Faculty Bibliography

OBJECTIVE: The objective of this study was to investigate the role of vascular ATP-binding cassette transporter G1 (ABCG1) in atherogenesis without a confounding difference in macrophage ABCG1 expression. ABCG1 is highly expressed in macrophages and endothelial cells. ABCG1 preserves endothelial function by maintaining endothelial NO synthase activity and by reducing adhesion molecule expression and monocyte adhesion. METHODS AND RESULTS: To investigate the role of vascular ABCG1 in atherosclerosis in vivo Abcg1(-/-)/Ldlr(-/-) and Ldlr(-/-) mice were transplanted with wild-type bone marrow and fed a Western-type diet for 12 or 23 weeks. The atherosclerotic lesion area was similar in both groups after 12 weeks but was increased in Abcg1(-/-)/Ldlr(-/-) recipients after 23 weeks, especially in the aortic arch (2.2-fold; P<0.01). Endothelial NO synthase-mediated vascular relaxation was impaired in male Abcg1(-/-)/Ldlr(-/-) recipients. CONCLUSIONS: Our data show an atheroprotective role of vascular ABCG1, especially in the aortic arch, likely related to its role in the preservation of endothelial NO synthase activity.

Excess lipid accumulation in the heart is associated with decreased cardiac function in humans and in animal models. The reasons are unclear, but this is generally believed to result from either toxic effects of intracellular lipids or excessive fatty acid oxidation (FAO). PPARgamma expression is increased in the hearts of humans with metabolic syndrome, and use of PPARgamma agonists is associated with heart failure. Here, mice with dilated cardiomyopathy due to cardiomyocyte PPARgamma overexpression were crossed with PPARalpha-deficient mice. Surprisingly, this cross led to enhanced expression of several PPAR-regulated genes that mediate fatty acid (FA) uptake/oxidation and triacylglycerol (TAG) synthesis. Although FA oxidation and TAG droplet size were increased, heart function was preserved and survival improved. There was no marked decrease in cardiac levels of triglyceride or the potentially toxic lipids diacylglycerol (DAG) and ceramide. However, long-chain FA coenzyme A (LCCoA) levels were increased, and acylcarnitine content was decreased. Activation of PKCalpha and PKCdelta, apoptosis, ROS levels, and evidence of endoplasmic reticulum stress were also reduced. Thus, partitioning of lipid to storage and oxidation can reverse cardiolipotoxicity despite increased DAG and ceramide levels, suggesting a role for other toxic intermediates such as acylcarnitines in the toxic effects of lipid accumulation in the heart.

We previously observed that treatment of mice with a dominant negative form of cJun (dn-cJun) increased the expression of genes involved in lipid metabolism and modulated the expression of nine microRNAs (miR). To investigate the potential effect of these miRs on the expression of the genes of lipid metabolism, we performed studies in cultured HepG2 cells. Transfection of HepG2 cells with sense or antisense miR-370 or miR-122 upregulated and downregulated, respectively, the transcription factor sterol-regulatory element binding protein 1c (SREBP-1c) and the enzymes diacylglycerol acyltransferase-2 (DGAT2), fatty acid synthase (FAS), and acyl-CoA carboxylase 1 (ACC1) that regulate fatty acid and triglyceride biosynthesis. The other seven miRs identified by the miR array screening did not affect the expression of lipogenic genes. miR-370 upregulated the expression of miR-122. Furthermore, the effect of miR-370 on the expression of the lipogenic genes was abolished by antisense miR-122. miR-370 targets the 3' untranslated region (UTR) of Cpt1alpha, and it downregulated the expression of the carnitine palmitoyl transferase 1alpha (Cpt1alpha) gene as well as the rate of beta oxidation. Our data suggest that miR-370 acting via miR-122 may have a causative role in the accumulation of hepatic triglycerides by modulating initially the expression of SREBP-1c, DGAT2, and Cpt1alpha and, subsequently, the expression of other genes that affect lipid metabolism.

Hyperglycemia and reduced insulin actions affect many biological processes. One theory is that aberrant metabolism of glucose via several pathways including the polyol pathway causes cellular toxicity. Aldose reductase (AR) is a multifunctional enzyme that reduces aldehydes. Under diabetic conditions AR converts glucose into sorbitol, which is then converted to fructose. This article reviews the biology and pathobiology of AR actions. AR expression varies considerably among species. In humans and rats, the higher level of AR expression is associated with toxicity. Flux via AR is increased by ischemia and its inhibition during ischemia reperfusion reduces injury. However, similar pharmacological effects are not observed in mice unless they express a human AR transgene. This is because mice have much lower levels of AR expression, probably insufficient to generate toxic byproducts. Human AR expression in LDL receptor knockout mice exacerbates vascular disease, but only under diabetic conditions. In contrast, a recent report suggests that genetic ablation of AR increased atherosclerosis and increased hydroxynonenal in arteries. It was hypothesized that AR knockout prevented reduction of toxic aldehydes. Like many in vivo effects found in genetically manipulated animals, interpretation requires the reproduction of human-like physiology. For AR, this will require tissue specific expression of AR in sites and at levels that approximate those in humans

Retinoids are absolutely required for normal growth and development during the postnatal period. We studied the delivery of retinoids to milk, availing of mouse models modified for proteins thought to be essential for this process. Milk retinyl esters were markedly altered in mice lacking the enzyme lecithin:retinol acyltransferase (Lrat(-/-)), indicating that this enzyme is normally responsible for the majority of retinyl esters incorporated into milk and not an acyl-CoA dependent enzyme, as proposed in the literature. Unlike wild-type milk, much of the retinoid in Lrat(-/-) milk is unesterified retinol, not retinyl ester. The composition of the residual retinyl ester present in Lrat(-/-) milk was altered from predominantly retinyl palmitate and stearate to retinyl oleate and medium chain retinyl esters. This was accompanied by increased palmitate and decreased oleate in Lrat(-/-) milk triglycerides. In other studies, we investigated the role of retinol-binding protein in retinoid delivery for milk formation. We found that Rbp(-/-) mice maintain milk retinoid concentrations similar to those in matched wild-type mice. This appears to arise due to greater postprandial delivery of retinoid, a lipoprotein lipase (LPL)-dependent pathway. Importantly, LPL also acts to assure delivery of long-chain fatty acids (LCFA) to milk. The fatty acid transporter CD36 also facilitated LCFA but not retinoid incorporation into milk. Our data show that compensatory pathways for the delivery of retinoids ensure their optimal delivery and that LRAT is the most important enzyme for milk retinyl ester formation.

PURPOSE OF REVIEW: Diseases associated with ectopic disposition of lipids are becoming an increasingly important medical problem as the incidence of type 2 diabetes and obesity increases. One of the organs affected by lipotoxicity is the heart and this review presents an update on human and animal studies of this problem. RECENT FINDINGS: Human studies have clearly correlated heart dysfunction with the content of triglyceride. More recently human heart samples have been used to assess gene changes associated with altered lipid accumulation. Genetically altered mice have been created that develop lipotoxic cardiomyopathies and newer investigations are attempting to delineate curative therapies. SUMMARY: Human studies will confirm the metabolic changes associated with lipotoxic cardiomyopathy and, hopefully, animal studies will guide treatment options.

BACKGROUND: Emerging evidence in obesity and diabetes mellitus demonstrates that excessive myocardial fatty acid uptake and oxidation contribute to cardiac dysfunction. Transgenic mice with cardiac-specific overexpression of the fatty acid-activated nuclear receptor peroxisome proliferator-activated receptor-alpha (myosin heavy chain [MHC]-PPARalpha mice) exhibit phenotypic features of the diabetic heart, which are rescued by deletion of CD36, a fatty acid transporter, despite persistent activation of PPARalpha gene targets involved in fatty acid oxidation. METHODS AND RESULTS: To further define the source of fatty acid that leads to cardiomyopathy associated with lipid excess, we crossed MHC-PPARalpha mice with mice deficient for cardiac lipoprotein lipase (hsLpLko). MHC-PPARalpha/hsLpLko mice exhibit improved cardiac function and reduced myocardial triglyceride content compared with MHC-PPARalpha mice. Surprisingly, in contrast to MHC-PPARalpha/CD36ko mice, the activity of the cardiac PPARalpha gene regulatory pathway is normalized in MHC-PPARalpha/hsLpLko mice, suggesting that PPARalpha ligand activity exists in the lipoprotein particle. Indeed, LpL mediated hydrolysis of very-low-density lipoprotein activated PPARalpha in cardiac myocytes in culture. The rescue of cardiac function in both models was associated with improved mitochondrial ultrastructure and reactivation of transcriptional regulators of mitochondrial function. CONCLUSIONS: MHC-PPARalpha mouse hearts acquire excess lipoprotein-derived lipids. LpL deficiency rescues myocyte triglyceride accumulation, mitochondrial gene regulatory derangements, and contractile function in MHC-PPARalpha mice. Finally, LpL serves as a source of activating ligand for PPARalpha in the cardiomyocyte.

Mutations in SCO2, a protein required for the proper assembly and functioning of cytochrome c oxidase (COX; complex IV of the mitochondrial respiratory chain), cause a fatal infantile cardioencephalomyopathy with COX deficiency. We have generated mice harboring a Sco2 knock-out (KO) allele and a Sco2 knock-in (KI) allele expressing an E-->K mutation at position 129 (E129K), corresponding to the E140K mutation found in almost all human SCO2-mutated patients. Whereas homozygous KO mice were embryonic lethals, homozygous KI and compound heterozygous KI/KO mice were viable, but had muscle weakness; biochemically, they had respiratory chain deficiencies as well as complex IV assembly defects in multiple tissues. There was a concomitant reduction in mitochondrial copper content, but the total amount of copper in examined tissues was not reduced. These mouse models should be of use in further studies of Sco2 function, as well as in testing therapeutic approaches to treat the human disorder.