Andrea Wan

FoxO1 is crucial for sustaining cardiomyocyte metabolism and cell survival

Diabetic cardiomyopathy is a term used to describe cardiac muscle damage-induced heart failure. Multiple structural and biochemical reasons have been suggested to induce this disorder. The most prominent feature of the diabetic myocardium is attenuated insulin signalling that reduces survival kinases (Akt), potentially switching on protein targets like FoxOs, initiators of cell death. FoxO1, a prominent member of the forkhead box family and subfamily O of transcription factors and produced from the FKHR gene, is involved in regulating metabolism, cell proliferation, oxidative stress response, immune homeostasis, pluripotency in embryonic stem cells, and cell death. In this review we describe distinctive functions of FoxOs, specifically FoxO1 under conditions of nutrient excess, insulin resistance and diabetes, and its manipulation to restore metabolic equilibrium to limit cardiac damage due to cell death. Because FoxO1 helps cardiac tissue to combat a variety of stress stimuli, it could be a major determinant in regulating diabetic cardiomyopathy. In this regard, we highlight studies from our group and others who illustrate how cardiac tissue-specific FoxO1 deletion protects the heart against cardiomyopathy and how its down-regulation in endothelial tissue could prevent against atherosclerotic plaques. In addition, we also describe studies that show FoxO1s beneficial qualities by highlighting their role in inducing anti-oxidant, autophagic, and anti-apoptotic genes under stress conditions of ischaemia-reperfusion and myocardial infarction. Thus, the aforementioned FoxO1 traits could be useful in curbing cardiac tissue-specific impairment of function following diabetes.

Diabetes triggers a PARP1 mediated death pathway in the heart through participation of FoxO1

Cardiomyocyte cell death is a major contributing factor for diabetic cardiomyopathy, and multiple mechanisms have been proposed for its development. We hypothesized that following diabetes, an increased nuclear presence of the Forkhead transcription factor, FoxO1, could turn on cardiac cell death through mediation of nitrosative stress. Streptozotocin (100 mg/kg) was used to induce irreversible hyperglycemia in Wistar rats, and heart tissues and blood samples extracted starting from 1 to 4 days. Diazoxide (100 mg/kg), which produced acute reversible hyperglycemia, were followed for up to 12 h. In both animal models of hyperglycemia, attenuation of survival signals was accompanied by increased nuclear FoxO1. This was accompanied by a simultaneous increase in iNOS expression and iNOS induced protein nitrosylation of GAPDH, increased GAPDH binding to Siah1 and facilitated nuclear translocation of the complex. Even though caspase-3 was cleaved during diabetes, its nitrosylation modification affected its ability to inactivate PARP. As a result, there was PARP activation followed by nuclear compartmentalization of AIF, and increased phosphatidyl serine externalization. Our data suggests a role for FoxO1 mediated iNOS induced S-nitrosylation of target proteins like GAPDH and caspase-3 in initiating cardiac cell death following hyperglycemia, and could explain the impact of glycemic control in preventing cardiovascular disease in patients with diabetes.

Endothelial Heparanase Regulates Heart Metabolism by Stimulating Lipoprotein Lipase Secretion From Cardiomyocytes

Objective—After diabetes mellitus, transfer of lipoprotein lipase (LPL) from cardiomyocytes to the coronary lumen increases, and this requires liberation of LPL from the myocyte surface heparan sulfate proteoglycans with subsequent replenishment of this reservoir. At the lumen, LPL breaks down triglyceride to meet the increased demand of the heart for fatty acid. Here, we examined the contribution of coronary endothelial cells (ECs) toward regulation of cardiomyocyte LPL secretion. Approach and Results—Bovine coronary artery ECs were exposed to high glucose, and the conditioned medium was used to treat cardiomyocytes. EC-conditioned medium liberated LPL from the myocyte surface, in addition to facilitating its replenishment. This effect was attributed to the increased heparanase content in EC-conditioned medium. Of the 2 forms of heparanase secreted from EC in response to high glucose, active heparanase released LPL from the myocyte surface, whereas latent heparanase stimulated reloading of LPL from an intracellular pool via heparan sulfate proteoglycan–mediated RhoA activation. Conclusions—Endothelial heparanase is a participant in facilitating LPL increase at the coronary lumen. These observations provide an insight into the cross-talk between ECs and cardiomyocytes to regulate cardiac metabolism after diabetes mellitus.

Hyperglycemia-Induced Secretion of Endothelial Heparanase Stimulates a Vascular Endothelial Growth Factor Autocrine Network in Cardiomyocytes That Promotes Recruitment of Lipoprotein Lipase

Objective—During diabetes mellitus, coronary lipoprotein lipase increases to promote the predominant use of fatty acids. We have reported that high glucose stimulates active heparanase secretion from endothelial cells to cleave cardiomyocyte heparan sulfate and release bound lipoprotein lipase for transfer to the vascular lumen. In the current study, we examined whether heparanase also has a function to release cardiomyocyte vascular endothelial growth factor (VEGF), and whether this growth factor influences cardiomyocyte fatty acid delivery in an autocrine manner. Approach and Results—Acute, reversible hyperglycemia was induced in rats, and a modified Langendorff heart perfusion was used to separate the coronary perfusate from the interstitial effluent. Coronary artery endothelial cells were exposed to high glucose to generate conditioned medium, and VEGF release from isolated cardiomyocytes was tested using endothelial cell conditioned medium or purified active and latent heparanase. Autocrine signaling of myocyte-derived VEGF on cardiac metabolism was studied. High glucose promoted latent and active heparanase secretion into endothelial cell conditioned medium, an effective stimulus for releasing cardiomyocyte VEGF. Intriguingly, latent heparanase was more efficient than active heparanase in releasing VEGF from a unique cell surface pool. VEGF augmented cardiomyocyte intracellular calcium and AMP-activated protein kinase phosphorylation and increased heparin-releasable lipoprotein lipase. Conclusions—Our data suggest that the heparanase-lipoprotein lipase-VEGF axis amplifies fatty acid delivery, a rapid and adaptive mechanism that is geared to overcome the loss of glucose consumption by the diabetic heart. If prolonged, the resultant lipotoxicity could lead to cardiovascular disease in humans.

Endothelial cell–cardiomyocyte crosstalk in diabetic cardiomyopathy

The incidence of diabetes is increasing globally, with cardiovascular disease accounting for a substantial number of diabetes-related deaths. Although atherosclerotic vascular disease is a primary reason for this cardiovascular dysfunction, heart failure in patients with diabetes might also be an outcome of an intrinsic heart muscle malfunction, labelled diabetic cardiomyopathy. Changes in cardiomyocyte metabolism, which encompasses a shift to exclusive fatty acid utilization, are considered a leading stimulus for this cardiomyopathy. In addition to cardiomyocytes, endothelial cells (ECs) make up a significant proportion of the heart, with the majority of ATP generation in these cells provided by glucose. In this review, we will discuss the metabolic machinery that drives energy metabolism in the cardiomyocyte and EC, its breakdown following diabetes, and the research direction necessary to assist in devising novel therapeutic strategies to prevent or delay diabetic heart disease.

Cardiomyocyte VEGF Regulates Endothelial Cell GPIHBP1 to Relocate Lipoprotein Lipase to the Coronary Lumen During Diabetes Mellitus

Objective— Lipoprotein lipase (LPL)–mediated triglyceride hydrolysis is the major source of fatty acid for cardiac energy. LPL, synthesized in cardiomyocytes, is translocated across endothelial cells (EC) by its transporter glycosylphosphatidylinositol-anchored high-density lipoprotein–binding protein 1 (GPIHBP1). Previously, we have reported an augmentation in coronary LPL, which was linked to an increased expression of GPIHBP1 following moderate diabetes mellitus. We examined the potential mechanism by which hyperglycemia amplifies GPIHBP1. Approach and Results— Exposure of rat aortic EC to high glucose induced GPIHBP1 expression and amplified LPL shuttling across these cells. This effect coincided with an elevated secretion of heparanase. Incubation of EC with high glucose or latent heparanase resulted in secretion of vascular endothelial growth factor (VEGF). Primary cardiomyocytes, being a rich source of VEGF, when cocultured with EC, restored EC GPIHBP1 that is lost because of cell passaging. Furthermore, recombinant VEGF induced EC GPIHBP1 mRNA and protein expression within 24 hours, an effect that could be prevented by a VEGF neutralizing antibody. This VEGF-induced increase in GPIHBP1 was through Notch signaling that encompassed Delta-like ligand 4 augmentation and nuclear translocation of the Notch intracellular domain. Finally, cardiomyocytes from severely diabetic animals exhibiting attenuation of VEGF were unable to increase EC GPIHBP1 expression and had lower LPL activity at the vascular lumen in perfused hearts. Conclusion— EC, as the first responders to hyperglycemia, can release heparanase to liberate myocyte VEGF. This growth factor, by activating EC Notch signaling, is responsible for facilitating GPIHBP1-mediated translocation of LPL across EC and regulating LPL-derived fatty acid delivery to the cardiomyocytes.

Endothelial Cell Heparanase Taken Up by Cardiomyocytes Regulates Lipoprotein Lipase Transfer to the Coronary Lumen After Diabetes

After diabetes, the heart has a singular reliance on fatty acid (FA) for energy production, which is achieved by increased coronary lipoprotein lipase (LPL) that breaks down circulating triglycerides. Coronary LPL originates from cardiomyocytes, and to translocate to the vascular lumen, the enzyme requires liberation from myocyte surface heparan sulfate proteoglycans (HSPGs), an activity that needs to be sustained after chronic hyperglycemia. We investigated the mechanism by which endothelial cells (EC) and cardiomyocytes operate together to enable continuous translocation of LPL after diabetes. EC were cocultured with myocytes, exposed to high glucose, and uptake of endothelial heparanase into myocytes was determined. Upon uptake, the effect of nuclear entry of heparanase was also investigated. A streptozotocin model of diabetes was used to expand our in vitro observations. In high glucose, EC-derived latent heparanase was taken up by cardiomyocytes by a caveolae-dependent pathway using HSPGs. This latent heparanase was converted into an active form in myocyte lysosomes, entered the nucleus, and upregulated gene expression of matrix metalloproteinase-9. The net effect was increased shedding of HSPGs from the myocyte surface, releasing LPL for its onwards translocation to the coronary lumen. EC-derived heparanase regulates the ability of the cardiomyocyte to send LPL to the coronary lumen. This adaptation, although acutely beneficial, could be catastrophic chronically because excess FA causes lipotoxicity. Inhibiting heparanase function could offer a new strategy for managing cardiomyopathy observed after diabetes.

Loss of VEGFB and its signaling in the diabetic heart is associated with increased cell death signaling

Vascular endothelial growth factor B (VEGFB) is highly expressed in metabolically active tissues, such as the heart and skeletal muscle, suggesting a function in maintaining oxidative metabolic and contractile function in these tissues. Multiple models of heart failure have indicated a significant drop in VEGFB. However, whether there is a role for decreased VEGFB in diabetic cardiomyopathy is currently unknown. Of the VEGFB located in cardiomyocytes, there is a substantial and readily releasable pool localized on the cell surface. The immediate response to high glucose and the secretion of endothelial heparanase is the release of this surface-bound VEGFB, which triggers signaling pathways and gene expression to influence endothelial cell (autocrine action) and cardiomyocyte (paracrine effects) survival. Under conditions of hyperglycemia, when VEGFB production is impaired, a robust increase in vascular endothelial growth factor receptor (VEGFR)-1 expression ensues as a possible mechanism to enhance or maintain VEGFB signaling. However, even with an increase in VEGFR1 after diabetes, cardiomyocytes are unable to respond to VEGFB. In addition to the loss of VEGFB production and signaling, evaluation of latent heparanase, the protein responsible for VEGFB release, also showed a significant decline in expression in whole hearts from animals with chronic or acute diabetes. Defects in these numerous VEGFB pathways were associated with an increased cell death signature in our models of diabetes. Through this bidirectional interaction between endothelial cells (which secrete heparanase) and cardiomyocytes (which release VEGFB), this growth factor could provide the diabetic heart protection against cell death and may be a critical tool to delay or prevent cardiomyopathy.NEW & NOTEWORTHY We discovered a bidirectional interaction between endothelial cells (which secrete heparanase) and cardiomyocytes [which release vascular endothelial growth factor B (VEGFB)]. VEGFB promoted cell survival through ERK and cell death gene expression. Loss of VEGFB and its downstream signaling is an early event following hyperglycemia, is sustained with disease progression, and could explain diabetic cardiomyopathy.

Lipoprotein lipase and angiopoietin-like 4 – Cardiomyocyte secretory proteins that regulate metabolism during diabetic heart disease

Abstract Cardiac diseases have been extensively studied following diabetes and altered metabolism has been implicated in its initiation. In this context, there is a shift from glucose utilization to predominantly fatty acid metabolism. We have focused on the micro- and macro-environments that the heart uses to provide fatty acids to the cardiomyocyte. Specifically, we will discuss the cross talk between endothelial cells, smooth muscles and cardiomyocytes, and their respective secretory products that allows for this shift in metabolism. These changes will then be linked to alterations in the cardiovascular system and the augmented heart disease observed during diabetes. Traditionally, the heart was only thought of as an organ that supplies oxygen and nutrients to the body through its function as a pump. However, the heart as an endocrine organ has also been suggested. Secreted products from the cardiomyocytes include the natriuretic peptides atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). Both have been shown to have vasodilatory, diuretic and antihypertensive effects. These peptides have been extensively studied and their deficiency is considered to be a major cause for the initiation of cardiovascular and cardiometabolic disorders. Another secretory enzyme, lipoprotein lipase (LPL), has been implicated in diabetic heart disease. LPL is a triglyceride-hydrolyzing enzyme that is synthesized within the cardiomyocyte and secreted towards the lumen under various conditions. For example, moderate or short-term hyperglycemia stimulates the release of LPL from the cardiomyocytes towards the endothelial cells. This process allows LPL to contact lipoprotein triglycerides, initiating their break down, with the product of lipolysis (free fatty acids, FA) translocating towards the cardiomyocytes for energy consumption. This mechanism compensates for the lack of glucose availability following diabetes. Under prolonged, chronic conditions of hyperglycemia, there is a need to inhibit this mechanism to avoid the excess delivery of FA to the cardiomyocytes, an effect that is known to induce cardiac cell death. Thus, LPL inhibition is made possible by a FA-induced activation of PPAR β/δ, which augments angiopoietin-like 4 (Angptl4), an inhibitor of LPL activity. In the current review, we will focus on the mediators and conditions that regulate LPL and Angptl4 secretion from the cardiomyocyte, which are critical for maintaining cardiac metabolic homeostasis.

The Function of Heparanase in Diabetes and its Complications

Heparan sulfate proteoglycans are ubiquitous glycoproteins that contain several heparan sulfate polysaccharide side chains attached to a core protein. They function not only as a primary structural component of the extracellular matrix, but also provide a storage depot for bioactive molecules, such as basic fibroblast growth factor, vascular endothelial growth factor and lipoprotein lipase. Heparanase is an endoglycosidase that specifically hydrolyzes heparan sulfate into oligosaccharides. Recent studies have indicated that heparanase is engaged in the initiation and progression of diabetes, in addition to its associated complications. This review focuses on the participation of heparanase in the cleavage of heparan sulfate proteoglycans in pancreatic islets promoting beta cell death, promotion of atherosclerosis, and its role in cardiac metabolic switching in the early stage of cardiomyopathy during diabetes. Understanding the mechanisms by which heparanase is regulated in diabetes could provide a drug target to prevent diabetes and its complications.