Autophagy guided interventions to modify the cardiac phenotype of Danon disease.

Danon disease is a lethal X-linked genetic syndrome resulting from radical mutations in the LAMP2 gene. LAMP2 protein deficiency results in defective lysosomal function, autophagy arrest and a multisystem disorder primarily involving the heart, skeletal muscle and the central nervous system. Cardiomyopathy is the main cause of morbidity and mortality. To investigate the mechanisms of and develop therapies for cardiac Danon disease we engineered a mouse model carrying an exon 6 deletion human mutation in LAMP2, which recapitulates the human cardiac disease phenotype. Mice develop cardiac hypertrophy followed by left ventricular dilatation and systolic dysfunction, in association with progressive fibrosis, oxidative stress, accumulation of autophagosomes and activation of proteasome. Stimulation of autophagy in Danon mice (by exercise training, caloric restriction, and rapamycin) aggravate the disease phenotype, promoting dilated cardiomyopathy. Inhibiting autophagy (by high fat diet or hydroxychloroquine) is better tolerated by Danon mice compared to wild type but is not curative. Inhibiting proteasome by Velcade was found to be highly toxic to Danon mice, suggesting that proteasome is activated to compensate for defective autophagy. In conclusion, activation of autophagy should be avoided in Danon patients. Since Danon's is a lifelong disease, we suggest that lifestyle interventions to decrease cardiac stress may be useful to slow progression of Danon's cardiomyopathy. While Danon mice better tolerate high fat diet and sedentary lifestyle, the benefit regarding cardiomyopathy in humans needs to be balanced against other health consequences of such interventions.

Heme-oxygenase and lipid mediators in obesity and associated cardiometabolic diseases: Therapeutic implications.

Obesity-mediated metabolic syndrome remains the leading cause of death worldwide. Among many potential targets for pharmacological intervention, a promising strategy involves the heme oxygenase (HO) system, specifically its inducible form, HO-1. This review collects and updates much of the current knowledge relevant to pharmacology and clinical medicine concerning HO-1 in metabolic diseases and its effect on lipid metabolism. HO-1 has pleotropic effects that collectively reduce inflammation, while increasing vasodilation and insulin and leptin sensitivity. Recent reports indicate that HO-1 with its antioxidants via the effect of bilirubin increases formation of biologically active lipid metabolites such as epoxyeicosatrienoic acid (EET), omega-3 and other polyunsaturated fatty acids (PUFAs). Similarly, HO-1and bilirubin are potential therapeutic targets in the treatment of fat-induced liver diseases. HO-1-mediated upregulation of EET is capable not only of reversing endothelial dysfunction and hypertension, but also of reversing cardiac remodeling, a hallmark of the metabolic syndrome. This process involves browning of white fat tissue (i.e. formation of healthy adipocytes) and reduced lipotoxicity, which otherwise will be toxic to the heart. More importantly, this review examines the activity of EET in biological systems and a series of pathways that explain its mechanism of action and discusses how these might be exploited for potential therapeutic use. We also discuss the link between cardiac ectopic fat deposition and cardiac function in humans, which is similar to that described in obese mice and is regulated by HO-1-EET-PGC1α signaling, a potent negative regulator of the inflammatory adipokine NOV.

Pharmacological and Clinical Significance of Heme Oxygenase-1.

This Special Issue collates and updates the current knowledge of the pharmacology and clinical applications concerning the enzyme heme oxygenase (HO) [...].

The pivotal role of heme Oxygenase-1 in reversing the pathophysiology and systemic complications of NAFLD.

The pathogenesis and molecular pathways involved in non-alcoholic fatty liver disease (NAFLD) are reviewed, as well as what is known about mitochondrial dysfunction that leads to heart disease and the progression to steatohepatitis and hepatic fibrosis. We focused our discussion on the role of the antioxidant gene heme oxygenase-1 (HO-1) and its nuclear coactivator, peroxisome proliferator-activated receptor-gamma coactivator (PGC1-α) in the regulation of mitochondrial biogenesis and function and potential therapeutic benefit for cardiac disease, NAFLD as well as the pharmacological effect they have on the chronic inflammatory state of obesity. The result is increased mitochondrial function and the conversion of white adipocyte tissue to beige adipose tissue ("browning of white adipose tissue") that leads to an improvement in signaling pathways and overall liver function. Improved mitochondrial biogenesis and function is essential to preventing the progression of hepatic steatosis to NASH and cirrhosis as well as preventing cardiovascular complications.

Can charcoal improve outcomes in COVID-19 infections?

COVID-19 infection causes considerable morbidity and mortality, especially to those who are aged, have impaired renal function and are obese. We propose to examine the potential utility of oral activated charcoal with the hypothesis that such treatment would lower absorption of microbiome derived toxins and ameliorate systemic oxidant stress and inflammation.

The Pivotal Role of Adipocyte-Na K peptide in Reversing Systemic Inflammation in Obesity and COVID-19 in the Development of Heart Failure.

This review summarizes data from several laboratories that have demonstrated a role of the Na/K-ATPase, specifically its α1 subunit, in the generation of reactive oxygen species (ROS) via the negative regulator of Src. Together with Src and other signaling proteins, the Na/K-ATPase forms an oxidant amplification loop (NKAL), amplifies ROS, and participates in cytokines storm in obesity. The development of a peptide fragment of the α1 subunit, NaKtide, has been shown to negatively regulate Src. Several groups showed that the systemic administration of the cell permeable modification of NaKtide (pNaKtide) or its selective delivery to fat tissue-adipocyte specific expression of NaKtide-ameliorate the systemic elevation of inflammatory cytokines seen in chronic obesity. Severe acute respiratory syndrome - coronavirus 2 (SARS-CoV-2), the RNA Coronavirus responsible for the COVID-19 global pandemic, invades cells via the angiotensin converting enzyme 2 (ACE-2) receptor (ACE2R) that is appended in inflamed fat tissue and exacerbates the formation of the cytokines storm. Both obesity and heart and renal failure are well known risks for adverse outcomes in patients infected with COVID-19. White adipocytes express ACE-2 receptors in high concentration, especially in obese patients. Once the virus invades the white adipocyte cell, it creates a COVID19-porphyrin complex which degrades and produces free porphyrin and iron and increases ROS. The increased formation of ROS and activation of the NKAL results in a further potentiated formation of ROS production, and ultimately, adipocyte generation of more inflammatory mediators, leading to systemic cytokines storm and heart failure. Moreover, chronic obesity also results in the reduction of antioxidant genes such as heme oxygenase-1 (HO-1), increasing adipocyte susceptibility to ROS and cytokines. It is the systemic inflammation and cytokine storm which is responsible for many of the adverse outcomes seen with COVID-19 infections in obese subjects, leading to heart failure and death. This review will also describe the potential antioxidant drugs and role of NaKtide and their demonstrated antioxidant effect used as a major strategy for improving obesity and epicardial fat mediated heart failure in the context of the COVID pandemic.

Author Correction: The Adipocyte Na/K-ATPase Oxidant Amplification Loop is the Central Regulator of Western Diet-Induced Obesity and Associated Comorbidities.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

OX-HDL: A Starring Role in Cardiorenal Syndrome and the Effects of Heme Oxygenase-1 Intervention.

In this review, we will evaluate how high-density lipoprotein (HDL) and the reverse cholesterol transport (RCT) pathway are critical for proper cardiovascular-renal physiology. We will begin by reviewing the basic concepts of HDL cholesterol synthesis and pathway regulation, followed by cardiorenal syndrome (CRS) pathophysiology. After explaining how the HDL and RCT pathways become dysfunctional through oxidative processes, we will elaborate on the potential role of HDL dysfunction in CRS. We will then present findings on how HDL function and the inducible antioxidant gene heme oxygenase-1 (HO-1) are interconnected and how induction of HO-1 is protective against HDL dysfunction and important for the proper functioning of the cardiovascular-renal system. This will substantiate the proposal of HO-1 as a novel therapeutic target to prevent HDL dysfunction and, consequently, cardiovascular disease, renal dysfunction, and the onset of CRS.

Milk thistle seed cold press oil attenuates markers of the metabolic syndrome in a mouse model of dietary-induced obesity.

Milk thistle cold press oil (MTO) is an herbal remedy derived from Silybum marianum which contains a low level of silymarin and mixture of polyphenols and flavonoids. The effect of MTO on the cardiovascular and metabolic complications of obesity was studied in mice that were fed a high-fat diet (HFD) for 20 weeks and treated with MTO for the final 8 weeks of the diet. MTO treatment attenuated HFD-induced obesity, fasting hyperglycemia, hypertension, and induced markers of mitochondrial fusion and browning of white adipose. Markers of inflammation were also attenuated in both adipose and the liver of MTO-treated mice. In addition, MTO resulted in the improvement of liver fibrosis. These results demonstrate that MTO has beneficial actions to attenuate dietary obesity-induced weight gain, hyperglycemia, hypertension, inflammation, and suggest that MTO supplementation may prove beneficial to patients exhibiting symptoms of metabolic syndrome. PRACTICAL APPLICATIONS: Natural supplements are increasingly being considered as potential therapies for many chronic cardiovascular and metabolic diseases. Milk thistle cold press oil (MTO) is derived from Silybum marianum which is used as a dietary supplement in different parts of the world. The results of the present study demonstrate that MTO supplementation normalizes several metabolic and cardiovascular complications arising from dietary-induced obesity. MTO supplementation also had anti-inflammatory actions in the adipose as well as the liver. These results suggest that supplementation of MTO into the diet of obese individuals may afford protection against the worsening of cardiovascular and metabolic disease and improve inflammation and liver fibrosis.

Relationship between dietary sodium and sugar intake: A cross-sectional study of the National Health and Nutrition Examination Survey 2001-2016.

Dietary sodium intake and cardiovascular outcomes have a reported J-shaped curve relationship. This study analyzes the relationship between dietary sodium and sugar intake as a potential mechanism to explain this association. The authors examined cross-sectional data from the National Health and Nutrition Examination Survey (NHANES) 2001-2016 where dietary sodium, carbohydrate, fat, cholesterol, and sugar intakes were assessed by 24-hour dietary recall and were standardized to a total daily intake of 2000 calories. Sodium intake was categorized into sodium quintiles (SQ) as follows: SQ1(0.06-2.6 g/d); SQ2(2.6-3.0 g/d); SQ3(3.0-3.4 g/d); SQ4(3.4-4.0 g/d); and SQ5(4.0-29.3 g/d). Simple and multivariate linear regression using SQ3 as reference were used to assess associations between daily sodium intake and the other nutrients. Our results showed that among 38 722 participants that met our study criteria, the mean age was 43.6 years (SD 16.8 years) and sex was equally distributed (48.8% male vs 51.2% female). Sugar intake went down across increasing SQs and was significantly higher in SQ1 (141.2 g/d) and SQ2 (118.6 g/d) and significantly lower in SQ4 (97.9 g/d) and SQ5 (85.6 g/d) compared to SQ3 (108.6 g/d; all P < .01). These same trends remained unchanged and significant in the fully adjusted multivariate model. In conclusion, NHANES study participants reporting low sodium intake on 24-hour dietary recall have a higher consumption of sugar. The negative impact of low sodium diet on cardiovascular health may be explained at least partially by the associated high sugar intake.

Oxidant-Induced Alterations in the Adipocyte Transcriptome: Role of the Na,K-ATPase Oxidant Amplification Loop.

(1) Background: Recently we have noted that adipocyte specific expression of the peptide, NaKtide, which was developed to attenuate the Na,K-ATPase oxidant amplification loop, could ameliorate the phenotypical features of uremic cardiomyopathy. We performed this study to better characterize the cellular transcriptomes that are involved in various biological pathways associated with adipocyte function occurring with renal failure. (2) Methods: RNAseq was performed on the visceral adipose tissue of animals subjected to partial nephrectomy. Specific expression of NaKtide in adipocytes was achieved using an adiponectin promoter. To better understand the cause of gene expression changes in vivo, 3T3L1 adipocytes were exposed to indoxyl sulfate (IS) or oxidized low density lipoprotein (oxLDL), with and without pNaKtide (the cell permeant form of NaKtide). RNAseq was also performed on these samples. (3) Results: We noted a large number of adipocyte genes were altered in experimental renal failure. Adipocyte specific NaKtide expression reversed most of these abnormalities. High correlation with some cardiac specific phenotypical features was noted amongst groups of these genes. In the murine adipocytes, both IS and oxLDL induced similar pathway changes as were noted in vivo, and pNaKtide appeared to reverse these changes. Network analysis demonstrated tremendous similarities between the network revealed by gene expression analysis with IS compared with oxLDL, and the combined in vitro dataset was noted to also have considerable similarity to that seen in vivo with experimental renal failure. (4) Conclusions: This study suggests that the myriad of phenotypical features seen with experimental renal failure may be fundamentally linked to oxidant stress within adipocytes.

Cold Press Pomegranate Seed Oil Attenuates Dietary-Obesity Induced Hepatic Steatosis and Fibrosis through Antioxidant and Mitochondrial Pathways in Obese Mice.

AIM: Obesity is associated with metabolic syndrome, hypertension, dyslipidemia, nonalcoholic fatty liver disease (NAFLD), and type 2 diabetes. In this study, we investigated whether the dietary supplementation of pomegranate seed oil (PSO) exerted a protective effect on liver lipid uptake, fibrosis, and mitochondrial function in a mouse model of obesity and insulin resistance. METHOD: In this in vivo study, eight-week-old C57BL/6J male mice were fed with a high fat diet (HFD) for 24 weeks and then were divided into three groups as follows: group (1) Lean; group (n = 6) (2) HF diet; group (n = 6) (3) HF diet treated with PSO (40 mL/kg food) (n = 6) for eight additional weeks starting at 24 weeks. Physiological parameters, lipid droplet accumulation, inflammatory biomarkers, antioxidant biomarkers, mitochondrial biogenesis, insulin sensitivity, and hepatic fibrosis were determined to examine whether PSO intervention prevents obesity-associated metabolic syndrome. RESULTS: The PSO group displayed an increase in oxygen consumption, as well as a decrease in fasting glucose and blood pressure (p < 0.05) when compared to the HFD-fed mice group. PSO increased both the activity and expression of hepatic HO-1, downregulated inflammatory adipokines, and decreased hepatic fibrosis. PSO increased the levels of thermogenic genes, mitochondrial signaling, and lipid metabolism through increases in Mfn2, OPA-1, PRDM 16, and PGC1α. Furthermore, PSO upregulated obesity-mediated hepatic insulin receptor phosphorylation Tyr-972, p-IRB tyr1146, and pAMPK, thereby decreasing insulin resistance. CONCLUSIONS: These results indicated that PSO decreased obesity-mediated insulin resistance and the progression of hepatic fibrosis through an improved liver signaling, as manifested by increased insulin receptor phosphorylation and thermogenic genes. Furthermore, our findings indicate a potential therapeutic role for PSO in the prevention of obesity-associated NAFLD, NASH, and other metabolic disorders.

Genetic Polymorphisms Complicate COVID-19 Therapy: Pivotal Role of HO-1 in Cytokine Storm.

Coronaviruses are very large RNA viruses that originate in animal reservoirs and include severe acute respiratory distress syndrome (SARS) and Middle East respiratory syndrome (MERS) and other inconsequential coronaviruses from human reservoirs like the common cold. SARS-CoV-2, the virus that causes COVID-19 and is believed to originate from bat, quickly spread into a global pandemic. This RNA virus has a special affinity for porphyrins. It invades the cell at the angiotensin converting enzyme-2 (ACE-2) receptor and binds to hemoproteins, resulting in a severe systemic inflammatory response, particularly in high ACE-2 organs like the lungs, heart, and kidney, resulting in systemic disease. The inflammatory response manifested by increased cytokine levels and reactive oxygen species results in inhibition of heme oxygenase (HO-1), with a subsequent loss of cytoprotection. This has been seen in other viral illness like human immunodeficiency virus (HIV), Ebola, and SARS/MERS. There are a number of medications that have been tried with some showing early clinical promise. This illness disproportionately affects patients with obesity, a chronic inflammatory disease with a baseline excess of cytokines. The majority of the medications used in the treatment of COVID-19 are metabolized by cytochrome P450 (CYP) enzymes, primarily CYP2D6. This is further complicated by genetic polymorphisms of CYP2D6, HO-1, ACE, and ACE-2. There is a potential role for HO-1 upregulation to treat/prevent cytokine storm. Current therapy must focus on antivirals and heme oxygenase upregulation. Vaccine development will be the only magic bullet.

Cold-Pressed Nigella Sativa Oil Standardized to 3% Thymoquinone Potentiates Omega-3 Protection against Obesity-Induced Oxidative Stress, Inflammation, and Markers of Insulin Resistance Accompanied with Conversion of White to Beige Fat in Mice.

Excessive lipid accumulation in white adipose tissue (WAT) results in adipocyte hypertrophy and chronic low-grade inflammation, which is the major cause of obesity-associated insulin resistance and consequent metabolic disease. The development of beige adipocytes in WAT (browning of WAT) increases energy expenditure and has been considered as a novel strategy to counteract obesity. Thymoquinone (TQ) is the main bioactive quinone derived from the plant Nigella Sativa and has antioxidative and anti-inflammatory capacities. Fish oil omega 3 (ω3) enhances both insulin sensitivity and glucose homeostasis in obesity, but the involved mechanisms remain unclear. The aim of this study is to explore the effects of TQ and ω3 PUFAs (polyunsaturated fatty acids) on obesity-associated inflammation, markers of insulin resistance, and the metabolic effects of adipose tissue browning. 3T3-L1 cells were cultured to investigate the effects of TQ and ω3 on the browning of WAT. C57BL/6J mice were fed a high-fat diet (HFD), supplemented with 0.75% TQ, and 2% ω3 in combination for eight weeks. In 3T3-L1 cells, TQ and ω3 reduced lipid droplet size and increased hallmarks of beige adipocytes such as uncoupling protein-1 (UCP1), PR domain containing 16 (PRDM16), fibroblast growth factor 21 (FGF21), Sirtuin 1 (Sirt1), Mitofusion 2 (Mfn2), and heme oxygenase 1 (HO-1) protein expression, as well as increased the phosphorylation of Protein Kinase B (AKT) and insulin receptors. In the adipose tissue of HFD mice, TQ and ω3 treatment attenuated levels of inflammatory adipokines, Nephroblastoma Overexpressed (NOV/CCN3) and Twist related protein 2 (TWIST2), and diminished adipocyte hypoxia by decreasing HIF1α expression and hallmarks of beige adipocytes such as UCP1, PRDM16, FGF21, and mitochondrial biogenesis markers Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), Sirt1, and Mfn2. Increased 5' adenosine monophosphate-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) phosphorylation and HO-1 expression were observed in adipose with TQ and ω3 treatment, which led to increased pAKT and pIRS1 Ser307 expression. In addition to the adipose, TQ and ω3 also increased inflammation and markers of insulin sensitivity in the liver, as demonstrated by increased phosphorylated insulin receptor (pIR tyr972), insulin receptor beta (IRß), UCP1, and pIRS1 Ser307 and reduced NOV/CCN3 expression. Our data demonstrate the enhanced browning of WAT from TQ treatment in combination with ω3, which may play an important role in decreasing obesity-associated insulin resistance and in reducing the chronic inflammatory state of obesity.

Targeting the Heme-Heme Oxygenase System to Prevent Severe Complications Following COVID-19 Infections.

SARS-CoV-2 is causing a pandemic resulting in high morbidity and mortality. COVID-19 patients suffering from acute respiratory distress syndrome (ARDS) are often critically ill and show lung injury and hemolysis. Heme is a prosthetic moiety crucial for the function of a wide variety of heme-proteins, including hemoglobin and cytochromes. However, injury-derived free heme promotes adhesion molecule expression, leukocyte recruitment, vascular permeabilization, platelet activation, complement activation, thrombosis, and fibrosis. Heme can be degraded by the anti-inflammatory enzyme heme oxygenase (HO) generating biliverdin/bilirubin, iron/ferritin, and carbon monoxide. We therefore postulate that free heme contributes to many of the inflammatory phenomena witnessed in critically ill COVID-19 patients, whilst induction of HO-1 or harnessing heme may provide protection. HO-activity not only degrades injurious heme, but its effector molecules possess also potent salutary anti-oxidative and anti-inflammatory properties. Until a vaccine against SARS-CoV-2 becomes available, we need to explore novel strategies to attenuate the pro-inflammatory, pro-thrombotic, and pro-fibrotic consequences of SARS-CoV-2 leading to morbidity and mortality. The heme-HO system represents an interesting target for novel "proof of concept" studies in the context of COVID-19.

Central Role for Adipocyte Na,K-ATPase Oxidant Amplification Loop in the Pathogenesis of Experimental Uremic Cardiomyopathy.

BACKGROUND: Oxidative stress in adipocyte plays a central role in the pathogenesis of obesity as well as in the associated cardiovascular complications. The putative uremic toxin indoxyl sulfate induces oxidative stress and dramatically alters adipocyte phenotype in vitro. Mice that have undergone partial nephrectomy serve as an experimental model of uremic cardiomyopathy. This study examined the effects on adipocytes of administering a peptide that reduces oxidative stress to the mouse model. METHODS: A lentivirus vector introduced the peptide NaKtide with an adiponectin promoter into the mouse model of experimental uremic cardiomyopathy, intraperitoneally. Then adipocyte-specific expression of the peptide was assessed for mice fed a standard diet compared with mice fed a western diet enriched in fat and fructose. RESULTS: Partial nephrectomy induced cardiomyopathy and anemia in the mice, introducing oxidant stress and an altered molecular phenotype of adipocytes that increased production of systemic inflammatory cytokines instead of accumulating lipids, within 4 weeks. Consumption of a western diet significantly worsened the adipocyte oxidant stress, but expression of NaKtide in adipocytes completely prevented the worsening. The peptide-carrying lentivirus achieved comparable expression in skeletal muscle, but did not ameliorate the disease phenotype. CONCLUSIONS: Adipocyte-specific expression of NaKtide, introduced with a lentiviral vector, significantly ameliorated adipocyte dysfunction and uremic cardiomyopathy in partially nephrectomized mice. These data suggest that the redox state of adipocytes controls the development of uremic cardiomyopathy in mice subjected to partial nephrectomy. If confirmed in humans, the oxidative state of adipocytes may be a therapeutic target in chronic renal failure.

Therapeutic approaches to diabetic cardiomyopathy: Targeting the antioxidant pathway.

The global epidemic of cardiovascular disease continues unabated and remains the leading cause of death both in the US and worldwide. We hereby summarize the available therapies for diabetes and cardiovascular disease in diabetics. Clearly, the current approaches to diabetic heart disease often target the manifestations and certain mediators but not the specific pathways leading to myocardial injury, remodeling and dysfunction. Better understanding of the molecular events determining the evolution of diabetic cardiomyopathy will provide insight into the development of specific and targeted therapies. Recent studies largely increased our understanding of the role of enhanced inflammatory response, ROS production, as well as the contribution of Cyp-P450-epoxygenase-derived epoxyeicosatrienoic acid (EET), Peroxisome Proliferator-Activated Receptor Gamma Coactivator-1α (PGC-1α), Heme Oxygenase (HO)-1 and 20-HETE in pathophysiology and therapy of cardiovascular disease. PGC-1α increases production of the HO-1 which has a major role in protecting the heart against oxidative stress, microcirculation and mitochondrial dysfunction. This review describes the potential drugs and their downstream targets, PGC-1α and HO-1, as major loci for developing therapeutic approaches beside diet and lifestyle modification for the treatment and prevention of heart disease associated with obesity and diabetes.