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Summary: Adiponectin-based therapeutic strategies are promising for managing metabolic diseases and reducing inflammation, prompting the development of adiponectin receptor agonists. However, monitoring their pharmacodynamic actions in clinical applications is challenging. This study aimed to identify peripheral biomarkers to monitor adiponectin actions using ALY688, an adiponectin receptor agonist peptide. RNA sequencing analysis of whole blood identified a cluster of genes that were significantly increased in the ALY688-treated group compared to the control. This gene cluster was validated by qPCR and further confirmed in human peripheral blood mononuclear cells treated with ALY688 ex vivo. We also confirmed a functional outcome of ALY688 action in mice as our study also demonstrated the anti-inflammatory effect of ALY688 in a sublethal LPS mouse model. In summary, a newly identified gene cluster signature is suitable for assessing the pharmacodynamic action of adiponectin or its mimetics in blood samples.

ABSTRACTMyocardial ischemia/reperfusion (I/R) injury is exacerbated in diabetic individuals and animal models. We tested whether autophagy is an important cellular determinant of cell death. First, we utilized a cellular model of hypoxia reoxygenation (H/R) in H9c2 cells cultured in low or high glucose (HG) and tested cell death using flow cytometry to detect Annexin-V and propidium iodide, imaging cell viability ReadyProbe and lactate dehydrogenase release. We observed that cell death induced by H/R was enhanced by HG. Kinetic analysis of caspase-3 activity using a fluorescence reporter probe, stable expression of the VC3AI biosensor and western blotting indicated that H/R induced activation of caspase-3 was enhanced by HG. Temporal autophagy flux analysis using DapRed and DalGreen probes indicated an initial increase in response to H/R that was reduced upon prolonged (24h) R. HG suppressed this induction of autophagy. This was verified using LC3 HiBiT reporter assay, tandem-fluorescent LC3, and western blotting. Lysosomal cathepsin activity was also elevated at 6h and suppressed at 24h R. Autophagy-deficient cells were generated via CRISPR-mediated knockout of atg7 and the effect of combined HG and H/R treatment on caspase activation and cell death was elevated in comparison with wild type cells. We then performed coronary artery ligation surgery to induce ischemia, followed by reperfusion, in wild-type or streptozotocin (STZ)-induced hyperglycemic mice. Non-invasive 3-dimensional imaging using fluorescence molecular tomography combined with computerized tomography was employed to monitor spatio-temporal activation of cardiac autophagy and apoptosis. Upon systemic injection of a near infra-red cathepsin activatable probe we found that hyperglycemic mice had lower activity in the infarct region after I/R versus wild type. In parallel, we observed a higher extent of I/R-induced apoptosis, detected with an annexin-V probe, in hyperglycemic mice. Collectively, these results revealed that impaired autophagic flux in the presence of high glucose levels exacerbates I/R injury.Abbreviation: satg7, autophagy-related 7; FMT, fluorescence molecular tomography; HG, high glucose; H/R, hypoxia/reoxygenation; I/R, ischemia/reperfusion; LC3, MAP1LC3; N, normoxia; NG, normal glucose; NIR, near-infrared; p62, SQSTM1; STZ, streptozotocin.

ABSTRACTHeart failure, a leading driver of global mortality, remains a topic of intense contemporary research interest due to the prevailing unmet need in cardiometabolic therapeutics. Numerous mechanisms with the potential to influence the onset and development of heart failure remain incompletely understood. Firstly, myocardial autophagy, which involves lysosomal degradation of damaged cellular components, confers context-dependent beneficial and detrimental effects. Secondly, sterile inflammation may arise following cardiac stress and exacerbate the progression of heart failure. Inflammation changes in a temporal manner and its onset must be adequately resolved to limit progression of heart failure. Mitochondria are an important factor in contributing to sterile inflammation by releasing damage associated molecular patterns (DAMPs) including mitochondrial DNA (mtDNA). Accordingly, this is one reason why the selective autophagy of mitochondria to maintain optimal function is important in determining cardiac function. In this review, we examine the increasing evidence suggesting crosstalk between autophagy and sterile inflammation together with their role in the development of heart failure. In particular, this is exemplified in the preclinical models of ischaemia/reperfusion injury and pressure overload induced heart failure. We also highlight potential therapeutic approaches focusing on autophagy and addressing sterile inflammation, aiming to enhance outcomes in heart failure.

Abstract Metabolic dysfunction‐associated steatohepatitis (MASH) is the severe form of non‐alcoholic fatty liver disease which has a high potential to progress to cirrhosis and hepatocellular carcinoma, yet adequate effective therapies are lacking. Hypoadiponectinemia is causally involved in the pathogenesis of MASH. This study investigated the pharmacological effects of adiponectin replacement therapy with the adiponectin‐derived peptide ALY688 (ALY688‐SR) in a mouse model of MASH. Human induced pluripotent stem (iPS) cell‐derived hepatocytes were used to test cytotoxicity and signaling of unmodified ALY688 in vitro. High‐fat diet with low methionine and no added choline (CDAHF) was used to induce MASH and test the effects of ALY688‐SR in vivo. Histological MASH activity score (NAS) and fibrosis score were determined to assess the effect of ALY688‐SR. Transcriptional characterization of mice through RNA sequencing was performed to indicate potential molecular mechanisms involved. In cultured hepatocytes, ALY688 efficiently induced adiponectin‐like signaling, including the AMP‐activated protein kinase and p38 mitogen‐activated protein kinase pathways, and did not elicit cytotoxicity. Administration of ALY688‐SR in mice did not influence body weight but significantly ameliorated CDAHF‐induced hepatic steatosis, inflammation, and fibrosis, therefore effectively preventing the development and progression of MASH. Mechanistically, ALY688‐SR treatment markedly induced hepatic expression of genes involved in fatty acid oxidation, whereas it significantly suppressed the expression of pro‐inflammatory and pro‐fibrotic genes as demonstrated by transcriptomic analysis. ALY688‐SR may represent an effective approach in MASH treatment. Its mode of action involves inhibition of hepatic steatosis, inflammation, and fibrosis, possibly via canonical adiponectin‐mediated signaling.

Identification of new mechanisms mediating insulin sensitivity is important to allow validation of corresponding therapeutic targets. In this study, we first used a cellular model of skeletal muscle cell iron overload and found that endoplasmic reticulum (ER) stress and insulin resistance occurred after iron treatment. Insulin sensitivity was assessed using cells engineered to express an Akt biosensor, based on nuclear FoxO localization, as well as western blotting for insulin signaling proteins. Use of salubrinal to elevate eIF2α phosphorylation and promote the unfolded protein response (UPR) attenuated iron-induced insulin resistance. Salubrinal induced autophagy flux and its beneficial effects on insulin sensitivity were not observed in autophagy-deficient cells generated by overexpressing a dominant-negative ATG5 mutant or via knockout of ATG7. This indicated the beneficial effect of salubrinal-induced UPR activation was autophagy-dependent. We translated these observations to an animal model of systemic iron overload-induced skeletal muscle insulin resistance where administration of salubrinal as pretreatment promoted eIF2α phosphorylation, enhanced autophagic flux in skeletal muscle and improved insulin responsiveness. Together, our results show that salubrinal elicited an eIF2α-autophagy axis leading to improved skeletal muscle insulin sensitivity both in vitro and in mice.

Abstract Strategies to enhance autophagy flux have been suggested to improve outcomes in cardiac ischemic models. We explored the role of adiponectin in mediating cardiac autophagy under ischemic conditions induced by permanent coronary artery ligation. We studied the molecular mechanisms underlying adiponectin's cardio‐protective effects in adiponectin knockout (Ad‐KO) compared with wild‐type (WT) mice subjected to ischemia by coronary artery ligation and H9c2 cardiomyocyte cell line exposed to hypoxia. Systemic infusion of a cathepsin‐B activatable near‐infrared probe as a biomarker for autophagy and detection via noninvasive three‐dimensional fluorescence molecular tomography combined with computerized tomography to quantitate temporal changes, indicated increased activity in the myocardium of WT mice after myocardial infarction which was attenuated in Ad‐KO. Seven days of ischemia increased myocardial adiponectin accumulation and elevated ULK1/AMPK phosphorylation and autophagy assessed by Western blotting for LC3 and p62, an outcome not observed in Ad‐KO mice. Cell death, assessed by TUNEL analysis and the ratio of Bcl‐2:Bax, plus cardiac dysfunction, measured using echocardiography with strain analysis, were exacerbated in Ad‐KO mice. Using cellular models, we observed that adiponectin stimulated autophagy flux in isolated primary adult cardiomyocytes and increased basal and hypoxia‐induced autophagy in H9c2 cells. Real‐time temporal analysis of caspase‐3/7 activation and caspase‐3 Western blot indicated that adiponectin suppressed activation by hypoxia. Hypoxia‐induced mitochondrial reactive oxygen species production and cell death were also attenuated by adiponectin. Importantly, the ability of adiponectin to reduce caspase‐3/7 activation and cell death was not observed in autophagy‐deficient cells generated by CRISPR‐mediated deletion of Atg7. Collectively, our data indicate that adiponectin acts in an autophagy‐dependent manner to attenuate cardiomyocyte caspase‐3/7 activation and cell death in response to hypoxia in vitro and ischemia in mice.

Aims: Adiponectin has been shown to mediate cardioprotective effects and levels are typically reduced in patients with cardiometabolic disease. Hence, there has been intense interest in developing adiponectin-based therapeutics. The aim of this translational research study was to examine the functional significance of targeting adiponectin signaling with the adiponectin receptor agonist ALY688 in a mouse model of heart failure with reduced ejection fraction (HFrEF), and the mechanisms of cardiac remodeling leading to cardioprotection. Methods and results: Wild-type mice were subjected to transverse aortic constriction (TAC) to induce left ventricular pressure overload (PO), or sham surgery, with or without daily subcutaneous ALY688-SR administration. Temporal analysis of cardiac function was conducted via weekly echocardiography for 5 weeks and we observed that ALY688 attenuated the PO-induced dysfunction. ALY688 also reduced cardiac hypertrophic remodeling, assessed via LV mass, heart weight to body weight ratio, cardiomyocyte cross sectional area, ANP and BNP levels. ALY688 also attenuated PO-induced changes in myosin light and heavy chain expression. Collagen content and myofibroblast profile indicated that fibrosis was attenuated by ALY688 with TIMP1 and scleraxis/periostin identified as potential mechanistic contributors. ALY688 reduced PO-induced elevation in circulating cytokines including IL-5, IL-13 and IL-17, and the chemoattractants MCP-1, MIP-1β, MIP-1alpha and MIP-3α. Assessment of myocardial transcript levels indicated that ALY688 suppressed PO-induced elevations in IL-6, TLR-4 and IL-1β, collectively indicating anti-inflammatory effects. Targeted metabolomic profiling indicated that ALY688 increased fatty acid mobilization and oxidation, increased betaine and putrescine plus decreased sphingomyelin and lysophospholipids, a profile indicative of improved insulin sensitivity. Conclusion: These results indicate that the adiponectin mimetic peptide ALY688 reduced PO-induced fibrosis, hypertrophy, inflammation and metabolic dysfunction and represents a promising therapeutic approach for treating HFrEF in a clinical setting.

Abstract Memory impairments have been well documented in people with Duchenne muscular dystrophy (DMD). However, the underlying mechanisms are poorly understood, and there is an unmet need to develop new therapies to treat this condition. Using a novel object recognition test, we show that recognition memory impairments in D2.mdx mice are completely prevented by daily treatment with the new adiponectin receptor agonist ALY688 from day 7 to 28 of age. In comparison to age‐matched wild‐type mice, untreated D2.mdx mice demonstrated lower hippocampal mitochondrial respiration (carbohydrate substrate), greater serum interleukin‐6 cytokine content and greater hippocampal total tau and Raptor protein contents. Each of these measures was partly or fully preserved after treatment with ALY688. Collectively, these results indicate that adiponectin receptor agonism improves recognition memory in young D2.mdx mice.

Duchenne muscular dystrophy (DMD) is caused by genetic mutations in the cytoskeletal-sarcolemmal anchor protein dystrophin. Repeated cycles of sarcolemmal tearing and repair lead to a variety of secondary cellular and physiological stressors that are thought to contribute to weakness, atrophy, and fibrosis. Collectively, these stressors can contribute to a pro-inflammatory milieu in locomotor, cardiac, and respiratory muscles. Given the many unwanted side effects that accompany current anti-inflammatory steroid-based approaches for treating DMD (e.g., glucocorticoids), there is a need to develop new therapies that address inflammation and other cellular dysfunctions. Adiponectin receptor (AdipoR) agonists, which stimulate AdipoR1 and R2 isoforms on various cell types, have emerged as therapeutic candidates for DMD due to their anti-inflammatory, anti-fibrotic, and pro-myogenic properties in pre-clinical human and rodent DMD models. Although these molecules represent a new direction for therapeutic intervention, the mechanisms through which they elicit their beneficial effects are not yet fully understood, and DMD-specific data is limited. The overarching goal of this review is to investigate how adiponectin signaling may ameliorate pathology associated with dystrophin deficiency through inflammatory-dependent and -independent mechanisms and to determine if current data supports their future progression to clinical trials.

Abstract Iron overload is associated with various pathological changes which contribute to metabolic syndrome, many of which have been proposed to occur via damaging tissue through an excessive amount of reactive oxygen species (ROS) production. In this study, we established a model of iron overload in L6 skeletal muscle cells and observed that iron enhanced cytochrome c release from depolarized mitochondria, assayed by immunofluorescent colocalization of cytochrome c with Tom20 and the use of JC-1, respectively. This subsequently elevated apoptosis, determined via use of a caspase-3/7 activatable fluorescent probe and western blotting for cleaved caspase-3. Using CellROX deep red and mBBr, we observed that iron increased generation of reactive oxygen species (ROS), and that pretreatment with the superoxide dismutase mimetic MnTBAP reduced ROS production and attenuated iron-induced intrinsic apoptosis and cell death. Furthermore, using MitoSox Red we observed that iron enhanced mROS and the mitochondria-targeted anti-oxidant SKQ1 reduced iron-induced ROS generation and cell death. Western blotting for LC3-II and P62 levels as well as immunofluorescent detection of autophagy flux with LC3B and P62 co-localization indicated that iron acutely (2–8 h) activated and later (12–24 h) attenuated autophagic flux. We used autophagy-deficient cell models generated by overexpressing a dominant-negative Atg5 mutant or CRISPR-mediated ATG7 knock out to test the functional significance of autophagy and observed that autophagy-deficiency exacerbated iron-induced ROS production and apoptosis. In conclusion, our study showed that high iron levels promoted ROS production, blunted the self-protective autophagy response and led to cell death in L6 skeletal muscle cells.

Inflammation plays an important role in the pathophysiology of cardiometabolic diseases. Sterile inflammation, a non-infectious and damage-associated molecular pattern (DAMP)-induced innate response, is now well-established to be closely associated with development and progression of cardiometabolic diseases. The NOD-like receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome is well-established as a major player in sterile inflammatory responses. It is a multimeric cytosolic protein complex which regulates the activation of caspase-1 and subsequently promotes cleavage and release of interleukin (IL)-1 family cytokines, which have a deleterious impact on the development of cardiometabolic diseases. Therefore, targeting NLRP3 itself or the downstream consequences of NLRP3 activation represent excellent potential therapeutic targets in inflammatory cardiometabolic diseases. Here, we review our current understanding of the role which NLRP3 inflammasome regulation plays in cardiometabolic diseases such as obesity, diabetes, non-alcoholic steatohepatitis (NASH), atherosclerosis, ischemic heart disease and cardiomyopathy. Finally, we highlight the potential of targeting NLPR3 or related signaling molecules as a therapeutic approach.

Cardiometabolic diseases exert a significant health impact, leading to a considerable economic burden globally. The metabolic syndrome, characterized by a well-defined cluster of clinical parameters, is closely linked to an elevated risk of cardiovascular disease. Current treatment strategies often focus on addressing individual aspects of metabolic syndrome. We propose that exploring novel therapeutic approaches that simultaneously target multiple facets may prove more effective in alleviating the burden of cardiometabolic disease. There is a growing body of evidence suggesting that mitochondria can serve as a pivotal target for the development of therapeutics aimed at resolving both metabolic and vascular dysfunction. MitoNEET was identified as a binding target for the thiazolidinedione (TZD) class of antidiabetic drugs and is now recognized for its role in regulating various crucial cellular processes. Indeed, mitoNEET has demonstrated promising potential as a therapeutic target in various chronic diseases, encompassing cardiovascular and metabolic diseases. In this review, we present a thorough overview of the molecular mechanisms of mitoNEET, with an emphasis on their implications for cardiometabolic diseases in more recent years. Furthermore, we explore the potential impact of these findings on the development of novel therapeutic strategies and discuss potential directions for future research.

While inflammation is an important immune response for protection from infections, excessive or prolonged inflammation can lead to a variety of debilitating diseases including skin disease, diabetes, heart disease, stroke, autoimmune diseases and cancer. Inflammation is a graded response that is typically initiated when resident macrophages sense the presence of pathogens or damage in the tissue and produce inflammatory cytokines and chemokines to kill the pathogen, clear debris and dead tissue, and initiate tissue repair. Here we show that copper-infused fabrics can prevent inflammation by blocking the production of inflammatory cytokines from macrophages after being exposed to LPS, a component of bacterial cell wall. Mechanistically, we show that copper-infused fabrics can significantly reduce the NF-κB and IRF3 activation in LPS-stimulated macrophages. Given the importance of excessive inflammation in diabetes, we show that copper can reduce insulin resistance mediated by inflammatory cytokines in muscle cells. Our data show that copper infused fabrics may be useful to reduce excessive inflammation in macrophages and improve insulin sensitivity in skeletal muscles.

The Scintillating Bubble Chamber (SBC) collaboration is developing liquid-noble bubble chambers for the detection of sub-keV nuclear recoils. These detectors benefit from the electron recoil rejection inherent in moderately-superheated bubble chambers with the addition of energy reconstruction provided from the scintillation signal. The ability to measure low-energy nuclear recoils allows the search for GeV-scale dark matter and the measurement of coherent elastic neutrino-nucleus scattering on argon from MeV-scale reactor antineutrinos. The first physics-scale detector, SBC-LAr10, is in the commissioning phase at Fermilab, where extensive engineering and calibration studies will be performed. In parallel, a functionally identical low-background version, SBC-SNOLAB, is being built for a dark matter search underground at SNOLAB. SBC-SNOLAB, with a 10 kg-yr exposure, will have sensitivity to a dark matter–nucleon cross section of 2×10−42 cm2 at 1 GeV/c2 dark matter mass, and future detectors could reach the boundary of the argon neutrino fog with a tonne-yr exposure. In addition, the deployment of an SBC detector at a nuclear reactor could enable neutrino physics investigations including measurements of the weak mixing angle and searches for sterile neutrinos, the neutrino magnetic moment, and the light Z’ gauge boson.

Although metabolic complications are common in thalassemia patients, there is still an unmet need to better understand underlying mechanisms. We used unbiased global proteomics to reveal molecular differences between the th3/+ mouse model of thalassemia and wild-type control animals focusing on skeletal muscles at 8 weeks of age. Our data point toward a significantly impaired mitochondrial oxidative phosphorylation. Furthermore, we observed a shift from oxidative fibre types toward more glycolytic fibre types in these animals, which was further supported by larger fibre-type cross-sectional areas in the more oxidative type fibres (type I/type IIa/type IIax hybrid). We also observed an increase in capillary density in th3/+ mice, indicative of a compensatory response. Western blotting for mitochondrial oxidative phosphorylation complex proteins and PCR analysis of mitochondrial genes indicated reduced mitochondrial content in the skeletal muscle but not the hearts of th3/+ mice. The phenotypic manifestation of these alterations was a small but significant reduction in glucose handling capacity. Overall, this study identified many important alterations in the proteome of th3/+ mice, amongst which mitochondrial defects leading to skeletal muscle remodelling and metabolic dysfunction were paramount.

Abstract Background The prevalence of type 2 diabetes, obesity and their various comorbidities have continued to rise. In skeletal muscle lipotoxicity is well known to be a contributor to the development of insulin resistance. Here it was examined if the small molecule adiponectin receptor agonist AdipoRon mimicked the effect of adiponectin to attenuate palmitate induced reactive oxygen species (ROS) production and cell death in L6 skeletal muscle cells. Methods L6 cells were treated ±0.1 mM PA, and ± AdipoRon, then assays analyzing reactive oxygen species (ROS) production and cell death, and intracellular and extracellular levels of sphingosine-1 phosphate (S1P) were conducted. To determine the mechanistic role of S1P gain (using exogenous S1P or using THI) or loss of function (using the SKI-II) were conducted. Results Using both CellROX and DCFDA assays it was found that AdipoRon reduced palmitate-induced ROS production. Image-IT DEAD, MTT and LDH assays all indicated that AdipoRon reduced palmitate-induced cell death. Palmitate significantly increased intracellular accumulation of S1P, whereas in the presence of AdipoRon there was increased release of S1P from cells to extracellular medium. It was also observed that direct addition of extracellular S1P prevented palmitate-induced ROS production and cell death, indicating that S1P is acting in an autocrine manner. Pharmacological approaches to enhance or decrease S1P levels indicated that accumulation of intracellular S1P correlated with enhanced cell death. Conclusion This data indicates that increased extracellular levels of S1P in response to adiponectin receptor activation can activate S1P receptor-mediated signaling to attenuate lipotoxic cell death. Taken together these findings represent a possible novel mechanism for the protective action of adiponectin.

Adiponectin regulates white adipose tissue (WAT) metabolism and promotes insulin-sensitizing and anti-atherosclerotic effects in vivo. In this context, small molecule adiponectin receptor agonists have become of great therapeutic value for the treatment of metabolic diseases. Here, we investigated the effects of the adiponectin mimetic compound ALY688 on WAT metabolism. To accomplish this, rat epididymal (Epid) and subcutaneous inguinal (Sc Ing) adipocytes were isolated and incubated with ALY688. Subsequently, several parameters of glucose and fat metabolism were assessed. ALY688 promoted AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) phosphorylation, enhanced glucose oxidation, and suppressed fat oxidation in adipocytes from both fat depots. ALY688 did not affect basal and insulin-stimulated rates of glucose uptake, glucose incorporation into lipids, and AKTSer473 and p38 mitogen-activated protein kinase (MAPK) phosphorylations in either Epid or Sc Ing adipocytes. ALY688 did not alter basal lipolysis in Epid and Sc Ing adipocytes, but it enhanced isoproterenol-induced lipolysis in Epid adipocytes. Adiponectin receptor 2 (AdipoR2) mRNA was the prevalent isoform expressed in all adipocytes, and Epid adipocytes displayed significantly higher AdipoR2 mRNA expression than Sc Ing adipocytes. In conclusion, ALY688 can regulate adiposity and affect glycaemic control by altering substrate portioning in the WAT in a fat depot-specific manner.

Iron overload (IO) is associated with cardiovascular diseases, including heart failure. Our study’s aim was to examine the mechanism by which IO triggers cell death in H9c2 cells. IO caused accumulation of intracellular and mitochondrial iron as shown by the use of iron-binding fluorescent reporters, FerroOrange and MitoFerroFluor. Expression of cytosolic and mitochondrial isoforms of Ferritin was also induced by IO. IO-induced iron accumulation and cellular ROS was rapid and temporally linked. ROS accumulation was detected in the cytosol and mitochondrial compartments with CellROX, DCF-DA and MitoSOX fluorescent dyes and partly reversed by the general antioxidant N-acetyl cysteine or the mitochondrial antioxidant SkQ1. Antioxidants also reduced the downstream activation of apoptosis and lytic cell death quantified by Caspase 3 cleavage/activation, mitochondrial Cytochrome c release, Annexin V/Propidium iodide staining and LDH release of IO-treated cells. Finally, overexpression of MitoNEET, an outer mitochondrial membrane protein involved in the transfer of Fe-S clusters between mitochondrial and cytosol, was observed to lower iron and ROS accumulation in the mitochondria. These alterations were correlated with reduced IO-induced cell death by apoptosis in MitoNEET-overexpressing cells. In conclusion, IO mediates H9c2 cell death by causing mitochondrial iron accumulation and subsequent general and mitochondrial ROS upregulation.

Abstract Background Adiponectin exerts several beneficial cardiovascular effects, however their specific molecular mechanisms require additional understanding. This study investigated the mechanisms of adiponectin action in the heart during high fat diet (HFD) feeding or in palmitate (PA) treated H9c2 cardiomyoblasts. Methods 6-week-old male adiponectin knock out (Ad-KO) mice were fed chow or 60% HFD for 6 weeks then received saline or recombinant adiponectin (3μg/g body weight) for an additional 2 weeks. After acute insulin stimulation (4 U/kg), tissue and serum samples were collected for analysis. H9c2 cardiomyocytes were treated ±0.1 mM PA, the adiponectin receptor agonist AdipoRon, or the antioxidant MnTBAP then assays to analyze reactive oxygen species (ROS) production and cell death were conducted. To specifically determine the mechanistic role of S1P, gain and loss of function studies were conducted with adding S1P to cells or the inhibitors THI and SKI-II, respectively. Results HFD feeding induced cardiac insulin resistance in Ad-KO mice, which was reversed following replenishment of normal circulating adiponectin levels. In addition, myocardial total triglyceride was elevated by HFD and lipidomic analysis showed increased levels of ceramides and sphingosine-1-phosphate (S1P), with only the latter being corrected by adiponectin administration. Similarly, treatment of H9C2 cardiomyoblasts with PA led to a significant increase of intracellular S1P but not in conditioned media whereas AdipoRon significantly increased S1P production and secretion from cells. AdipoRon or the antioxidant MnTBAP significantly reduced PA-induced cell death. Gain and loss of function studies suggested S1P secretion and autocrine receptor activation mediated the effect of AdipoRon to attenuate PA-induced ROS production and cell death. Conclusion Our data establish adiponectin signaling-mediated increase in S1P secretion as a mechanism via which HFD or PA induced cardiomyocyte lipotoxicity, leading to insulin resistance and cell death, is attenuated.

Both iron overload and deficiency can promote development of cardiomyopathy. Advances in our knowledge from recent research have indicated numerous potential cellular mechanisms. Regulation of myocardial autophagy by iron is of particular interest and will be reviewed here. Autophagy is already well established to play a significant role in regulating the development of heart failure. This review will focus on regulation of autophagy by iron, crosstalk between autophagy and other cellular process which have also already been implicated in heart failure (oxidative stress, mitochondrial dysfunction, endoplasmic reticulum stress, ferroptosis) and the therapeutic potential of targeting these interactions.

The Meal Balance Index (MBI) assesses the nutritional quality and balance of meals. It is a score between 0 and 100 that takes into account both shortfall and excess nutrients, adjusted for the energy content of the meal. In the present study the score was applied to 147849 meals reported in the National Health and Nutrition Examination Surveys (NHANES) 2005-2014 in order to evaluate its validity and compare against exemplary meals designed as part of 24h diets that meet US dietary guidelines. Meals from exemplary menu plans developed by nutrition experts scored on average 76±14 (mean ± standard deviation) whereas those of NHANES participants scored 45±14. Scores of breakfast, lunch, dinner, snack, considered jointly as independent variables, were moderately but positively and significantly associated with the Healthy Eating Index (Pearson correlation 0.6). MBI scores were significantly associated with the density of positive micronutrients (e.g. Vit A, Vit C) and favorable food groups (e.g. fruits, whole grains) not directly included in the MBI algorithm. The MBI is a valid tool to assess the nutritional quality of meals reported in the US population and if applied to culinary recipe websites could potentially help users to understand which meals are nutritionally balanced. Choice of more balanced individual meals can guide healthier cooking and eating.

Cardiometabolic diseases encompass those affecting the heart and vasculature as well as other metabolic problems, such as insulin resistance, diabetes, and non-alcoholic fatty liver disease. These diseases tend to have common risk factors, one of which is impaired adiponectin action. This may be due to reduced bioavailability of the hormone or resistance to its effects on target tissues. A strong negative correlation between adiponectin levels and cardiometabolic diseases has been well-documented and research shown that adiponectin has cardioprotective, insulin sensitizing and direct beneficial metabolic effects. Thus, therapeutic approaches to enhance adiponectin action are widely considered to be desirable. The complexity of adiponectin structure and function has so far made progress in this area less than ideal. In this article we will review the effects and mechanism of action of adiponectin on cardiometabolic tissues, identify scenarios where enhancing adiponectin action would be of clinical value and finally discuss approaches via which this can be achieved.

Despite advances in our knowledge and attempts to improve therapies, β-thalassemia remains a prevalent disorder with increased risk for the development of cardiomyopathy. Using an untargeted discovery-based lipidomic workflow, we uncovered that transfusion-dependent thalassemia (TDT) patients had a unique circulating lipidomic signature consisting of 387 lipid features, allowing their significant discrimination from healthy controls (Q-value < 0.01). In particular, TDT patients had elevated triacylglycerols and long-chain acylcarnitines, albeit lower ether phospholipids or plasmalogens, sphingomyelins, and cholesterol esters, reminiscent of that previously characterized in cardiometabolic diseases resulting from mitochondrial and peroxisomal dysfunction. Discriminating lipid (sub)classes correlated differentially with clinical parameters, reflecting blood (ether phospholipids) and iron (cholesterol ester) status or heart function (triacylglycerols). We also tested 15 potential serum biomarkers related to cardiometabolic disease and found that both lipocalin-2 and, for the first time, endocan-1 levels were significantly elevated in TDT patients and showed a strong correlation with blood parameters and three ether diacylglycerophosphatidylcholine species. In conclusion, this study identifies new characteristics of TDT patients which may have relevance in developing biomarkers and therapeutics.

Iron is essential for energy metabolism, and states of iron deficiency or excess are detrimental for organisms and cells. Therefore, iron and carbohydrate metabolism are tightly regulated. Serum iron and glucose levels are subjected to hormonal regulation by hepcidin and insulin, respectively. Hepcidin is a liver-derived peptide hormone that inactivates the iron exporter ferroportin in target cells, thereby limiting iron efflux to the bloodstream. Insulin is a protein hormone secreted from pancreatic β-cells that stimulates glucose uptake and metabolism via insulin receptor signaling. There is increasing evidence that systemic, but also cellular iron and glucose metabolic pathways are interconnected. This review article presents relevant data derived primarily from mouse models and biochemical studies. In addition, it discusses iron and glucose metabolism in the context of human disease.

The vascular endothelium is a dynamic structure responsible for the separation and regulated movement of biological material between circulation and interstitial fluid. Hormones and nutrients can move across the endothelium either via a transcellular or paracellular route. Transcellular endothelial transport is well understood and broadly acknowledged to play an important role in the normal and abnormal physiology of endothelial function. However, less is known about the role of the paracellular route. Although the concept of endothelial dysfunction in diabetes is now widely accepted, we suggest that alterations in paracellular transport should be studied in greater detail and incorporated into this model. In this review we provide an overview of endothelial paracellular permeability and discuss its potential importance in contributing to the development of diabetes and associated complications. Accordingly, we also contend that if better understood, altered endothelial paracellular permeability could be considered as a potential therapeutic target for diabetes.

Adiponectin, circulating levels of which are reduced in obesity and diabetes, mediates cardiac extracellular matrix (ECM) remodeling in response to pressure overload (PO). Here, we performed a detailed temporal analysis of progressive cardiac ECM remodelling in adiponectin knockout (AdKO) and wild-type (WT) mice at 3 days and 1, 2, 3 and 4 weeks following the induction of mild PO via minimally invasive transverse aortic banding. We first observed that myocardial adiponectin gene expression was reduced after 4 weeks of PO, whereas increased adiponectin levels were detected in cardiac homogenates at this time despite decreased circulating levels of adiponectin. Scanning electron microscopy and Masson's trichrome staining showed collagen accumulation increased in response to 2 and 4 weeks of PO in WT mice, while fibrosis in AdKO mice was notably absent after 2 weeks but highly apparent after 4 weeks of PO. Time and intensity of fibroblast appearance after PO was not significantly different between AdKO and WT animals. Gene array analysis indicated that MMP2, TIMP2, collagen 1α1 and collagen 1α3 were induced after 2 weeks of PO in WT but not AdKO mice. After 4 weeks MMP8 was induced in both genotypes, MMP9 only in WT mice and MMP1α only in AdKO mice. Direct stimulation of primary cardiac fibroblasts with adiponectin induced a transient increase in total collagen detected by picrosirius red staining and collagen III levels synthesis, as well as enhanced MMP2 activity detected via gelatin zymography. Adiponectin also enhanced fibroblast migration and attenuated angiotensin-II induced differentiation to a myofibroblast phenotype. In conclusion, these data indicate that increased myocardial bioavailability of adiponectin mediates ECM remodeling following PO and that adiponectin deficiency delays these effects.

Diabetes is associated with higher incidence of myocardial infarction (MI) and increased propensity for subsequent events post-MI. Here we conducted a temporal analysis of the influence of diabetes on cardiac dysfunction and remodeling after ischemia reperfusion (IR) injury in mice. Diabetes was induced using streptozotocin and IR performed by ligating the left anterior descending coronary artery for 30 min followed by reperfusion for up to 42 days. We first evaluated changes in cardiac function using echocardiography after 24 hours reperfusion and observed IR injury significantly decreased the systolic function, such as ejection fraction, fractional shortening and end systolic left ventricular volume (LVESV) in both control and diabetic mice. The longitudinal systolic and diastolic strain rate were altered after IR, but there were no significant differences between diabetic mice and controls. However, a reduced ability to metabolize glucose was observed in the diabetic animals as determined by PET-CT scanning using 2-deoxy-2-((18)F)fluoro-D-glucose. Interestingly, after 24 hours reperfusion diabetic mice showed a reduced infarct size and less apoptosis indicated by TUNEL analysis in heart sections. This may be explained by increased levels of autophagy detected in diabetic mice hearts. Similar increases in IR-induced macrophage infiltration detected by CD68 staining indicated no change in inflammation between control and diabetic mice. Over time, control mice subjected to IR developed mild left ventricular dilation whereas diabetic mice exhibited a decrease in both end diastolic left ventricular volume and LVESV with a decreased intraventricular space and thicker left ventricular wall, indicating concentric hypertrophy. This was associated with marked increases in fibrosis, indicted by Masson trichrome staining, of heart sections in diabetic IR group. In summary, we demonstrate that diabetes principally influences distinct IR-induced chronic changes in cardiac function and remodeling, while a smaller infarct size and elevated levels of autophagy with similar cardiac function are observed in acute phase.

BACKGROUND: This study aimed to determine the association of lipoprotein-associated phospholipase A(2) (Lp-PLA(2)) activity in circulation and peripheral blood mononuclear cells (PBMCs) with inflammatory and oxidative stress markers in nonobese women and according to menopausal status. Lp-PLA(2) activity, a marker for cardiovascular risk is associated with inflammation and oxidative stress. METHODOLOGY/PRINCIPAL FINDINGS: Eighty postmenopausal women (53.0±4.05 yr) and 96 premenopausal women (39.7±9.25 yr) participated in this study. Lp-PLA(2) activities, interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1β in plasma as well as in PBMCs were measured. Plasma ox-LDL was also measured. Postmenopausal women demonstrated higher circulating levels of ox-LDL and IL-6, as well as IL-6, TNF-α, and IL-1β in PBMCs, than premenopausal women. In both groups, plasma Lp-PLA(2) activity positively correlated with Lp-PLA(2) activity in PBMCs and plasma ox-LDL. In premenopausal women, Lp-PLA(2) activities in plasma and PBMCs positively correlated with IL-6, TNF-α, and IL-1β in PBMCs. In postmenopausal women, plasma ox-LDL positively correlated with PBMC cytokine production. In subgroup analysis of postmenopausal women according to plasma ox-LDL level (median level: 48.715 U/L), a significant increase in Lp-PLA(2) activity in the plasma but not the PBMCs was found in the high ox-LDL subgroup. Plasma Lp-PLA(2) activity positively correlated with unstimulated PBMC Lp-PLA(2) activity in the low ox-LDL subgroup (r = 0.627, P

Cardiomyocyte apoptosis is an important remodeling event contributing to heart failure and adiponectin may mediate cardioprotective effects at least in part via attenuating apoptosis. Here we used hypoxia-reoxygenation (H/R) induced apoptosis in H9c2 cells to examine the effect of adiponectin and cellular mechanisms of action. We first used TUNEL labeling in combination with laser scanning cytometry to demonstrate that adiponectin prevented H/R-induced DNA fragmentation. The anti-apoptotic effect of adiponectin was also verified via attenuation of H/R-induced phosphatidylserine exposure using annexin V binding. H/R-induced apoptosis via the mitochondrial-mediated intrinsic pathway of apoptosis as assessed by cytochrome c release into cytosol and caspase-3 activation, both of which were attenuated by adiponectin. Mechanistically, we demonstrated that adiponectin enhanced anti-oxidative potential in these cells which led to attenuation of the increase in intracellular reactive oxygen species (ROS) caused by H/R. To further address the mechanism of adiponctins anti-apoptotic effects we used siRNA to efficiently knockdown adiponectin receptor (AdipoR1) expression and found that this attenuated the protective effects of adiponectin on ROS production and caspase 3 activity. Knockdown of APPL1, an important intracellular binding partner for AdipoR, also significantly reduced the ability of adiponectin to prevent H/R-induced ROS generation and caspase 3 activity. In summary, H/R-induced ROS generation and activation of the intrinsic apoptotic pathway was prevented by adiponectin via AdipoR1/APPL1 signaling and increased anti-oxidant potential.