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Glucose transport in the heart
Abstract
The heart is a unique organ in many ways. It consists of specialized muscle cells (cardiomyocytes), which are adapted to contract constantly in a coordinated fashion. This is vital to the survival of the organism given the central role of the heart in the maintenance of the cardiovascular system that delivers oxygen, metabolic substrates and hormones to the rest of the body. In order for the heart to maintain its function it must receive a constant supply of metabolic substrates, to generate ATP to maintain contractile function, without fatigue. Thus the heart is capable of utilizing a variety of metabolic substrates and is able to rapidly adapt its substrate utilization in the face of changes in substrate supply. The major metabolic substrate for the heart is fatty acids. However, up to 30% of myocardial ATP is generated by glucose and lactate, with smaller contributions from ketones and amino acids. Although glucose is not the major metabolic substrate in the heart at rest, there are many circumstances in which it assumes greater importance such as during ischemia, increased workload and pressure overload hypertrophy. Like all other cells, glucose is transported into cardiac myocytes by members of the family of facilitative glucose transporters (GLUTs). In this regard, cardiomyocytes bear many similarities to skeletal muscle, but there are also important differences. For example, the most abundant glucose transporter in the heart is the GLUT4 transporter, in which translocation to the plasma membrane represents an important mechanism by which the net flux of glucose into the cell is regulated. Because cardiomyocytes are constantly contracting it is likely that contraction mediated GLUT4 translocation represents an important mechanism that governs the entry of glucose into the heart. While this is also true in skeletal muscle, because many muscles are often at rest, insulin mediated GLUT4 translocation represents a quantitatively more important mechanism regulating skeletal muscle glucose uptake than is the case in the heart. In contrast to skeletal muscle, where most GLUT1 is in perineural sheaths (1), in the heart there is significant expression of GLUT1 (2), which under certain circumstances is responsible for a significant component of basal cardiac glucose uptake. This review will summarize the current state of knowledge regarding the regulation of glucose transporter expression, and the regulation of glucose transport into myocardial cells.
... The distribution of glucose throughout the body to be used as fuel to create energy is made by the blood and mediated/controlled/impacted mainly by organs such as liver, intestine, pancreas, kidney, heart, and Brain/HPA axis [25] [26] [27]. The glucose circulating level is impacted by external and internal factors such as infections, stress, fears and phobias, physical and intellectual activities, etc. Internally, the largest customers for glucose consumption are the brain (in its multiple cognitive and emotional functions) and the heart, and of course, the skeletal muscles, most of them with binary functions of customer and "service" provider [28] [29]. ...
... In this scenario, the triad glucose -insulin-ATP has to work synergetically and perfectly well with cells in order to permit the creation, distribution, and storage of energy, which is of paramount importance for normal life dynamics [6] [25]. ...
... Glucose is the fuel for the cells energy generation, but it only turns to ATP/ energy inside the cells. ATP can be used to store energy for future reactions or be taken out to pay for reactions when energy is required by the cells [25]. In this scenario, glucose and insulin are distributed throughout the body to power each and every cell of our body giving us the sensation of energy and well-being. ...
... Specifically, insulin causes GLUT4 translocation and increases the activation of GLUT4 transporters in the heart. Insulin also stimulates the translocation of GLUT1 in cardiomyocytes [52]. ...
... GLUT1 is mainly dominant in the embryonic heart, but the ratio of GLUT1/GLUT4 becomes level shortly after birth [52]. In the heart, the main regulators of GLUT1 are the SP1 and SP3 transcription factors. ...
... At the same time, the GLUT4 expression levels remain the same, which indicates that GLUT4 is the heart's main regulator of insulin-mediated glucose uptake. However, there have been circumstances when GLUT1 levels increase, as, for example, in rats with chronic left ventricular hypertrophy (LVH) and during inflammatory myocarditis [52]. ...
This year, 2022, marks the 100th anniversary of the isolation of human insulin and its administration to patients suffering from diabetes mellitus (DM). Insulin exerts many effects on the human body, including the cardiac tissue. The pathways implicated include the PKB/Akt signaling pathway, the Janus kinase, and the mitogen-activated protein kinase pathway and lead to normal cardiac growth, vascular smooth muscle regulation, and cardiac contractility. This review aims to summarize the existing knowledge and provide new insights on insulin pathways of cardiac tissue, along with the role of left ventricular assist devices on insulin regulation and cardiac function.
... Introduction Diabetes mellitus is a condition of metabolic imbalance, indicated by a high level of blood glucose (hyperglycemia) resulting from a reduction of insulin secretion, action, or both. [1][2][3] Generally, diabetes mellitus is divided into two types based on the etiology: Type 1 (T1D) and Type 2 diabetes (T2D). In type 1 diabetes, the pancreas cannot produce insulin as the pancreatic β-cells of the islets are destructed; therefore, regular insulin injections are essential in maintaining blood glucose levels in the body. ...
... In type 1 diabetes, the pancreas cannot produce insulin as the pancreatic β-cells of the islets are destructed; therefore, regular insulin injections are essential in maintaining blood glucose levels in the body. [1,3] In T2D, insufficient insulin is produced by the pancreas, or the tissues have become insulin resistant and do not adequately maintain glucose homeostasis. The management of T2D includes decreased calorie intake, increased exercises, oral medication, and insulin administration. ...
... The glucose absorption in the cardiac muscle is facilitated by GLUT 1 and GLUT 4 proteins. [1,25] Therefore, there is a hypoglycemic effect contained in the aqueous ginger extract. ...
Diabetes mellitus is a condition of metabolic imbalance, indicated by a high level of blood glucose (hyperglycemia) resulting from a reduction of insulin secretion, action, or both. People with diabetes suffer from a lack or deficiency of insulin or insulin resistance. The metabolic imbalances are often not satisfactorily corrected using conventional medicines and even cause some side effects, which can be detrimental. Research on herbal medicines for the treatment of diabetes is urged by the need to reduce unwanted side effects common with conventional medicines/treatments used in glucose regulation. This study aims to investigate the antidiabetic effect of Ginger ( Zingiber officinale ) aqueous extract in improving the glucose uptake in mouse tissues in vitro. This study is a true experimental research design with a posttest-only control group design. There were three groups of mice in this study: the control group, which were only given plain water; the second group of mice with 5% aqueous ginger extract and the last group were given 25% aqueous ginger extract. All groups were given treatment for four consecutive weeks, then dissected their cardiac muscle, skeletal muscle, pancreas, and liver tissues to analyze the glucose uptake. The result showed that both the ginger aqueous extract groups were able to increase the glucose uptake of the mice. In conclusion, this research has shown that aqueous ginger extract may have improved the glucose uptake in most tissues of the mice in the groups. Therefore, ginger could have great potential as an alternative way in the treatment of diabetes type 2.
... In the heart, glucose enters cardiomyocytes via glucose transporters (GLUTs) which are expressed by various cell types. Among 14 members of the GLUT family (53), the most abundant GLUTs in the human heart are the insulin-sensitive glucose transporter GLUT4 (54,55), and the insulin-independent glucose transporter GLUT1 (54,56). ...
... In the heart, glucose enters cardiomyocytes via glucose transporters (GLUTs) which are expressed by various cell types. Among 14 members of the GLUT family (53), the most abundant GLUTs in the human heart are the insulin-sensitive glucose transporter GLUT4 (54,55), and the insulin-independent glucose transporter GLUT1 (54,56). ...
Heart failure is the leading cause of death worldwide. The inability of the adult mammalian heart to regenerate following injury results in the development of systolic heart failure. Thus, identifying novel approaches toward regenerating the adult heart has enormous therapeutic potential for adult heart failure. Mitochondrial metabolism is an essential homeostatic process for maintaining growth and survival. The emerging role of mitochondrial metabolism in controlling cell fate and function is beginning to be appreciated. Recent evidence suggests that metabolism controls biological processes including cell proliferation and differentiation, which has profound implications during development and regeneration. The regenerative potential of the mammalian heart is lost by the first week of postnatal development when cardiomyocytes exit the cell cycle and become terminally differentiated. This inability to regenerate following injury is correlated with the metabolic shift from glycolysis to fatty acid oxidation that occurs during heart maturation in the postnatal heart. Thus, understanding the mechanisms that regulate cardiac metabolism is key to unlocking metabolic interventions during development, disease, and regeneration. In this review, we will focus on the emerging role of metabolism in cardiac development and regeneration and discuss the potential of targeting metabolism for treatment of heart failure.
... The remaining ATP production is accounted for by glycolysis and the oxidation of ketone bodies, lactate, and amino acids (Opie, 2014;Pascual & Coleman, 2016;Stanley et al., 2005;Szablewski, 2017). Glucose and lactate metabolism may account for up to 30% of myocardial ATP during basal state (Abel, 2004;Opie, 2014). Glucose enters the cardiomyocyte through specific glucose transmembrane transporters GLUT1 and GLUT4 (Shao & Tian, 2015;Szablewski, 2017). ...
... In a dog model of myocardial ischemia, GLUT-1 mRNA and polypeptide expression were increased in ischemic cardiomyocytes (Feldhaus & Liedtke, 1998). GLUT-1 upregulation has also been shown to occurs during critical illness, induced by cytokines and by the hypoxic-ischemic environment through the possible upregulation of the hypoxia-inducible factor-1α in the heart (Abel, 2004;Brosius et al., 1997;Chen et al., 2001;Dungan et al., 2009;Feldhaus & Liedtke, 1998;Shao & Tian, 2015). In the diabetic cardiomyopathy model, glucose overload results in the accumulation of glycolysis substrates that carry several metabolic fates. ...
Stress hyperglycemia is a transient increase in blood glucose during acute physiological stress in the absence of glucose homeostasis dysfunction. Its's presence has been described in critically ill patients who are subject to many physiological insults. In this regard, hyperglycemia and impaired glucose tolerance are also frequent in patients who are admitted to the intensive care unit for heart failure and cardiogenic shock. The hyperglycemia observed at the beginning of these cardiac disorders appears to be related to a variety of stress mechanisms. The release of major stress and steroid hormones, catecholamine overload, and glucagon all participate in generating a state of insulin resistance with increased hepatic glucose output and glycogen breakdown. In fact, the observed pathophysiological response, which appears to regulate a stress situation, is harmful because it induces mitochondrial impairment, oxidative stress‐related injury to cells, endothelial damage, and dysfunction of several cellular channels. Paradigms are now being challenged by growing evidence of a phenomenon called glucotoxicity, providing an explanation for the benefits of lowering glucose levels with insulin therapy in these patients. In the present review, the authors present the data published on cardiac glucotoxicity and discuss the benefits of lowering plasma glucose to improve heart function and to positively affect the course of critical illness. A review of the pathophysiological and clinical evidence surrounding the impact of stress hyperglycemia and cardiac glucotoxicity on cardiac physiology.
... The 1 st syllogism: Hyperglycaemia, insulinemia and intracellular glucose availability. Both GLUT1 and GLUT4 are present in cardiac myocytes (Abel, 2004;Zheng et al., 2021). The expression of the latter is dependent on insulin availability. ...
... Glucose metabolism contributes to the energy supply required for blood and oxygen delivery from the heart to other organs. Glucose uptake in cardiomyocytes is regulated by GLUTs, mainly GLUT1 and GLUT4 isoforms, which predominate in the fetal and adult heart, respectively [54][55][56][57][58][59]. In cardiomyocytes, glucose can activate different metabolic pathways: 1) polyol, 2) glycolysis, 3) pentose phosphate, and 4) hexosamine biosynthetic pathway [60]. ...
The main function of the renin-angiotensin-aldosterone system (RAAS) is the regulation of blood pressure; therefore, researchers have focused on its study to treat cardiovascular and renal diseases. One of the most widely used treatments derived from the study of RAAS, is the use of angiotensin-converting enzyme inhibitors (ACEi). Since it was discovered, the main target of ACEi has been the cardiovascular and renal systems. However, being the RAAS expressed locally in several specialized tissues and cells such as pneumocytes, hepatocytes, spleenocytes, enterocytes, adipocytes, and neurons the effect of inhibitors has expanded, because it is expected that RAAS has a role in the specific function of those cells. Many chronic degenerative diseases compromise the correct function of those organs, and in most of them, the RAAS is overactivated. Therefore, the use of ACEi must exert a benefit on an impaired system. Accordingly, the objective of this review is to present a brief overview of the cardiovascular and renal actions of ACEi and its effects in organs that are not the classic targets of ACEi that carry on glucose and lipid metabolism.
... 35 Glucose transporters 1 and 4 are the main isoforms involved in the transport of glucose into the cardiomyocytes during neonatal and adult stages, respectively. 36 Overexpression of GLUT1 promoted glycolysis and nucleotide synthesis and increased heart regeneration upon cryoinjury in juvenile and adult mice. 37 Apart from the role of glucose uptake and glycolysis in cardiomyocyte proliferation, glycolytic enzymes can play a direct role in heart regeneration. ...
Current therapies for heart failure aim to prevent the deleterious remodeling that occurs after MI injury, but currently no therapies are available to replace lost cardiomyocytes. Several organisms now being studied are capable of regenerating their myocardium by the proliferation of existing cardiomyocytes. In this review, we summarize the main metabolic pathways of the mammalian heart and how modulation of these metabolic pathways through genetic and pharmacological approaches influences cardiomyocyte proliferation and heart regeneration.
... 42 GLUT4 plays a key role in glucose uptake inducing elevation of cardiac contractility as well as increasing the amount of circulating insulin through the sarcolemma of cardiomyocytes. 43 ...
Background: Low availability of Glut-4 transporters in the sarcolemma of cardiac cells characterizes myocardial insulin resistance (MIR), which is triggered separately from generalized insulin resistance. Insulin receptors are quite evident in the heart muscle and vessels, and mitochondrial activity performs a significant role in MIR preserving cellular homeostasis through cell reproduction, cells livelihoods, and energy generation. Objective: To evaluate the MIR mechanism and its association with hypertension by signaling pathways design. Methods: PubMed database was employed to search for reviews publications with MIR. The referenced data of the signaling pathway was chosen by aggregating references from the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. A signaling pathway was designed based on MIR research manuscripts, where we show several mechanisms included in the MIR. The KEGG server was employed to exploit the interrelationship protein-protein, and elaborate signaling pathway diagram. The signaling pathway mapping was carried out with PathVisio software. Results: We selected 42 articles from a total of 450 articles in the PubMed database that presented a significant association between the terms “insulin resistance myocardial” AND “signaling pathway” AND “systemic arterial hypertension”. Founded on database-validated research papers, we chose well-founded pathways and we succeeded in representative description of these pathways. The reproduction contigs taken from the KEGG database designed the signaling pathway of the bio-molecules that lead to MIR. Thus, the acting among multiple mechanisms releases factors that participate in the development of MIR. Conclusion: The interaction among various mechanisms and molecular interactions are important factors in developing MIR.
... The process of glucose entry from the blood into the cell is the rate-limiting step in myocardial glucose metabolism; glucose enters cardiomyocytes through GLUTs and then undergoes glycolysis (Luiken and Glatz 2015;Till et al. 1997). The main Gluts in the myocardium are GLUT1 and GLUT4, and GLUT4 is the most abundant glucose transporter in the myocardium, the level of which is approximately 4× higher than that of GLUT1; GLUT4 is the main Gluts by which cardiomyocytes regulate glucose uptake (Abel 2004). Under physiological conditions, more than 90% of GLUT4 is distributed in intracellular vesicle-like structures, such as microsomes, Golgi complexes, and tubular vesicles (Richter and Hargreaves 2013;Luiken and Glatz 2015). ...
Exercise preconditioning (EP) is a line of scientific inquiry into the short-term biochemical mediators of cardioprotection in the heart. This study examined the involvement of autophagy induced by energy metabolism in myocardial remodelling by EP and myocardial protection. A total of 120 healthy male Sprague Dawley (SD) rats were randomly divided into six groups. Plasma cTnI, HBFP staining and electrocardiographic indicators were examined in the context of myocardial ischemic/hypoxic injury and protection. Western blotting and fluorescence double labelling were used to investigate the relationship between energy metabolism and autophagy in EP-resistant myocardial injury caused by exhaustive exercise. Compared with those in the C group, the levels of myocardial ischemic/hypoxic injury were significantly increased in the EE group. Compared with those in the EE group, the levels of myocardial ischemic/hypoxic injury were significantly decreased in the EEP + EE and LEP + EE groups. Compared with that in the EE group, the level of GLUT4 in the sarcolemma was significantly increased, and the colocalization of GLUT4 with the sarcolemma was significantly increased in the EEP + EE and LEP + EE groups (P < 0.05). LC3-II and LC3-II/LC3-I levels of the EEP + EE group were significantly elevated compared with those in the EE group (P < 0.05). The levels of p62 were significantly decreased in the EEP + EE and LEP + EE groups compared with the EE group (P < 0.05). EP promotes GLUT4 translocation and induced autophagy to alleviate exhaustive exercise-induced myocardial ischemic/hypoxic injury.
... The cardiomyocyte plasma membrane is impermeable to glucose; as such, glucose uptake is mediated by glucose transporters (GLUTs) of which 14 members have been identified in various tissues to date. GLUT1 and GLUT4 are the most abundantly expressed isoforms in the heart [31], and notably, both transporters display an expression profile coinciding with different stages of cardiac development, with GLUT1 and GLUT4 being predominantly expressed in foetal and adult hearts, respectively [32]. The upregulation of GLUT1 has been found to play important roles in cardiac development and in response to cardiac stress [33,34]. ...
Ischemic heart disease (IHD) is the leading cause of heart failure (HF) and is a significant cause of morbidity and mortality globally. An ischemic event induces cardiomyocyte death, and the ability for the adult heart to repair itself is challenged by the limited proliferative capacity of resident cardiomyocytes. Intriguingly, changes in metabolic substrate utilisation at birth coincide with the terminal differentiation and reduced proliferation of cardiomyocytes, which argues for a role of cardiac metabolism in heart regeneration. As such, strategies aimed at modulating this metabolism-proliferation axis could, in theory, promote heart regeneration in the setting of IHD. However, the lack of mechanistic understanding of these cellular processes has made it challenging to develop therapeutic modalities that can effectively promote regeneration. Here, we review the role of metabolic substrates and mitochondria in heart regeneration, and discuss potential targets aimed at promoting cardiomyocyte cell cycle re-entry. While advances in cardiovascular therapies have reduced IHD-related deaths, this has resulted in a substantial increase in HF cases. A comprehensive understanding of the interplay between cardiac metabolism and heart regeneration could facilitate the discovery of novel therapeutic targets to repair the damaged heart and reduce risk of HF in patients with IHD.
... As glucose undergoes glycolysis and facilitates the anaerobic generation of ATP, glucose is considered the most efficient energy substrate of the heart. Glucose uptake by cardiomyocytes occurs along a steep concentration gradient by isoforms of specific transmembrane glucose transporters, namely, GLUT1 and GLUT4 [46,47]. After cardiomyocytes take up glucose, the cytosolic glucose is rapidly phosphorylated to glucose 6phosphate (G6P) by hexokinase II, the cardiac isoform of hexokinase. ...
The aberrant increase in cardio-metabolic diseases over the past couple of decades has drawn researchers’ attention to explore and unveil the novel mechanisms implicated in cardiometabolic diseases. Recent evidence disclosed that the derangement of cardiac energy substrate metabolism plays a predominant role in the development and progression of chronic cardiometabolic diseases. Hence, in-depth comprehension of the novel molecular mechanisms behind impaired cardiac metabolism-mediated diseases is crucial to expand treatment strategies. The complex and dynamic pathways of cardiac metabolism are systematically controlled by the novel executor, microRNAs (miRNAs). miRNAs regulate target gene expression by either mRNA degradation or translational repression through base pairing between miRNA and the target transcript, precisely at the 3’ seed sequence and conserved heptametrical sequence in the 5’ end, respectively. Multiple miRNAs are involved throughout every cardiac energy substrate metabolism and play a differential role based on the variety of target transcripts. Novel theoretical strategies have even entered the clinical phase for treating cardiometabolic diseases, but experimental evidence remains inadequate. In this review, we identify the potent miRNAs, their direct target transcripts, and discuss the remodeling of cardiac metabolism to cast light on further clinical studies and further the expansion of novel therapeutic strategies. This review is categorized into four sections which encompass (i) a review of the fundamental mechanism of cardiac metabolism, (ii) a divulgence of the regulatory role of specific miRNAs on cardiac metabolic pathways, (iii) an understanding of the association between miRNA and impaired cardiac metabolism, and (iv) summary of available miRNA targeting therapeutic approaches.
... The turnover of high-energy phosphates in the myocardium is very dynamic and efficientalthough cardiac ATP stores amount to less than 1.0 g, the heart utilizes as much as a few kilograms daily [19]. Regarding biofuels, myocardial tissue is considered 'omnivorous', and cardiomyocytes utilize predominantly fatty acids (FAs), glucose and lactate, with the limited use of amino acids and ketones [17,20,21]. Under physiological conditions, the β-oxidation of FAs accounts for the majority of produced cardiac acetyl-coenzyme A, a common compound synthesized also from other energy substrates [17]. ...
Among different pathomechanisms involved in the development of heart failure, adverse metabolic myocardial remodeling closely related to ineffective energy production, constitutes the fundamental feature of the disease and translates into further progression of both cardiac dysfunction and maladaptations occurring within other organs. Being the component of key enzymatic machineries, iron plays a vital role in energy generation and utilization, hence the interest in whether, by correcting systemic and/or cellular deficiency of this micronutrient, we can influence the energetic efficiency of tissues, including the heart. In this review we summarize current knowledge on disturbed energy metabolism in failing hearts as well as we analyze experimental evidence linking iron deficiency with deranged myocardial energetics.
... GLUT1 activation typically exists in a basal condition, whereas GLUT4 is insulin stimulated, and it expresses in tissues such as adipose, skeletal muscle, and cardiac muscle. 15,16 In T2DM, the insulin-regulated GLUT4 is inefficient to redistribute to cell membranes due to insulin resistance. 17 Wright et al. found reduced GLUT content in mice after only 2 weeks of high-fat diet (HFD), 18 and that concomitantly impacts the subsequent glucose oxidation and glycolysis. ...
Diabetes mellitus (DM) is a serious epidemic around the globe, and cardiovascular diseases account for the majority of deaths in patients with DM. Diabetic cardiomyopathy (DCM) is defined as a cardiac dysfunction derived from DM without the presence of coronary artery diseases and hypertension. Patients with either type 1 or type 2 DM are at high risk of developing DCM and even heart failure. Metabolic disorders of obesity and insulin resistance in type 2 diabetic environments result in dyslipidaemia and subsequent lipid‐induced toxicity (lipotoxicity) in organs including the heart. Although various mechanisms have been proposed underlying DCM, it remains incompletely understood how lipotoxicity alters cardiac function and how DM induces clinical heart syndrome. With recent progress, we here summarize the latest discoveries on lipid‐induced cardiac toxicity in diabetic hearts and discuss the underlying therapies and controversies in clinical DCM.
... Contrarily, GLUT4 is primarily found in intracellular vesicles during the resting state and is transferred to the plasma membrane in response to insulin stimulation or environmental changes [46]. In healthy patients, insulin increases the translocation of GLUT4 to the plasma membrane through AMPK via activating PI3K and Akt [47,48], while studies in diabetic glomeruli have shown that the expression of GLUT1 and glucose flux both were increased as mTOR activity was increased and vice versa, involving a feed-forward mechanism [49,50]. ...
Patients with type two diabetes mellitus (T2DM) are at increased risk for cardiovascular diseases. Impairments of endothelin-1 (ET-1) signaling and mTOR pathway have been implicated in diabetic cardiomyopathies. However, the molecular interplay between the ET-1 and mTOR pathway under high glucose (HG) conditions in H9c2 cardiomyoblasts has not been investigated. We employed MTT assay, qPCR, western blotting, fluorescence assays, and confocal microscopy to assess the oxidative stress and mitochondrial damage under hyperglycemic conditions in H9c2 cells. Our results showed that HG-induced cellular stress leads to a significant decline in cell survival and an impairment in the activation of ETA-R/ETB-R and the mTOR main components, Raptor and Rictor. These changes induced by HG were accompanied by a reactive oxygen species (ROS) level increase and mitochondrial membrane potential (MMP) loss. In addition, the fragmentation of mitochondria and a decrease in mitochondrial size were observed. However, the inhibition of either ETA-R alone by ambrisentan or ETA-R/ETB-R by bosentan or the partial blockage of the mTOR function by silencing Raptor or Rictor counteracted those adverse effects on the cellular function. Altogether, our findings prove that ET-1 signaling under HG conditions leads to a significant mitochondrial dysfunction involving contributions from the mTOR pathway.
... The immunohistochemical analysis revealed that, compared to the control, BPA treatment reduced GLUT1 mainly in the epicardial region of the heart ( Figure 1A), while the increase in CD36 affected all heart tissue. In addition to GLUT1, the prevalent glucose transporter in the fetal heart, we performed protein expression analysis on GLUT4 (the primary transporter in the postnatal/adult heart) and GLUT3 known for its higher affinity for hexose compared to GLUT1 and GLUT4 [42]. As can be observed in Supplementary Figure S1, the levels of GLUT4 were significantly decreased in the fetal heart of BPA-fed animals, and GLUT3 tended to be increased (Supplementary Figure S1). ...
Dietary exposure to Bisphenol A (BPA), an industrial chemical present in food containers, affects nutrient metabolism in the myocardium of offspring during intrauterine life. Using a murine model, we observed that fetal hearts from mothers exposed to BPA (2.5 μg/kg/day) for 20 days before mating and for all of the gestation had decreased expression of glucose transporter-1 (GLUT1), the principal sugar transporter in the fetal heart, and increased expression of fatty acid cluster of differentiation 36 transporter (CD36), compared to control fetuses from vehicle-treated mothers. We confirmed the suppression of GLUT1 by exposing fetal heart organotypic cultures to BPA (1 nM) for 48 h but did not detect changes in CD36 compared to controls. During pregnancy, the placenta continuously releases extracellular vesicles such as exosomes into fetal circulation. These vesicles influence the growth and development of fetal organs. When fetal heart cultures were treated with cord blood-derived exosomes isolated from BPA-fed animals, GLUT1 expression was increased by approximately 40%. Based on our results, we speculate that exosomes from cord blood, in particular placenta-derived nanovesicles, could contribute to the stabilization of the fetal heart metabolism by ameliorating the harmful effects of BPA on GLUT1 expression.
... A distinction is made between the passive concentrationgradient induced diffusion transport through the membrane (for fat-soluble substances) [13][14][15] , and the transport against the gradient, which is possible only by a certain carrier with the expenditure of energy. For example, there are carriers that bind to the transported molecules and move with them through the membrane (such as glucose) 16,17 . There are stationary carriers that form a pore in the membrane, i.e. a transfer channel (for example, Fig. 1a, inset) [18][19][20] . ...
In this work demostrates a unique method for determining the absolute value of the friction force of a nanoobject on the surface of a cell membrane using atomic force microscopy. The tribological properties of membranes of adult human buccal epithelium cells in the presence of a protective adsorption buffer layer of ~ 100 nm on their surface were studied using atomic force microscopy in the contact scanning mode. Local mapping of the tribological characteristics of the surface was carried out, viz. friction FL = FL(x, y) and adhesion Fadh = Fadh(x, y) forces were measured. Studies of the friction force Ffr on the membrane surface at the nanolevel showed that its value varies discretely with an interval equal to lLF ≈ 100 nm. It was shown that such discreteness is determined by the interval lLF of the action of adhesive forces Fadh and indicates the fractal nature of the functional dependence of the friction force on the coordinate Ffr = Ffr(x). Thus, for nano-objects with dimensions ≤ lLF, the absolute value of Ffr decreases according to a power law with an increase in the size of the object, which contradicts the similar dependence of the friction force for macro-objects in the global approximation.
... Many metabolic approaches have been tried in the management of myocardial ischemia with variable success. Increasing glucose metabolism through infusion of insulin, pyruvate infusion, use of glucagon-like peptide-1 (GLP-1)-based therapies, and AMPactivated protein kinase (AMPK) activation using metformin and cardiac-specific gain of function of glucose transporter GLUT 1 have all been shown to decrease infarct size in IR models through stimulation of myocardial glucose metabolism [7,8] and suppression of FFA release from adipocytes [5,9,10]. In the glucose-insulin-potassium (GIK) clinical trials, even though there were no changes in 30-day mortality or progression to MI, there was a decrease in infarct size and in-house mortality, as well as an improvement in composite outcome following cardiac arrest [11]. ...
Acute myocardial infarction (MI) is one of the leading causes of death worldwide. Early identification of ischemia and establishing reperfusion remain cornerstones in the treatment of MI, as mortality and morbidity can be significantly reduced by establishing reperfusion to the affected areas. The aim of the current study was to investigate the metabolomic changes in the serum in a swine model of MI induced by ischemia and reperfusion (I/R) injury, and to identify circulating metabolomic biomarkers for myocardial injury at different phases. Female Yucatan minipigs were subjected to 60 min of ischemia followed by reperfusion, and serum samples were collected at baseline, 60 min of ischemia, 4 h of reperfusion, and 24 h of reperfusion. Circulating metabolites were analyzed using an untargeted metabolomic approach. A bioinformatic approach revealed that serum metabolites show distinct profiles during ischemia and during early and late reperfusion. Some notable changes during ischemia include accumulation of metabolites that indicate impaired mitochondrial function and N-terminally modified amino acids. Changes in branched-chain amino-acid metabolites were noted during early reperfusion, while bile acid pathway derivatives and intermediates predominated in the late reperfusion phases. This indicates a potential for such an approach toward identification of the distinct phases of ischemia and reperfusion in clinical situations.
... In the heart, the most common glucose transport isoforms are GLUT1 and GLUT4. Other less common isoforms also exist (21). GLUT1 is predominately expressed in the developing embryonic heart but then declines postnatally. ...
Energy metabolism in the heart is affected during states of dysfunction. Understanding how the heart utilizes substrates in cardiomyopathy may be key to the development of alternative treatment modalities. Myocardial insulin resistance has been proposed as a possible barrier to effective glucose metabolism in the heart. Extensive literature on the topic in adult individuals exists; however, review in the pediatric population is sparse. The pathophysiology of disease in children and adolescents is unique. The aim of this paper is to review the current knowledge on insulin resistance in dilated cardiomyopathy while also filling the gap when considering care in the pediatric population.
... While glucose uptake into cardiomyocytes is dependent on insulin activity, the uptake of FA and HB is not hormonally regulated 9,10 . Glucose enters cardiomyocytes mostly via the insulin-dependent glucose transporter 4 (GLUT4) 11 and is directed through multiple metabolic pathways such as glycolysis, glycogen synthesis, polyol, hexosamine biosynthetic or pentose phosphate pathways. The end product of glycolysis, mal heart to respond properly to the energy demand. ...
Objectives
We will review current concepts regarding bioenergetic decline in heart failure (HF). In the heart, the high energy demand must be met by continuous ATP generation. Cardiac energetic machinery orchestrates the ATP production by using oxidation of multiple energetic substrates including fatty acids (FA), glucose, amino acids and ketone bodies. The normal heart is metabolically flexible and able to use different energetic fuels during physiologic or pathologic circumstances to better match the energy demand. Mitochondria have critical role in maintaining cardiac metabolic flexibility.
Methods
We analyzed the scientific literature pertinent to HF and mitochondrial dysfunction.
Results
The general consent is that metabolic flexibility is lost in HF with either preserved or reduced ejection fraction (HFpEF and HFrEF, respectively). The prototype of HFpEF is the metabolic heart disease that is characterized by increased reliance on FA oxidation for ATP production and decreased glucose oxidation, while HFrEF presents a decreased FA oxidation. Both types of HF are associated with a decline in mitochondrial function leading to increased oxidative stress, abnormalities in the redox status and energy deficit.
Conclusion
Current research is committed to find novel metabolically targeted therapeutic approaches to improve energetic metabolism and alleviate HF progression.
... To our knowledge, the impact of diabetes or diabetic conditions on insulin signaling pathways in 2D or 3D models of CAVD has not been described in the literature so far. However, studies for classical insulin signaling have been reported for cardiac fibroblasts [25] and cardiomyocytes [37,38]. Binding of insulin to the insulin receptor triggers a phosphorylation cascade of downstream proteins, whereby the insulin signal is transmitted intracellularly [12]. ...
Type 2 diabetes mellitus (T2D) is one of the prominent risk factors for the development and progression of calcific aortic valve disease. Nevertheless, little is known about molecular mechanisms of how T2D affects aortic valve (AV) remodeling. In this study, the influence of hyperinsulinemia and hyperglycemia on degenerative processes in valvular tissue is analyzed in intact AV exposed to an either static or dynamic 3D environment, respectively. The complex native dynamic environment of AV is simulated using a software-governed bioreactor system with controlled pulsatile flow. Dynamic cultivation resulted in significantly stronger fibrosis in AV tissue compared to static cultivation, while hyperinsulinemia and hyperglycemia had no impact on fibrosis. The expression of key differentiation markers and proteoglycans were altered by diabetic conditions in an environment-dependent manner. Furthermore, hyperinsulinemia and hyperglycemia affect insulin-signaling pathways. Western blot analysis showed increased phosphorylation level of protein kinase B (AKT) after acute insulin stimulation, which was lost in AV under hyperinsulinemia, indicating acquired insulin resistance of the AV tissue in response to elevated insulin levels. These data underline a complex interplay of diabetic conditions on one hand and biomechanical 3D environment on the other hand that possesses an impact on AV tissue remodeling.
... There were decreased protein abundances of several rate-limiting enzymes critical for the regulation of glucose uptake, glucose phosphorylation, and the conversion of pyruvate to acetyl-CoA (Fig. 1j, k). Levels of the insulin-responsive glucose transporter that regulates glucose entry in the beating heart, glucose transporter type 4 [GLUT4; also known as solute carrier family 2 (SLC2A4)] 14 (FC = 0.80, p = 6.5 × 10 -3 ), and downstream hexokinase 1 and 2 (HK1, FC = 0.75, p = 9.1 × 10 -3 and HK2, FC = 0.69, p = 4.7 × 10 -4 ), were signi cantly decreased in HFpEF myocardium (Fig. 1j). HFpEF hearts had over 2.5-fold increase in pyruvate dehydrogenase kinase 4 (PDK4) expression (FC = 2.60, p = 2.3 × 10 -5 ), which can reduce glucose oxidation by deactivating the pyruvate dehydrogenase complex (PDHC), a key mitochondrial enzyme that catalyzes the oxidative decarboxylation of pyruvate to acetyl-CoA, linking glycolysis to the tricarboxylic acid cycle 15 . ...
Heart Failure with preserved Ejection Fraction or ‘HFpEF’ is now the most common type of heart failure worldwide, with considerable morbidity and mortality, and with no effective pharmacotherapies. For the first time, we show that the human and murine HFpEF heart have impaired uptake of ketone bodies, and reveal an intrinsic capacity of human and murine hearts to generate their own ketones via the canonical rate-limiting ketogenic enzyme, 3-hydroxy-3-methylglutaryl CoA synthase 2 (HMGCS2). We demonstrate that protein levels of HMGCS2 are dramatically elevated in HFpEF myocardium that does not overcome reduced specific activity of this enzyme. This appeared to be due to hyperacetylation caused by depletion of the de-acetylating enzyme sirtuin 3 and decreased oxidized/reduced nicotinamide adenine dinucleotide (NAD+/NADH) ratio, upon which the deacetylation activity of sirtuins is dependent. Also for the first time, we show that the canonical stimuli that serve to turn on ketogenesis in the liver are present locally in the HFpEF heart, augmented by a profound energy deficiency as determined by depleted phosphocreatine/ adenosine triphosphate (PCr/ATP) ratio. Thus, metabolically inflexible HFpEF hearts try to auto-regulate their energy supply by upregulating ketogenesis as an apparent “rescue strategy”. These results serve to reframe our understanding of cardiac ketone metabolism and open opportunities for novel therapeutic strategies for HFpEF, urgently needed.
... In the chemogenetic heart failure model, we previously observed a metabolic switch in fuel preference from FA oxidation toward higher glucose utilization, demonstrated by in vivo 18 F-FDG-PET imaging in the hearts of DAAO-infected/D-alanine-treated animals (20). Glucose is transported into the adult heart mainly by the insulin-dependent glucose transporter Glut4, and to a lesser extent by the insulin-independent transporter Glut1 (33,34). Interestingly, our transcriptomic data showed a decrease in Glut4 transcript abundance and an increase of Glut1 transcripts (Supplemental Fig. S4). ...
... 45 47 Glucose uptake in cardiomyocytes is mediated by GLUT1 and GLUT4. 48,49 There is evidence to indicate that metabolic remodeling precedes most other pathological alterations and likely plays an essential role in cardiac hypertrophy and heart failure. 50 At the same time, acute phase of drug-induced cardiomyopathy may not manifest in considerable changes in myocardial perfusion, unless perfusion deficit is exaggerated by stress. ...
Background
Drug‐induced cardiomyopathy is a significant medical problem. Clinical diagnosis of myocardial injury is based on initial electrocardiogram, levels of circulating biomarkers, and perfusion imaging with single photon emission computed tomography (SPECT). Positron emission tomography (PET) is an alternative imaging modality that provides better resolution and sensitivity than SPECT, improves diagnostic accuracy, and allows therapeutic monitoring. The objective of this study was to assess the detection of drug‐induced cardiomyopathy by PET using 2‐deoxy‐2‐[¹⁸F]fluoro‐D‐glucose (FDG) and compare it with the conventional SPECT technique with [99mTc]‐Sestamibi (MIBI).
Methods
Cardiomyopathy was induced in Sprague Dawley rats using high‐dose isoproterenol. Nuclear [¹⁸F]FDG/PET and [99mTc]MIBI/SPECT were performed before and after isoproterenol administration. [¹⁸F]FDG (0.1 mCi, 200‐400 µL) and [99mTc]MIBI (2 mCi, 200‐600 µL) were administered via the tail vein and imaging was performed 1 hour postinjection. Isoproterenol‐induced injury was confirmed by the plasma level of cardiac troponin and triphenyltetrazolium chloride (TTC) staining.
Results
Isoproterenol administration resulted in an increase in circulating cardiac troponin I and showed histologic damage in the myocardium. Visually, preisoproterenol and postisoproterenol images showed alterations in cardiac accumulation of [¹⁸F]FDG, but not of [99mTc]MIBI. Image analysis revealed that myocardial uptake of [¹⁸F]FDG reduced by 60% after isoproterenol treatment, whereas that of [99mTc]MIBI decreased by 45%.
Conclusion
We conclude that [¹⁸F]FDG is a more sensitive radiotracer than [99mTc]MIBI for imaging of drug‐induced cardiomyopathy. We theorize that isoproterenol‐induced cardiomyopathy impacts cellular metabolism more than perfusion, which results in more substantial changes in [¹⁸F]FDG uptake than in [99mTc]MIBI accumulation in cardiac tissue.
... The GLUT-1 transporter's rate is influenced by glucose concentration (although practically kM is sufficiently low that it is almost always operating at Vmax), while GLUT-4 requires insulin to fulfil its role. 30 Insulin also has effects to increase activity of enzymes involved in glycolysis such as phosphofructokinase 2 and pyruvate dehydrogenase itself, via pyruvate dehydrogenase kinase. 31 It therefore follows that glucose can be taken up and metabolized by the heart in conditions of low insulin, but maximizing glucose utilization requires additional insulin. ...
Nicotinic acid receptor agonists have previously been shown to cause acute reductions in cardiac contractility. We sought to uncover the changes in cardiac metabolism underlying these alterations in function. In nine humans, we recorded cardiac energetics and function before and after a single oral dose of nicotinic acid using cardiac MRI to demonstrate contractile function and Phosphorus‐31 (³¹P) magnetic resonance spectroscopy to demonstrate myocardial energetics. Nicotinic Acid 400 mg lowered ejection fraction by 4% (64 ± 8% to 60 ± 7%, P = .03), and was accompanied by a fall in phosphocreatine/ATP ratio by 0.4 (2.2 ± 0.4 to 1.8 ± 0.1, P = .04). In four groups of eight Wistar rats, we used pyruvate dehydrogenase (PDH) flux studies to demonstrate changes in carbohydrate metabolism induced by the nicotinic acid receptor agonist, Acipimox, using hyperpolarized Carbon‐13 (¹³C) magnetic resonance spectroscopy. In rats which had been starved overnight, Acipimox caused a fall in ejection fraction by 7.8% (67.5 ± 8.9 to 60 ± 3.1, P = .03) and a nearly threefold rise in flux through PDH (from 0.182 ± 0.114 to 0.486 ± 0.139, P = .002), though this rise did not match pyruvate dehydrogenase flux observed in rats fed carbohydrate rich chow (0.726 ± 0.201). In fed rats, Acipimox decreased pyruvate dehydrogenase flux (to 0.512 ± 0.13, P = .04). Concentration of plasma insulin fell by two‐thirds in fed rats administered Acipimox (from 1695 ± 891 ng/L to 550 ± 222 ng/L, P = .005) in spite of glucose concentrations remaining the same. In conclusion, we demonstrate that nicotinic acid receptor agonists impair cardiac contractility associated with a decline in cardiac energetics and show that the mechanism is likely a combination of reduced fatty acid availability and a failure to upregulate carbohydrate metabolism, essentially starving the heart of fuel.
... Slc2a3, commonly known as Glut3, is expressed in tissues with heightened energy demands and high metabolic rates since it has the highest glucose affinity and greatest transport capacity in the glucose transporter protein family [40]. Hence, it is highly expressed in the brain, cardiomyocytes, and cardiomyoblasts [41]. Decreased expression of Slc2a3 was observed in the cardiac tissue of HFD mice, indicating dysregulated myocardial glucose transport in response to obesity. ...
Obesity is associated with an increased risk of developing cardiovascular disease (CVD), with limited alterations in cardiac genomic characteristics known. Cardiac transcriptome analysis was conducted to profile gene signatures in high-fat diet (HFD)-induced obese mice. A total of 184 differentially expressed genes (DEGs) were identified between groups. Based on the gene ontology (GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of DEGs, the critical role of closely interlocked glucose metabolism was determined in HFD-induced cardiac remodeling DEGs, including Nr4a1, Fgf21, Slc2a3, Pck1, Gck, Hmgcs2, and Bpgm. Subsequently, the expression levels of these DEGs were evaluated in both the myocardium and palmitic acid (PA)-stimulated H9c2 cardiomyocytes using qPCR. Nr4a1 was highlighted according to its overexpression resulting from the HFD. Additionally, inhibition of Nr4a1 by siRNA reversed the PA-induced altered expression of glucose metabolism-related DEGs and hexokinase 2 (HK2), the rate-limiting enzyme in glycolysis, thus indicating that Nr4a1 could modulate glucose metabolism homeostasis by regulating the expression of key enzymes in glycolysis, which may subsequently influence cardiac function in obesity. Overall, we provide a comprehensive understanding of the myocardium transcript molecular framework influenced by HFD and propose Nr4a1 as a key glucose metabolism target in obesity-induced CVD.
... Both GLUT1 and GLUT4 are the main glucose transporters in the heart. Increased GLUT1expression suggests a shift from an oxidative to a glycolytic metabolism [38]. In high fat diet-fed transgenic mice overexpressing GLUT1 specifically in the heart, cardiac dysfunction associated to excessive lipid accumulation was observed [39]. ...
Background:
In metabolic disorders, myocardial fatty infiltration is critically associated with lipotoxic cardiomyopathy.
Methods:
Twenty Psammomys obesus gerbils were randomly assigned to normal plant or high fat diet. Sixteen weeks later, myocardium was sampled for pathobiological evaluation.
Results:
A sixteen-week high fat diet resulted in myocardial structure disorganization, with collagen deposits, lipid accumulation, cardiomyocyte apoptosis and inflammatory cell infiltration. Myocardial expressions of glucose transporter GLUT1 and pyruvate dehydrogenase (PDH) inhibitor, PDH kinase (PDK)4 increased, while insulin-regulated GLUT4 expression remained unchanged. Myocardial expressions of molecules regulating fatty acid transport, CD36 and fatty acid binding protein (FABP)3, were increased, while expression of rate-controlling fatty acid β-oxidation, carnitine palmitoyl transferase (CPT)1B decreased. Myocardial expression of AMP-activated protein kinase (AMPK), decreased, while expression of peroxisome proliferator activated receptors (PPAR)-α and -γ did not change.
Conclusion:
In high fat diet fed Psammomys obesus, an original experimental model of nutritionally induced metabolic syndrome mixing genetic predisposition and environment interactions, a short period of high fat feeding was sufficient to induce myocardial structural alterations, associated with altered myocardial metabolic gene expression in favor of lipid accumulation.
Congenital heart defects (CHDs) are the most common form of birth defects in humans. They occur in 9 out of 1000 live births and are defined as structural abnormalities of the heart. Understanding CHDs is difficult due to the heterogeneity of the disease and its multifactorial etiology. Advances in genomic sequencing have made it possible to identify the genetic factors involved in CHDs. However, genetic origins have only been found in a minority of CHD cases, suggesting the contribution of non-inherited (environmental) risk factors to the etiology of CHDs. Maternal pregestational diabetes is associated with a three- to five-fold increased risk of congenital cardiopathies, but the underlying molecular mechanisms are incompletely understood. According to current hypotheses, hyperglycemia is the main teratogenic agent in diabetic pregnancies. It is thought to induce cell damage, directly through genetic and epigenetic dysregulations and/or indirectly through production of reactive oxygen species (ROS). The purpose of this review is to summarize key findings on the molecular mechanisms altered in cardiac development during exposure to hyperglycemic conditions in utero. It also presents the various in vivo and in vitro techniques used to experimentally model pregestational diabetes. Finally, new approaches are suggested to broaden our understanding of the subject and develop new prevention strategies.
Curcumin, the active compound found in turmeric, is believed to delay the development of diabetes through several mechanisms. This study aimed to investigate if an aqueous extract of turmeric can improve glucose uptake and uric acid in mouse tissues in vitro, after inclusion of turmeric in the diet for four weeks. Fourteen adult male Swiss mice were divided into three groups. The first group was the control (n=6) that was given clean water, the second group of mice (n=4) was given 5% autoclaved turmeric extract in drinking water, and the third group (n=4) was given 5% non-autoclaved turmeric extract in drinking water. After four weeks, the cardiac muscle, skeletal muscle, pancreas, and liver tissues were dissected and used for analysis. The results showed that the aqueous 5% turmeric extract reduced glucose in cardiac tissues while the plasma glucose was not changed. Cardiac muscle, liver, pancreas, and skeletal muscle showed glucose absorption after the 5% turmeric treatment. This research shows that turmeric did improve glucose uptake in most tissues, although it was not significant due to the limitations of this study. Tissues may need to be cultured longer and media processed quicker to prevent evaporation. Turmeric continues to show great potential in the treatment of type 2 diabetes and may present an alternative way of treating diabetes.
Cardiovascular diseases are among the primary life-threatening conditions affecting human society. Intermittent fasting is shown to be functional in the prevention of cardiovascular diseases, however, the information on fasting-associated modifications in myocardial biomolecules is limited. This study aimed to determine the impact of 18-h intermittent fasting administered for five weeks on 12 months-old rats using supervised linear discriminant analysis and support vector machine algorithms constructed on spectrochemical data obtained from myocardial tissues. These algorithms revealed gross biomolecular modifications, while quantitative analyses demonstrated higher amounts of saturated lipids (19%), triglycerides (11%), and lipids (56%), in addition to enhancement in membrane dynamics (18%). The concentrations of nucleic acids and glucose are increased by 52%, while the glycogen content is diminished by 61%. The protein carbonylation/oxidation is reduced by 38%, whereas a 35% increase in protein content was measured. Phosphorylated proteins have been calculated to be at higher concentrations in the 13–62% range. The study findings demonstrated significant molecular changes in the myocardium of rats subjected to intermittent fasting.
Hypertension is an important risk factor in the pathogenesis of diastolic dysfunction. Growing evidence indicates that glucose metabolism plays an essential role in diastolic dysfunction. TP53‐induced glycolysis and apoptosis regulator (TIGAR) has been shown to regulate glucose metabolism and heart failure (HF). In the present study, we investigated the role of TIGAR in diastolic function and cardiac fibrosis during pressure overload (PO)‐induced HF. WT mice subjected to transverse aortic constriction (TAC), a commonly used method to induce diastolic dysfunction, exhibited diastolic dysfunction as evidenced by increased E/A ratio and E/E′ ratio when compared to its sham controls. This was accompanied by increased cardiac interstitial fibrosis. In contrast, the knockout of TIGAR attenuated PO‐induced diastolic dysfunction and interstitial fibrosis. Mechanistically, the levels of glucose transporter Glut‐1, Glut‐4, and key glycolytic enzyme phosphofructokinase 1 (PFK‐1) were significantly elevated in TIGAR KO subjected to TAC as compared to that of WT mice. Knockout of TIGAR significantly increased fructose 2,6‐bisphosphate levels and phosphofructokinase activity in mouse hearts. In addition, PO resulted in a significant increase in perivascular fibrosis and endothelial activation in the WT mice, but not in the TIGAR KO mice. Our present study suggests a necessary role of TIGAR‐mediated glucose metabolism in PO‐induced cardiac fibrosis and diastolic dysfunction.
Our group previously demonstrated that an excess of nutrients, as observed in diabetes, provokes an increase in cardiac protein acetylation responsible for a reduced insulin-stimulated translocation of the glucose transporter GLUT4 to the plasma membrane. The acetylated proteins involved in this event have yet not been identified. α-Tubulin is a promising candidate as a major cytoskeleton component involved, among other things, in the translocation of GLUT4-containing vesicles from their intracellular pools towards the plasma membrane. Moreover, α-tubulin is known to be acetylated, Lys40 (K40) being its best characterized acetylated residue. The present work sought to evaluate the impact of α-tubulin K40 acetylation on cardiac glucose entry, with a particular interest in GLUT4 translocation. First, we observed that a mouse model of high-fat diet-induced obesity presented an increase in cardiac α-tubulin K40 acetylation level. Next, we showed that treatment of insulin-sensitive primary cultured adult rat cardiomyocytes with tubacin, a specific tubulin acetylation inducer, reduced insulin-stimulated glucose uptake and GLUT4 translocation. Conversely, decreasing α-tubulin K40 acetylation by expressing a non-acetylable dominant form of α-tubulin (mCherry α-tubulin K40A mutant) remarkably intensified insulin-induced glucose transport. Finally, mCherry α-tubulin K40A expression similarly improved glucose transport in insulin-resistant cardiomyocytes or after AMP-activated protein kinase activation. Taken together, our study demonstrates that modulation of α-tubulin K40 acetylation level affects glucose transport in cardiomyocytes, offering new putative therapeutic insights regarding modulation of glucose metabolism in insulin-resistant and diabetic hearts.
The failing heart is characterized by elevated levels of reactive oxygen species. We have developed an animal model of heart failure induced by chemogenetic production of oxidative stress in the heart using a recombinant adeno-associated virus (AAV9) expressing yeast D-amino acid oxidase (DAAO) targeted to cardiac myocytes. When DAAO-infected animals are fed the DAAO substrate D-alanine, the enzyme generates hydrogen peroxide (H 2 O 2 ) in the cardiac myocytes, leading to dilated cardiomyopathy. However, the underyling mechanisms of oxidative stress induced heart failure remain incompletely understood. Therefore, we investigated the effects of chronic oxidative stress on the cardiac transcriptome and metabolome. Rats infected with recombinant cardiotropic AAV9 expressing DAAO or control AAV9 were treated for 7 weeks with D-alanine to stimulate chemogenetic H 2 O 2 production by DAAO and generate dilated cardiomyopathy. After hemodynamic assessment, left and right ventricular tissues were processed for RNA sequencing and metabolomic profiling. DAAO induced dilated cardiomyopathy was characterized by marked changes in the cardiac transcriptome and metabolome both in the left and right ventricle. Downregulated transcripts related to energy metabolism and mitochondrial function, accompanied by striking alterations in metabolites involved in cardiac energetics redox homeostasis, and amino acid metabolism. Upregulated transcripts involved cytoskeletal organization and extracellular matrix. Finally, we noted increased metabolite levels of antioxidants glutathione and ascorbate. These findings provide evidence that chemogenetic generation of oxidative stress leads to a robust heart failure model with distinct transcriptomic and metabolomic signatures and set the basis for understanding the underlining pathophysiology of chronic oxidative stress in the heart.
A high global prevalence of diabetes and its implications on the heart in vivo and in vitro tools have been pursued to alleviate the complications of high glucose. This chapter oulines the methods used for maintaining H9C2 cardiomyoblasts in vitro and for stimulating hyperglycemic situation. In addition, we present a method to assess cellular GLUT-4 expression using qRT-PCR. This cellular model also allows us to examine the therapeutic approach of an antioxidant, Trolox, for upregulating GLUT-4 and uptake of glucose under hyperglycemic condition.
Insulin is the peptide hormone. It exerts anabolic effect on the metabolism of carbohydrates, lipids, and proteins. Insulin controls the plasma glucose concentration. It activates insulin receptors on the insulin sensitive cells to promote uptake of glucose. Insulin regulates the glycogenesis in hepatocytes. Insulin suppresses the activity of alpha cells in pancreas and restricts the release of glucagon hormone, thus insulin suppresses the glycogenolysis and gluconeogenesis in hepatocytes. Insulin promotes lipogenesis in the adipose tissues and suppresses the mobilization of triglycerides in adipocytes. Insulin hormone promotes the uptake of amino acids by cells and induces polypeptide synthesis through its regulatory action at the level of transcription.
Several hormones and their receptors have a critical role in the homeostasis of cardiomyocytes. They also modulate pathophysiological alterations in the cells and are therapeutic targets in heart diseases. Insulin controls substrate utilization in cardiomyocytes. Insulin promotes glucose uptake and its utilization via glycolysis and also regulates uptake of long-chain fatty acid and protein synthesis. There is evidence that varied thyroid hormone levels alter the growth patterns of both neonatal and adult cardiomyocytes. Oestrogen and its receptors influence the complex network of genomic and nongenomic pathways that govern cardiac metabolism, cytoprotection, cardiomyocyte regeneration and the electrophysiological and contractile function of the heart. Growth hormone is a participant in stimulating cardiomyocyte growth during development of the heart as well as in the maintenance of the structure and function of the normal adult heart. Aldosterone-mediated effects on the heart include increased oxidative stress, apoptosis, cardiac fibrosis, as well as left-ventricular hypertrophy.
Skeletal muscle wasting has been well-documented among hemodialysis patients. This catabolic condition can be induced by numerous factors, including low-grade inflammation, and is associated with impairments in functional capacity and quality of life, as well as an increased mortality risk. We previously showed that 12 weeks of intradialytic resistance training increases lean mass, functional capacity, and the quality of life of hemodialysis patients. This chapter provides the details of a protocol of intradialytic exercise that leads to improvements in inflammatory status, body composition, and functional capacity.
Background: Metabolic remodeling precedes most alterations during cardiac hypertrophic growth under hemodynamic stress. The elevation of glucose utilization has been recognized as a hallmark of metabolic remodeling. However, its role in cardiac hypertrophic growth and heart failure in response to pressure overload remains to be fully illustrated. Here, we aimed to dissect the role of cardiac PKM1 (pyruvate kinase muscle isozyme 1) in glucose metabolic regulation and cardiac response under pressure overload.
Methods: Cardiac specific deletion of PKM1 was achieved by crossing the floxed PKM1 mouse model with the cardiomyocyte-specific Cre transgenic mouse. PKM1 transgenic mice were generated under the control of tetracycline response elements, and cardiac specific overexpression of PKM1 was induced by doxycycline administration in adult mice. Pressure overload was triggered by transverse aortic constriction (TAC). Primary neonatal rat ventricular myocytes were used to dissect molecular mechanisms. Moreover, metabolomics and NMR spectroscopy analyses were conducted to determine cardiac metabolic flux in response to pressure overload.
Results: We found that PKM1 expression is reduced in failing human and mouse hearts. Importantly, cardiomyocyte-specific deletion of PKM1 exacerbates cardiac dysfunction and fibrosis in response to pressure overload. Inducible overexpression of PKM1 in cardiomyocytes protects the heart against TAC-induced cardiomyopathy and heart failure. At the mechanistic level, PKM1 is required for the augmentation of glycolytic flux, mitochondrial respiration, and ATP production under pressure overload. Furthermore, deficiency of PKM1 causes a defect in cardiomyocyte growth and a decrease in pyruvate dehydrogenase complex activity at both in vitro and in vivo levels.
Conclusions: These findings suggest that PKM1 plays an essential role in maintaining a homeostatic response in the heart under hemodynamic stress.
Insulin receptors are highly expressed in the heart and vasculature. Insulin signaling regulates cardiac growth, survival, substrate uptake, utilization and mitochondrial metabolism. Insulin signaling modulates the cardiac responses to physiological and pathological stressors. Altered insulin signaling in the heart may contribute to the pathophysiology of ventricular remodeling and heart failure progression. Myocardial insulin signaling adapts rapidly to changes in the systemic metabolic milieu. What may initially represent an adaptation to protect the heart from carbo-toxicity, may contribute to amplifying the risk of heart failure in obesity and diabetes. This review article presents the multiple roles of insulin signaling in cardiac physiology and pathology and discusses the potential therapeutic consequences of modulating myocardial insulin signaling.
The global incidence, associated mortality rates and economic burden of diabetes are now such that it is considered one of the most pressing worldwide public health challenges. Considerable research is now devoted to better understanding the mechanisms underlying the onset and progression of this disease, with an ultimate aim of improving the array of available preventive and therapeutic interventions. One area of particular unmet clinical need is the significantly elevated rate of cardiomyopathy in diabetic patients, which in part contributes to cardiovascular disease being the primary cause of premature death in this population. This review will first consider the role of metabolism and more specifically the insulin sensitive glucose transporter GLUT4 in diabetic cardiac disease, before addressing how we may use exercise to intervene in order to beneficially impact key functional clinical outcomes.
Background
Under normal conditions, the heart obtains ATP through the oxidation of fatty acids, glucose, and ketones. While fatty acids are the main source of energy in the heart, under certain conditions, the main source of energy shifts to glucose where pyruvate converts into lactate, to meet the energy demand. The Warburg effect is the energy shift from oxidative phosphorylation to glycolysis in the presence of oxygen. This effect is observed in tumors as well as in diseases, including cardiovascular diseases. If glycolysis is more dominant than glucose oxidation, the two pathways uncouple, contributing to the severity of the heart condition. Recently, several studies have documented changes in metabolism in several cardiovascular diseases; however, the specific mechanisms remain unclear.
Methods
This literature review was conducted by an electronic database of Pub Med, Google Scholar, and Scopus published until 2020. Relevant papers are selected based on inclusion and exclusion criteria.
Results
A total of 162 potentially relevant articles after the title and abstract screening were screened for full-text. Finally, 135 papers were included for the review article.
Discussion
This review discusses the effects of alterations in glucose metabolism, particularly the Warburg effect, on cardiovascular diseases, including heart failure, atrial fibrillation, and cardiac hypertrophy.
Conclusion
Reversing the Warburg effect could become a potential treatment option for cardiovascular diseases.
Cells display distinct metabolic characteristics depending on its differentiation stage. The fuel type of the cells serves not only as a source of energy but also as a driver of differentiation. Glucose, the primary nutrient to the cells, is a critical regulator of rapidly growing embryos. This metabolic change is a consequence as well as a cause of changes in genetic program. Disturbance of fetal glucose metabolism such as diabetic pregnancy is associated with congenital heart disease. In utero hyperglycemia impacts the left-right axis establishment, migration of cardiac neural crest cells, conotruncal formation and mesenchymal formation of the cardiac cushion during early embryogenesis and causes cardiac hypertrophy in late fetal stages. In this review, we focus on the role of glucose in cardiogenesis and the molecular mechanisms underlying heart diseases associated with hyperglycemia.
Although sodium-glucose co-transporter 1 (SGLT1) has been identified as one of the major SGLT isoforms expressed in the heart, its exact role remains elusive. Evidences using phlorizin, the most common inhibitor of SGLTs, suggested its role in glucose transport. However, phlorizin could also affect classical facilitated diffusion via glucose transporters (GLUTs), bringing into question the relevance of SGLT1 in overall cardiac glucose uptake. Accordingly, we assessed the contribution of SGLT1 in cardiac glucose uptake using the SGLT1 knock-out mouse model, which lacks exon 1. Glucose uptake was similar in cardiomyocytes isolated from SGLT1 knock-out ( D ex1 KO) and control littermate (WT) mice, either under basal state, insulin, or hyperglycemia. Similarly, in vivo basal and insulin-stimulated cardiac glucose transport measured by micro-PET scan technology did not differ between WT and D ex1 KO mice. Micromolar concentrations of phlorizin had no impact on glucose uptake in either isolated WT or D ex1 KO-derived cardiomyocytes. However, higher concentrations (1mM) completely inhibited insulin-stimulated glucose transport without affecting insulin signaling nor GLUT4 translocation, independently from cardiomyocyte genotype. Interestingly, we discover that mouse and human hearts expressed a shorter slc5a1 transcript, leading to SGLT1 protein lacking transmembrane domains and residues involved in glucose and sodium bindings. In conclusion, cardiac SGLT1 does not contribute to overall glucose uptake, probably due to the expression of slc5a1 transcript variant. The inhibitory effect of phlorizin on cardiac glucose uptake is SGLT1-independent and can be explained by GLUT transporter inhibition. These data open new perspectives in understanding the role of SGLT1 in the heart.
As a constantly contracting muscular pump, the heart has an incessant need for energy to fuel contractile function. As such, the heart is highly adapted to metabolize multiple substrates such as fatty acids, glucose, lactate, and ketone bodies in mitochondria to generate many times its weight each day in adenosine triphosphate (ATP). However, in addition to generating ATP, cardiac metabolites also generate important signaling molecules that have profound effects on cardiac structure and function that are independent of their role in energy generation. This chapter provides an overview of cardiac energy metabolism and its regulation, and discusses the crosstalk between cardiac metabolism and signaling, and the significance of cardiac energetics in heart disease. The chapter focuses mainly on heart failure (HF) and discusses metabolic dysfunction in the failing heart, metabolic remodeling, and risk for HF development, methods to evaluate defects in cardiac energetics, and therapeutic approaches to modulate cardiac energetics.
Insulin impairs β2-adrenergic receptor (β2AR) function through G protein-coupled receptor kinase 2 (GRK2) by phosphorylation but less is known about dephosphorylation mechanisms mediated by protein phosphatase 2A (PP2A). Pharmacologic or genetic inhibition of phosphoinositide 3-kinase γ (PI3Kγ) unexpectedly resulted in significant reduction of insulin-mediated β2AR phosphorylation. Interestingly, β2AR-associated phosphatase activity was inhibited by insulin but was reversed by knock-down of PI3Kγ showing negative regulation of PP2A by PI3Kγ. Co-immunoprecipitation and surface plasmon resonance studies using purified proteins showed that GRK2 and PI3Kγ form a complex and could be recruited to β2ARs as GRK2 interacts with insulin receptor substrate following insulin treatment. Consistently, β-blocker pretreatment did not reduce insulin-mediated β2AR phosphorylation indicating agonist- and Gβγ-independent non-canonical regulation of receptor function. Mechanistically, PI3Kγ inhibits PP2A activity at the βAR complex by phosphorylating an intracellular inhibitor of PP2A (I2PP2A). Knock-down or CRISPR ablation of endogenous I2PP2A unlocked PP2A inhibition mediating β2AR dephosphorylation showing an unappreciated acute regulation of PP2A in mediating insulin-β2AR cross-talk.
Summary
Insulin impairs β2-adrenergic receptor (β2AR) function through G protein-coupled receptor kinase 2 (GRK2). We show that insulin simultaneously inhibits protein phosphatase 2A (PP2A) sustaining β2AR functional impairment. Unexpectedly, releasing PP2A inhibition by PI3Kγ preserves β2AR function despite intact insulin-driven GRK2-mechanisms.
Unlabelled:
Using isolated rat cardiomyocytes we have examined: 1) the effect of insulin on the cellular distribution of glucose transporter 4 (GLUT4) and GLUT1, 2) the total amount of these transporters, and 3) the co-localization of GLUT4, GLUT1, and secretory carrier membrane proteins (SCAMPs) in intracellular membranes. Insulin induced 5.7- and 2.7-fold increases in GLUT4 and GLUT1 at the cell surface, respectively, as determined by the nonpermeant photoaffinity label [3H]2-N-[4(1-azi-2,2,2-trifluoroethyl)benzoyl]-1, 3-bis-(D-mannos-4-yloxy)propyl-2-amine. The total amount of GLUT1, as determined by quantitative Western blot analysis of cell homogenates, was found to represent a substantial fraction ( approximately 30%) of the total glucose transporter content. Intracellular GLUT4-containing vesicles were immunoisolated from low density microsomes by using monoclonal anti-GLUT4 (1F8) or anti-SCAMP antibodies (3F8) coupled to either agarose or acrylamide. With these different immunoisolation conditions two GLUT4 membrane pools were found in nonstimulated cells: one pool with a high proportion of GLUT4 and a low content in GLUT1 and SCAMP 39 (pool 1) and a second GLUT4 pool with a high content of GLUT1 and SCAMP 39 (pool 2). The existence of pool 1 was confirmed by immunotitration of intracellular GLUT4 membranes with 1F8-acrylamide. Acute insulin treatment caused the depletion of GLUT4 in both pools and of GLUT1 and SCAMP 39 in pool 2.
In conclusion:
1) GLUT4 is the major glucose transporter to be recruited to the surface of cardiomyocytes in response to insulin; 2) these cells express a high level of GLUT1; and 3) intracellular GLUT4-containing vesicles consist of at least two populations, which is compatible with recently proposed models of GLUT4 trafficking in adipocytes.
Glucose enters the heart via GLUT1 and GLUT4 glucose transporters. GLUT4-deficient mice develop striking cardiac hypertrophy and die prematurely. Whether their cardiac changes are caused primarily by GLUT4 deficiency in cardiomyocytes or by metabolic changes resulting from the absence of GLUT4 in skeletal muscle and adipose tissue is unclear. To determine the role of GLUT4 in the heart we used cre-loxP recombination to generate G4H(-/-) mice in which GLUT4 expression is abolished in the heart but is present in skeletal muscle and adipose tissue. Life span and serum concentrations of insulin, glucose, FFAs, lactate, and beta-hydroxybutyrate were normal. Basal cardiac glucose transport and GLUT1 expression were both increased approximately 3-fold in G4H(-/-) mice, but insulin-stimulated glucose uptake was abolished. G4H(-/-) mice develop modest cardiac hypertrophy associated with increased myocyte size and induction of atrial natriuretic and brain natriuretic peptide gene expression in the ventricles. Myocardial fibrosis did not occur. Basal and isoproterenol-stimulated isovolumic contractile performance was preserved. Thus, selective ablation of GLUT4 in the heart initiates a series of events that results in compensated cardiac hypertrophy.
Because GLUT-4 expression is decreased whereas GLUT-1 expression is increased in denervated skeletal muscle, we examined the effects of denervation on GLUT-4 and GLUT-1 gene transcription. The right hindlimb skeletal muscle of male transgenic mice containing sequential truncations (2,400, 1,639, 1,154, and 730 bp) of the human GLUT-4 promoter linked to the chloramphenacol acyl transferase (CAT) gene was denervated, and the contralateral hindlimb was sham operated. RNase protection analysis revealed that after 72 h denervation decreased CAT mRNA and GLUT-4 mRNA levels 64-85%, respectively (P < 0.05), in the gastrocnemius muscles. In contrast, denervation of the right hindlimb of male rats increased GLUT-1 gene transcription and GLUT-1 mRNA levels by 94 and 213%, respectively (P < 0.05). In conclusion, GLUT-4 transcription is decreased but GLUT-1 transcription is increased in denervated skeletal muscle, suggesting that the effects of denervation on GLUT-4 and GLUT-1 expression are, in part, transcriptionally mediated. Furthermore, these data indicate that a DNA sequence regulated by denervation is located within 730 bp of the 5'-flanking promoter region of the human GLUT-4 gene.
The insulin-regulated glucose transporter GLUT4 was immunolocalized in rat cardiac muscle under conditions of basal and stimulated glucose uptake, achieved by fasting and a combined exercise/insulin stimulus, respectively. In basal myocytes there was very little (less than 1%) GLUT4 in the different domains of the plasma membrane (sarcolemma, intercalated disk, and transverse tubular system). GLUT4 was localized in small tubulo-vesicular elements that occur predominantly near the sarcolemma and the transverse tubular system and in the trans-Golgi region. Upon stimulation approximately 42% of GLUT4 was found in the plasma membrane. Each domain of the plasma membrane contributed equally to this effect. GLUT4-positive, clathrin-coated pits were also present at each cell surface domain. The remainder of the labeling was in tubulo-vesicular elements at the same sites as in basal cells and in the intercalated disk areas. The localization of GLUT4 in cardiac myocytes is essentially the same as in brown adipocytes, skeletal muscle, and white adipocytes. We conclude that increased glucose transport in muscle and fat is accounted for by translocation of GLUT4 from the intracellular tubulo-vesicular elements to the plasma membrane. The labeling of coated pits indicates that in stimulated myocytes, as in adipocytes, GLUT4 recycles constantly between the endosomal compartment and the plasma membrane and that stimulation of the exocytotic rate constant is likely the major mechanism for GLUT4 translocation.
In the present study we examined mRNA and protein levels for the muscle/adipose tissue glucose transporter (GLUT-4) in various tissues of spontaneously obese mice (C57BL/KsJ, db/db) and their lean littermates (db/+). Obese (db/db) mice were studied at 5 wk of age, when they were rapidly gaining weight and were severely insulin resistant, evidenced by hyperglycemia (plasma glucose 683 +/- 60 vs. 169 +/- 4 mg/dl in db/+, P less than 0.05) and hyperinsulinemia (plasma insulin 14.9 +/- 0.53 vs. 1.52 +/- 0.08 ng/ml in db/+, P less than 0.05). The GLUT-4 mRNA was reduced in quadriceps muscle (67.5 +/- 8.5%, P = 0.02), but unaltered in adipose tissue (120 +/- 19%, NS), heart (95.7 +/- 6.1%, NS), or diaphragm (75.2 +/- 12.1%, NS) in obese (db/db) mice relative to levels in lean littermates. The GLUT-4 protein, measured by quantitative immunoblot analysis using two different GLUT-4 specific antibodies, was not different in five insulin-sensitive tissues including diaphragm, heart, red and white quadriceps muscle, and adipose tissue of obese (db/db) mice compared with tissue levels in lean littermates; these findings were consistent when measured relative to tissue DNA levels as an index of cell number. These data suggest that the marked defect in glucose utilization previously described in skeletal muscle of these young obese mice is not due to a decrease in the level of the major muscle glucose transporter. An alternate step in insulin-dependent activation of the glucose transport process is probably involved.
This study deals with the effect of 5-hydroxytryptamine (5-HT; serotonin) on glucose transport in isolated rat cardiac myocytes. In these cells, 5-HT (10-300 microns), as well as tryptamine, 5-methoxytryptamine and dopamine, elicited a 3-5 fold increase in glucose transport, as compared with control. This effect was maximal after 90 min, and was concomitant with a 1.8- and 1.5-fold increase in the amounts of glucose transporters GLUT1 and GLUT4 at the cell surface of the cardiomyocytes, as determined by using the photoaffinity label 3H-2-N-[4-(1-azi-2,2,2-trifluoroethyl)benzoyl]-1,3-bis-(D-manno s-4-yl) propyl-2-amine (3H-ATB-BMPA). In contrast, 3-3000 microM of the selective 5-HT receptor agonists 5-carboxyamido-tryptamine, alpha-methyl-serotonin, 2-methyl-serotonin or renzapride failed to stimulate glucose transport. The effect of 5-HT was not affected by (i) the 5-HT receptor antagonists methysergide (1 microM), ketanserin (1 microM), cyproheptadine (1 microM), MDL 72222 (1 microM) or ICS 205-930 (3 microM), nor by (ii) the adrenergic receptor antagonists prazosin (1 microM), yohimbine (1 microM) or propranolol (5 microM), nor by (iii) the dopaminergic antagonists SCH 23390 (1 microM) or haloperidol (1 microM). The monoamine oxidase inhibitors clorgyline (1 microM) and tranylcypromine (1 microM) completely suppressed the effect of 5-HT, whereas the control and insulin-stimulated rates of glucose transport were unaffected. Addition of catalase or glutathione diminished the 5-HT-dependent stimulation of glucose transport by 50%; these two factors are known to favour the degradation of H2O2 (which can be formed during the deamination of amines by monoamine oxidases). Glutathione also depressed the stimulatory action of exogenously added H2O2 (20 microM) by 30%. Furthermore, in cells treated with 5_HT, a time-dependent accumulation of 5-hydroxy-1H-indol-3-ylacetic acid (a product of 5-HT metabolism via monoamine oxidases) was observed, which paralleled the changes in glucose transport. In conclusion, the stimulation of glucose transport by 5-HT in cardiomyocytes is not mediated by a 5-HT1, 5-HT2, 5-HT3 or 5-HT4 receptor, nor by an adrenergic or dopaminergic receptor, but is likely to occur through the degradation of by a monoamine oxidase and concomitant formation of H2O2.
We previously reported that 2400 base pairs (bp) of 5′-flanking DNA is sufficient for tissue-specific and hormonal/metabolic
regulation of the human GLUT4 gene in transgenic mice (Liu, M.-L., Olson, A. L., Moye-Rowley, W. S., Buse, J. B., Bell, G. I., and Pessin, J. E.(1992)
J. Biol. Chem. 267, 11673-11676). To further define the DNA sequences required for GLUT4 expression, we generated transgenic mice carrying 1975, 1639, 1154, 730, and 412 bp of the GLUT4 5′-flank (hG4) fused to the chloramphenicol acetyltransferase (CAT) reporter gene. The 1975-hG4-CAT, 1639-hG4-CAT, and 1154-hG4-CAT
constructs were expressed in a tissue-specific manner identical to the endogenous murine GLUT4 mRNA. Regulation of these reporter gene constructs in insulin-deficient diabetes also paralleled the endogenous gene. In
contrast, 730-hG4-CAT was expressed at high levels only in skeletal muscle and at low levels in all of the other tissues examined.
Additionally, expression of 412-hG4-CAT was completely unrestricted. Neither the 730-hG4-CAT nor the 412-hG4-CAT reporter
genes displayed any insulin-dependent regulation. These data demonstrate that a skeletal muscle-specific DNA element is located
within 730 bp of the GLUT4 5′-flanking DNA but that 1154 bp is necessary to direct the full extent of tissue-specific and insulin-dependent regulation
of the human GLUT4 gene in transgenic mice.
GLUT1 and GLUT4 glucose transporter expression is highly regulated in muscle and adipose tissue during perinatal life. Here we have investigated the role of thyroid hormones in the regulation of GLUT4 induction and GLUT1 repression associated to neonatal development. Perinatal hypothyroidism markedly impaired GLUT4 protein induction in heart. This effect was heart specific, and a greater expression of GLUT4 was detected in brown adipose tissue from neonatal hypothyroid rats compared with controls. These changes in GLUT4 protein expression were not detected in brown adipose tissue or heart when hypothyroidism was induced in adult rats. These results indicate that GLUT4 induction during perinatal life is highly sensitive to thyroid hormones in both heart and adipose tissue. Perinatal hypothyroidism was characterized by decreased cardiac GLUT4 mRNA concentrations. T3 injection caused a marked increase in cardiac levels of GLUT4 mRNA in hypothyroid neonates. Thus, in 13-day-old hypothyroid rats, GLUT4 mRNA levels increased 3-fold 1 h after T3 injection. Under these conditions, retinoic acid also caused a rapid increase in cardiac GLUT4 mRNA levels from hypothyroid neonates. In addition, cardiac levels of GLUT4 protein markedly increased in fetuses and in neonates 24 h after T3 injection. These findings suggest that a direct effect of thyroid hormones is the promotion of cardiac GLUT4 gene expression. GLUT1 protein expression was markedly enhanced in brown adipose tissue and heart during neonatal hypothyroidism as well as in hypothyroidism induced in adult rats. This was concomitant to greater levels of GLUT1 mRNA in hearts from hypothyroid neonates. Immunofluorescence analysis indicated that cardiomyocytes from hypothyroid pups contained an enhanced level of GLUT1 protein. Furthermore, T3 injection caused a decrease in cardiac levels of GLUT1 mRNA in hypothyroid neonates. These results indicate that thyroid hormone manipulation leads to inverse regulation of GLUT1 and GLUT4 glucose transporter gene expression in the neonatal heart. We conclude that thyroid hormones play a pivotal role controlling the transition of glucose transporter carriers from fetal to neonatal levels in heart and brown adipose tissue.
Our aim was to study glucose transporters GLUT1 and GLUT4 in relation to in vivo glucose uptake in rat cardiac and skeletal muscle. The levels of both transporters were of a similar order of magnitude in whole muscle tissue (GLUT1/GLUT4 ratio varied from 0.1 to 0.6), suggesting that both may have an important physiological role in regulating muscle glucose metabolism. GLUT4 correlated very strongly (r2 = 0.97) with maximal insulin-stimulated glucose uptake (Rg' max., estimated using the glucose clamp plus 2-deoxy[3H]glucose bolus technique) in six skeletal muscles and heart. A distinct difference in regulation of the two transporters was evident in heart: in 5 h-fasted rats, basal glucose uptake and GLUT1 levels in heart were very high and both were reduced, by 90 and 60% respectively, by 48 h fasting. However, in heart (and in red skeletal muscle), neither GLUT4 levels nor Rg' max. were reduced by 48 h fasting. GLUT1 was shown to be specifically expressed in cardiac myocytes, because intracellular vesicles enriched in GLUT4 contained significant levels of GLUT1. In conclusion, the high association of muscle GLUT4 content with insulin responsiveness in different muscles, and the preservation of both with fasting, supports a predominant role of GLUT4 in insulin-mediated glucose uptake. GLUT1 may play an important role in mediating cardiac muscle glucose uptake in the basal metabolic state. Marked changes in GLUT1 expression with alterations in the metabolic state, such as prolonged fasting, may play an important role in cardiac glucose metabolism.
The insulin-regulated aminopeptidase (IRAP) is a zinc-dependent membrane aminopeptidase. It is the homologue of the human placental leucine aminopeptidase. In fat and muscle cells, IRAP colocalizes with the insulin-responsive glucose transporter GLUT4 in intracellular vesicles and redistributes to the cell surface in response to insulin, as GLUT4 does. To address the question of the physiological function of IRAP, we generated mice with a targeted disruption of the IRAP gene (IRAP-/-). Herein, we describe the characterization of these mice with regard to glucose homeostasis and regulation of GLUT4. Fed and fasted blood glucose and insulin levels in the IRAP-/- mice were normal. Whereas IRAP-/- mice responded to glucose administration like control mice, they exhibited an impaired response to insulin. Basal and insulin-stimulated glucose uptake in extensor digitorum. longus muscle, and adipocytes isolated from IRAP-/- mice were decreased by 30-60% but were normal for soleus muscle from male IRAP-/- mice. Total GLUT4 levels were diminished by 40-85% in the IRAP-/- mice in the different muscles and in adipocytes. The relative distribution of GLUT4 in subcellular fractions of basal and insulin-stimulated IRAP-/- adipocytes was the same as in control cells. We conclude that IRAP-/- mice maintain normal glucose homeostasis despite decreased glucose uptake into muscle and fat cells. The absence of IRAP does not affect the subcellular distribution of GLUT4 in adipocytes. However, it leads to substantial decreases in GLUT4 expression.
Freshly isolated adult rat ventricular cardiomyocytes have been used to characterize the action profile of the new thiazolidinedione antidiabetic drug MCC-555. Preincubation of cells with the compound (100 μm for 30 min or 10 μm for 2 h) did not modify basal 3-O-methylglucose transport, but produced a marked sensitizing effect (2- to 3-fold increase in insulin action at 3 × 10⁻¹¹m insulin) and a further enhancement of maximum insulin action (1.8-fold). MCC-555 did not modulate autophosphorylation of the insulin receptor and tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1). However, insulin action (10⁻¹⁰ and 10⁻⁷m) on IRS-1-associated phosphatidylinositol (PI) 3-kinase activity was enhanced 2-fold in the presence of MCC-555. Association of the p85 adapter subunit of PI 3-kinase to IRS-1 was not modified by the drug. Immunoblotting experiments demonstrated expression of the peroxisomal proliferator-activated receptor-γ in cardiomyocytes reaching about 30% of the abundance observed in adipocytes. The insulin-sensitizing effect of MCC-555 was lost after inhibition of protein synthesis by preincubation of the cells with cycloheximide (1 mm; 30 min). Cardiomyocytes from obese Zucker rats exhibited a completely blunted response of glucose transport at 3 × 10⁻¹¹m insulin. MCC-555 ameliorates this insulin resistance, producing a 2-fold stimulation of glucose transport, with maximum insulin action being 1.6-fold higher than that in control cells. This drug effect was paralleled by a significant dephosphorylation of IRS-1 on Ser/Thr. In conclusion, MCC-555 rapidly sensitizes insulin-stimulated cardiac glucose uptake by enhancing insulin signaling resulting from increased intrinsic activity of PI 3-kinase. Acute activation of protein expression leading to a modulation of the Ser/Thr phosphorylation state of signaling proteins such as IRS-1 may be underlying this process. It is suggested that MCC-555 may provide a causal therapy of insulin resistance by targeted action on the defective site in the insulin signaling cascade.
Previously we have demonstrated that striated muscle GLUT4 gene expression decreased following streptozotocin-induced diabetes due to a loss of MEF2A transcription factor expression without any significant effect on the MEF2D isoform (Mora, S. and J. E. Pessin (2000) J Biol Chem, 275:16323–16328). In contrast to both cardiac and skeletal muscle, adipose tissue displays a selective decrease in MEF2D expression in diabetes without any significant alteration in MEF2A protein content. Adipose tissue also expresses very low levels of the MEF2 transcription factors and nuclear extracts from white adipose tissue exhibit poor in vitro binding to the MEF2 element. However, addition of in vitro synthesized MEF2A to adipose nuclear extracts results in the formation of the expected MEF2/DNA complex. More importantly, binding to the MEF2 element was also compromised in the diabetic condition. Furthermore, in vivo overexpression of MEF2A selectively in adipose tissue did not affect GLUT4 or MEF2D expression and was not sufficient to prevent GLUT4 down-regulation that occurred in insulin-deficient states.
Insulin stimulates glucose transport in muscle and adipose tissue by triggering the translocation of the glucose transporter GLUT-4 from intracellular vesicles to the cell surface. In the present study we have attempted to characterize the intracellular GLUT-4 compartment using vesicle immunoadsorption. Silver staining of this fraction indicates that this compartment contains numerous polypeptides that exhibit a marked change in mobility upon treatment with reducing agents. The polypeptide composition of GLUT-4-containing vesicles isolated from a variety of insulin-sensitive cell types, including heart, adipose tissue, skeletal muscle and 3T3-L1 adipocytes, is similar. In addition, the polypeptide composition of the GLUT-4 compartment isolated from CHO cells transfected with GLUT-4 resembles that observed in insulin-sensitive cells. Two major proteins in this vesicle fraction isolated from all cell types are the transferrin receptor (TfR) and the mannose 6-phosphate/IGF II receptor (MPR). Furthermore, vesicles immunoadsorbed from adipocytes, with antibodies specific for GLUT-4 and the TfR, also show conservation in their overall polypeptide composition. Protein micro sequencing of a major 80 kDa polypeptide enriched in the GLUT-4 compartment isolated from skeletal muscle revealed this protein to be rat transferrin, These data indicate that there is a close relationship between the intracellular GLUT-4 compartment and the endosomal system. Future studies will be required to determine if it is possible to isolate subcompartments within this system to determine if GLUT-4 is targeted to a specialized secretory compartment in insulin-sensitive cells or simply a subdomain within recycling endosomes.
Using isolated rat cardiomyocytes we have examined: 1) the effect of insulin on the cellular distribution of glucose transporter 4 (GLUT4) and GLUT1, 2) the total amount of these transporters, and 3) the co-localization of GLUT4, GLUT1, and secretory carrier membrane proteins (SCAMPs) in intracellular membranes. Insulin induced 5.7- and 2.7-fold increases in GLUT4 and GLUT1 at the cell surface, respectively, as determined by the nonpermeant photoaffinity label [H-3]2-N-[4(1-azi-2,2,2-trifluoroethyl)benzoyl] -1,3-bis-(D-mannos-4-yloxy)propyl-2-amine. The total amount of GLUT1, as determined by quantitative Western blot analysis of cell homogenates, was found to represent a substantial fraction (similar to 30%) of the total glucose transporter content. Intracellular GLUT4-containing vesicles were immunoisolated from low density microsomes by using monoclonal anti-GLUT4 (1F8) or anti-SCAMP antibodies (3FS) coupled to either agarose or acrylamide. With these different immunoisolation conditions two GLUT4 membrane pools were found in nonstimulated cells: one pool with a high proportion of GLUT4 and a low content in GLUT1 and SCAMP 39 (pool 1) and a second GLUT4 pool with a high content of GLUT1 and SCAMP 39 (pool 2). The existence of pool 1 was confirmed by immunotitration of intracellular GLUT4 membranes with 1F8-acrylamide. Acute insulin treatment caused the depletion of GLUT4 in both pools and of GLUT1 and SCAMP 39 in pool 2. In conclusion: 1) GLUT4 is the major glucose transporter to be recruited to the surface of cardiomyocytes in response to insulin; 2) these cells express a high level of GLUT1; and 3) intracellular GLUT4-containing vesicles consist of at least two populations, which is compatible with recently proposed models of GLUT4 trafficking in adipocytes.
Myocardial ischemia elicits translocation of the insulin-sensitive glucose transporter GLUT-4 from intracellular membrane stores to the sarcolemma. Because glucose metabolism is of crucial importance for post-ischemic recovery of the heart, myocardial uptake of [3H]-labeled 2-deoxyglucose and subcellular localization of GLUT-4 were determined during reperfusion in isolated rat hearts perfused with medium containing 0.4 mmpalmitate and 8 mmglucose. Hearts were subjected to 20 min of no-flow ischemia, followed by reperfusion for up to 60 min. Subcellular localization of GLUT-4 was determined by cell fractionation followed by immunoblotting. After 15 and 60 min of reperfusion uptake of 2-deoxyglucose was significantly higher (91±9 and 96±8 nmol/min/g wet weight, respectively) as compared to control values (65±1 nmol/min/g wet weight). Ischemia elicited translocation of GLUT-4 to the sarcolemma, which persisted after 15 min of reperfusion. However, after 60 min of reperfusion the subcellular distribution of GLUT-4 was similar to control hearts. In conclusion, reversal of ischemia-induced translocation of GLUT-4 to the sarcolemma is rather slow, possibly facilitating glucose uptake early during reperfusion. However, myocardial uptake and phosphorylation of 2-deoxyglucose remains enhanced late during reperfusion, when pre-ischemic distribution of GLUT-4 is almost completely restored, indicating that additional mechanisms are likely to be involved in post-ischemic stimulation of glucose uptake.
Exposure of perfused rat hearts to the second-generation sulfonylurea glyburide led to a dramatic increase in glycolytic flux and lactate production. Maximal response in the absence of insulin occurred at a concentration of 2.8 μM and resulted in a 45 percent increase in the glucose utilization rate. When insulin was included in the buffer, glyburide response was significantly increased. Similarly, glyburide potentiated the metabolic effects of insulin. Since glyburide did not promote glycogenolysis, the increase in glycolysis was caused solely by the rise in glucose utilization. The classic cross-over plot for glycolytic intermediates and transport showed that glyburide stimulates glycolysis by activating the rate-limiting enzyme phosphofructokinase and promoting glucose transport. Glycolytic intermediate data also suggested that the sulfonylurea promotes oxidation of pyruvate via the citric acid cycle. Since the drug does not alter oxygen consumption, the contribution of glucose to overall adenosine triphosphate production rises while that of fatty acids falls. These metabolic changes aid the heart in resisting an ischemic insult.
Two groups of hypothyroid rats were used; one group was given 2-mercapto-1-methylimidazole (MMI) treatment in the drinking water of the mothers and was killed at 2 and 4 days of life, and the other group was given similar MMI treatment and then was thyroidectomized at 5 days of life and killed at 8 or 20 days. Serum insulin, growth hormone (GH), and insulin-like growth factor I (IGF-I) were decreased in MMI-treated rats but increased in MMI-treated plus thyroidectomized rats. No significant reduction of thyroid hormones was observed in 2-day-old MMI rats. Protein and mRNA expression of GLUT-1 increased, and those of GLUT-4 decreased, in the heart in all populations independent of changes in insulin, GH, and IGF-I levels. However, GLUT-4 protein and mRNA expression in quadriceps and gastrocnemius skeletal muscles decreased at 4 days and increased at 8 and 20 days of life in parallel with insulin, GH, and IGF-I levels. GLUT-1 in the skeletal muscles seemed regulated posttranscriptionally and presented a decrease of mRNA expression in all stages studied. A differential sensitivity to insulin regulation of GLUT-1 and GLUT-4 glucose transporters seems to be one of the causes for the tissue-specific regulation of these glucose transporters in heart and skeletal muscles during the perinatal period.
Previous studies have shown that chronic cardiac denervation impairs myocardial glucose oxidation. To investigate this further we tested whether the tissue content of glucose transporters, activity of glycolytic enzymes or metabolic capacity of pyruvate dehydrogenase were altered. Moreover, we investigated whether the decline in glucose utilization was associated with an upregulation of proteins and enzymes involved in fatty acid handling. Chronic cardiac denervation results also in decreased left ventricular efficiency. We explored whether alterations in mitochondrial properties could be held responsible for this phenomenon.
Twelve adult dogs were included in the study. In 6 of them chronic cardiac denervation was accomplished by surgical ablation of the extrinsic nerve fibers. The other 6 dogs were sham-operated. Biopsies were obtained from the left ventricle after 4-5 weeks of denervation. The content or enzymatic activity of proteins involved in fatty acid and glucose handling was assessed. Features of glutamate oxidation were measured in freshly isolated mitochondria.
The content or activity of a set of fatty acid handling proteins did not change during chronic cardiac denervation. In contrast GLUT1 content significantly increased in the chronically denervated left ventricle, while the active form of pyruvate dehydrogenase declined (p < 0.05). Glutamate oxidation characteristics in freshly isolated mitochondria were not affected by chronic denervation.
The impairment of glucose oxidation in the chronically denervated myocardium is most likely caused by a decline of pyruvate dehydrogenase in its active form. It is unlikely that the decrease in work efficiency is caused by alterations in mitochondrial properties.
The expression of GLUT-1 (erythroid/Hep G2) and GLUT-4 (muscle/fat) glucose transporters was assessed during development in rat heart, skeletal muscle, and brown adipose tissue. GLUT-4 protein expression was detectable in fetal heart by day 21 of pregnancy; it increased progressively after birth, attaining levels close to those of adults at day 15 post natal. In contrast, GLUT-4 messenger RNA (mRNA) was already present in hearts from 17 day-old fetuses. GLUT-4 mRNA stayed low during early postnatal life in heart and brown adipose tissue and only increased after day 10 post natal. The expression pattern for GLUT-4 protein in skeletal muscle during development was comparable to that observed in heart. In contrast to heart and skeletal muscle, GLUT-4 protein in brown adipose tissue was detected in high levels (30% of adult) during late fetal life. During fetal life, GLUT-1 presented a very high expression level in brown adipose tissue, heart, and skeletal muscle. Soon after birth, GLUT-1 protein diminished progressively, attaining adult levels at day 10 in heart and skeletal muscle. GLUT-1 mRNA levels in heart followed a similar pattern to the GLUT-1 protein, being very high during fetal life and decreasing early in post natal life. GLUT-1 protein showed a complex pattern in brown adipose tissue: fetal levels were high, decreased after birth, and increased subsequently in post natal life, reaching a peak by day 9. Progesterone-induced postmaturity protected against the decrease in GLUT-1 protein associated with post natal life in skeletal muscle and brown adipose tissue. However, GLUT-4 induction was not blocked by postmaturity in any of the tissues subjected to study. These results indicate that: 1) during fetal and early post natal life, GLUT-1 is a predominant glucose transporter isotype expressed in heart, skeletal muscle, and brown adipose tissue; 2) during early post natal life there is a generalized GLUT-1 repression; 3) during development, there is a close correlation between protein and mRNA levels for GLUT-1, and therefore regulation at a pretranslational level plays a major regulatory role; 4) the onset of GLUT-4 protein induction occurs between days 20-21 of fetal life; based on data obtained in rat heart and brown adipose tissue, there is a dissociation during development between mRNA and protein levels for GLUT-4, suggesting modifications at translational or posttranslational steps; and 5) postmaturity blocks the decrease in GLUT-1 expression but not the induction of GLUT-4, observed soon after birth.(ABSTRACT TRUNCATED AT 400 WORDS)
Since thyroid hormone stimulates cardiac metabolism, for which glucose is an important fuel, we examined the effects of thyroid hormone on glucose transporter gene expression in rat heart. Treatment of hypothyroid animals with T3 for up to 6 days caused a 2-fold increase in GLUT4 mRNA at 1 day, and a 4-fold increase at 6 days (per unit DNA). GLUT4 protein, however, was not increased. In contrast, GLUT1 mRNA was transiently decreased by T3 treatment at 1 and 3 days, but returned to normal by 6 days. Concomitantly, GLUT1 protein decreased to 18% of the control value at 6 days. In chronically hypothyroid or hyperthyroid rats, GLUT4 mRNA varied directly with thyroid status, but GLUT4 protein was invariant, consistent with the acute effects of T3 treatment. Moreover, hypothyroidism increased GLUT1 mRNA and protein expression. We conclude that, in contrast to previous observations in skeletal muscle, GLUT4 protein content in rat heart is not altered by thyroid hormone. Cardiac GLUT1 expression is increased in hypothyroidism, and suppressed by thyroid hormone, at both the protein and mRNA levels. The observed discrepancy between GLUT4 mRNA and protein levels suggests that post-transcriptional regulatory events play a major role in GLUT4 expression in rat heart.
To investigate the mechanism by which cardiac glucose utilization increases during hypoxia and increased work load, we studied the effect of 2 and 14 days of hypobaric hypoxia on the expression of two subtypes of the facilitative D-glucose transporter, the GLUT-4 or "insulin-regulatable" isoform and the GLUT-1 isoform thought to mediate basal transport. Rats lose weight when exposed to hypobaric hypoxia, so fasting controls were used in the 2-day studies and pair-fed controls in the 14-day experiments. Hypobaric hypoxia (PO2 69 mmHg) resulted in right ventricular (RV), but not left ventricular (LV), hypertrophy. RV and LV GLUT-1 mRNA levels increased 2- to 3-fold after 2 days and 1.5- to 2-fold after 14 days of hypobaric hypoxia compared with both fasted rats and normal controls. RV GLUT-1 protein increased approximately 3-fold and LV GLUT-1 protein increased 1.5-fold after 14 days of hypobaric hypoxia vs. both pair-fed and normal controls. RV GLUT-4 mRNA decreased to 26% and RV GLUT-4 protein decreased to 54% of normal control levels as a result of 2 days of hypobaric hypoxia. RV GLUT-4 mRNA decreased to 64% of normal control levels with no change in RV GLUT-4 protein as a result of 2 days of fasting. We conclude that hypobaric hypoxia increases cardiac GLUT-1 expression at the pretranslational level in both ventricles. The greater increase in GLUT-1 protein on the right suggests an additive effect of pressure overload. GLUT-4 expression is reduced early in the development of RV hypertrophy.
Field stimulation of isolated adult ventricular cardiomyocytes was used to study the effect of contractile activity on 3-O-methylglucose transport and the subcellular distribution of Glut4. Cells contracting at a frequency of 1 Hz for 30 min exhibited unaltered basal and insulin-stimulated rates of glucose transport when compared to resting cells. However, at 5 Hz 3-O-methylglucose transport increased to 224% of control after 5 min. Under these conditions insulin was unable to produce a significant additional stimulation of glucose transport. Immunoblotting with an anti-Glut4 polyclonal antibody showed that both insulin and contraction (5 Hz) increased the amount of Glut4 in a plasma membrane fraction by about 8-fold with a parallel decrease in an intracellular membrane fraction by 60-65%. These data suggest the existence of an identical insulin- and contraction-recruitable Glut4 transporter pool in cardiomyocytes.
Electrical stimulation of the ventromedial hypothalamus (VMH) increased the rate constant of glucose uptake in rat heart and brown adipose tissue (BAT), as measured in vivo by the 2-deoxy-D-[3H]glucose method. The increase in glucose uptake in BAT was abolished by local sympathetic denervation. To analyze the mechanism of this hypothalamic modulation, the effects of VMH stimulation and insulin treatment on the number and dissociation constant (Kd) of glucose transporters in the plasma and microsomal membranes were examined by means of [3H]cytochalasin B binding. VMH stimulation did not alter either the number or Kd value of glucose transporters in plasma and microsomal membranes prepared from heart and BAT, whereas insulin treatment increased the number of glucose transporters in the plasma membranes and decreased those in the microsomal membranes. D-Glucose transport activity was also measured with the same plasma membrane vesicles. An apparent functional activity of transporters was detected to be increased in the heart and BAT plasma membranes after VMH stimulation but not after insulin treatment. These results suggest that VMH stimulation enhances glucose utilization in heart and BAT via sympathetic innervation and that the mechanism by which VMH stimulation increases tissue glucose uptake is different from that of insulin, possibly causing an activation of glucose transporters present in the plasma membrane.
Starvation (48 h) decreases fructose 2,6-bisphosphate (Fru-2,6-P2) concentrations and the ratio of free to acylated carnitine in hearts of euthyroid rats. These decreases, which are indicative of increased lipid fuel oxidation, are accompanied by decreased rates of glucose uptake and phosphorylation, assessed by using radioactive 2-deoxyglucose. Cardiac concentrations of acylated carnitines were increased at the expense of free carnitine even in the fed state in response to experimental hyperthyroidism, but neither Fru-2,6-P2 concentrations nor rates of glucose utilization were suppressed. Starvation (48 h) did not further increase the proportion of acylated carnitine in the heart in hyperthyroidism, and suppression of Fru-2,6-P2 concentrations and glucose utilization rates by starvation was attenuated. Although glucose utilization rates were decreased, starvation did not decrease immunoreactive GLUT 4 protein concentrations. Furthermore, although hyperthyroidism was associated with a statistically significant (30-40%) increase in relative abundance of GLUT 4 mRNA, the amount of GLUT 4 protein was not increased by hyperthyroidism in either the fed or the starved state. The results demonstrate a significant effect of hyperthyroidism to enhance cardiac glucose utilization in starvation by a mechanism which does not involve changes in GLUT 4 expression but may be secondary to changes in glucose-lipid interactions at the tissue level.
The insulin-regulatable glucose transporter (GLUT-4) is expressed in adipose tissue and in cardiac and skeletal muscle (D. E. James, R. Brown, J. Navarro, and P. F. Pilch. Nature Lond. 333: 183-185, 1988). We examined GLUT-4 development between postnatal days 1 and 41 (P1-P41) in male and female rats in these tissues by quantitative immunoblotting. GLUT-4 was detectable in each tissue at comparable levels at P1. However, the subsequent patterns of GLUT-4 development were distinctive. GLUT-4 increased in the diaphragm after P7, peaked at P20, and then declined. GLUT-4 expression in the heart increased rapidly after P7 to plateau on P41 at levels four times greater than the diaphragm. In sharp contrast, adipose tissue expression was highest between P3 and P5 but declined to a nadir at P20 before rebounding at P34. These patterns were observed for both sexes within each tissue, but female GLUT-4 expression was higher in diaphragm and heart and lower in adipose tissue. The expression of GLUT-4 appears to be regulated in a tissue-specific manner by a developmental program that may coordinate the expression of other proteins of metabolic importance.
The uptake of 2-deoxyglucose by perfused rat hearts was compared to the distribution of the insulin-regulatable glucose transporter (GLUT4) in membrane preparations from the same hearts. The hearts were treated with the alpha-adrenergic combination of epinephrine + propranolol, the beta-adrenergic agonist isoproterenol, high (8 mM) Ca2+ concentrations, insulin and the alpha adrenergic combination or insulin alone. Epinephrine (1 microM) + propranolol (10 microM), isoproterenol (10 microM), high Ca2+, insulin (1 microM) + epinephrine (1 microM) + propranolol (10 microM) and insulin (1 microM) each led to an increase in 2-deoxyglucose uptake and a shift in the recovery of the GLUT4 from a high-speed pellet membrane fraction (putatively intracellular) to a low-speed pellet membrane fraction (putatively sarcolemmal). There were significant correlations (r = -0.673, P less than 0.001) between the stimulation of 2-deoxyglucose uptake and the loss of GLUT4 from the intracellular membrane fraction, or the increase in the sarcolemmal fraction. The data provide evidence that the GLUT4 is translocated by agents that stimulate glucose transport in heart, and therefore this mechanism is not restricted to insulin.
The effect of insulin on glucose transport and glucose transporters was studied in perfused rat heart. Glucose transport was measured by the efflux of labelled 3-O-methylglucose from hearts preloaded with this hexose. Insulin stimulated 3-O-methylglucose transport by: (a) doubling the maximal velocity (Vmax); (b) decreasing the Kd from 6.9 to 2.7 mM; (c) increasing the Hill coefficient toward 3-O-methylglucose from 1.9 to 3.1; (d) increasing the efficiency of the transport process (k constant). Glucose transporters in enriched plasma and microsomal membranes from heart were quantified by the [3H]cytochalasin-B-binding assay. When added to normal hearts, insulin produced the following changes in the glucose transporters: (a) it increased the translocation of transporters from an intracellular pool to the plasma membranes; (b) it increased (from 1.6 to 2.7) the Hill coefficient of the transporters translocated into the plasma membranes toward cytochalasin B, suggesting the existence of a positive co-operativity among the transporters appearing in these membranes; (c) it increased the affinity of the transporters (and hence, possibly, of glucose) for cytochalasin B. The data provide evidence that the stimulatory effect of insulin on glucose transport may be due not to the sole translocation of intracellular glucose transporters to the plasma membrane, but to changes in the functional properties thereof.
Aspects of the regulation of the glucose transport by perfused hearts of normal rats have been studied by measuring glucose transport (via the efflux of labelled 3-O-methyl-D-glucose) and glucose transporters (via the labelled cytochalasin B binding assay). Similarly to what is observed with insulin, increasing workload (by raising perfusion pressure from 50 to 100 mm Hg) stimulated glucose transport 7 to 8-fold. Glucose (via its analog 3-O-methylglucose, used at 15 mmol/l) stimulated its own transport 4-fold. The three stimuli favored the translocation of glucose transporters from an intracellular pool (microsomes) to the plasma membrane. Insulin increased the apparent affinity (decreased dissociation constant values) of plasma membrane transporters for cytochalasin, as well as the Hill coefficient, indicating the occurrence of a positive cooperativity amongst plasma membrane transporters. Workload increased only the Hill coefficient, glucose only the apparent affinity for cytochalasin of plasma membrane transporters. This study shows that insulin, workload and glucose itself stimulate glucose transport by favouring the translocation process of glucose transporter as well as by changing, albeit by a different mechanism, the functional properties of the transporters once translocated to the plasma membrane.
Isolated cardiac myocytes from lean and genetically obese (fa/fa) Zucker rats were used to study cellular alterations related to the obesity syndrome in this tissue. Scatchard analysis of insulin binding data suggested a reduction in the number of low affinity sites in cells from obese rats; in contrast, an unaltered high affinity segment with Kd values of 5.7 +/- 0.6 and 4.5 +/- 0.7 X 10(-10) mol/liter (n = 4) in lean and obese rats, respectively, has been observed. Insulin internalization, as estimated from the amount of increased cell-associated radioactivity in chloroquine-treated cells, was decreased by 70% from 12.8 fmol insulin/10(6) cells X 120 min in lean rats to 3.8 fmol/10(6) cells X 120 min in obese rats. Determinations of initial velocities of 3-O-methylglucose influx were used for assessing glucose transport activity. Basal activity of the glucose transport system was reduced in cells from obese animals. This was found to be due to a decreased maximum velocity of the carrier with corresponding values of 69.8 +/- 5.2 and 38.3 +/- 3.2 nmol/10 sec X 10(6) cells (n = 3) in cardiocytes from lean and obese rats, respectively. Glucose transport exhibited an unaltered sensitivity toward stimulation by insulin, but an impaired responsiveness in cardiocytes from obese rats. The data suggest involvement of both receptor and postreceptor defects in the development of an insulin-resistant state in cardiac muscle.
Exposure of perfused rat hearts to the second-generation sulfonylurea glyburide led to a dramatic increase in glycolytic flux and lactate production. Maximal response in the absence of insulin occurred at a concentration of 2.8 microM and resulted in a 45 percent increase in the glucose utilization rate. When insulin was included in the buffer, glyburide response was significantly increased. Similarly, glyburide potentiated the metabolic effects of insulin. Since glyburide did not promote glycogenolysis, the increase in glycolysis was caused solely by the rise in glucose utilization. The classic cross-over plot for glycolytic intermediates and transport showed that glyburide stimulates glycolysis by activating the rate-limiting enzyme phosphofructokinase and promoting glucose transport. Glycolytic intermediate data also suggested that the sulfonylurea promotes oxidation of pyruvate via the citric acid cycle. Since the drug does not alter oxygen consumption, the contribution of glucose to overall adenosine triphosphate production rises while that of fatty acids falls. These metabolic changes aid the heart in resisting an ischemic insult.
Overall D-glucose metabolism and 3-0-methylglucose transport were measured in the perfused heart preparation of lean and genetically obese (fa/fa) rats. Absolute values of basal and insulin-stimulated glucose metabolism were decreased in hearts of 15-week-old obese rats when compared to lean age-matched controls. Basal and maximally stimulated (i.e., by the combined addition of insulin and increasing perfusion pressure) 3-0-methylglucose transport was normal in hearts from young obese rats (5-week-old). However, when only one stimulus was used (insulin or increasing perfusion pressure alone), 3-0-methylglucose transport was stimulated to values that were lower than those of lean rats. Basal 3-0-methylglucose transport was four times lower in hearts from older obese rats (15-week-old) than in lean ones of the same age. At this age, stimulation of 3-0-methylglucose transport by insulin alone, by increasing perfusion pressure alone or by the combination of both stimuli, reached values in obese rats that were only half those of lean animals. It is concluded that: (a) in the early phase of the syndrome, the basal glucose transport system in hearts of obese rats is normal, but its response to stimulation becomes abnormal and; (b) at a later phase of obesity, the glucose transport system becomes abnormal even under basal conditions and its responsiveness to various stimuli is markedly impaired.
Cardiac ventricular tissue of lean and genetically obese (fa/fa) Zucker rats was used to study the expression, subcellular distribution and insulin-induced recruitment of the glucose transporter GLUT4 and to elucidate possible molecular alterations of the translocation process. Hearts were removed from basal and insulin-treated (20 min) lean and obese Zucker rats, and processed for subcellular fractionation and Western blotting of proteins. In obese rats, the total GLUT4 content in a crude membrane fraction was reduced to 75 +/- 8% (P = 0.019) of lean controls. In contrast, GLUT4 abundance in plasma membranes was not significantly different between lean and obese rats with a concomitant decrease (47 +/- 3%) in the microsomal fraction of obese animals. In plasma membranes of lean animals insulin was found to increase the GLUT4 abundance to 294 +/- 43% of control with a significantly (P = 0.009) reduced effect in the obese group (139 +/- 10% of control). In these animals insulin failed to recruit GLUT4 from the microsomal fraction, whereas the hormone induced a significant decrease (41 +/- 4%) of microsomal GLUT4 in lean controls. In GLUT4-enriched membrane vesicles, obtained from cardiac microsomes of lean rats, a 24 kDa GTP-binding protein could be detected, whereas no significant labelling of this species was observed in GLUT4 vesicles prepared from obese animals. In addition to the translocation of GLUT4, insulin was found to promote the movement of the small GTP-binding protein rab4A from the cytosol (decrease to 61 +/- 13% of control) to the plasma membrane (increase to 177 +/- 19% of control) in lean rats with no effect of the hormone on rab4A redistribution in the obese group. In conclusion, cardiac glucose uptake of insulin-resistant obese Zucker rats is subject to multiple cellular abnormalities involving a reduced expression, altered redistribution and defective recruitment of GLUT4. We show here an association of the latter defect with alterations at the level of small GTP-binding proteins possibly leading to an impaired trafficking of GLUT4 in the insulin-resistant state.
Insulin resistance in skeletal muscle and adipose tissue often accompanies hypertension; however, it has not been shown that heart muscle is similarly affected. The aims of this study were to determine whether basal and insulin-stimulated glucose transport and glucose transporter mRNA content are altered in the spontaneously hypertensive rat (SHR) heart.
Hearts from 16-18-month-old SHRs were compared to their normotensive (WKY) controls. The accumulation of 2-deoxyglucose-6-phosphate (2DG6P), detected using 31P nuclear magnetic resonance spectroscopy, was used to assess glucose uptake before and during insulin stimulation in the isolated perfused heart. The mRNA levels of both the insulin-sensitive glucose transporter (GLUT-4) and the transporter responsible for basal glucose uptake (GLUT-1) were quantified by Northern blot analysis.
The hypertensive rat hearts exhibited hypertrophy in that the heart/body weight ratio was increased by 59%. In these hearts, the basal rate of glucose uptake was 3-fold greater and hexokinase activity was 1.6 fold greater than that of the control rat hearts. On exposure to insulin, accumulation of 2DG6P increased 5-fold in the control hearts, but only 1.4-fold in the SHR hearts. Thus, in the presence of insulin, the rate of glucose uptake by the hypertensive rat heart was significantly (P < 0.05) reduced, being 82% of control. GLUT-4 mRNA content was decreased was no significant difference in the GLUT-1 mRNA content.
We have demonstrated insulin resistance in the hypertrophied heart of the hypertensive rat that may have a molecular basis in a lower GLUT-4 content.
The insulin-sensitive glucose transporter, GLUT4, is the most abundant facilitative glucose transporter in muscle and adipose tissue, the major sites for postprandial glucose disposal. To assess the role of GLUT4 in glucose homeostasis, we have disrupted the murine GLUT4 gene. Because GLUT4 has been shown to be dysregulated in pathological states such as diabetes and obesity, it was expected that genetic ablation of GLUT4 would result in abnormal glucose homeostasis. The mice deficient in GLUT4 (GLUT4-null) are growth-retarded and exhibit decreased longevity associated with cardiac hypertrophy and severely reduced adipose tissue deposits. Blood glucose levels in female GLUT4-null mice are not significantly elevated in either the fasting or fed state; in contrast, male GLUT4-null mice have moderately reduced glycaemias in the fasted state and increased glycaemias in the fed state. However, both female and male GLUT4-null mice exhibit postprandial hyperinsulinaemia, indicating possible insulin resistance. Increased expression of other glucose transporters is observed in the liver (GLUT2) and heart (GLUT1) but not skeletal muscle. Oral glucose tolerance tests show that both female and male GLUT4-null mice clear glucose as efficiently as controls, but insulin tolerance tests indicate that these mice are less sensitive to insulin action. The GLUT4-null mice demonstrate that functional GLUT4 protein is not required for maintaining nearly normal glycaemia but that GLUT4 is absolutely essential for sustained growth, normal cellular glucose and fat metabolism, and expected longevity.
The purpose of this study was to determine the interactive effects of 10-12 wk of streptozotocin-induced diabetes (65 mg/kg) and moderate-intensity exercise training on total myocardial GLUT-4 and GLUT-1 proteins. Sprague-Dawley rats (n = 52) were randomly divided into sedentary control (SC), exercise-trained control (ETC), sedentary diabetic (SD), and exercise-trained control (ETD) groups. Diabetes (SD), and exercise-trained diabetic (ETD) groups. Diabetes resulted in a 70% reduction in myocardial GLUT-4 (28.3+/- 3.1 and 94.6 +/- 3.4% for SD and SC, respectively; P < 0.0001) and an 18.5% decrease in GLUT-1 (62.5 +/- 4.7 and 76.8 +/- 4.5% for SD and SC, respectively; P = 0.06). Exercise training increased citrate synthase activity in the medial and long heads of the triceps brachii in both groups (P < 0.001). Fasting blood glucose improved with training in diabetic animals (348 +/- 27 and 569 +/- 28 mg/dl for ETD and SD, respectively; P < 0.05). The diabetes-induced reduction in GLUT-4 was attenuated with exercise training (46.8 +/- 9.3% for ETD; P < 0.02 compared with SD). In contrast, training resulted in a further 25% decrease compared with SD in GLUT-1 in ETD (46.8 +/- 9.3%; P < 0.03 compared with SD). Exercise training had no effect on either GLUT-4 (87.2 +/- 4.0%) or GLUT-1 (75.4 +/- 5.1%) in ETC. GLUT-4 inversely correlated (r = -0.81; P < or = 0.001) with fasting blood glucose. In conclusion, diabetes resulted in a 70% reduction in myocardial GLUT-4 and an 18% decrease in GLUT-1.(ABSTRACT TRUNCATED AT 250 WORDS)
The effects of the antidiabetic drug metformin on glucose transport were investigated in freshly isolated heart muscle cells from healthy and streptozotocin-diabetic rats. In vivo treatment of diabetic rats with metformin failed to affect the basal and insulin-stimulated rate of glucose transport measured in isolated cells. In vitro exposure to therapeutic concentrations (< or = 10(-4) M) of metformin did not influence glucose transport, even upon incubation times up to 5 h or in the presence of high glucose (20 nM). In contrast, higher metformin concentrations produced an 8- to 12-fold increase in glucose uptake (with a lag of 90 min, and a maximum at 180 min and approximately 5 mM). In the presence of submaximal insulin concentrations (< or = 3.10(-10) M), the effects of metformin (5 mM) and of insulin were more than additive, whereas, at saturating insulin concentrations (10(-8) M), partial additivity was observed. Like insulin, metformin caused an approximately 1.6-fold increase in the content of both glucose transporter isoforms GLUT1 and GLUT4 in the plasma membrane of cardiac myocytes, with a corresponding decrease in an intracellular membrane fraction. cAMP-elevating treatments depressed the metformin-, but not the insulin-dependent glucose uptake, by 20-30%. In myocytes from diabetic rats, the rate of metformin-activated glucose transport was similar to that of cells from control animals, whereas basal and insulin-stimulated transport were substantially diminished. Finally, metformin (5 mM) induced a slight depression of oxygen consumption and energy metabolism of myocytes (as determined by measuring their level of energy-rich phosphates) comparable to the effects of hypoxia in rat hearts. In conclusion, these data do not provide evidence in favor of the hypothesis that glucose uptake by muscle tissue represents the site of metformin's therapeutic action in vivo. On the other hand, the large, insulin-independent effect of metformin at high concentrations (approximately mM) in vitro may be related to the action of hypoxia and occurs through a redistribution of glucose carriers from an intracellular locus to the plasma membrane. The mechanism (or signal) involved in metformin's action is likely to differ from that triggered by insulin and is not impaired in the diabetic state.
Our previous studies on the acute regulation of glucose transport in perfused rat hearts were extended to explore further the mechanism of regulation by anoxia; to test the effects of palmitate, a transport inhibitor; and to compare the translocation of two glucose transporter isoforms (GLUT1 and GLUT4). Following heart perfusions under various conditions, glucose transporters in intracellular membranes were quantitated by reconstitution of transport activity and by Western blotting. Rotenone stimulated glucose uptake and decreased the intracellular contents of glucose transporters. This indicates that it activates glucose transport via net outward translocation, similarly to anoxia. However, two uncouplers of oxidative phosphorylation produced little or no effect. Increased workload (which stimulates glucose transport) reduced the intracellular contents of transporters, while palmitate increased the contents, indicating that these factors cause net translocation from or to the intracellular pool, respectively. Relative changes in GLUT1 were similar to those in GLUT4 for most factors tested. A plot of changes in total intracellular transporter content vs. changes in glucose uptake was roughly linear, with a slope of -0.18. This indicates that translocation accounts for most of the changes in glucose transport, and the basal pool of intracellular transporters is five times as large as the plasma membrane pool.
Subcellular fractions obtained from rat cardiac ventricular tissue were used to elucidate a possible functional relationship between small-molecular-mass G-proteins and the insulin-responsive glucose transporter GLUT4. Proteins were separated by SDS/PAGE and transferred to nitrocellulose membranes. Incubation with [alpha-32P]GTP revealed the presence of two major distinct GTP-binding protein bands of 24 and 26 kDa in both plasma and microsomal membranes. Immunoadsorption of microsomal membranes to anti-GLUT4 antibodies was used to isolate GLUT4-enriched membrane vesicles. This material was found to contain a much decreased amount of small G-proteins, with the exclusive presence of the 24 kDa species. Insulin treatment in vivo had no effect on the microsomal membrane content of small GTP-binding proteins, but significantly decreased the 24 kDa species in GLUT4-enriched vesicles by 36 +/- 5% (n = 3). This correlated with a decreased (30-40%) recovery of GLUT4-enriched vesicles from insulin-treated animals. Western-blot analysis of microsomal membranes with a panel of antisera against rab GTP-binding proteins indicated the presence of rab4A, with a molecular mass of 24 kDa, whereas rab1A, rab2 and rab6 were not observed. rab4A was barely detectable in GLUT4-enriched vesicles; however, insulin produced an extensive shift of rab4A from the cytosol and the microsomal fraction to the plasma membrane with a parallel increase in GLUT4. These data show that a small GTP-binding protein is co-localized with GLUT4 in an insulin-responsive intracellular compartment, and strongly suggest that this protein is involved in the exocytosis of GLUT4 in cardiac muscle. Furthermore, the observed translocation of rab4A is compatible with insulin-induced endosome recycling processes, possibly including the glucose transporters.
The effects of the fatty acid inhibitor 4-bromocrotonic acid (4-BCA) on glucose utilization was studied in isolated rat myocytes. In contrast to its potent inhibition of [1-14C]palmitate oxidation, 4-BCA strongly stimulated the oxidation of [1-14C]glucose and [2-14C]-pyruvate in a concentration-dependent manner. At a concentration of 300 microM, 4-BCA increased glucose oxidation threefold and that of pyruvate oxidation twofold. The rate of transport of [U-14C]-2-deoxyglucose was significantly stimulated by 4-BCA. The transport of 2-deoxyglucose was increased sevenfold with 200 microM 4-BCA, whereas insulin (10 microU)/ml enhanced 2-deoxyglucose transport twofold. The addition of insulin to myocytes preincubated with 4-BCA did not further increase glucose transport. Cytochalasin B and anti-GLUT 4 antibody decreased the 4-BCA-induced stimulation of glucose transport. These results suggest that the stimulation of 2-deoxy-glucose transport by 4-BCA occurs through an increase in the activity of insulin-responsive glucose transporters, GLUT 4, in the sarcolemmal membrane.
The study of the regulation of glucose utilization by inhibition of fatty acid oxidation is greatly enhanced by the availability of specific inhibitors of fatty acid oxidation. This study examines the regulation of cardiac glucose utilization by inhibition of fatty acid oxidation at different sites. The effects of Etomoxir and 4-bromocrotonic acid (4-BCA) on the oxidation of [1-14C]palmitate, [1-14C]-octanoate and [U-14C]glucose were studied in isolated rat myocytes. Fifty percent inhibition of palmitate oxidation was achieved at 8 microM Etomoxir and 40 microM 4-BCA. Octanoate oxidation was inhibited only by 4-BCA. In contrast to their effect on palmitate oxidation, these inhibitors significantly stimulated the oxidation of glucose in a concentration-dependent manner. Moreover, the oxidation of [2-14C]pyruvate was increased two-fold by these compounds. The rate of utilization of [U-14C]-2-deoxyglucose was also stimulated 2-3 times by these inhibitors. These studies suggest that the stimulation of glucose utilization via the inhibition of fatty acid oxidation may be mediated through the stimulation of both glucose transport and the oxidation of pyruvate by the pyruvate dehydrogenase complex.
We compared the expression and cell-type localization of GLUT-1 mRNA and protein between cardiac and skeletal muscle of normal rats. Also, since we recently showed that cardiac GLUT-1 is upregulated in rats exposed to hypobaric hypoxia, we examined the cellular localization of GLUT-1 in cardiac tissue of normal and hypoxic rats. Confocal light microscopy and double immunofluorescent labeling revealed intense localization of GLUT-1 around neurofilament immunoreactivity within gastrocnemius muscle consistent with the previously described localization of large amounts of GLUT-1 in perineurial sheaths of skeletal muscle. However, using the same methods, we were unable to visualize GLUT-1 adjacent to nerve fibers in numerous sections of right or left ventricles or atria. Compared with skeletal myoctes, however, GLUT-1 immunofluorescence among cardiomyocytes was much more intense, particularly along the plasma membrane and especially intercalated discs. GLUT-1 immunofluorescence was also seen within the walls of arterioles within the heart. The predominant localization of GLUT-1 expression to cardiomyocytes in heart tissue was confirmed by in situ mRNA hybridization to digoxigenin-conjugated GLUT-1 cDNA. Northern blot analysis demonstrated that GLUT-1 mRNA was increased severalfold in the cardiac tissues compared with skeletal muscle. Although we detected GLUT-1 protein by immunoblotting of detergent extracts of the heart, we could not detect GLUT-1 in similar extracts of skeletal muscle. The cell type distribution of GLUT-1 in hearts of hypoxic rats was not different by immunohistochemistry from normals. These data indicate that 1) the cell-type distribution of GLUT-1 in the heart differs markedly from that in skeletal muscle. GLUT-1 in cardiac tissue, unlike skeletal muscle, is predominantly expressed within myocytes. 2) Cardiac GLUT-1 is not located along nerve fibers. 3) GLUT-1 mRNA and protein levels in cardiac tissue are considerably greater than in skeletal muscle. 4) The hypoxia-induced increase in cardiac GLUT-1 that we previously reported must occur within cardiomyocytes.
The in vivo glucose uptake and the levels of two glucose transporter proteins (GLUT1 and GLUT4) were measured in heart and in various types of skeletal muscle from streptozotocin-diabetic rats. Diabetes (12-16 weeks) reduced the in vivo glucose uptake (glucose metabolic index, GMI), and the levels of GLUT1 and GLUT4 in heart by 75%, 60% and 70%, respectively. In diaphragm consisting of approximately equal amounts of type I (slow-contracting oxidative), IIa (fast-contracting oxidative) and IIb (fast-contracting glycolytic) fibers, GMI and GLUT4 levels were reduced by 60% and 40%, respectively, with no change in GLUT1 levels. In muscle consisting mainly of type I fibers (e.g., m. soleus), GMI and GLUT4 levels were reduced by 60% and 30%, respectively, whereas GLUT1 levels were unaltered. In mixed-type muscle consisting of type IIa and IIb fibers (e.g., m. plantaris and red part of m. gastrocnemius), GMI and GLUT1 levels were unchanged, whereas GLUT4 levels were decreased by 45%. In contrast, GMI was increased by 100% in type IIb fibers (e.g., the white part of m. gastrocnemius), probably reflecting the 4-fold increase in blood glucose levels, whereas GLUT4 levels were lowered by 55% with no change in GLUT1 levels. These data demonstrate a marked difference in the response of in vivo glucose uptake to long-term hypoinsulinemia between oxidative (type I) and glycolytic (type IIb) fibers. Furthermore, in contrast to the GLUT4, GLUT1 levels are regulated differentially in heart and skeletal muscle in response to streptozotocin-induced diabetes.
Acute myocardial ischemia is accompanied by an increase in glucose uptake and metabolism, which appears to be important in protecting myocardial cells from irreversible ischemic injury. Because insulin augments myocardial glucose uptake by inducing the translocation of glucose transporters from an intracellular compartment to the plasma membrane, we hypothesized that acute ischemia would trigger a similar translocation.
We used a subcellular fractionation method to separate intracellular membrane and plasma membranes from control, ischemic, and hypoxic Langendorff-isolated perfused rat hearts and determined the expression of the major myocardial glucose transporter, GLUT4, in these separated membrane fractions. We found that translocation of GLUT4 molecules occurred in ischemic, hypoxic, and insulin-treated hearts and in hearts that underwent ischemia plus insulin treatment. The percentages of GLUT4 molecules present on the plasma membrane in the different conditions were as follows: control, 18.0 +/- 2.8%; ischemia, 41.3 +/- 9.4%; hypoxia, 31.1 +/- 2.9%; insulin, 61.1 +/- 2.6%; and ischemia plus insulin, 66.8 +/- 5.7%. Among the statistically significant differences in these values were the difference between control and ischemia and the difference between ischemia alone and insulin plus ischemia.
Ischemia causes substantial translocation of GLUT4 molecules to the plasma membrane of cardiac myocytes. A combination of insulin plus ischemia stimulates an even greater degree of GLUT4 translocation. GLUT4 translocation is likely to mediate at least part of the increased glucose uptake of ischemic myocardium and may be a mechanism for the cardioprotective effect of insulin during acute myocardial ischemia.
Age-related changes in glucose metabolism and glucose transporter protein content have been described in adipose tissue and
skeletal muscle, two tissues that express the GLUT-4 isoform of the glucose transporter protein. I studied the effect of age
on the levels of GLUT-4 protein in a third insulin-sensitive tissue: the heart. Cardiac ventricles were sampled from male
Fischer 344/Brown Norway F1 hybrid (F344/BNNia) rats. The total protein concentration of the left ventricle did not change
with age. GLUT-4 levels per mg of protein declined by 15% between 3.5 and 13 months of age, and by another 12% during adulthood
(between 13 and 25 months of age); when expressed per g wet weight, the decreases were 13% and 17%, respectively. Linear regression
analysis revealed a significant (p < .0001) relationship (r2 = .634) between age and myocardial GLUT-4. These results demonstrate
that the GLUT-4 levels in the left ventricle decrease in an age-related fashion and suggest that the capacity for glucose
transport might also be reduced.
Extrapancreatic activity of the sulfonylurea, glipizide, was evaluated in the neonatal streptozotocin-induced rat model of noninsulin-dependent diabetes. Two day old Wistar rats were given a bolus of streptozotocin (90 mg/kg i.p.) to cause noninsulin-dependent diabetes; these animals became severely glucose intolerant and eventually developed a cardiomyopathy characterized by reduced heart rate, contractility and cardiac output. Male littermates injected with citrate buffer served as nondiabetic controls. At four weeks of age, the nondiabetic and NIDD rats were administered by gavage either glipizide (2.5 mg/kg) or the methyl cellulose vehicle. Throughout the treatment protocol, no difference in the degree of glucose intolerance was observed between the glipizide-treated and vehicle-treated animals. Glipizide therapy also was ineffective in improving plasma insulin levels, which were significantly depressed in the diabetic group. Yet, animals treated with glipizide for one year exhibited improved myocardial contractile function relative to the vehicle-fed or ad lib fed diabetic animals. Heart rate was significantly elevated and there was a tendency for both the rate of relaxation and contractility to be elevated in sulfonylurea-treated group. Glipizide also reduced the degree of insulin resistance in the heart. Since these changes occur in the absence of changes in glucose tolerance or insulin levels, the heart appears to be very sensitive to the direct effects of the sulfonylureas.
Biochemical mechanisms underlying impaired myocardial glucose utilization in diabetes mellitus have not been elucidated. We studied sarcolemmal vesicles (SL) in control, streptozotocin-induced diabetic (D), and insulin-treated diabetic (Tx) rats and found that 3-O-methylglucose transport rates were decreased 53% in D rats and were normalized by insulin therapy. Immunoblot analyses of SL revealed that GLUT4 glucose transporters were decreased 56% in D and were normal in Tx rats. Thus diminished transport rates could be fully explained by reduced numbers of SL GLUT4 with normal functional activity. To determine whether SL GLUT4 were decreased due to tissue depletion or abnormal subcellular distribution, we measured GLUT4 in total membranes (SL plus intracellular fractions). Total GLUT4 (per mg membrane protein or per DNA) was decreased 45-51% in D [half time = 3.5 days after streptozotocin], and these values were restored to normal in Tx rats. Also, diabetes decreased GLUT4 mRNA levels by 43%, and this effect was reversed by insulin therapy. We conclude that, in diabetes, 1) impaired myocardial glucose utilization is the result of a decrease in glucose transport activity, and 2) transport rates are reduced due to pretranslational suppression of GLUT4 gene expression and can be corrected by insulin therapy. GLUT4 depletion could limit glucose availability under conditions of increased workload and anoxia and could cause myocardial dysfunction.
The tissue distribution of the GLUT3 glucose transporter protein was examined in human tissues using a rabbit antiserum directed against the C-terminal peptide sequence of human GLUT3. This anti-serum was shown to recognize the human GLUT3 protein in Chinese hamster ovary cells transfected with GLUT3 cDNA and to immunoprecipitate an authentic glucose transport protein in brain and testis membranes, as assessed by glucose-inhibitable photolabeling with [3H] cytochalasin-B. The GLUT3 protein, migrating with an apparent mol wt of approximately 48 kilodaltons, was strongly expressed in brain and testis membranes as well as in spermatozoa. It was not detectable in membranes from erythrocytes, adipocytes, heart, skeletal muscle, liver, kidney, spleen, thyroid, and prostate. Very low levels may be present in placenta. In brain, GLUT3 protein was strongly expressed in grey matter regions and was only weakly expressed in white matter, suggesting that it may be important in providing glucose to regions of high metabolic activity, i.e. to areas associated with synaptic transmission. None was found in peripheral (femoral) nerve. It appeared to be stable for up to 47 h in autopsy brain tissue kept at 4 C. The tissue distribution of human GLUT3 protein thus appears to be highly restricted (brain and testis/spermatozoa), in contrast with a previous report. Its function may be to provide a high affinity glucose transport system in cells that are highly dependent on glucose as a fuel source.
Insulin stimulates glucose transport in muscle and adipose tissue by triggering the translocation of the glucose transporter GLUT-4 from intracellular vesicles to the cell surface. In the present study we have attempted to characterize the intracellular GLUT-4 compartment using vesicle immunoadsorption. Silver staining of this fraction indicates that this compartment contains numerous polypeptides that exhibit a marked change in mobility upon treatment with reducing agents. The polypeptide composition of GLUT-4-containing vesicles isolated from a variety of insulin-sensitive cell types, including heart, adipose tissue, skeletal muscle and 3T3-L1 adipocytes, is similar. In addition, the polypeptide composition of the GLUT-4 compartment isolated from CHO cells transfected with GLUT-4 resembles that observed in insulin-sensitive cells. Two major proteins in this vesicle fraction isolated from all cell types are the transferrin receptor (TfR) and the mannose 6-phosphate/IGF II receptor (MPR). Furthermore, vesicles immunoadsorbed from adipocytes, with antibodies specific for GLUT-4 and the TfR, also show conservation in their overall polypeptide composition. Protein micro sequencing of a major 80 kDa polypeptide enriched in the GLUT-4 compartment isolated from skeletal muscle revealed this protein to be rat transferrin. These data indicate that there is a close relationship between the intracellular GLUT-4 compartment and the endosomal system. Future studies will be required to determine if it is possible to isolate subcompartments within this system to determine if GLUT-4 is targeted to a specialized secretory compartment in insulin-sensitive cells or simply a subdomain within recycling endosomes.
There is some evidence that inhibition of angiotensin-converting enzyme (ACE) activity can improve the uptake and conversion of glucose by heart and skeletal muscle in diabetes. To study the underlying mechanisms, we treated streptozotocin-induced diabetic rats with the angiotensin type 1 receptor (AT1) antagonist ICI D8731 and the ACE inhibitor fosinopril for 4 months and determined the expression of the myocardial glucose transporter proteins. In diabetic rats, the expression of the insulin-regulated glucose transporter (GLUT4) was strongly diminished as shown by Western and Northern blots. ICI D8731 prevented the decrease of GLUT4 protein in diabetes but had no influence on the amount of mRNA encoding for GLUT1 and GLUT4. GLUT1 protein was hardly detected in the rat heart and was affected neither by diabetes nor by treatment with the AT1 antagonist. Additionally, ICI D8731 influenced the translocation of GLUT4 from the intracellular pool to the plasma membrane, because treatment increased the amount of GLUT4 protein in the plasma membranes as well as in intracellular membrane fractions compared with membranes of untreated diabetic control rats. In contrast, inhibition of ACE by fosinopril influenced neither the expression nor the translocation of the glucose transporter proteins. These observations indicate that angiotensin II has a distinct influence on the post-transcriptional regulation of the GLUT4 transporter protein, presumably indirectly as a consequence of hemodynamic effects and structural alterations of the vessel wall.