Faculty Bibliography

OBJECTIVE: Patients with diabetes often have dyslipidemia and increased postprandial lipidmia. Induction of diabetes in LDL receptor (Ldlr(-/-)) knockout mice also leads to marked dyslipidemia. The reasons for this are unclear. RESEARCH DESIGN AND METHODS: We placed Ldlr(-/-) and heterozygous LDL receptor knockout (Ldlr(+/-)) mice on a high-cholesterol (0.15%) diet, induced diabetes with streptozotocin (STZ), and assessed reasons for differences in plasma cholesterol. RESULTS: STZ-induced diabetic Ldlr(-/-) mice had plasma cholesterol levels more than double those of nondiabetic controls. Fast-performance liquid chromatography and ultracentrifugation showed an increase in both VLDL and LDL. Plasma VLDL became more cholesterol enriched, and both VLDL and LDL had a greater content of apolipoprotein (apo)E. In LDL the ratio of apoB48 to apoB100 was increased. ApoB production, assessed using [(35)S]methionine labeling in Triton WR1339-treated mice, was not increased in fasting STZ-induced diabetic mice. Similarly, postprandial lipoprotein production was not increased. Reduction of cholesterol in the diet to normalize the amount of cholesterol intake by the control and STZ-induced diabetic animals reduced plasma cholesterol levels in STZ-induced diabetic mice, but plasma cholesterol was still markedly elevated compared with nondiabetic controls. LDL from STZ-induced diabetic mice was cleared from the plasma and trapped more rapidly by livers of control mice. STZ treatment reduced liver expression of the proteoglycan sulfation enzyme, heparan sulfate N-deacetylase/N-sulfotrasferase-1, an effect that was reproduced in cultured hepatocytyes by a high glucose-containing medium. CONCLUSIONS: STZ-induced diabetic, cholesterol-fed mice developed hyperlipidemia due to a non-LDL receptor defect in clearance of circulating apoB-containing lipoproteins.

The intestine and other tissues are able to synthesize retinyl esters in an acyl-CoA-dependent manner involving an acyl-CoA:retinol acyltransferase (ARAT). However, the molecular identity of this ARAT has not been established. Recent studies of lecithin:retinol acyltransferase (LRAT)-deficient mice indicate that LRAT is responsible for the preponderance of retinyl ester synthesis in the body, aside from in the intestine and adipose tissue. Our present studies, employing a number of mutant mouse models, identify diacylglycerol acyltransferase 1 (DGAT1) as an important intestinal ARAT in vivo. The contribution that DGAT1 makes to intestinal retinyl ester synthesis becomes greater when a large pharmacologic dose of retinol is administered by gavage to mice. Moreover, when large retinol doses are administered another intestinal enzyme(s) with ARAT activity becomes apparent. Surprisingly, although DGAT1 is expressed in adipose tissue, DGAT1 does not catalyze retinyl ester synthesis in adipose tissue in vivo. Our data also establish that cellular retinol-binding protein, type II (CRBPII), which is expressed solely in the adult intestine, in vivo channels retinol to LRAT for retinyl ester synthesis. Contrary to what has been proposed in the literature based on in vitro studies, CRBPII does not directly prevent retinol from being acted upon by DGAT1 or other intestinal ARATs in vivo.

Hepatic steatosis is often associated with insulin resistance and obesity and can lead to steatohepatitis and cirrhosis. In this study, we have demonstrated that hormone-sensitive lipase (HSL) and adipose triglyceride lipase (ATGL), two enzymes critical for lipolysis in adipose tissues, also contribute to lipolysis in the liver and can mobilize hepatic triglycerides in vivo and in vitro. Adenoviral overexpression of HSL and/or ATGL reduced liver triglycerides by 40-60% in both ob/ob mice and mice with high fat diet-induced obesity. However, these enzymes did not affect fasting plasma triglyceride and free fatty acid levels or triglyceride and apolipoprotein B secretion rates. Plasma 3-beta-hydroxybutyrate levels were increased 3-5 days after infection in both HSL- and ATGL-overexpressing male mice, suggesting an increase in beta-oxidation. Expression of genes involved in fatty acid transport and synthesis, lipid storage, and mitochondrial bioenergetics was unchanged. Mechanistic studies in oleate-supplemented McA-RH7777 cells with adenoviral overexpression of HSL or ATGL showed that reduced cellular triglycerides could be attributed to increases in beta-oxidation as well as direct release of free fatty acids into the medium. In summary, hepatic overexpression of HSL or ATGL can promote fatty acid oxidation, stimulate direct release of free fatty acid, and ameliorate hepatic steatosis. This study suggests a direct functional role for both HSL and ATGL in hepatic lipid homeostasis and identifies these enzymes as potential therapeutic targets for ameliorating hepatic steatosis associated with insulin resistance and obesity.

OBJECTIVE: Although epidemiologic data suggest that hypertriglyceridemia and elevated plasma levels of fatty acids are toxic to arteries, in vitro correlates have been inconsistent. To investigate whether increased endothelial cell expression of lipoprotein lipase (LpL), the primary enzyme creating free fatty acids from circulating triglycerides (TG), affects vascular function, we created transgenic mice that express human LpL (hLpL) driven by the promoter and enhancer of the Tie2 receptor. METHODS AND RESULTS: Mice expressing this transgene, denoted EC-hLpL and L for low and H for high expression, had decreased plasma TG levels compared with wild-type mice (WT): 106+/-31 in WT, 37+/-17 (line H), and 63+/-31 mg/dL (line L) because of a reduction in VLDL TG; plasma cholesterol and HDL levels were unaltered. Crossing a high expressing EC-hLpL transgene onto the LpL knockout background allowed for survival of the pups; TG in these mice was approximately equal to that of heterozygous LpL knockout mice. Surprisingly, under control conditions the EC-hLpL transgene did not alter arterial function or endothelial cell gene expression; however, after tumor necrosis factor (TNF)-alpha treatment, arterial vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and endogenous TNF-alpha mRNA levels were increased and arteries had impaired endothelium-dependent vasodilatation. This was associated with reduced eNOS dimers. CONCLUSIONS: Therefore, we hypothesize that excess vascular wall LpL augments vascular dysfunction in the setting of inflammation.

Hypertriglyceridemia is one of the known causes of pancreatitis. Estrogen treatment can aggravate hypertriglyceridemia by increasing very low density lipoprotein secretion and reducing hepatic triglyceride lipase. In this paper, we present 3 patients who developed severe hypertriglyceridemia with conditions that increased estrogen. Two patients were found to have genetic lipoprotein lipase deficiency and were treated with birth control pills. The third was a patient with polycystic ovary disease who was receiving ovulation induction therapy for in vitro fertilization.

A panel was convened on February 18, 2008, to discuss the challenges to the diagnosis, evaluation, treatment, and management of clustered cardiometabolic risk factors. Peter W.F. Wilson, MD of Emory University School of Medicine, Atlanta, GA, moderated the panel. Henry R. Black, MD, New York University School of Medicine, New York, NY, Anthony N. Fabricatore, PhD, University of Pennsylvania, Philadelphia, PA, and Ira J. Goldberg, MD, Columbia University, New York, NY, participated in the discussion. This expert panel discussion was supported by and each author received an honorarium from Pfizer Inc for time and effort spent participating in the discussion and reviewing the transcript for important intellectual content before publication. The authors maintained full control of the discussion and the resulting content of this article.

Under normal circumstances, most energy substrate used for heart contraction derives from fatty acids in the form of nonesterified fatty acids bound to albumin or fatty acids derived from lipolysis of lipoprotein-bound triglyceride by lipoprotein lipase (LpL). By creating LpL knockout mice (hLpL0), we learned that loss of cardiac LpL leads to myocardial dysfunction; therefore, neither nonesterified fatty acids nor increased glucose metabolism can replace LpL actions. hLpL0 mice do not survive abdominal aortic constriction and they develop more heart failure with hypertension. Conversely, we created a mouse overexpressing cardiomyocyte-anchored LpL. This transgene produced cardiac lipotoxicity and dilated cardiomyopathy. Methods to alter this phenotype and the causes of other models of lipotoxicity are currently being studied and will provide further insight into the physiology of lipid metabolism in the heart.

The creation of mouse models that recapitulate human diabetic cardiovascular disease remains a significant challenge. Part of the problem relates to the lack of a clear understanding of the human phenotype. Although improved insulin-treat of hyperglycemia reduces cardiovascular events in patients with type 1 diabetes, similar data are not available in type 2 diabetes. Moreover, whether human vascular disease is increased by hyperglycemia, defective insulin actions, or other factors is not known. Significant progress has been made in developing models of both type 1 and type 2 diabetes in mouse that can be used to study the relationship between hyperglycemia and atherosclerosis. This review describes mouse models that recapitulate specific aspects of diabetic dyslipidemia, hyperglycemia/insulin resistance, and diabetic vascular disease. Overall, the studies have clearly demonstrated that hyperlipidemia is a major driver of atherosclerotic vascular disease in the mouse. The effects of hyperglycemia and insulin resistance on murine atherosclerosis remain uncertain.

Three forms of PPARs are expressed in the heart. In animal models, PPARgamma agonist treatment improves lipotoxic cardiomyopathy; however, PPARgamma agonist treatment of humans is associated with peripheral edema and increased heart failure. To directly assess effects of increased PPARgamma on heart function, we created transgenic mice expressing PPARgamma1 in the heart via the cardiac alpha-myosin heavy chain (alpha-MHC) promoter. PPARgamma1-transgenic mice had increased cardiac expression of fatty acid oxidation genes and increased lipoprotein triglyceride (TG) uptake. Unlike in cardiac PPARalpha-transgenic mice, heart glucose transporter 4 (GLUT4) mRNA expression and glucose uptake were not decreased. PPARgamma1-transgenic mice developed a dilated cardiomyopathy associated with increased lipid and glycogen stores, distorted architecture of the mitochondrial inner matrix, and disrupted cristae. Thus, while PPARgamma agonists appear to have multiple beneficial effects, their direct actions on the myocardium have the potential to lead to deterioration in heart function.

PURPOSE OF REVIEW: How do lipids arrive in the heart and other tissues? This review focuses on new information on pathways of lipid uptake into the heart. RECENT FINDINGS: Fatty acids, the major cardiac fuel, are obtained from either lipoproteins or free fatty acids associated with albumin. The heart is the tissue with the most robust expression of lipoprotein lipase, and recent data attest to the importance of this enzyme in supplying optimal amounts of fatty acids for the heart. Genetic deletion of CD36 also shows that this transporter is important for cardiac uptake of lipids. Retinoid acquisition by the heart involves pathways parallel to those used for fatty acid uptake: a pathway for acquisition of core lipoprotein retinyl ester and another for nonlipoprotein retinol. Dilated lipotoxic cardiomyopathy is the consequence of excess lipid uptake. SUMMARY: Genetic modifications that affect lipid uptake, oxidation, and storage are being exploited to elucidate the pathophysiology of cardiomyopathies and to discover how lipids relate to heart failure in humans with obesity and diabetes mellitus. This information is likely to lead to new diagnostic categories of cardiomyopathy and more pathophysiologically appropriate treatments.