deficient mice display late onset leanness associated with elevated energy expenditure and improved glucose control but develop severe NASH-like liver pathology.

The beneficial effects of n-3 polyunsaturated fatty acids on diet induced obesity and impaired glucose control do not require Gpr120.

GPR120 (Ffar4) has been postulated to represent an important receptor mediating the improved metabolic profile seen upon ingestion of a diet enriched in polyunsaturated fatty acids (PUFAs). GPR120 is highly expressed in the digestive system, adipose tissue, lung and macrophages and also present in the endocrine pancreas. A new Gpr120 deficient mouse model on pure C57bl/6N background was developed to investigate the importance of the receptor for long-term feeding with a diet enriched with fish oil. Male Gpr120 deficient mice were fed two different high fat diets (HFDs) for 18 weeks. The diets contained lipids that were mainly saturated (SAT) or mainly n-3 polyunsaturated fatty acids (PUFA). Body composition, as well as glucose, lipid and energy metabolism, was studied. As expected, wild type mice fed the PUFA HFD gained less body weight and had lower body fat mass, hepatic lipid levels, plasma cholesterol and insulin levels and better glucose tolerance as compared to those fed the SAT HFD. Gpr120 deficient mice showed a similar improvement on the PUFA HFD as was observed for wild type mice. If anything, the Gpr120 deficient mice responded better to the PUFA HFD as compared to wild type mice with respect to liver fat content, plasma glucose levels and islet morphology. Gpr120 deficient animals were found to have similar energy, glucose and lipid metabolism when fed HFD PUFA compared to wild type mice. Therefore, GPR120 appears to be dispensable for the improved metabolic profile associated with intake of a diet enriched in n-3 PUFA fatty acids.

Low dose of the liver X receptor agonist, AZ876, reduces atherosclerosis in APOE*3Leiden mice without affecting liver or plasma triglyceride levels

BACKGROUND AND PURPOSE Liver X receptor (LXR) agonists are atheroprotective but often induce hypertriglyceridaemia and liver steatosis. We investigated the effect of a novel high‐affinity LXR activator, AZ876, on plasma lipids, inflammation and atherosclerosis, and compared the effects with another LXR agonist, GW3965.

The role of mitochondrial glycerol-3-phosphate acyltransferase-1 in regulating lipid and glucose homeostasis in high-fat diet fed mice.

Glycerol-3-phosphate acyltransferase (GPAT) is involved in triacylglycerol (TAG) and phospholipid synthesis, catalyzing the first committed step. In order to further investigate the in vivo importance of the dominating mitochondrial variant, GPAT1, a novel GPAT1(-/-) mouse model was generated and studied. Female GPAT1(-/-) mice had reduced body weight-gain and adiposity when fed chow diet compared with littermate wild-type controls. Furthermore, GPAT1(-/-) females on chow diet showed decreased liver TAG content, plasma cholesterol and TAG levels and increased ex vivo liver fatty acid oxidation and plasma ketone bodies. However, these beneficial effects were abolished and the glucose tolerance tended to be impaired when GPAT1(-/-) females were fed a long-term high-fat diet (HFD). GPAT1-deficiency was not associated with altered whole body energy expenditure or respiratory exchange ratio. In addition, there were no changes in male GPAT1(-/-) mice fed either diet except for increased plasma ketone bodies on chow diet, indicating a gender-specific phenotype. Thus, GPAT1-deficiency does not protect against HFD-induced obesity, hepatic steatosis or whole body glucose intolerance.

Decreased expression of peroxisome proliferator-activated receptor α and liver fatty acid binding protein after partial hepatectomy of rats and mice

Abstract: Background Aims: Marked changes in metabolism, including liver steatosis and hypoglycemia, occur after partial hepatectomy. Peroxisome proliferator‐activated receptor α (PPARα) is a nuclear hormone receptor that is activated by fatty acids and involved in hepatic fatty acid metabolism and regeneration. Liver fatty acid binding protein (LFABP) is an abundant protein in liver cytosol whose expression is regulated by PPARα. It is involved in fatty acid uptake and diffusion and in PPARα signaling. The aim of this study was to investigate the expression of PPARα and LFABP during liver regeneration.

Discovery of (3-(4-(2-Oxa-6-azaspiro[3.3]heptan-6-ylmethyl)phenoxy)azetidin-1-yl)(5-(4-methoxyphenyl)-1,3,4-oxadiazol-2-yl)methanone (AZD1979), a Melanin Concentrating Hormone Receptor 1 (MCHr1) Antagonist with Favorable Physicochemical Properties

A novel series of melanin concentrating hormone receptor 1 (MCHr1) antagonists were the starting point for a drug discovery program that culminated in the discovery of 103 (AZD1979). The lead optimization program was conducted with a focus on reducing lipophilicity and understanding the physicochemical properties governing CNS exposure and undesired off-target pharmacology such as hERG interactions. An integrated approach was taken where the key assay was ex vivo receptor occupancy in mice. The candidate compound 103 displayed appropriate lipophilicity for a CNS indication and showed excellent permeability with no efflux. Preclinical GLP toxicology and safety pharmacology studies were without major findings and 103 was taken into clinical trials.