Downregulation Of Fatty Acid Oxidation Research Articles

Urine metabolite changes after cardiac surgery predict acute kidney injury.

Acute kidney injury (AKI) is a serious complication in patients undergoing cardiac surgery, with the underlying mechanism remaining elusive and a lack of specific biomarkers for cardiac surgery-associated AKI (CS-AKI). We performed an untargeted metabolomics analysis of urine samples procured from a cohort of patients with or without AKI at 6 and 24 h following cardiac surgery. Based on the differential urinary metabolites discovered, we further examined the expressions of the key metabolic enzymes that regulate these metabolites in kidney during AKI using a mouse model of ischemia-reperfusion injury (IRI) and in hypoxia-treated tubular epithelial cells (TECs). The urine metabolomic profiles in AKI patients were significantly different from those in non-AKI patients, including upregulation of tryptophan metabolism- and aerobic glycolysis-related metabolites, such as l-tryptophan and d-glucose-1-phosphate, and downregulation of fatty acid oxidation (FAO) and tricarboxylic acid (TCA) cycle-related metabolites. Spearman correlation analysis showed that serum creatinine was positively correlated with urinary l-tryptophan and indole, which had high accuracy for predicting AKI. In animal experiments, we demonstrated that the expression of rate-limiting enzymes in glycolysis, such as hexokinase II (HK2), was significantly upregulated during renal IRI. However, the TCA cycle-related key enzyme citrate synthase was significantly downregulated after IRI. In vitro, hypoxia induced downregulation of citrate synthase in TECs. In addition, FAO-related gene peroxisome proliferator-activated receptor alpha (PPARα) was remarkably downregulated in kidney during renal IRI. This study presents urinary metabolites related to CS-AKI, indicating the rewiring of the metabolism in kidney during AKI, identifying potential AKI biomarkers.

Integrating Clinical Phenotype With Multiomics Analyses of Human Cardiac Tissue Unveils Divergent Metabolic Remodeling in Genotype-Positive and Genotype-Negative Patients With Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is caused by sarcomere gene mutations (genotype-positive HCM) in ≈50% of patients and occurs in the absence of mutations (genotype-negative HCM) in the other half of patients. We explored how alterations in the metabolomic and lipidomic landscape are involved in cardiac remodeling in both patient groups. We performed proteomics, metabolomics, and lipidomics on myectomy samples (genotype-positive N=19; genotype-negative N=22; and genotype unknown N=6) from clinically well-phenotyped patients with HCM and on cardiac tissue samples from sex- and age-matched and body mass index-matched nonfailing donors (N=20). These data sets were integrated to comprehensively map changes in lipid-handling and energy metabolism pathways. By linking metabolomic and lipidomic data to variability in clinical data, we explored patient group-specific associations between cardiac and metabolic remodeling. HCM myectomy samples exhibited (1) increased glucose and glycogen metabolism, (2) downregulation of fatty acid oxidation, and (3) reduced ceramide formation and lipid storage. In genotype-negative patients, septal hypertrophy and diastolic dysfunction correlated with lowering of acylcarnitines, redox metabolites, amino acids, pentose phosphate pathway intermediates, purines, and pyrimidines. In contrast, redox metabolites, amino acids, pentose phosphate pathway intermediates, purines, and pyrimidines were positively associated with septal hypertrophy and diastolic impairment in genotype-positive patients. We provide novel insights into both general and genotype-specific metabolic changes in HCM. Distinct metabolic alterations underlie cardiac disease progression in genotype-negative and genotype-positive patients with HCM.

Energy metabolic reprogramming regulates programmed cell death of renal tubular epithelial cells and might serve as a new therapeutic target for acute kidney injury.

Acute kidney injury (AKI) induces significant energy metabolic reprogramming in renal tubular epithelial cells (TECs), thereby altering lipid, glucose, and amino acid metabolism. The changes in lipid metabolism encompass not only the downregulation of fatty acid oxidation (FAO) but also changes in cell membrane lipids and triglycerides metabolism. Regarding glucose metabolism, AKI leads to increased glycolysis, activation of the pentose phosphate pathway (PPP), inhibition of gluconeogenesis, and upregulation of the polyol pathway. Research indicates that inhibiting glycolysis, promoting the PPP, and blocking the polyol pathway exhibit a protective effect on AKI-affected kidneys. Additionally, changes in amino acid metabolism, including branched-chain amino acids, glutamine, arginine, and tryptophan, play an important role in AKI progression. These metabolic changes are closely related to the programmed cell death of renal TECs, involving autophagy, apoptosis, necroptosis, pyroptosis, and ferroptosis. Notably, abnormal intracellular lipid accumulation can impede autophagic clearance, further exacerbating lipid accumulation and compromising autophagic function, forming a vicious cycle. Recent studies have demonstrated the potential of ameliorating AKI-induced kidney damage through calorie and dietary restriction. Consequently, modifying the energy metabolism of renal TECs and dietary patterns may be an effective strategy for AKI treatment.

Increased acylcarnitines in infant heart failure indicate fatty acid oxidation inhibition: towards therapeutic options?

Heart failure in adults is characterized by reduction of long-chain fatty acid oxidation in favour of carbohydrate metabolism. This adaptive phenomenon becomes maladaptive because energy conversion decreases and lipid toxic derivatives known to impair cardiac function are accumulating. No data are available concerning metabolic modification in heart failure in children. In order to evaluate the fatty acid oxidation in children suffering from heart failure, acylcarnitine profiles on dried blood spots were obtained from children under 16years old with dilated cardiomyopathy and clinical heart failure (DCM-HF) and control children. Nine children were included in the DCM-HF group and eight in the control group. Acylcarnitine profiles revealed a significant 3.1-fold increase of total acylcarnitines (sum of C3 to C18 acylcarnitine species) in DCM-HF children compared with controls. This result persisted considering the sum of long-chain acylcarnitines (sum of C14 to C18 species), medium-chain acylcarnitines (sum of C8 to C12 species), and short-chain acylcarnitines (sum of C3 to C6 species), respectively, 2.0-, 2.6-, and 1.9-fold increase compared with the control group. A significant linear correlation was found between left ventricular dilatation or ejection fraction and acylcarnitines accumulation. Finally, acylcarnitine ratio C16OH/C16 and C18OH/C18 enhanced in the DCM-HF group, suggesting a diminution of the long-chain hydroxyl acyl-CoA dehydrogenase activity. Our results suggest down-regulation of fatty acid oxidation in children with heart failure. Such lipidomic alteration could worsen heart function and may suggest considering a metabolic treatment of heart failure in children.

Downregulation of fatty acid oxidation led by Hilpda increases G2/M arrest/delay-induced kidney fibrosis

Hypoxia-regulated proximal tubular epithelial cells (PTCs) G2/M phase arrest/delay was involved in production of renal tubulointerstitial fibrosis (TIF). TIF is a common pathological manifestation of progression in patients with chronic kidney disease (CKD), and is often accompanied by lipid accumulation in renal tubules. However, cause-effect relationship between hypoxia-inducible lipid droplet-associated protein (Hilpda), lipid accumulation, G2/M phase arrest/delay and TIF remains unclear. Here we found that overexpression of Hilpda downregulated adipose triglyceride lipase (ATGL) promoted triglyceride overload in the form of lipid accumulation, leading to defective fatty acid β-oxidation (FAO), ATP depletion in a human PTC cell line (HK-2) under hypoxia and in mice kidney tissue treated with unilateral ureteral obstruction (UUO) and unilateral ischemia-reperfusion injury (UIRI). Hilpda-induced lipid accumulation caused mitochondrial dysfunction, enhanced expression of profibrogenic factors TGF-β1, α-SMA and Collagen I elevation, and reduced expression of G2/M phase-associated gene CDK1, as well as increased CyclinB1/D1 ratio, resulted in G2/M phase arrest/delay and profibrogenic phenotypes. Hilpda deficiency in HK-2 cell and kidney of mice with UUO had sustained expression of ATGL and CDK1 and reduced expression of TGF-β1, Collagen I and CyclinB1/D1 ratio, resulting in the amelioration of lipid accumulation and G2/M arrest/delay and subsequent TIF. Expression of Hilpda correlated with lipid accumulation, was positively associated with tubulointerstitial fibrosis in tissue samples from patients with CKD. Our findings suggest that Hilpda deranges fatty acid metabolism in PTCs, which leads to G2/M phase arrest/delay and upregulation of profibrogenic factors, and consequently promote TIF which possibly underlie pathogenesis of CKD.

Pressure overload and volume overload-induced chronic heart failure are associated with characteristic left ventricular myocardial proteomic alterations

Abstract Introduction Hemodynamic overload induces pathological remodeling of the left ventricle (LV) and eventually heart failure (HF). The two types of chronic hemodynamic stress, namely pressure overload (PO) and volume overload (VO) evoke characteristically different functional and structural alterations in the myocardium. Nevertheless, whether PO- and VO-induced HF are also associated with distinct LV proteomic alterations has not been investigated yet. Aim Hence, we thought to perform a proteomic analysis on LV myocardial samples from rat models of PO- and VO-induced HF. Methods PO–induced HF was evoked by transverse aortic constriction (TAC). VO–induced HF was established by creating an aortocaval fistula (ACF). Age-matched sham-operated animals served as controls for TAC (ShamT) and ACF (ShamA), respectively. Pressure-volume (P-V) analysis, echocardiography, histology and quantitative real-time PCR were carried out to provide a detailed characterization of the two HF models. Peptides obtained via the digestion of myocardial proteins with trypsin and LysC were labeled with isobaric tags (TMT16) and measured with LC-MS/MS in a bottom-up explorative proteomic approach. Differential expression and gene ontology enrichment analysis (GO:BP) was carried out on summarized protein reporter ion intensities. Results In both the TAC and ACF groups, presence of typical signs and symptoms of HF (dyspnea at rest, fatigue, ascites) increased lung-to-tibial length ratio and elevated LV natriuretic peptide mRNA expression levels confirmed the development of advanced HF. Furthermore, the TAC model was associated with massive wall thickening, concentric LV hypertrophy (LVH), marked interstitial fibrosis and substantially impaired active relaxation and passive filling (slope of end-diastolic P-V relationship: 0.103±0.015 vs. 0.023±0.003mmHg/μl, TAC vs. ShamT, P<0.001). In contrast, the ACF model was predominantly characterized by LV dilatation, eccentric LVH, moderate fibrosis and severely reduced LV contractility (slope of end-systolic P-V relationship: 0.5±0.1 vs. 2.3±0.3mmHg/μl, ACF vs. ShamA, P<0.001). Proteomic analysis revealed that out of the 4691 identified and quantified proteins, 1404 and 913 have shown upregulation, while 1359 and 886 downregulation in the TAC and ACF groups respectively compared to their corresponding sham groups. GO:BP analysis has indicated that the downregulation of mitochondrion organization, ATP metabolic processes and oxidative phosphorylation and the upregulation of actin cytoskeleton organization were the most profound alterations in the TAC model. In contrast, the ACF model was associated with robust downregulation of fatty acid oxidation and upregulation of endocytosis, defense and immune response on the proteomic level. Conclusions PO and VO-induced advanced HF are not only associated with characteristically different functional and structural remodeling but also with distinct LV proteomic alterations. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Research, Development and Innovation Office (NKFIH) of Hungary

UHPLC-MS/MS-Based Metabolomics and Clinical Phenotypes Analysis Reveal Broad-Scale Perturbations in Early Pregnancy Related to Gestational Diabetes Mellitus.

Gestational diabetes mellitus (GDM) is the most common metabolic disturbance during pregnancy, with adverse effects on both mother and fetus. The establishment of early diagnosis and risk assessment model is of great significance for preventing and reducing adverse outcomes of GDM. In this study, the broad-scale perturbations related to GDM were explored through the integration analysis of metabolic and clinical phenotypes. Maternal serum samples from the first trimester were collected for targeted metabolomics analysis by using ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Statistical analysis was conducted based on the levels of the 184 metabolites and 76 clinical indicators from GDM women (n =60) and matched healthy controls (n =90). Metabolomics analysis revealed the down-regulation of fatty acid oxidation in the first trimester of GDM women, which was supposed to be related to the low serum level of dehydroepiandrosterone.While the significantly altered clinical phenotypes were mainly related to the increased risk of cardiovascular disease, abnormal iron metabolism, and inflammation. A phenotype panel established from the significantly changed serum indicators can be used for the early prediction of GDM, with the area under the receiver-operating characteristic curve (ROC) 0.83. High serum uric acid and C-reaction protein levels were risk factors for GDM independent of body mass indexes, with ORs 4.76 (95% CI: 2.08-10.90) and 3.10 (95% CI: 1.38-6.96), respectively. Predictive phenotype panel of GDM, together with the risk factors of GDM, will provide novel perspectives for the early clinical warning and diagnosis of GDM.

Life Cycle Exposure to Environmentally Relevant Concentrations of Diphenyl Phosphate (DPhP) Inhibits Growth and Energy Metabolism of Zebrafish in a Sex-Specific Manner.

Due to commercial uses and environmental degradation of aryl phosphate esters, diphenyl phosphate (DPhP) is frequently detected in environmental matrices and is thus of growing concern worldwide. However, information on potential adverse effects of chronic exposure to DPhP at environmentally realistic concentrations was lacking. Here, we investigated the effects of life cycle exposure to DPhP on zebrafish at environmentally relevant concentrations of 0.8, 3.9, or 35.6 μg/L and employed a dual-omics approach (metabolomics and transcriptomics) to characterize potential modes of action. Exposure to DPhP at 35.6 μg/L for 120 days resulted in significant reductions in body mass and length of male zebrafish, but did not cause those same effects to females. Predominant toxicological mechanisms, including inhibition of oxidative phosphorylation, down-regulation of fatty acid oxidation, and up-regulation of phosphatidylcholine degradation, were revealed by integrated dual-omics analysis and successfully linked to adverse outcomes. Activity of succinate dehydrogenase and protein content of carnitine O-palmitoyltransferase 1 were significantly decreased in livers of male fish exposed to DPhP, which further confirmed the proposed toxicological mechanisms. This study is the first to demonstrate that chronic, low-level exposure to DPhP can retard growth via inhibiting energy output in male zebrafish.

The Role of Metabolism in Modulating Radiation Fibrosis & Lymphedema

With the improvements in cancer diagnosis and therapy, clinical outcome has increased substantially, to the extent that by 2026, there will be >20M cancer survivors in the United States alone. The majority of survivors fare well; however, there is a significant proportion of patients who continue to experience the aftermath of their treatments such as radiation fibrosis (RF) or lymphedema. Until recently, there has been little understanding of the biological basis of RF, which is defined by the aberrant accumulation of extracellular matrix (ECM), leading to reduced tissue elasticity, and potential loss of organ function. The regulation of ECM production and degradation is a complex process, mediated through the production of a myriad of cytokines such as TGFB, and environmental conditions such as hypoxia. We have recently identified that metabolic dysregulation, and in particular, downregulation of fatty acid oxidation (FAO) is a key pathway driving the production and reducing the degradation of collagen, the predominant component of the ECM. Conversely, upregulation of PPAR signaling, a major mediator of FAO, reduced transcription of key ECM genes; whilst increasing internalization and degradation of extracellular collagen. Furthermore, CD36, a membrane transporter of long chain fatty acids, was discovered to play a critical role in regulating the trafficking of collagen and lysosomal degradation. Finally, using a pharmacogenomics approach, caffeic acid was identified to be a compound which can be systemically administered, and reduced RF in pre-clinical models. These data, along with other reports in the literature, strongly point towards the use of metabolic drugs as a therapeutic strategy by which we can mitigate the deleterious effects of RF on cancer patients. More recently, we have also uncovered that metabolic dysregulation likely plays a similar role in the development of lymphedema, a common long-term toxicity in women after breast cancer therapy. Finally, these findings have broad application to the increasing burden of fibrosis beyond cancer, wherein end-stage renal, liver, cardiac, and pulmonary fibrosis accounts for up to 1/3 of the deaths around the world.

Detrimental Role of High Dietary Phosphate Intake on Skeletal Muscle ATP Synthesis in Healthy Humans

Background Inorganic phosphate (Pi) is found in large quantities in processed food to increase palatability and longevity on the shelf. Prior work from our group using animal models suggests that increased dietary Pi is associated with lower resting metabolic rate as well as downregulation of fatty acid oxidation and upregulation of carbohydrate metabolism. We hypothesized that higher dietary Pi would be associated with lower resting mitochondrial function as measured by in-vivo, 31P MR spectroscopy. Methods The study sample included participants from the Dallas Heart Study without history of cardiovascular vascular disease or kidney disease. Dietary Pi (in mg/kcal/day) was measured from a 24-hour dietary recall (ASA24®). 31P MR spectroscopy (7T, Philips) was used for the assessment of ATP synthesis production at rest in the calf muscle using wideband inversion transfer technology. We calculated the correlation coefficient (Pearson's) to quantify the association between both dietary Pi intake normalized to total energy intake (mg/Kcal/day) and urinary Pi (mg/gram of creatinine, Cr) with resting ATP synthesis (mM/min). Results We studied 12 individuals (46% female) with a mean age and BMI of 59 and 28, respectively. Within this population, 15% were African American and 46% were non-white. Hypertension was present in 46% and diabetes in 23%. All but one participant had normal renal function (mean GFR 80 mL/min). The average dietary Pi intake was 1408 mg and the average urinary Pi was 44 mg/L. Results are shown in figure 1 and 2. Figure 1 demonstrates an inverse association between dietary Pi and resting ATP synthesis (r = -0.62, p= 0.03). We observed a directionally similar pattern of association between urinary Pi/Cr and resting ATP synthesis (Figure 2). Conclusions Our study suggests a deleterious role of inorganic phosphate on resting ATP synthesis in skeletal muscle. Given the importance of muscle mitochondrial function for exercise capacity, these data suggest that excess dietary Pi intake, associated with the Western diet, may explain reduced physical activity, which may predispose the general population to increased cardiovascular risk.

Downregulation of fatty acid oxidation by involvement of HIF-1α and PPARγ in human gastric adenocarcinoma and related clinical significance.

Lipid metabolism rewiring in gastric adenocarcinoma (GA) pathogenesis is still not clearly elucidated. This study aimed to describe the role of lipid catabolism in GA patient outcomes and possible therapeutic targets by analyzing the effect of hypoxia-inducible factor-1α (HIF-1α) on fatty acid oxidation (FAO). AGS cell line was cultured in normoxic and hypoxic conditions, and FAO-related genes were analyzed by real-time-PCR and Western-blot. The study group comprised 108 newly diagnosed GA patients and 152 control cases. Serum concentrations of medium and long-chain acyl-CoA dehydrogenases (MCAD and LCAD) proteins were measured using ELISA, and local expression of HIF-1α, carnitine palmitoyl transferase 1 (CPT1A) and peroxisome proliferator-activated receptor γ (PPARγ) was evaluated by immunohistochemistry. In addition, gene expression of PPARγ, CPT1A, LCAD, and MCAD was assessed by real-time-PCR. In vitro findings indicate HIF-1α upregulation and FAO-related genes and proteins reduction in the hypoxic culture of AGS cells. GA patients had significantly lower circulating levels of LCAD compared to controls. Higher protein expression of HIF-1α and downregulated CPT1A and PPARγ were observed in GA tissues versus controls. Gene expression of CPT1A, PPARγ, LCAD, and MCAD were repressed in GA tissues compared to controls. Moreover, reduced expression of CPT1A, PPARγ, and MCAD were correlated with HIF-1α upregulation in GA. Poor patient outcome was associated with lower PPARγ and LCAD expression in GA. HIF-1α upregulation in human GA patients and AGS cells was paralleled by downregulation of lipid catabolism genes potentially via reduced PPARγ-mediated FAO. This metabolic adaptation to hypoxic condition may play a role in GA pathogenesis and might have clinical and therapeutic value in GA patients.

The Role of Metabolism in Modulating Radiation Fibrosis & Lymphedema

With the improvements in cancer diagnosis and therapy, clinical outcome has increased substantially, to the extent that by 2026, there will be >20M cancer survivors in the United States alone. The majority of survivors fare well; however, there is a significant proportion of patients who continue to experience the aftermath of their treatments such as radiation fibrosis (RF) or lymphedema. Until recently, there has been little understanding of the biological basis of RF, which is defined by the aberrant accumulation of extracellular matrix (ECM), leading to reduced tissue elasticity, and potential loss of organ function. The regulation of ECM production and degradation is a complex process, mediated through the production of a myriad of cytokines such as TGFB, and environmental conditions such as hypoxia. We have recently identified that metabolic dysregulation, and in particular, downregulation of fatty acid oxidation (FAO) is a key pathway driving the production and reducing the degradation of collagen, the predominant component of the ECM. Conversely, upregulation of PPAR signaling, a major mediator of FAO, reduced transcription of key ECM genes; whilst increasing internalization and degradation of extracellular collagen. Furthermore, CD36, a membrane transporter of long chain fatty acids, was discovered to play a critical role in regulating the trafficking of collagen and lysosomal degradation. Finally, using a pharmacogenomics approach, caffeic acid was identified to be a compound which can be systemically administered, and reduced RF in pre‐clinical models.These data, along with other reports in the literature, strongly point towards the use of metabolic drugs as a therapeutic strategy by which we can mitigate the deleterious effects of RF on cancer patients. More recently, we have also uncovered that metabolic dysregulation likely plays a similar role in the development of lymphedema, a common long‐term toxicity in women after breast cancer therapy. Finally, these findings have broad application to the increasing burden of fibrosis beyond cancer, wherein end‐stage renal, liver, cardiac, and pulmonary fibrosis accounts for up to 1/3 of the deaths around the world.Support or Funding InformationThe work is funded in part by the Canadian Institutes of Health Research, Genome Canada, the Canadian Cancer Society Research Institute, the Physicians Services Incorporated Foundation, the Harry Barberian Research Scholarship, the Mariano Elia Chair in Head and Neck Cancer Research, the Peter and Shelagh Godsoe Chair in Radiation Medicine, the Princess Margaret Cancer Centre Head and Neck Translational Program, the Princess Margaret Cancer Centre Radiation Medicine Program and the Ottawa Heart Institute Research Corporation, as well by philanthropic funding from the Jesse Rasch Foundation, the Wharton family, Joe’s Team and the Ministry of Health and Long‐Term Care.

Pyruvate dehydrogenase activation precedes the down-regulation of fatty acid oxidation in monocrotaline-induced myocardial toxicity in mice.

Fatty acid (FA) oxidation is impaired and glycolysis is promoted in the damaged heart. However, the factor(s) in the early stages of myocardial metabolic impairment remain(s) unclear. C57B6 mice were subcutaneously administered monocrotaline (MCT) in doses of 0.3mg/g body weight twice a week for 3 or 6weeks. Right and left ventricles at 3 and 6weeks after administration were subjected to capillary electrophoresis-mass spectrometry metabolomic analysis. We also examined mRNA and protein levels of key metabolic molecules. Although no evidence of PH and right ventricular failure was found in the MCT-administered mice by echocardiographic and histological analyzes, the expression levels of stress markers such as TNFα and IL-6 were increased in right and left ventricles even at 3weeks, suggesting that there was myocardial damage. Metabolites in the tricarboxylic acid (TCA) cycle were decreased and those in glycolysis were increased at 6weeks. The expression levels of FA oxidation-related factors were decreased at 6weeks. The phosphorylation level of pyruvate dehydrogenase (PDH) was significantly decreased at 3weeks. FA oxidation and the TCA cycle were down-regulated, whereas glycolysis was partially up-regulated by MCT-induced myocardial damage. PDH activation preceded these alterations, suggesting that PDH activation is one of the earliest events to compensate for a subtle metabolic impairment from myocardial damage.

Nanotized PPARα Overexpression Targeted to Hypertrophied Myocardium Improves Cardiac Function by Attenuating the p53-GSK3β-Mediated Mitochondrial Death Pathway.

Metabolic remodeling of cardiac muscles during pathological hypertrophy is characterized by downregulation of fatty acid oxidation (FAO) regulator, peroxisome proliferator-activated receptor alpha (PPARα). Thereby, we hypothesized that a cardiac-specific induction of PPARα might restore the FAO-related protein expression and resultant energy deficit. In the present study, consequences of PPARα augmentation were evaluated for amelioration of chronic oxidative stress, myocyte apoptosis, and cardiac function during pathological cardiac hypertrophy. Nanotized PPARα overexpression targeted to myocardium was done by a stearic acid-modified carboxymethyl-chitosan (CMC) conjugated to a 20-mer myocyte-targeted peptide (CMCP). Overexpression of PPARα ameliorated pathological hypertrophy and improved cardiac function. Augmented PPARα in hypertrophied myocytes revealed downregulated p53 acetylation (lys 382), leading to reduced apoptosis. Such cells showed increased binding of PPARα with p53 that in turn reduced interaction of p53 with glycogen synthase kinase-3β (GSK3β), which upregulated inactive phospho-GSK3β (serine [Ser]9) expression within mitochondrial protein fraction. Altogether, the altered molecular milieu in PPARα-overexpressed hypertrophy groups restored mitochondrial structure and function both in vitro and in vivo. Cardiomyocyte-targeted overexpression of a protein of interest (PPARα) by nanotized plasmid has been described for the first time in this study. Our data provide a novel insight towards regression of pathological hypertrophy by ameliorating mitochondrial oxidative stress in targeted PPARα-overexpressed myocardium. PPARα-overexpression during pathological hypertrophy showed substantial betterment of mitochondrial structure and function, along with downregulated apoptosis. Myocardium-targeted overexpression of PPARα during pathological cardiac hypertrophy led to an overall improvement of cardiac energy deficit and subsequent cardiac function, thereby, opening up a potential avenue for cardiac tissue engineering during hypertrophic cardiac pathophysiology.

The Regulation of Lipid Deposition by Insulin in Goose Liver Cells Is Mediated by the PI3K-AKT-mTOR Signaling Pathway.

BackgroundWe previously showed that the fatty liver formations observed in overfed geese are accompanied by the activation of the PI3K-Akt-mTOR pathway and an increase in plasma insulin concentrations. Recent studies have suggested a crucial role for the PI3K-Akt-mTOR pathway in regulating lipid metabolism; therefore, we hypothesized that insulin affects goose hepatocellular lipid metabolism through the PI3K-Akt-mTOR signaling pathway.MethodsGoose primary hepatocytes were isolated and treated with serum-free media supplemented with PI3K-Akt-mTOR pathway inhibitors (LY294002, rapamycin, and NVP-BEZ235, respectively) and 50 or 150 nmol/L insulin.ResultsInsulin induced strong effects on lipid accumulation as well as the mRNA and protein levels of genes involved in lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in primary goose hepatocytes. The stimulatory effect of insulin on lipogenesis was significantly decreased by treatment with PI3K-Akt-mTOR inhibitors. These inhibitors also rescued the insulin-induced down-regulation of fatty acid oxidation and VLDL-TG assembly and secretion.ConclusionThese findings suggest that the stimulatory effect of insulin on lipid deposition is mediated by PI3K-Akt-mTOR regulation of lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in goose hepatocytes.

Nontargeted Urinary Metabolite Profiling of a Mouse Model of Crohn’s Disease

Crohn's disease is an inflammatory disorder of the bowel, believed to arise from the dysregulation of intestinal mucosal immunity. The interleukin-10-deficient (IL10-/-) mouse, which develops intestinal inflammation in the presence of gut microflora, serves as a mouse model of Crohn's disease. Nontargeted urinary metabolite profiling was carried out to identify systemic metabolic changes associated with the development of intestinal inflammation caused by IL10-deficiency. Spot urine samples, collected from IL10-/- and wildtype mice at ages 5.5, 7, 8.5, and 10.5 weeks old were analyzed by gas chromatography-mass spectrometry (GCMS). The data were analyzed using XCMS software, multiple t tests, and ANOVA. Among the key metabolic differences detected were elevated urinary levels of xanthurenic acid and fucose in IL10-/- mice relative to wildtype, indicating upregulation of tryptophan catabolism and perturbed fucosylation in IL10-/- mice. Three short-chain dicarboxylic acid metabolites were decreased in urine of IL10-/- mice relative to wildtype, suggesting the downregulation of fatty acid oxidation in IL10-/- mice. These metabolic differences were reproducible in an independent set of mice. This study demonstrates that nontargeted GCMS metabolite profiling of IL10-/- mice can provide insights into the metabolic effects of IL10-deficiency and identify potential markers of intestinal inflammation.

Fasting induces a biphasic adaptive metabolic response in murine small intestine.

BackgroundThe gut is a major energy consumer, but a comprehensive overview of the adaptive response to fasting is lacking. Gene-expression profiling, pathway analysis, and immunohistochemistry were therefore carried out on mouse small intestine after 0, 12, 24, and 72 hours of fasting.ResultsIntestinal weight declined to 50% of control, but this loss of tissue mass was distributed proportionally among the gut's structural components, so that the microarrays' tissue base remained unaffected. Unsupervised hierarchical clustering of the microarrays revealed that the successive time points separated into distinct branches. Pathway analysis depicted a pronounced, but transient early response that peaked at 12 hours, and a late response that became progressively more pronounced with continued fasting. Early changes in gene expression were compatible with a cellular deficiency in glutamine, and metabolic adaptations directed at glutamine conservation, inhibition of pyruvate oxidation, stimulation of glutamate catabolism via aspartate and phosphoenolpyruvate to lactate, and enhanced fatty-acid oxidation and ketone-body synthesis. In addition, the expression of key genes involved in cell cycling and apoptosis was suppressed. At 24 hours of fasting, many of the early adaptive changes abated. Major changes upon continued fasting implied the production of glucose rather than lactate from carbohydrate backbones, a downregulation of fatty-acid oxidation and a very strong downregulation of the electron-transport chain. Cell cycling and apoptosis remained suppressed.ConclusionThe changes in gene expression indicate that the small intestine rapidly looses mass during fasting to generate lactate or glucose and ketone bodies. Meanwhile, intestinal architecture is maintained by downregulation of cell turnover.

Spatial distribution of blood flow predicts metabolic phenotypes and electrophysiological properties in the heart

High resolution analysis (300 μl) reveals a temporally stable, patchy pattern of perfusion in the LV free wall. Low flow areas (LowF) receive <60% of the average flow and display decreased energy turnover and enhanced fatty acid content, while the mechanism of this spatial heterogeneity is unknown. Lipid profile analysis of canine LowF showed an elevated triglyceride content (+212%) when compared to High flow samples (HighF). In LowF, mRNA expression of PPARα was reduced to ¼ and the same was true for 5 key enzymes or transporters in lipid metabolism (p<0.05 – 0.001), suggesting preferential glycolysis in LowF. Potassium ion channel expression also displayed flow-related differences, with enhanced expression of ERG (3.5-fold, p<0.001) and KChIP2 (2.1-fold) in LowF, mediating resp. governing IKr and Ito. Model analysis predicted a shorter local APD in LowF, which was confirmed by epicardial mapping of local QT intervals. This revealed a patchy, temporally stable pattern and a QT prolongation upon ERG blockade predominantly in areas of short basal QT (p<0.01). In short QT areas, ERG expression was increased and basal perfusion decreased. Thus, within the apparently homogenous left ventricular wall, distinct metabolic and electrophysiological phenotypes exist. In LowF, both downregulation of fatty acid oxidation and APD shortening will reduce O2 consumption in these regions and may thus partially explain the spatial heterogeneity of perfusion. Supported by DFG.

Nuclear Factor-κB Activation Leads to Down-regulation of Fatty Acid Oxidation during Cardiac Hypertrophy

Little is known about the mechanisms responsible for the fall in fatty acid oxidation during the development of cardiac hypertrophy. We focused on the effects of nuclear factor (NF)-kappaB activation during cardiac hypertrophy on the activity of peroxisome proliferator-activated receptor (PPAR) beta/delta, which is the predominant PPAR subtype in cardiac cells and plays a prominent role in the regulation of cardiac lipid metabolism. Phenylephrine-induced cardiac hypertrophy in neonatal rat cardiomyocytes caused a reduction in the expression of pyruvate dehydrogenase kinase 4 (Pdk4), a target gene of PPARbeta/delta involved in fatty acid utilization, and a fall in palmitate oxidation that was reversed by NF-kappaB inhibitors. Lipopolysaccharide stimulation of NF-kappaB in embryonic rat heart-derived H9c2 myotubes, which only express PPARbeta/delta, caused both a reduction in Pdk4 expression and DNA binding activity of PPARbeta/delta to its response element, effects that were reversed by NF-kappaB inhibitors. Coimmunoprecipitation studies demonstrated that lipopolysaccharide strongly stimulated the physical interaction between the p65 subunit of NF-kappaB and PPARbeta/delta, providing an explanation for the reduced activity of PPARbeta/delta. Finally, we assessed whether this mechanism was present in vivo in pressure overload-induced cardiac hypertrophy. In hypertrophied hearts of banded rats the reduction in the expression of Pdk4 was accompanied by activation of NF-kappaB and enhanced interaction between p65 and PPARbeta/delta. These results indicate that NF-kappaB activation during cardiac hypertrophy down-regulates PPARbeta/delta activity, leading to a fall in fatty acid oxidation, through a mechanism that involves enhanced protein-protein interaction between the p65 subunit of NF-kappaB and PPARbeta/delta.

Moderate severity heart failure does not involve a downregulation of myocardial fatty acid oxidation.

Recent human and animal studies have demonstrated that in severe end-stage heart failure (HF), the cardiac muscle switches to a more fetal metabolic phenotype, characterized by downregulation of free fatty acid (FFA) oxidation and an enhancement of glucose oxidation. The goal of this study was to examine myocardial substrate metabolism in a model of moderate coronary microembolization-induced HF. We hypothesized that during well-compensated HF, FFA oxidation would predominate as opposed to a more fetal metabolic phenotype of greater glucose oxidation. Cardiac substrate uptake and oxidation were measured in normal dogs (n = 8) and in dogs with microembolization-induced HF (n = 18, ejection fraction = 28%) by infusing three isotopic tracers ([9,10-(3)H]oleate, [U-(14)C]glucose, and [1-(13)C]lactate) in anesthetized open-chest animals. There were no differences in myocardial substrate metabolism between the two groups. The total activity of pyruvate dehydrogenase, the key enzyme regulating myocardial pyruvate oxidation (and hence glucose and lactate oxidation) was not affected by HF. We did not observe any difference in the activity of carnitine palmitoyl transferase I (CPT-I) and its sensitivity to inhibition by malonyl-CoA between groups; however, malonyl-CoA content was decreased by 22% with HF, suggesting less in vivo inhibition of CPT-I activity. The differences in malonyl-CoA content cannot be explained by changes in the Michaelis-Menten constant and maximal velocity for malonyl-CoA decarboxylase because neither were affected by HF. These results support the concept that there is no decrease in fatty acid oxidation during compensated HF and that the downregulation of fatty acid oxidation enzymes and the switch to carbohydrate oxidation observed in end-stage HF is only a late-stage phenomenon.