Search

LB

Objective: Fatty acid uptake and oxidation characterize the metabolism of alternatively activated macrophage polarization in vitro, but the in vivo biology is less clear. We assessed the roles of LpL (lipoprotein lipase)-mediated lipid uptake in macrophage polarization in vitro and in several important tissues in vivo. Approach and Results: We created mice with both global and myeloid-cell specific LpL deficiency. LpL deficiency in the presence of VLDL (very low-density lipoproteins) altered gene expression of bone marrow–derived macrophages and led to reduced lipid uptake but an increase in some anti- and some proinflammatory markers. However, LpL deficiency did not alter lipid accumulation or gene expression in circulating monocytes nor did it change the ratio of Ly6C high /Ly6C low . In adipose tissue, less macrophage lipid accumulation was found with global but not myeloid-specific LpL deficiency. Neither deletion affected the expression of inflammatory genes. Global LpL deficiency also reduced the numbers of elicited peritoneal macrophages. Finally, we assessed gene expression in macrophages from atherosclerotic lesions during regression; LpL deficiency did not affect the polarity of plaque macrophages. Conclusions: The phenotypic changes observed in macrophages upon deletion of Lpl in vitro is not mimicked in tissue macrophages.

Objective— SGLT2 (sodium-glucose cotransporter 2) inhibition in humans leads to increased levels of LDL (low-density lipoprotein) cholesterol and decreased levels of plasma triglyceride. Recent studies, however, have shown this therapy to lower cardiovascular mortality. In this study, we aimed to determine how SGLT2 inhibition alters circulating lipoproteins. Approach and Results— We used a mouse model expressing human CETP (cholesteryl ester transfer protein) and human ApoB100 (apolipoprotein B100) to determine how SGLT2 inhibition alters plasma lipoprotein metabolism. The mice were fed a high-fat diet and then were made partially insulin deficient using streptozotocin. SGLT2 was inhibited using a specific antisense oligonucleotide or canagliflozin, a clinically available oral SGLT2 inhibitor. Inhibition of SGLT2 increased circulating levels of LDL cholesterol and reduced plasma triglyceride levels. SGLT2 inhibition was associated with increased LpL (lipoprotein lipase) activity in the postheparin plasma, decreased postprandial lipemia, and faster clearance of radiolabeled VLDL (very-LDL) from circulation. Additionally, SGLT2 inhibition delayed turnover of labeled LDL from circulation. Conclusions— Our studies in diabetic CETP-ApoB100 transgenic mice recapitulate many of the changes in circulating lipids found with SGLT2 inhibition therapy in humans and suggest that the increased LDL cholesterol found with this therapy is because of reduced clearance of LDL from the circulation and greater lipolysis of triglyceride-rich lipoproteins. Most prominent effects of SGLT2 inhibition in the current mouse model were seen with antisense oligonucleotides-mediated knockdown of SGLT2.

Dual peroxisome proliferator-activated receptor (PPAR)α/γ agonists that were developed to target hyperlipidemia and hyperglycemia in type 2 diabetes patients, caused cardiac dysfunction or other adverse effects. We studied the mechanisms that underlie the cardiotoxic effects of a dual PPARα/γ agonist, tesaglitazar, in wild type and diabetic (leptin receptor deficient - db/db) mice. Mice treated with tesaglitazar-containing chow or high fat diet developed cardiac dysfunction despite lower plasma triglycerides and glucose levels. Expression of cardiac peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), which promotes mitochondrial biogenesis, had the most profound reduction among various fatty acid metabolism genes. Furthermore, we observed increased acetylation of PGC1α, which suggests PGC1α inhibition and lowered sirtuin 1 (SIRT1) expression. This change was associated with lower mitochondrial abundance. Combined pharmacological activation of PPARα and PPARγ in C57BL/6 mice reproduced the reduction of PGC1α expression and mitochondrial abundance. Resveratrol-mediated SIRT1 activation attenuated tesaglitazar-induced cardiac dysfunction and corrected myocardial mitochondrial respiration in C57BL/6 and diabetic mice but not in cardiomyocyte-specific Sirt1-/- mice. Our data shows that drugs, which activate both PPARα and PPARγ lead to cardiac dysfunction associated with PGC1α suppression and lower mitochondrial abundance likely due to competition between these two transcription factors.

Objective: Although Sodium glucose cotransporter 2 (SGLT2) inhibition in humans sometimes leads to increased levels of low density lipoprotein (LDL) cholesterol, this therapy is associated with marked reduction in cardiovascular disease. In this study, we aimed to determine how SGLT2 inhibition alters circulating lipoproteins. Approach and Results: We used a mouse model in which plasma lipoprotein profiles were humanized by expression of human cholesteryl ester transfer protein and human apolipoprotein B100 to determine how SGLT2 inhibition alters lipoprotein profiles. The mice were fed a high fat diet and then were made partially insulin deficient using streptozotocin. SGLT2 was inhibited using a specific anti-sense oligonucleotide or canagliflozin, a clinically available oral SGLT2 inhibitor. Inhibition of SGLT2 increased circulating levels of LDL cholesterol and reduced plasma triglyceride levels. SGLT2 inhibition was associated with increased lipoprotein lipase activity in the post heparin plasma and decreased postprandial lipemia. The changes in lipoprotein metabolism were more significant in mice treated with SGLT2 ASO compared to canagliflozin. Conclusions: Our studies in mice recapitulate many of the changes in circulating lipids found with SGLT2 inhibition therapy in humans and suggest that the increased LDL cholesterol found with this therapy is due to greater lipolysis of triglyceride-rich lipoproteins. This change in lipoprotein physiology is likely, in part, to explain the reduced cardiovascular disease found with SGLT2 inhibition.

Cardiac metabolism affects systemic energetic balance. Previously, we showed that Krüppel-like factor (KLF)-5 regulates cardiomyocyte PPARα and fatty acid oxidation-related gene expression in diabetes. We surprisingly found that cardiomyocyte-specific KLF5 knockout mice (αMHC-KLF5(−/−)) have accelerated diet-induced obesity, associated with increased white adipose tissue (WAT). Alterations in cardiac expression of the mediator complex subunit 13 (Med13) modulates obesity. αMHC-KLF5(−/−) mice had reduced cardiac Med13 expression likely because KLF5 upregulates Med13 expression in cardiomyocytes. We then investigated potential mechanisms that mediate cross-talk between cardiomyocytes and WAT. High fat diet-fed αMHC-KLF5(−/−) mice had increased levels of cardiac and plasma FGF21, while food intake, activity, plasma leptin, and natriuretic peptides expression were unchanged. Consistent with studies reporting that FGF21 signaling in WAT decreases sumoylation-driven PPARγ inactivation, αMHC-KLF5(−/−) mice had less SUMO-PPARγ in WAT. Increased diet-induced obesity found in αMHC-KLF5(−/−) mice was absent in αMHC-[KLF5(−/−);FGF21(−/−)] double knockout mice, as well as in αMHC-FGF21(−/−) mice that we generated. Thus, cardiomyocyte-derived FGF21 is a component of pro-adipogenic crosstalk between heart and WAT.

Movement of circulating fatty acids (FAs) to parenchymal cells requires their transfer across the endothelial cell (EC) barrier. The multiligand receptor cluster of differentiation 36 (CD36) facilitates tissue FA uptake and is expressed in ECs and parenchymal cells such as myocytes and adipocytes. Whether tissue uptake of FAs is dependent on EC or parenchymal cell CD36, or both, is unknown. Using a cell-specific deletion approach, we show that EC, but not parenchymal cell, CD36 deletion increased fasting plasma FAs and postprandial triglycerides. EC-Cd36-KO mice had reduced uptake of radiolabeled long-chain FAs into heart, skeletal muscle, and brown adipose tissue; these uptake studies were replicated using [11C]palmitate PET scans. High-fat diet-fed EC-CD36-deficient mice had improved glucose tolerance and insulin sensitivity. Both EC and cardiomyocyte (CM) deletion of CD36 reduced heart lipid droplet accumulation after fasting, but CM deletion did not affect heart glucose or FA uptake. Expression in the heart of several genes modulating glucose metabolism and insulin action increased with EC-CD36 deletion but decreased with CM deletion. In conclusion, EC CD36 acts as a gatekeeper for parenchymal cell FA uptake, with important downstream effects on glucose utilization and insulin action.

Peroxisome proliferator-activated receptor (PPAR) agonists are used in metabolic diseases. PPARα agonists target hyperlipidemia and PPARγ agonists correct hyperglycemia. Dual PPARα/γ agonists were developed to combine the benefits in type 2 diabetes patients. Despite their beneficial effect, PPARα/γ agonists caused cardiac dysfunction. We studied the mechanisms that underlie this toxic effect aiming to improve a failed therapy. Wild type mice were fed with chow and high fat diets containing Tesaglitazar (TESA) a dual PPARα/γ agonist for 6 weeks. Although, TESA lowered plasma triglyceride and glucose levels compared to controls, 2D-echo revealed severe cardiac dysfunction. Assessment of the expression of cardiac fatty acid metabolism genes that are regulated by PPARs showed that mRNA and protein levels of PPARγ-coactivator (PGC1α), a regulator of mitochondrial biogenesis, had the most profound reduction. Furthermore, we observed increased acetylation of PGC1α levels, which indicates lower activation and lower mitochondrial respiration. We then examined the mechanisms that underlie lower expression of PGC1α upon activation of PPARα and PPARγ. Thus, we administered C57BL/6 mice with both single PPARα and PPARγ agonists (WY14643 and rosiglitazone). This treatment reproduced the reduction in PGC1α expression that we observed with TESA. Moreover, mitochondria abundance was lower. Luciferase promoter analysis in a human cardiomyocyte cell line showed that PPARα and PPARγ compete for binding on PPAR elements of the human Pgc1a promoter, as well as that PPARα binding compromises PPARγ-mediated activation of the Pgc1α promoter. Aiming to decipher the mechanism via which dual PPARα/γ activation increases acetylation and thus inhibits PGC1α, we measured cardiac protein levels of the deacetylase sirtuin 1 (SIRT1), which were lower in primary cardiomyocytes from TESA-treated mice. We then treated mice with diet containing TESA and Resveratrol (RSV), which stimulates SIRT1. RSV attenuated TESA-driven cardiac dysfunction and corrected mitochondrial respiratory rates in primary cardiomyocytes. We propose that SIRT1-mediated activation of PGC1α blunts the cardiotoxic effect of dual PPARα/γ agonists and improves their therapeutic potential.

Lipid accumulation is a pathological feature of every type of kidney injury. Despite this striking histological feature, physiological accumulation of lipids in the kidney is poorly understood. We studied whether the accumulation of lipids in the fasted kidney are derived from lipoproteins or NEFAs. With overnight fasting, kidneys accumulated triglyceride, but had reduced levels of ceramide and glycosphingolipid species. Fasting led to a nearly 5-fold increase in kidney uptake of plasma [14C]oleic acid. Increasing circulating NEFAs using a β adrenergic receptor agonist caused a 15-fold greater accumulation of lipid in the kidney, while mice with reduced NEFAs due to adipose tissue deficiency of adipose triglyceride lipase had reduced triglycerides. Cluster of differentiation (Cd)36 mRNA increased 2-fold, and angiopoietin-like 4 (Angptl4), an LPL inhibitor, increased 10-fold. Fasting-induced kidney lipid accumulation was not affected by inhibition of LPL with poloxamer 407 or by use of mice with induced genetic LPL deletion. Despite the increase in CD36 expression with fasting, genetic loss of CD36 did not alter fatty acid uptake or triglyceride accumulation. Our data demonstrate that fasting-induced triglyceride accumulation in the kidney correlates with the plasma concentrations of NEFAs, but is not due to uptake of lipoprotein lipids and does not involve the fatty acid transporter, CD36.

A prominent histological feature of chronic kidney disease, including that due to diabetes, is the accumulation of neutral lipids in the nephron. However, existing research on kidney lipid metabolism is limited. Therefore we characterized lipid accumulation in mouse models. We hypothesized that the major source of lipids in the kidney is circulating triglyceride (TG)-rich lipoproteins brought into the cells of the kidney via the coordinated action of lipoprotein lipase (LPL) and cluster of differentiation 36 (CD36). To test this hypothesis, we fasted mice overnight and inhibited LPL and CD36. We did not observe a change in TG accumulation in the kidney when LPL action was blocked using with poloxmer 407. We then used a microsomal TG transfer protein inhibitor, which inhibits TG-rich lipoprotein assembly, to lower circulating TG-rich lipoproteins. Intracellular kidney TG did not decrease but in fact increased up to 40% in the fasted state after the inhibitor was given, mirroring what was seen in the liver. There was no difference in TG accumulation in the kidneys of fasted Cd36 -/- mice. Furthermore, we did not observe a change in uptake of radiolabeled oleic acid in the kidneys of these mice. Genes which regulate fatty acid oxidation or de novo lipogenesis were also unchanged in Cd36 -/- compared to Cd36 +/+ . Surprisingly, apoC-III deficiency increased renal uptake of a TG-emulsion, as had been found in the liver. Finally, we measured TG in kidneys from streptozotocin-treated and Akita diabetic mice without renal failure. Neither of these models accumulated higher amounts of TG compared to non-diabetic controls; however, Akita mice down regulated fatty acid oxidation genes and had twice the intracellular free fatty acids. These findings provide novel insight into the lipid biochemistry of the kidney, showing it differs from other high energy requiring peripheral organs such as heart.

Objective— Diabetic hypertriglyceridemia is thought to be primarily driven by increased hepatic de novo lipogenesis. However, experiments in animal models indicated that insulin deficiency should decrease hepatic de novo lipogenesis and reduce plasma triglyceride levels. Approach and Results— To address the discrepancy between human data and genetically altered mouse models, we investigated whether insulin-deficient diabetic mice had triglyceride changes that resemble those in diabetic humans. Streptozotocin-induced insulin deficiency increased plasma triglyceride levels in mice. Contrary to the mouse models with impaired hepatic insulin receptor signaling, insulin deficiency did not reduce hepatic triglyceride secretion and de novo lipogenesis-related gene expression. Diabetic mice had a marked decrease in postprandial triglycerides clearance, which was associated with decreased lipoprotein lipase and peroxisome proliferator-activated receptor α mRNA levels in peripheral tissues and decreased lipoprotein lipase activity in skeletal muscle, heart, and brown adipose tissue. Diabetic heterozygous lipoprotein lipase knockout mice had markedly elevated fasting plasma triglyceride levels and prolonged postprandial triglycerides clearance. Conclusions— Insulin deficiency causes hypertriglyceridemia by decreasing peripheral lipolysis and not by an increase in hepatic triglycerides production and secretion.