Faculty Bibliography

Guidelines to reduce cardiovascular risk in diabetes include aggressive LDL lowering, but benefits are attenuated compared to those in patients without diabetes. Consistent with this, we have reported in mice that hyperglycemia impaired atherosclerosis regression. Aldose reductase (AR) is thought to contribute to clinical complications of diabetes by directing glucose into pathways producing inflammatory metabolites. Mice have low levels of AR, thus, raising them to human levels would be a more clinically relevant model to study changes in diabetes under atherosclerosis regression conditions. Donor aortae from western diet-fed Ldlr

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.

During reduced energy intake, skeletal muscle maintains homeostasis by rapidly suppressing insulin-stimulated glucose utilization. Loss of this adaptation is observed with deficiency of the fatty acid transporter CD36. A similar loss is also characteristic of the insulin resistant state where CD36 is dysfunctional. To elucidate what links CD36 to muscle glucose utilization we examined whether CD36 signaling might influence insulin action. First, we show that CD36 deletion specific to skeletal muscle reduces expression of insulin signaling and glucose metabolism genes. It decreases muscle ceramides but impairs glucose disposal during a meal. Second, in primary-derived human myotubes depletion of CD36 suppresses insulin signaling and the mechanism is shown to involve functional CD36 interaction with the insulin receptor (IR). CD36 promotes tyrosine phosphorylation of IR by the Fyn kinase, enhances IR recruitment of P85 and downstream signaling. Third, pretreatment for 15 minutes with saturated fatty acids suppresses CD36-Fyn enhancement of IR phosphorylation, while unsaturated fatty acids are neutral or stimulatory. These findings define mechanisms important for muscle glucose metabolism and optimal insulin responsiveness. Potential human relevance is suggested by GWA RNA-Seq data that associate genetically determined low muscle CD36 expression to incidence of diabetes type 2.

Excess plasma triglycerides (TGs) are a key component of obesity-induced metabolic syndrome. We have shown that γ-secretase inhibitor (GSI) treatment improves glucose tolerance due to inhibition of hepatic Notch signaling but found additional Notch-independent reduction of plasma TG-rich lipoproteins (TRLs) in GSI-treated, as well as hepatocyte-specific, γ-secretase knockout (L-Ncst) mice, which suggested a primary effect on hepatocyte TRL uptake. Indeed, we found increased VLDL and LDL particle uptake in L-Ncst hepatocytes and Ncst-deficient hepatoma cells, in part through reduced γ-secretase-mediated low-density lipoprotein receptor (LDLR) cleavage and degradation. To exploit this novel finding, we generated a liver-selective Nicastrin ASO, which recapitulated glucose and lipid improvements of L-Ncst mice, with increased levels of hepatocyte LDLR. Collectively, these results identify the role of hepatic γ-secretase to regulate LDLR and suggest that liver-specific GSIs may simultaneously improve multiple aspects of the metabolic syndrome.

Cholesterol is not the only lipid that causes heart disease. Triglyceride supplies the heart and skeletal muscles with highly efficient fuel and allows for the storage of excess calories in adipose tissue. Failure to transport, acquire, and use triglyceride leads to energy deficiency and even death. However, overabundance of triglyceride can damage and impair tissues. Circulating lipoprotein-associated triglycerides are lipolyzed by lipoprotein lipase (LpL) and hepatic triglyceride lipase. We inhibited these enzymes and showed that LpL inhibition reduces high-density lipoprotein cholesterol by >50%, and hepatic triglyceride lipase inhibition shifts low-density lipoprotein to larger, more buoyant particles. Genetic variations that reduce LpL activity correlate with increased cardiovascular risk. In contrast, macrophage LpL deficiency reduces macrophage function and atherosclerosis. Therefore, muscle and macrophage LpL have opposite effects on atherosclerosis. With models of atherosclerosis regression that we used to study diabetes mellitus, we are now examining whether triglyceride-rich lipoproteins or their hydrolysis by LpL affect the biology of established plaques. Following our focus on triglyceride metabolism led us to show that heart-specific LpL hydrolysis of triglyceride allows optimal supply of fatty acids to the heart. In contrast, cardiomyocyte LpL overexpression and excess lipid uptake cause lipotoxic heart failure. We are now studying whether interrupting pathways for lipid uptake might prevent or treat some forms of heart failure.

BACKGROUND AND AIMS/OBJECTIVE:Leukocytosis, particularly monocytosis, has been shown to promote atherosclerosis in both diabetic and non-diabetic mouse models. We previously showed that hyperglycemia independently promotes monocytosis and impairs the resolution of atherosclerosis. Since patients with chronic diabetes often develop dyslipidemia and also have increased risk for atherosclerosis, we sought to examine how controlling blood glucose affects atherosclerosis development in the presence of severe hyperlipidemia. METHODS:) mice after which they were fed a high-cholesterol diet for 4 weeks. Control and diabetic mice were treated with vehicle or sodium glucose cotransporter inhibitor (SGLT2i, Phlorizin or Dapagliflozin) for the duration of the diet. RESULTS:Induction of diabetes resulted in a dramatic increase in plasma cholesterol (TC) and triglyceride (TG) levels. These mice also exhibited an increased number of circulating monocytes and neutrophils. Monocytosis was driven by increased proliferation of progenitor cells in the bone marrow. Tighter glycemic control by SGLT2i treatment not only reduced monocytosis and atherosclerosis but also improved plasma lipoprotein profile. Interestingly, improved lipoprotein profile was not due to decreased TG synthesis or clearance via low density lipoprotein receptor-related protein (Lrp) 1 or scavenger receptor class B member (Scarb1) pathways, but likely mediated by heparin sulfate proteoglycans (HSPG)-dependent clearance mechanisms in the liver. Further examination of the liver revealed an important role for bile acid transporters (Abcg5, Abcg8) and cytochrome P450 enzymes in the clearance of hepatic cholesterol. CONCLUSIONS:These data suggest that tighter glycemic control in diabetes can improve lipoprotein clearance exclusive of Ldlr, likely via HSPG and bile acid pathways, and has an overall net positive effect on atherosclerosis.

Insulin signaling in the central nervous system (CNS) influences satiety, counterregulation, and peripheral insulin sensitivity. However, the broad distribution of insulin receptors (InsR) within neurons and glia has hampered mapping of specific neuronal sub-populations that mediate specific effects of insulin. Neurons expressing glucose transporter Glut4 influence peripheral insulin sensitivity. Here, we analyzed the effects of InsR signaling in hypothalamic Glut4 neurons on glucose sensing as well as leptin and amino acid signaling. We show that InsR signaling in Glut4 neurons dampens the glucagon response to hypoglycemia and 2-deoxyglucose-induced neuroglycopenia. Using immunohistochemistry in chemically identified hypothalamic Glut4 neurons, we show that InsR signaling promotes Akt signaling in response to insulin, leptin, and amino acids in a cell-autonomous fashion (i.e., in Glut4 neurons), but also generates an inhibitory signal to reduce Akt function in non-Glut4-neurons. We conclude that hypothalamic Glut4 neurons modulate the glucagon counterregulatory response, and that InsR signaling in Glut4 neurons is required to integrate hormonal and nutritional cues for the regulation of glucose metabolism

OBJECTIVE:Tissue macrophages induce and perpetuate proinflammatory responses, thereby promoting metabolic and cardiovascular disease. Lipoprotein lipase (LpL), the rate-limiting enzyme in blood triglyceride catabolism, is expressed by macrophages in atherosclerotic plaques. We questioned whether LpL, which is also expressed in the bone marrow (BM), affects circulating white blood cells and BM proliferation and modulates macrophage retention within the artery. APPROACH AND RESULTS/UNASSIGNED:We characterized blood and tissue leukocytes and inflammatory molecules in transgenic LpL knockout mice rescued from lethal hypertriglyceridemia within 18 hours of life by muscle-specific LpL expression (MCKL0 mice). LpL-deficient mice had ≈40% reduction in blood white blood cell, neutrophils, and total and inflammatory monocytes (Ly6C/Ghi). LpL deficiency also significantly decreased expression of BM macrophage-associated markers (F4/80 and TNF-α), master transcription factors (PU.1 and C/EBPα), and colony-stimulating factors (CSFs) and their receptors, which are required for monocyte and monocyte precursor proliferation and differentiation. As a result, differentiation of macrophages from BM-derived monocyte progenitors and monocytes was decreased in MCKL0 mice. Furthermore, although LpL deficiency was associated with reduced BM uptake and accumulation of triglyceride-rich particles and macrophage CSF-macrophage CSF receptor binding, triglyceride lipolysis products (eg, linoleic acid) stimulated expression of macrophage CSF and macrophage CSF receptor in BM-derived macrophage precursor cells. Arterial macrophage numbers decreased after heparin-mediated LpL cell dissociation and by genetic knockout of arterial LpL. Reconstitution of LpL-expressing BM replenished aortic macrophage density. CONCLUSIONS:LpL regulates peripheral leukocyte levels and affects BM monocyte progenitor differentiation and aortic macrophage accumulation.