Metabolic Flexibility of the Heart: The Role of Fatty Acid Metabolism in Health, Heart Failure, and Cardiometabolic Diseases.

This comprehensive review explores the critical role of fatty acid (FA) metabolism in cardiac diseases, particularly heart failure (HF), and the implications for therapeutic strategies. The heart's reliance on ATP, primarily sourced from mitochondrial oxidative metabolism, underscores the significance of metabolic flexibility, with fatty acid oxidation (FAO) being a dominant source. In HF, metabolic shifts occur with an altered FA uptake and FAO, impacting mitochondrial function and contributing to disease progression. Conditions like obesity and diabetes also lead to metabolic disturbances, resulting in cardiomyopathy marked by an over-reliance on FAO, mitochondrial dysfunction, and lipotoxicity. Therapeutic approaches targeting FA metabolism in cardiac diseases have evolved, focusing on inhibiting or stimulating FAO to optimize cardiac energetics. Strategies include using CPT1A inhibitors, using PPARα agonists, and enhancing mitochondrial biogenesis and function. However, the effectiveness varies, reflecting the complexity of metabolic remodeling in HF. Hence, treatment strategies should be individualized, considering that cardiac energy metabolism is intricate and tightly regulated. The therapeutic aim is to optimize overall metabolic function, recognizing the pivotal role of FAs and the need for further research to develop effective therapies, with promising new approaches targeting mitochondrial oxidative metabolism and FAO that improve cardiac function.

Transcriptional analysis reveals that the intracellular lipid accumulation impairs gene expression profiles involved in insulin response-associated cardiac functionality.

Cardiovascular disease (CVD) is a multisystemic and multicellular pathology that is generally associated with high levels of atherogenic lipoproteins in circulation. These lipoproteins tend to be retained and modified, for example, aggregated low-density lipoprotein (aggLDL), in the extracellular matrix of different tissues, such as the vascular wall and heart. The uptake of aggLDL generates a significant increase in cholesteryl ester (CE) in these tissues. We previously found that the accumulation of CE generates alterations in the insulin response in the heart. Although the insulin response is mainly associated with the uptake and metabolism of glucose, other studies have shown that insulin would fulfill functions in this tissue, such as regulating the calcium cycle and cardiac contractility. Here, we found that aggLDL induced-lipid accumulation altered the gene expression profile involved in processes essential for cardiac functionality, including insulin response and glucose uptake (Insr, Ins1, Pik3ip1, Slc2a4 gene expression), calcium cycle (Cacna1s and Gjc2 gene expression) and calcium-dependent cardiac contractility (Myh3), and cholesterol efflux (Abca1), in HL-1 cardiomyocytes. These observations were recapitulated using an in vivo model of hypercholesterolemic ApoE-KO mice. Altogether, these results may explain the deleterious effect of lipid accumulation in the myocardium, with important implications for lipid-overloaded associated CVD, including impaired insulin response, disrupted lipid metabolism, altered cardiac structure, and increased susceptibility to cardiovascular events.

Inhibitory Effects of LRP1-Based Immunotherapy on Cardiac Extracellular Matrix Biophysical Alterations Induced by Hypercholesterolemia.

The accumulation of lipids in cardiomyocytes contributes to cardiac dysfunction. The specific blockage of cardiomyocyte cholesteryl ester (CE) loading by antibodies (Abs) against the P3 sequence (Gly1127-Cys1140) of the LRP1 receptor improves cardiac insulin sensitivity. The impact of anti-P3 Abs on high-fat diet (HFD)-induced cardiac extracellular matrix (ECM) biophysical alterations was analyzed. Both IrP (without Abs) and P3-immunized rabbits (with Abs) were randomized into groups fed either HFD or a standard chow diet. Cardiac lipids, proteins, and carbohydrates were characterized by Fourier transform infrared spectroscopy in the attenuated total reflectance mode. The hydric organization and physical structure were determined by differential scanning calorimetry. HFD increased the levels of esterified lipids, collagen, and α-helical structures and upregulated fibrosis, bound water, and ECM plasticization in the heart. The inhibitory effect of anti-P3 Abs on cardiac CE accumulation was sufficient to reduce the collagen-filled extracellular space, the level of fibrosis, and the amount of bound water but did not counteract ECM plasticization in the heart of hypercholesterolemic rabbits.

Decreased low-density lipoprotein receptor-related protein 1 expression in pro-inflammatory monocytes is associated with subclinical atherosclerosis.

Subclinical atherosclerosis (SCA) occurs in asymptomatic individuals. Blood peripheral monocytes are involved in the development of atherosclerosis. Circulating monocytes acquire pro-inflammatory profiles, and they are involved in the early stages of atherosclerosis development. Low-density lipoprotein Receptor-related Protein 1 (LRP1) is expressed in monocytes, mainly in classical and intermediate subsets. Although LRP1 is highly expressed in macrophages and vascular smooth muscle cells (VSMCs) in atherosclerotic plaque formation, its expression in circulating monocytes has not been studied in SCA. The aim of this study was to characterize the LRP1 expression level in circulating monocytes of individuals with SCA and compared with individuals with low (LR) and intermediate (IR) risk of cardiovascular diseases, both without evidence of atherosclerotic lesions in carotid and coronary arteries. LRP1 and additional markers (CD11b, CD11c, and CD36) at cell surface of monocytes were analyzed by flow cytometry assays, whereas LRP1 and pro-inflammatory factors gene expressions were measured in isolated monocytes by quantitative RT-PCRs. Both LRP1 protein and LRP1 mRNA were significantly reduced in monocytes in SCA and IR respect to LR. Conversely, CD36, CD11b, and CD11c monocytic markers showed no significant changes between the different study groups. Finally, increased gene expressions of TNF-α and IL-1ß were detected in monocytes of SCA, which were associated with decreased LRP1 expression at the cell surface in total monocytes. In summary, we propose that the decreased LRP1 expression at cell surface in total monocytes with pro-inflammatory profile is associated with the development of atherosclerosis in asymptomatic individuals.

Targeting cholesteryl ester accumulation in the heart improves cardiac insulin response.

BACKGROUND: Antibodies against the P3 sequence (Gly1127-Cys1140) of LRP1 (anti-P3 Abs) specifically block cholesteryl ester (CE) accumulation in vascular cells. LRP1 is a key regulator of insulin receptor (InsR) trafficking in different cell types. The link between CE accumulation and the insulin response are largely unknown. Here, the effects of P3 peptide immunization on the alterations induced by a high-fat diet (HFD) in cardiac insulin response were evaluated. METHODS: Irrelevant (IrP)- or P3 peptide-immunized rabbits were randomized into groups fed either HFD or normal chow. Cardiac lipid content was characterized by thin-layer chromatography, confocal microscopy, and electron microscopy. LRP1, InsR and glucose transporter type 4 (GLUT4) levels were determined in membranes and total lysates from rabbit heart. The interaction between InsR and LRP1 was analyzed by immunoprecipitation and confocal microscopy. Insulin signaling activity and glucose uptake were evaluated in HL-1 cells exposed to rabbit serum from the different groups. FINDINGS: HFD reduces cardiac InsR and GLUT4 membrane levels and the interactions between LRP1/InsR. Targeting the P3 sequence on LRP1 through anti-P3 Abs specifically reduces CE accumulation in the heart independently of changes in the circulating lipid profile. This restores InsR and GLUT4 levels in cardiac membranes as well as the LRP1/InsR interactions of HFD-fed rabbits. In addition, anti-P3 Abs restores the insulin signaling cascade and glucose uptake in HL-1 cells exposed to hypercholesterolemic rabbit serum. INTERPRETATION: LRP1-immunotargeting can block CE accumulation within the heart with specificity, selectivity, and efficacy, thereby improving the cardiac insulin response; this has important therapeutic implications for a wide range of cardiac diseases. FUNDING: Fundació MARATÓ TV3: grant 101521-10, Instiuto de Salud Carlos III (ISCIII) and ERDFPI18/01584, Fundación BBVA Ayudas a Equipos de Investigación 2019. SECyT-UNC grants PROYECTOS CONSOLIDAR 2018-2021; FONCyT, Préstamo BID PICT grant 2015-0807 and grant 2017-4497.

Activated Alpha-2 Macroglobulin Improves Insulin Response via LRP1 in Lipid-Loaded HL-1 Cardiomyocytes.

Activated alpha-2 Macroglobulin (α2M*) is specifically recognized by the cluster I/II of LRP1 (Low-density lipoprotein Receptor-related Protein-1). LRP1 is a scaffold protein for insulin receptor involved in the insulin-induced glucose transporter type 4 (GLUT4) translocation to plasma membrane and glucose uptake in different types of cells. Moreover, the cluster II of LRP1 plays a critical role in the internalization of atherogenic lipoproteins, such as aggregated Low-density Lipoproteins (aggLDL), promoting intracellular cholesteryl ester (CE) accumulation mainly in arterial intima and myocardium. The aggLDL uptake by LRP1 impairs GLUT4 traffic and the insulin response in cardiomyocytes. However, the link between CE accumulation, insulin action, and cardiac dysfunction are largely unknown. Here, we found that α2M* increased GLUT4 expression on cell surface by Rab4, Rab8A, and Rab10-mediated recycling through PI3K/Akt and MAPK/ERK signaling activation. Moreover, α2M* enhanced the insulin response increasing insulin-induced glucose uptake rate in the myocardium under normal conditions. On the other hand, α2M* blocked the intracellular CE accumulation, improved the insulin response and reduced cardiac damage in HL-1 cardiomyocytes exposed to aggLDL. In conclusion, α2M* by its agonist action on LRP1, counteracts the deleterious effects of aggLDL in cardiomyocytes, which may have therapeutic implications in cardiovascular diseases associated with hypercholesterolemia.

LRP1 mediates the IGF-1-induced GLUT1 expression on the cell surface and glucose uptake in Müller glial cells.

Insulin-like Growth Factor-1 (IGF-1) is involved in the normal development and survival of retinal cells. Low-density lipoprotein Receptor-related Protein-1 (LRP1) plays a key role on the regulation of several membrane proteins, such as the IGF-1 receptor (IGF-1R). In brain astrocytes, LRP1 interact with IGF-1R and the glucose transporter type 1 (GLUT1), regulating the glucose uptake in these cells. Although GLUT1 is expressed in retinal Müller Glial Cells (MGCs), its regulation is not clear yet. Here, we investigated whether IGF-1 modulates GLUT1 traffic to plasma membrane (PM) and glucose uptake, as well as the involvement of LRP1 in this process in the human Müller glial-derived cell line (MIO-M1). We found that IGF-1 produced GLUT1 translocation to the PM, in a time-dependent manner involving the intracellular signaling activation of MAPK/ERK and PI3K/Akt pathways, and generated a significant glucose uptake. Moreover, we found a molecular association between LRP1 and GLUT1, which was significantly reduced by IGF-1. Finally, cells treated with specific siRNA for LRP1 showed an impaired GLUT1 expression on PM and decreased glucose uptake induced by IGF-1. We conclude that IGF-1 regulates glucose homeostasis in MGCs involving the expression of LRP1.

LRP1-Mediated AggLDL Endocytosis Promotes Cholesteryl Ester Accumulation and Impairs Insulin Response in HL-1 Cells.

Abstract: The cardiovascular disease (CVD) frequently developed during metabolic syndrome and type-2 diabetes mellitus is associated with increased levels of aggregation-prone small LDL particles. Aggregated LDL (aggLDL) internalization is mediated by low-density lipoprotein receptor-related protein-1 (LRP1) promoting intracellular cholesteryl ester (CE) accumulation. Additionally, LRP1 plays a key function in the regulation of insulin receptor (IR) and glucose transporter type 4 (GLUT4) activities. Nevertheless, the link between LRP1, CE accumulation, and insulin response has not been previously studied in cardiomyocytes. We aimed to identify mechanisms through which aggLDL, by its interaction with LRP1, produce CE accumulation and affects the insulin-induced intracellular signaling and GLUT4 trafficking in HL-1 cells. We demonstrated that LRP1 mediates the endocytosis of aggLDL and promotes CE accumulation in these cells. Moreover, aggLDL reduced the molecular association between IR and LRP1 and impaired insulin-induced intracellular signaling activation. Finally, aggLDL affected GLUT4 translocation to the plasma membrane and the 2-NBDG uptake in insulin-stimulated cells. We conclude that LRP1 is a key regulator of the insulin response, which can be altered by CE accumulation through LRP1-mediated aggLDL endocytosis.

Activated α2-Macroglobulin Regulates LRP1 Levels at the Plasma Membrane through the Activation of a Rab10-dependent Exocytic Pathway in Retinal Müller Glial Cells.

Activated α2-macroglobulin (α2M*) and its receptor, low-density lipoprotein receptor-related protein 1 (LRP1), have been linked to proliferative retinal diseases. In Müller glial cells (MGCs), the α2M*/LRP1 interaction induces cell signaling, cell migration, and extracellular matrix remodeling, processes closely associated with proliferative disorders. However, the mechanism whereby α2M* and LRP1 participate in the aforementioned pathologies remains incompletely elucidated. Here, we investigate whether α2M* regulates both the intracellular distribution and sorting of LRP1 to the plasma membrane (PM) and how this regulation is involved in the cell migration of MGCs. Using a human Müller glial-derived cell line, MIO-M1, we demonstrate that the α2M*/LRP1 complex is internalized and rapidly reaches early endosomes. Afterward, α2M* is routed to degradative compartments, while LRP1 is accumulated at the PM through a Rab10-dependent exocytic pathway regulated by PI3K/Akt. Interestingly, Rab10 knockdown reduces both LRP1 accumulation at the PM and cell migration of MIO-M1 cells induced by α2M*. Given the importance of MGCs in the maintenance of retinal homeostasis, unravelling this molecular mechanism can potentially provide new therapeutic targets for the treatment of proliferative retinopathies.

The Role of Low-Density Lipoprotein Receptor-Related Protein 1 in Lipid Metabolism, Glucose Homeostasis and Inflammation.

Metabolic syndrome (MetS) is a highly prevalent disorder which can be used to identify individuals with a higher risk for cardiovascular disease and type 2 diabetes. This metabolic syndrome is characterized by a combination of physiological, metabolic, and molecular alterations such as insulin resistance, dyslipidemia, and central obesity. The low-density lipoprotein receptor-related protein 1 (LRP1­A member of the LDL receptor family) is an endocytic and signaling receptor that is expressed in several tissues. It is involved in the clearance of chylomicron remnants from circulation, and has been demonstrated to play a key role in the lipid metabolism at the hepatic level. Recent studies have shown that LRP1 is involved in insulin receptor (IR) trafficking and intracellular signaling activity, which have an impact on the regulation of glucose homeostasis in adipocytes, muscle cells, and brain. In addition, LRP1 has the potential to inhibit or sustain inflammation in macrophages, depending on its cellular expression, as well as the presence of particular types of ligands in the extracellular microenvironment. In this review, we summarize existing perspectives and the latest innovations concerning the role of tissue-specific LRP1 in lipoprotein and glucose metabolism, and examine its ability to mediate inflammatory processes related to MetS and atherosclerosis.

Insulin-induced exocytosis regulates the cell surface level of low-density lipoprotein-related protein-1 in Müller Glial cells.

Low-density lipoprotein (LDL) receptor-related protein-1 (LRP1) is expressed in retinal Müller glial cells (MGCs) and regulates intracellular translocation to the plasma membrane (PM) of the membrane proteins involved in cellular motility and activity. Different functions of MGCs may be influenced by insulin, including the removal of extracellular glutamate in the retina. In the present work, we investigated whether insulin promotes LRP1 translocation to the PM in the Müller glial-derived cell line MIO-M1 (human retinal Müller glial cell-derived cell line). We demonstrated that LRP1 is stored in small vesicles containing an approximate size of 100 nm (mean diameter range of 100-120 nm), which were positive for sortilin and VAMP2, and also incorporated GLUT4 when it was transiently transfected. Next, we observed that LRP1 translocation to the PM was promoted by insulin-regulated exocytosis through intracellular activation of the IR/PI3K/Akt axis and Rab-GTPase proteins such as Rab8A and Rab10. In addition, these Rab-GTPases regulated both the constitutive and insulin-induced LRP1 translocation to the PM. Finally, we found that dominant-negative Rab8A and Rab10 mutants impaired insulin-induced intracellular signaling of the IR/PI3K/Akt axis, suggesting that these GTPase proteins as well as the LRP1 level at the cell surface are involved in insulin-induced IR activation.