Faculty Bibliography

CD36 is a multifunctional immuno-metabolic receptor with many ligands. One of its physiological functions in the heart is the high-affinity uptake of long-chain fatty acids (FAs) from albumin and triglyceride rich lipoproteins. CD36 deletion markedly reduces myocardial FA uptake in rodents and humans. The protein is expressed on endothelial cells and cardiomyocytes and at both sites is likely to contribute to FA uptake by the myocardium. CD36 also transduces intracellular signaling events that influence how the FA is utilized and mediate metabolic effects of FA in the heart. CD36 mediated signaling regulates AMPK activation in a way that adjusts oxidation to FA uptake. It also impacts remodeling of myocardial phospholipids and eicosanoid production, effects exerted via influencing intracellular calcium (iCa2+) and the activation of phospholipases. Under excessive FA supply CD36 contributes to lipid accumulation, inflammation and dysfunction. However, it is also important for myocardial repair after injury via its contribution to immune cell clearance of apoptotic cells. This review describes recent progress regarding the multiple actions of CD36 in the heart and highlights those areas requiring future investigation. This article is part of a Special Issue entitled: Heart Lipid Metabolism edited by G.D. Lopaschuk.

The global obesity epidemic and its impact on cardiovascular outcomes is a topic of ongoing debate and investigation in the cardiology community. It is well known that obesity is associated with multiple cardiovascular risk factors. Although life-style changes are the first line of therapy, they are often insufficient in achieving weight loss goals. Liraglutide, naltrexone/bupropion, and phentermine/topiramate are new agents that have been recently approved to treat obesity, but their effects on cardiovascular risk factors and outcomes are not well described. This review summarizes data currently available for these novel agents regarding drug safety, effects on major cardiovascular risk factors, impact on cardiovascular outcomes, outcomes research that is currently in progress, and areas of uncertainty. Given the impact of obesity on cardiovascular health, there is a pressing clinical need to understand the effects of these agents beyond weight loss alone.

Hypertriglyceridemia is an independent risk factor for cardiovascular disease, and plasma triglycerides (TGs) correlate strongly with plasma apolipoprotein C-III (ApoC-III) levels. Antisense oligonucleotides (ASOs) for ApoC-III reduce plasma TGs in primates and mice, but the underlying mechanism of action remains controversial. We determined that a murine-specific ApoC-III-targeting ASO reduces fasting TG levels through a mechanism that is dependent on low-density lipoprotein receptors (LDLRs) and LDLR-related protein 1 (LRP1). ApoC-III ASO treatment lowered plasma TGs in mice lacking lipoprotein lipase (LPL), hepatic heparan sulfate proteoglycan (HSPG) receptors, LDLR, or LRP1 and in animals with combined deletion of the genes encoding HSPG receptors and LDLRs or LRP1. However, the ApoC-III ASO did not lower TG levels in mice lacking both LDLR and LRP1. LDLR and LRP1 were also required for ApoC-III ASO-induced reduction of plasma TGs in mice fed a high-fat diet, in postprandial clearance studies, and when ApoC-III-rich or ApoC-III-depleted lipoproteins were injected into mice. ASO reduction of ApoC-III had no effect on VLDL secretion, heparin-induced TG reduction, or uptake of lipids into heart and skeletal muscle. Our data indicate that ApoC-III inhibits turnover of TG-rich lipoproteins primarily through a hepatic clearance mechanism mediated by the LDLR/LRP1 axis.

Cholesterol reduction has markedly reduced major cardiovascular disease (CVD) events and shown regression of atherosclerosis in some studies. However, CVD has for decades also been associated with increased levels of circulating triglyceride (TG)-rich lipoproteins. Whether this is due to a direct toxic effect of these lipoproteins on arteries or whether this is merely an association is unresolved. More recent genetic analyses have linked genes that modulate TG metabolism with CVD. Moreover, analyses of subgroups of hypertriglyceridemic (HTG) subjects in clinical trials using fibric acid drugs have been interpreted as evidence that TG reduction reduces CVD events. This review will focus on how HTG might cause CVD, whether TG reduction makes a difference, what pathophysiological defects cause HTG, and what options are available for treatment.

Kruppel-like factors (KLF) have important roles in metabolism. We previously found that KLF5 is a positive transcriptional regulator of peroxisome proliferator-activated receptor alpha (Ppara), a central regulator of cardiac fatty acid oxidation (FAO). Mice with cardiomyocyte-specific Klf5 ablation (alphaMHCKlf5 ) had reduced cardiac Ppara expression and FAO. At age 6-12 months these mice develop distinct cardiac dysfunction. The role of PPARalpha activation in I/R injury is unclear as both beneficial and detrimental effects have been reported. We aimed to study if Ppara expression changes during I/R are driven by KLF5 and explore its protective or detrimental role. Wild type mice were subjected to in vivo I/R or sham surgery. I/R resulted in an initial increase in Ppara, and its target gene pyruvate dehydrogenase kinase 4 (Pdk4) mRNA after 2h reperfusion, followed by decreased expression after 24h reperfusion. The Ppara expression is associated with parallel changes in cardiac Klf5 mRNA expression. Concurrent, there was a decrease of cardiac FAO-related genes carnitine palmitoyl-transferase 1beta (Cpt1b), very long chain acyl-CoA dehydrogenase (Vlcad), and acyl-CoA oxidase (Aox) in mice with I/R. To define the cell type causing the temporal changes in Klf5 and Ppara after I/R we isolated primary cardiomyocytes and fibroblasts. Our data suggest a similar effect in primary isolated cardiomyocytes only. Klf5 mRNA expression is increased after 2 hour hypoxia and normalized after 4 hour re-oxygenation in cardiomyocytes, whereas there are no changes after hypoxia/normoxia in fibroblasts. To assess the importance of cardiomyocyte KLF5 in I/R we used alphaMHC-Klf5 mice. Interestingly, despite reduced FAO, 7 month old alphaMHC-Klf5 mice subjected to I/R had a marked increase in mortality; 4 of 7 alphaMHCKlf5 mice died within the first 24h of reperfusion while no mortality was observed in age-matched wild type mice that underwent I/R. In conclusion, I/R is associated with an increase in Klf5 and Ppara in the first hours of reperfusion followed by a decrease in Klf5 and Ppara, likely accounted for by cardiomyocytes. Increased mortality for alphaMHC-Klf5 mice with I/R injury suggests that the initial increase may be an adaptive response that is critical for survival

The heart utilizes large amounts of fatty acids as energy providing substrates. The physiological balance of lipid uptake and oxidation prevents accumulation of excess lipids. Several processes that affect cardiac function, including ischemia, obesity, diabetes mellitus, sepsis, and most forms of heart failure lead to altered fatty acid oxidation and often also to the accumulation of lipids. There is now mounting evidence associating certain species of these lipids with cardiac lipotoxicity and subsequent myocardial dysfunction. Experimental and clinical data are discussed and paths to reduction of toxic lipids as a means to improve cardiac function are suggested.

Pulmonary surfactant is synthesized by type II pneumocytes and is mainly composed of phospholipids and minor amounts of cholesterol and specific proteins. Surfactant allows the lungs to inflate by decreasing alveolar surface tension and the work needed for inspiration in each breathing cycle. A delicate balance between synthesis, secretion, recycling and degradation of the lipids is required to maintain surfactant function and lung performance. Single nucleotide polymorphisms in the LDL receptor associated protein 1 (LRP-1) are associated with decreased lung function in patients with chronic obstructive pulmonary disease (COPD). But the function of LRP1 in the lung is not known. Amongst other roles, LRP-1 participates in liver lipoprotein metabolism, in adipocyte cell differentiation and in macrophage inflammatory cascades. We deleted LRP-1 in type II pneumocyte-derived A549 cells using stable transfection. LRP1 shRNA-treated cells and scrambled shRNA-treated cells were cultured in collagen coated transwell inserts at the air-liquid interphase, exposing the cellular basal side to the culture media and the apical side to the air. LRP-1 shRNA decreased surfactant phospholipid, cholesterol and cholesteryl esters on the apical surface of the cells. LRP-1 shRNA cells showed reduced gene expression of the rate limiting enzymes for fatty acid synthesis FAS, ACAT1 and SCD1, and of phosphatidylcholine synthesis enzymes choline transferase and choline kinase. Surprisingly, triglycerides and cholesteryl esters accumulated in intracellular lipid droplets in LRP-1 shRNA cells, while phospholipids and unesterified cholesterol were not changed when compared to scrambled shRNA cells. LRP-1 shRNA cells showed >3 fold overexpression of the fatty acid transporter CD36 and reduced gene expression of acyl-CoA oxidase. After an overnight incubation with fresh media in the basal side, LRP-1 shRNA cells showed higher media depletion of phospholipid, cholesterol esters and unesterifed cholesterol. Uptake of LDL-derived phosphatidylcholine was not altered by LRP-1 deletion, but the gene expression of phospholipid and cholesterol exporters ABCA1 and ABCG1 was reduced. Insulin signaling was affected by LRP-1 deletion. LRP-1 shRNA cells showed decreased protein expression of insulin receptor substrates 1 and 2 (IRS-1 and IRS-2) and increased phospho-serine IRS-1/total IRS-1 ratio. After IGF-1 treatment, LRP-1 shRNA cells failed to Tyr-phosphorylate the IGF-1 receptor b, and phosphorylation of the downstream target Akt was lower than in scrambled shRNA cells These data suggest that LRP-1 in type II pneumocytes is involved in lipid export, intracellular surfactant lipid metabolism and insulin signaling

RATIONALE: Fatty acid oxidation is transcriptionally regulated by peroxisome proliferator-activated receptor (PPAR)alpha and under normal conditions accounts for 70% of cardiac ATP content. Reduced Ppara expression during sepsis and heart failure leads to reduced fatty acid oxidation and myocardial energy deficiency. Many of the transcriptional regulators of Ppara are unknown. OBJECTIVE: To determine the role of Kruppel-like factor 5 (KLF5) in transcriptional regulation of Ppara. METHODS AND RESULTS: We discovered that KLF5 activates Ppara gene expression via direct promoter binding. This is blocked in hearts of septic mice by c-Jun, which binds an overlapping site on the Ppara promoter and reduces transcription. We generated cardiac myocyte-specific Klf5 knockout mice that showed reduced expression of cardiac Ppara and its downstream fatty acid metabolism-related targets. These changes were associated with reduced cardiac fatty acid oxidation, ATP levels, increased triglyceride accumulation and cardiac dysfunction. Diabetic mice showed parallel changes in cardiac Klf5 and Ppara expression levels. CONCLUSIONS: Cardiac myocyte KLF5 is a transcriptional regulator of Ppara and cardiac energetics.

3,5,3'-triiodo-L-thyronine (T3) and 3,5-diiodo-L-thyronine (T2), when administered to a model of familial hypercholesterolemia, i.e., low density lipoprotein receptor (LDLr)-knockout (Ldlr-/-) mice fed with a Western type diet (WTD), dramatically reduce circulating total and very low-density lipoprotein/LDL cholesterol with decreased liver apolipoprotein B (ApoB) production. The aim of the study was to highlight putative molecular mechanisms to manage cholesterol levels in the absence of LDLr. A comprehensive comparative profiling of changes in expression of soluble proteins in livers from Ldlr-/- mice treated with either T3 or T2 was performed. From a total proteome of 450 liver proteins, 25 identified proteins were affected by both T2 and T3, 18 only by T3 and 9 only by T2. Using in silico analyses, an overlap was observed with 11/14 pathways common to both iodothyronines, with T2 and T3 preferentially altering sub-networks centered around hepatocyte nuclear factor 4 alpha (HNF4alpha) and peroxisome proliferator-activated receptor alpha (PPARalpha), respectively. Both T2 and T3 administration significantly reduced nuclear HNF4alpha protein content, while T2, but not T3, decreased the expression levels of the HNFalpha transcriptional coactivator PGC-1alpha. Lower PPARalpha levels were found only following T3 treatment while both T3 and T2 lowered liver X receptor alpha (LXRalpha) nuclear content. Overall, this study, although it was not meant to investigate the use of T2 and T3 as a therapeutic agent, provides novel insights into the regulation of hepatic metabolic pathways involved in T3- and T2-driven cholesterol reduction in Ldlr-/- mice.