Targeting autophagy with SAR405 alleviates doxorubicin-induced cardiotoxicity.

Anthracycline antitumor agents, such as doxorubicin (DOX), are effective in the treatment of solid tumors and hematological malignancies, but anthracycline-induced cardiotoxicity (AIC) limits their application as chemotherapeutics. Dexrazoxane (DEX) has been adopted to prevent AIC. Using a chronic AIC mouse model, we demonstrated that DEX is insufficient to reverse DOX-induced cardiotoxicity. Although therapies targeting autophagy have been explored to prevent AIC, but whether novel autophagy inhibitors could alleviate or prevent AIC in clinically relevant models needs further investigation. Here, we show that genetic ablation of Atg7, a key regulator in the early phase of autophagy, protected mice against AIC. We further demonstrated that SAR405, a novel autophagy inhibitor, attenuated DOX-induced cytotoxicity. Intriguingly, the combination of DEX and SAR405 protected cells against DOX-induced cardiotoxicity in vivo. Using the cardiomyocyte cell lines AC16 and H9c2, we determined that autophagy was initiated during AIC. Our results suggest that inhibition of autophagy at its early phase with SAR405 combined with DEX represents an effective therapeutic strategy to prevent AIC.

Deletion of Rap1 protects against myocardial ischemia/reperfusion injury through suppressing cell apoptosis via activation of STAT3 signaling.

Ischemic heart disease is a leading cause of morbidity and mortality. Repressor activator protein 1 (Rap1), an established telomere-associated protein, is a novel modulator of hypoxia-induced apoptosis. This study aimed to explore the potential direct role of Rap1 in myocardial ischemia/reperfusion injury (I/RI) and to determine the underlying molecular mechanism. In a mouse model of myocardial I/RI (30-min of left descending coronary artery ligation followed by 2-h reperfusion), Rap1 deficiency significantly reduced myocardial infarct size (IS) and improved cardiac systolic/diastolic function. This was associated with a reduction in apoptosis in the post-ischemic myocardium. In H9C2 and primary cardiomyocytes, Rap1 knockdown or knockout significantly suppressed hypoxia/reoxygenation (H/R)-induced cell injury and apoptosis through increasing the phosphorylation/activation of STAT3 at site Ser727 and translocation of STAT3 to the nucleus. We surmise this since Stattic (selective STAT3 inhibitor) pretreatment canceled the abovementioned protective effect. Furthermore, co-immunoprecipitation assay revealed a direct interaction between Rap1 and STAT3, but not JAK2, suggesting that the association of Rap1 with STAT3 may contribute to the reduced activity of STAT3 (Ser727 ) upon H/R stimulation. In conclusion, Rap1 deficiency protects the heart from ischemic damage through STAT3-dependent reduction of cardiomyocyte apoptosis, which may yield viable target for pharmacological intervention in ischemic heart disease.

Iron overload inhibits late stage autophagic flux leading to insulin resistance.

Iron overload, a common clinical occurrence, is implicated in the metabolic syndrome although the contributing pathophysiological mechanisms are not fully defined. We show that prolonged iron overload results in an autophagy defect associated with accumulation of dysfunctional autolysosomes and loss of free lysosomes in skeletal muscle. These autophagy defects contribute to impaired insulin-stimulated glucose uptake and insulin signaling. Mechanistically, we show that iron overload leads to a decrease in Akt-mediated repression of tuberous sclerosis complex (TSC2) and Rheb-mediated mTORC1 activation on autolysosomes, thereby inhibiting autophagic-lysosome regeneration. Constitutive activation of mTORC1 or iron withdrawal replenishes lysosomal pools via increased mTORC1-UVRAG signaling, which restores insulin sensitivity. Induction of iron overload via intravenous iron-dextran delivery in mice also results in insulin resistance accompanied by abnormal autophagosome accumulation, lysosomal loss, and decreased mTORC1-UVRAG signaling in muscle. Collectively, our results show that chronic iron overload leads to a profound autophagy defect through mTORC1-UVRAG inhibition and provides new mechanistic insight into metabolic syndrome-associated insulin resistance.

Holo-lipocalin-2-derived siderophores increase mitochondrial ROS and impair oxidative phosphorylation in rat cardiomyocytes.

Lipocalin-2 (Lcn2), a critical component of the innate immune response which binds siderophores and limits bacterial iron acquisition, can elicit spillover adverse proinflammatory effects. Here we show that holo-Lcn2 (Lcn2-siderophore-iron, 1:3:1) increases mitochondrial reactive oxygen species (ROS) generation and attenuates mitochondrial oxidative phosphorylation in adult rat primary cardiomyocytes in a manner blocked by N-acetyl-cysteine or the mitochondria-specific antioxidant SkQ1. We further demonstrate using siderophores 2,3-DHBA (2,3-dihydroxybenzoic acid) and 2,5-DHBA that increased ROS and reduction in oxidative phosphorylation are direct effects of the siderophore component of holo-Lcn2 and not due to apo-Lcn2 alone. Extracellular apo-Lcn2 enhanced the potency of 2,3-DHBA and 2,5-DHBA to increase ROS production and decrease mitochondrial respiratory capacity, whereas intracellular apo-Lcn2 attenuated these effects. These actions of holo-Lcn2 required an intact plasma membrane and were decreased by inhibition of endocytosis. The hearts, but not serum, of Lcn2 knockout (LKO) mice contained lower levels of 2,5-DHBA compared with wild-type hearts. Furthermore, LKO mice were protected from ischemia/reperfusion-induced cardiac mitochondrial dysfunction. Our study identifies the siderophore moiety of holo-Lcn2 as a regulator of cardiomyocyte mitochondrial bioenergetics.

Astragaloside â £ Protects Against Aß1-42-induced Oxidative Stress, Neuroinflammation and Cognitive Impairment in Rats.

Objective To investigate the neuroprotective action of astragaloside Ⅳ (AS-Ⅳ) on spatial learning and memory impairment induced by amyloid-beta 1-42 (Aß1-42) in rats and elucidate its underlying molecular mechanisms. Methods Adult-male Sprague-Dawley rats (230-250 g) were divided into six groups randomly: control, Aß1-42, AS-Ⅳ, Aß1-42 plus 5 mg/kg·d AS-Ⅳ, Aß1-42 plus 25 mg/kg·d AS-Ⅳ, and Aß1-42 plus 50 mg/kg·d AS-Ⅳ groups. Aß1-42 were delivered by intracerebroventricular injection under the guidance of a brain stereotaxic apparatus. The Morris water maze test (hidden platform test, probe trials, visible platform test) was performed one week after Aß1-42 injection to obtain the ability of rat spatial learning and memory. AS-Ⅳ (5, 25 and 50 mg/kg·d) was administrated intraperitoneally once per day from the 8th day after Aß1-42 injection for 5 consecutive days. Average escape latencies, distances for searching for the platform under water and the percentage of total time elapsed and distance swam in the right quadrant after removing platform were determined by behavior software system. The vision and swim speeds of rats were also determined to exclude the effect of these factors on the parameters of learning and memory. After behavioral tests, the rats were sacrificed immediately by decapitation. Hippocampus were collected. The enzyme activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-px) and catalase (CAT) in the hippocampus obtained from different-treated rat brain were measured by following the manufacturer's instructions. The levels of interleukin-1 beta (IL-1ß) and tumor necrosis factor-alpha (TNF-α) in tissue lysates were assayed with ELISA. Results The water maze test results indicated that chronic treatments with AS-Ⅳ effectively protected the rats from Aß1-42-induced spatial learning and memory impairment. Furthermore, the activities of SOD, GSH-px and CAT decreased by Aß1-42 were also restored by AS-Ⅳ treatment in the hippocampus of rats. In addition, AS-Ⅳ significantly decreased the levels of IL-1ß and TNF-α in the hippocampus of Aß1-42-induced amnesia's rats. Conclusion Our findings suggest that AS-Ⅳ might be a useful chemical in improving the spatial memory and relieving the oxidative stress and neuroinflammation in Alzheimer patients.

Lipocalin-2 (NGAL) Attenuates Autophagy to Exacerbate Cardiac Apoptosis Induced by Myocardial Ischemia.

Lipocalin-2 (Lcn2; also termed neutrophil gelatinase-associated lipocalin (NGAL)) levels correlate positively with heart failure (HF) yet mechanisms via which Lcn2 contributes to the pathogenesis of HF remain unclear. In this study, we used coronary artery ligation surgery to induce ischemia in wild-type (wt) mice and this induced a significant increase in myocardial Lcn2. We then compared wt and Lcn2 knockout (KO) mice and observed that wt mice showed greater ischemia-induced caspase-3 activation and DNA damage measured by TUNEL than Lcn2KO mice. Analysis of autophagy by LC3 and p62 Western blotting, LC3 immunohistochemistry and transmission electron microscopy (TEM) indicated that Lcn2 KO mice had a greater ischemia-induced increase in autophagy. Lcn2KO were protected against ischemia-induced cardiac functional abnormalities measured by echocardiography. Upon treating a cardiomyocyte cell line (h9c2) with Lcn2 and examining AMPK and ULK1 phosphorylation, LC3 and p62 by Western blot as well as tandem fluorescent RFP/GFP-LC3 puncta by immunofluorescence, MagicRed assay for lysosomal cathepsin activity and TEM we demonstrated that Lcn2 suppressed autophagic flux. Lcn2 also exacerbated hypoxia-induced cytochromc c release from mitochondria and caspase-3 activation. We generated an autophagy-deficient H9c2 cell model by overexpressing dominant-negative Atg5 and found significantly increased apoptosis after Lcn2 treatment. In summary, our data indicate that Lcn2 can suppress the beneficial cardiac autophagic response to ischemia and that this contributes to enhanced ischemia-induced cell death and cardiac dysfunction. J. Cell. Physiol. 232: 2125-2134, 2017. © 2016 Wiley Periodicals, Inc.

Mice lacking prostaglandin E receptor subtype 4 manifest disrupted lipid metabolism attributable to impaired triglyceride clearance.

Upon high-fat feeding, prostaglandin E receptor subtype 4 (EP4)-knockout mice gain less body weight than their EP4(+/+) littermates. We investigated the cause of the lean phenotype. The mice showed a 68.8% reduction in weight gain with diminished fat mass that was not attributable to reduced food intake, fat malabsorption, or increased energy expenditure. Plasma triglycerides in the mice were elevated by 244.9%. The increase in plasma triglycerides was independent of changes in hepatic very low density lipoprotein (VLDL)-triglyceride production or intestinal chylomicron-triglyceride synthesis. However, VLDL-triglyceride clearance was drastically impaired in the EP4-knockout mice. The absence of EP4 in mice compromised the activation of lipoprotein lipase (LPL), the key enzyme responsible for trafficking of plasma triglycerides into peripheral tissues. Deficiency in EP4 reduced hepatic mRNA expression of the transcriptional factor cAMP response element binding protein H (by 36.8%) and LPL activators, including apolipoprotein (Apo)a5 (by 40.2%) and Apoc2 (by 61.3%). In summary, the lean phenotype of EP4-deficient mice resulted from reduction in adipose tissue and accretion of other peripheral organs caused by impaired triglyceride clearance. The findings identify a new metabolic dimension in the physiologic role played by endogenous EP4.

Ischemia-induced cardiac dysfunction is exacerbated in adiponectin-knockout mice due to impaired autophagy flux.

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.

Distinct changes in serum metabolites and lipid species in the onset and progression of NAFLD in Obese Chinese.

Introduction: Metabolic disturbances are major contributors to the onset and progression of non-alcoholic fatty liver disease (NAFLD), which includes a histological spectrum ranging from single steatosis (SS) to non-alcoholic steatohepatitis (NASH). This study aimed to identify serum metabolites and lipids enriched in different histological stages of NAFLD and to explore metabolites/lipids as non-invasive biomarkers in risk prediction of NAFLD and NASH in obese Chinese. Methods: Serum samples and liver biopsies were obtained from 250 NAFLD subjects. Untargeted metabolomic and lipidomic profiling were performed using Liquid Chromatography-Mass Spectrometry. Significantly altered metabolites and lipids were identified by MaAsLin2. Pathway enrichment was conducted with MetaboAnalyst and LIPEA. WGCNA was implemented to construct the co-expression network. Logistic regression models were developed to classify different histological stages of NAFLD. Results: A total of 263 metabolites and 550 lipid species were detected in serum samples. Differential analysis and pathway enrichment analysis revealed the progressive patterns in metabolic mechanisms during the transition from normal liver to SS and to NASH, including N-palmitoyltaurine, tridecylic acid, and branched-chain amino acid signaling pathways. The co-expression network showed a distinct correlation between different triglyceride and phosphatidylcholine species with disease severity. Multiple models classifying NAFLD versus normal liver and NASH versus SS identified important metabolic features associated with significant improvement in disease prediction compared to conventional clinical parameters. Conclusion: Different histological stages of NAFLD are enriched with distinct sets of metabolites, lipids, and metabolic pathways. Integrated algorithms highlight the important metabolic and lipidomic features for diagnosis and staging of NAFLD in obese individuals.

Chinese medicine Linggui Zhugan formula protects against diabetic kidney disease in close association with inhibition of proteinase 3-mediated podocyte apoptosis in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Linggui-Zhugan (LGZG) comprises four herbs and is a classic formula in traditional Chinese medicine. There is strong clinical evidence of its pleiotropic effects in the prevention of diabetes and its related complications. Although several classes of drugs are currently available for clinical management of diabetic kidney disease (DKD), tight glycemic and/or hypertension control may not prevent disease progression. This study evaluated the therapeutic effect of the ethnopharmacological agent LGZG on DKD. AIM OF THE STUDY: This study aimed to investigate the effects of LGZG formula with standard quality control on experimental DKD and its related metabolic disorders in animal model. Meanwhile, the present study aimed to investigate regulatory effects of LGZG on renal proteinase 3 (PR3) to reveal mechanisms underlying renoprotective benefits of LGZG. MATERIALS AND METHODS: LGZG decoction was fingerprinted by high-performance liquid chromatography for quality control. An experimental model of DKD was induced in C57 BL/6J mice by a combination of high-fat diet feeding, uninephrectomy, and intraperitoneal injection of streptozocin. The LGZG decoction was administrated by daily oral gavage. RESULTS: Treatment with LGZG formula significantly attenuated DKD-like traits (including severe albuminuria, mesangial matrix expansion, and podocyte loss) and metabolic dysfunction (disordered body composition and dyslipidemia) in mice. RNA sequencing data revealed a close association of LGZG treatment with marked modulation of signaling pathways related to podocyte injury and cell apoptosis. Mechanistically, LGZG suppressed the DKD-triggered increase in renal PR3 and podocyte apoptosis. In-vitro incubation of mouse immortalized podocytes with LGZG-medicated serum attenuated PR3-mediated apoptosis. CONCLUSION: Our data demonstrated that the LGZG formula protected against DKD in mice and was closely associated with its inhibitory effects on PR3-mediated podocyte apoptosis.

Metabolic Dysfunction-Associated Steatohepatitis Detected by Neutrophilic Crown-Like Structures in Morbidly Obese Patients: A Multicenter and Clinicopathological Study.

Metabolic dysfunction-associated steatohepatitis (MASH) is the progressive form of metabolic dysfunction-associated steatotic liver disease (MASLD), and closely associated with a high risk of liver-related morbidity and mortality. Although enhanced neutrophil infiltration of the liver is a histological hallmark of MASH, the morphological pattern of hepatic neutrophils and their relevance to the definition of MASH remain unknown. This clinicopathological study aimed to determine the association of neutrophilic crown-like structures (CLSs) in liver biopsies and evaluate their relevance to the histological diagnosis of MASH. A total of 483 morbidly obese adults who underwent bariatric surgery were recruited. Neutrophilic CLSs in liver biopsies were detected by immunohistochemistry for neutrophil elastase and proteinase 3. All participants were classified into 4 histological subgroups: no MASLD (118, 24.4%), MASLD (76, 15.7%), borderline MASH (185, 38.3%), and definite MASH (104, 21.5%). In the discovery cohort (n = 379), the frequency of neutrophilic CLSs increased in line with the severity of liver disease. The number of neutrophilic CLSs was positively correlated with established histological characteristics of MASH. At a cutoff value of <0.3 per 20× microscopic field, the number of neutrophilic CLSs yielded a robust diagnostic accuracy to discriminate no MASLD and MASLD from borderline MASH and definite MASH; a cutoff at >1.3 per 20× microscopic field exhibited a statistically significant accuracy to distinguish definite MASH from other groups (no MASLD, MASLD, and borderline MASH). The significance of neutrophilic CLSs in identifying borderline MASH and definite MASH was confirmed in an external validation cohort (n = 104). The frequency of neutrophilic CLSs was significantly higher than that of macrophagic CLSs. In conclusion, neutrophilic CLSs in the liver represent a typical histological characteristic of MASH and may serve as a promising indicator to improve the diagnostic accuracy of MASH during histological assessment of liver biopsies.

Identification and multicentric validation of soluble CDCP1 as a robust serological biomarker for risk stratification of NASH in obese Chinese.

The definitive diagnosis of non-alcoholic steatohepatitis (NASH) currently relies on invasive and labor-intensive liver biopsy. Here, we identified soluble CUB domain-containing protein 1 (sCDCP1) as a top-ranked non-invasive biomarker for NASH using Olink-based proteomics in 238 obese individuals with liver biopsies. Both the circulating concentration and hepatic mRNA abundance of sCDCP1 were significantly elevated in patients with NASH and correlated closely with each histological feature of NASH. In the pooled multicenter validation cohort, sCDCP1 as a standalone biomarker achieved an area under the receiver operating characteristic (AUROC) of 0.838 (95% confidence interval [CI] 0.789-0.887) for diagnosing NASH, which is better than those achieved with cytokeratin-18 and other non-invasive tests. Furthermore, the C-DAG model established by the combination of sCDCP1 with diabetes, aspartate aminotransferase (AST), and gender accurately rules in and rules out both NASH and fibrotic NASH (gray zones <20%). Thus, sCDCP1-based non-invasive tests can be potentially implemented for screening and early diagnosis of NASH and for ruling out low-risk individuals to avoid unnecessary liver biopsies.

Copper fabric improves the metabolic profile of obese mice: Potential role of the gut microbiota.

Copper and copper alloys have antimicrobial activity through the rapid contact killing of viruses, bacteria and yeasts on their surface. Dysregulation of host microbiota can contribute to the pathogenesis of inflammatory diseases such as obesity, diabetes and cardiovascular disease. Anecdotal evidence noted improved overall well-being in individuals sleeping on copper-containing fabric bedding. We hypothesized that the beneficial effect of copper-infused fabric bedding on cardiometabolic health is linked to changes in gut microbiota composition. This study utilized a mouse model of diet-induced obesity to assess the beneficial effects of copper-infused fabric bedding on metabolic health. Body composition, inflammatory markers, metabolic and cardiovascular status and changes in the faecal microbiota composition were evaluated for up to 2 months in mice fed with a normal chow diet or high fat high cholesterol diet in the presence of bedding made with and without copper-infused fabric. Results showed that mice subjected to diet-induced obesity and housed in cages with copper-infused fabric liner displayed less body weight gain than mice in cages with control fabric. Mice housed with copper-infused fabric also displayed improved glucose tolerance and reduced inflammation biomarker lipocalin-2. We also observed a beneficial shift in gut bacterial composition of obese mice housed with copper fabric bedding. Taken in conjunction, our study provides direct animal-based evidence supporting the beneficial effects of copper fabric on metabolic health.

Caloric restriction-mimetics for the reduction of heart failure risk in aging heart: with consideration of gender-related differences.

The literature is full of claims regarding the consumption of polyphenol or polyamine-rich foods that offer some protection from developing cardiovascular disease (CVD). This is achieved by preventing cardiac hypertrophy and protecting blood vessels through improving the function of endothelium. However, do these interventions work in the aged human hearts? Cardiac aging is accompanied by an increase in left ventricular hypertrophy, along with diastolic and systolic dysfunction. It also confers significant cardiovascular risks for both sexes. The incidence and prevalence of CVD increase sharply at an earlier age in men than women. Furthermore, the patterns of heart failure differ between sexes, as do the lifetime risk factors. Do caloric restriction (CR)-mimetics, rich in polyphenol or polyamine, delay or reverse cardiac aging equally in both men and women? This review will discuss three areas: (1) mechanisms underlying age-related cardiac remodeling; (2) gender-related differences and potential mechanisms underlying diminished cardiac response in older men and women; (3) we select a few polyphenol or polyamine rich compounds as the CR-mimetics, such as resveratrol, quercetin, curcumin, epigallocatechin gallate and spermidine, due to their capability to extend health-span and induce autophagy. We outline their abilities and issues on retarding aging in animal hearts and preventing CVD in humans. We discuss the confounding factors that should be considered for developing therapeutic strategies against cardiac aging in humans.

Serum Thrombospondin-2 Levels Are Closely Associated With the Severity of Metabolic Syndrome and Metabolic Associated Fatty Liver Disease.

CONTEXT: Metabolic associated fatty liver disease (MAFLD) is the hepatic manifestation of obesity-related metabolic syndrome (MetS). Noninvasive biomarkers for monitoring the progression and severity of these metabolic comorbidities are needed. OBJECTIVES: To investigate the associations of serum thrombospondin-2 (TSP2) with MetS and MAFLD severity, and the potential diagnostic value of serum TSP2 for identifying at-risk metabolic associated steatohepatitis (MASH). METHODS: Blood samples, clinical data, and liver biopsies were collected from consecutively recruited 252 individuals with morbid obesity receiving bariatric surgery. Histopathology samples of liver biopsies were examined in a blinded fashion by 3 independent pathologists. Serum TSP2 levels were measured by enzyme-linked immunosorbent assay. RESULTS: Serum TSP2 levels were significantly elevated in MetS (1.58 [1.07-2.20] ng/mL) compared with non-MetS (1.28 [0.84-1.73] ng/mL; P = .006) in obese patients and positively correlated with increasing number of the MetS components, fasting glucose, glycated hemoglobin, fasting insulin, C-peptide, and homeostatic model assessment of insulin resistance after adjustment of conventional confounders. Serum TSP2 levels differentiated MASH (1.74 [1.32-3.09] ng/mL) from the other non-MASH less severe groups: normal liver (1.41 [1.04-1.63] ng/mL), simple steatosis (1.45 [0.89-1.92] ng/mL), and borderline MASH (1.30 [0.99-2.17] ng/mL) (P < .05). Elevated serum TSP2 was positively associated with the severity of hepatic steatosis, inflammation, fibrosis, and abnormal liver function independent of age, sex and adiposity. Furthermore, high serum TSP2 identified at-risk MASH with area under the operating curve of 0.84 (95% CI 0.70-0.98). CONCLUSION: Serum TSP2 is closely associated with severity and progression of MetS and MAFLD, and is a promising noninvasive biomarker for differentiating MASH from benign steatosis and identifying at-risk MASH patients among individuals with obesity.

Hepatocyte-Secreted Autotaxin Exacerbates Nonalcoholic Fatty Liver Disease Through Autocrine Inhibition of the PPARα/FGF21 Axis.

BACKGROUND & AIMS: The prevalence of nonalcoholic fatty liver disease (NAFLD) has reached epidemic proportions globally as a result of the rapid increase in obesity. However, there is no Food and Drug Administration-approved pharmacotherapy available for NAFLD. This study investigated the role of autotaxin, a secreted enzyme that hydrolyzes lysophosphatidylcholine to produce lysophosphatidic acid (LPA), in the pathogenesis of NAFLD and to explore whether genetic or pharmacologic interventions targeting autotaxin ameliorate NAFLD. METHODS: The clinical association of autotaxin with the severity of NAFLD was analyzed in 125 liver biopsy-proven NAFLD patients. C57BL/6N mice or fibroblast growth factor 21 (FGF21)-null mice were fed a high-fat diet or a choline-deficient diet to investigate the role of the autotaxin-FGF21 axis in NAFLD development by hepatic knockdown and antibody neutralization. Huh7 cells were used to investigate the autocrine effects of autotaxin. RESULTS: Serum autotaxin levels were associated positively with histologic scores and NAFLD severity. Hepatocytes, but not adipocytes, were the major contributor to increased circulating autotaxin in both patients and mouse models with NAFLD. In mice, knocking-down hepatic autotaxin or treatment with a neutralizing antibody against autotaxin significantly reduced high-fat diet-induced NAFLD and high fat- and choline-deficient diet-induced nonalcoholic steatohepatitis and fibrosis, accompanied by a marked increase of serum FGF21. Mechanistically, autotaxin inhibited the transcriptional activity of peroxisome proliferator-activated receptor α through LPA-induced activation of extracellular signal-regulated kinas, thereby leading to suppression of hepatic FGF21 production. The therapeutic benefit of anti-autotaxin neutralizing antibody against NAFLD was abrogated in FGF21-null mice. CONCLUSIONS: Liver-secreted autotaxin acts in an autocrine manner to exacerbate NAFLD through LPA-induced suppression of the peroxisome proliferator-activated receptor α-FGF21 axis and is a promising therapeutic target for NAFLD.

Hepatic MDM2 Causes Metabolic Associated Fatty Liver Disease by Blocking Triglyceride-VLDL Secretion via ApoB Degradation.

Dysfunctional triglyceride-very low-density lipoprotein (TG-VLDL) metabolism is linked to metabolic-associated fatty liver disease (MAFLD); however, the underlying cause remains unclear. The study shows that hepatic E3 ubiquitin ligase murine double minute 2 (MDM2) controls MAFLD by blocking TG-VLDL secretion. A remarkable upregulation of MDM2 is observed in the livers of human and mouse models with different levels of severity of MAFLD. Hepatocyte-specific deletion of MDM2 protects against high-fat high-cholesterol diet-induced hepatic steatosis and inflammation, accompanied by a significant elevation in TG-VLDL secretion. As an E3 ubiquitin ligase, MDM2 targets apolipoprotein B (ApoB) for proteasomal degradation through direct protein-protein interaction, which leads to reduced TG-VLDL secretion in hepatocytes. Pharmacological blockage of the MDM2-ApoB interaction alleviates dietary-induced hepatic steatohepatitis and fibrosis by inducing hepatic ApoB expression and subsequent TG-VLDL secretion. The effect of MDM2 on VLDL metabolism is p53-independent. Collectively, these findings suggest that MDM2 acts as a negative regulator of hepatic ApoB levels and TG-VLDL secretion in MAFLD. Inhibition of the MDM2-ApoB interaction may represent a potential therapeutic approach for MAFLD treatment.

PM20D1 is a circulating biomarker closely associated with obesity, insulin resistance and metabolic syndrome.

OBJECTIVE: Peptidase M20 domain containing 1 (PM20D1), a secreted enzyme catalysing condensation of fatty acids and amino acids into the bioactive lipids N-acyl amino acids (NAAA), induces uncoupling protein 1 (UCP1)-independent adaptive thermogenesis in brown/beige adipocytes in mice. This study aimed to explore the associations of the circulating levels of PM20D1 and major NAAA with obesity-related metabolic complications in humans. DESIGN AND METHODS: Serum concentrations of PM20D1 and NAAA (C18:1-Leu and C18:1-Phe) in 256 Chinese subjects, including 78 lean and 178 overweight/obese individuals with or without diabetes, were measured with immunoassays and liquid chromatography-mass spectrometry, respectively. The impact of sulfonylurea and rosiglitazone on their circulating levels was examined in 62 patients with type 2 diabetes. RESULTS: Serum PM20D1 level was significantly elevated in overweight/obese individuals and was closely associated with circulating levels of C18:1-Leu and C18:1-Phe. Furthermore, serum PM20D1, C18:1-Leu and C18:1-Phe concentrations correlated positively with several parameters of adiposity as well as fasting and 2 h postprandial glucose, HbA1c, fasting insulin and HOMA-IR independent of BMI and age. Moreover, a significant elevation in PM20D1, C18:1-Leu and C18:1-Phe concentrations corresponding with increases in the number of components of the metabolic syndrome (MetS) was observed. Treatment with sulfonylurea significantly decreased circulating PM20D1, C18:1-Leu and C18:1-Phe in patients with type 2 diabetes. CONCLUSIONS: Increased serum levels of PM20D1 and its catalytic products NAAA are closely associated with obesity-related glucose dysregulation, insulin resistance and MetS and can be potentially used as clinical biomarkers for diagnosing and monitoring these disorders.

Cardiac Autophagy Deficiency Attenuates ANP Production and Disrupts Myocardial-Adipose Cross Talk, Leading to Increased Fat Accumulation and Metabolic Dysfunction.

Increased myocardial autophagy has been established as an important stress-induced cardioprotective response. Three weeks after generating cardiomyocyte-specific autophagy deficiency, via inducible deletion of autophagy-related protein 7 (Atg7), we found that these mice (AKO) had increased body weight and fat mass without altered food intake. Glucose and insulin tolerance tests indicated reduced insulin sensitivity in AKO mice. Metabolic cage analysis showed reduced ambulatory activity and oxygen consumption with a trend of elevated respiratory exchange ratio in AKO mice. Direct analysis of metabolism in subcutaneous and visceral adipocytes showed increased glucose oxidation and reduced ATGL expression and HSL phosphorylation with no change in lipid synthesis or fatty acid oxidation. Importantly, we found AKO mice had reduced myocardial and circulating levels of atrial natriuretic peptide (ANP), an established mediator of myocardial-adipose cross talk. When normal ANP levels were restored to AKO mice with use of osmotic pump, the metabolic dysfunction evident in AKO mice was corrected. We conclude that cardiac autophagy deficiency alters myocardial-adipose cross talk via decreased ANP levels with adverse metabolic consequences.