Amphiregulin Induces iNOS and COX-2 Expression through NF-κB and MAPK Signaling in Hepatic Inflammation.

Background: Inflammation is a major cause of hepatic tissue damage and accelerates the progression of nonalcoholic fatty liver disease (NAFLD). Amphiregulin (AREG), an epidermal growth factor receptor ligand, is associated with human liver cirrhosis and hepatocellular carcinoma. We aimed to investigate the effects of AREG on hepatic inflammation during NAFLD progression, in vivo and in vitro. Methods: AREG gene expression was measured in the liver of mice fed a methionine choline-deficient (MCD) diet for 2 weeks. We evaluated inflammatory mediators and signaling pathways in HepG2 cells after stimulation with AREG. Nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) were analyzed using an enzyme-linked immunosorbent assay and western blotting. Nuclear transcription factor kappa-B (NF-κB) and mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase, were analyzed using western blotting. Results: Proinflammatory cytokines (interleukin (IL)-6, IL-1ß, and IL-8) and immune cell recruitment (as indicated by L3T4, F4/80, and ly6G mRNA expression) increased, and expression of AREG increased in the liver of mice fed the MCD diet. AREG significantly increased the expression of IL-6 and IL-1ß and the production of NO, PGE2, and IL-8 in HepG2 cells. It also activated the protein expression of iNOS and COX-2. AREG-activated NF-κB and MAPKs signaling, and together with NF-κB and MAPKs inhibitors, AREG significantly reduced the protein expression of iNOS and COX-2. Conclusion: AREG plays a role in hepatic inflammation by increasing iNOS and COX-2 expression via NF-κB and MAPKs signaling.

Mitochondrial protease ClpP supplementation ameliorates diet-induced NASH in mice.

BACKGROUND & AIMS: Mitochondrial dysfunction is considered a pathogenic linker in the development of non-alcoholic steatohepatitis (NASH). Inappropriate mitochondrial protein-quality control, possibly induced by insufficiency of the mitochondrial matrix caseinolytic protease P (ClpP), can potentially cause mitochondrial dysfunction. Herein, we aimed to investigate hepatic ClpP levels in a diet-induced model of NASH and determine whether supplementation of ClpP can ameliorate diet-induced NASH. METHODS: NASH was induced by a high-fat/high-fructose (HF/HFr) diet in C57BL/6J mice. Stress/inflammatory signals were induced in mouse primary hepatocytes (MPHs) by treatment with palmitate/oleate (PA/OA). ClpP levels in hepatocytes were reduced using the RNAi-mediated gene knockdown technique but increased through the viral transduction of ClpP. ClpP activation was induced by administering a chemical activator of ClpP. RESULTS: Hepatic ClpP protein levels in C57BL/6J mice fed a HF/HFr diet were lower than the levels in those fed a normal chow diet. PA/OA treatment also decreased the ClpP protein levels in MPHs. Overexpression or activation of ClpP reversed PA/OA-induced mitochondrial dysfunction and stress/inflammatory signal activation in MPHs, whereas ClpP knockdown induced mitochondrial dysfunction and stress/inflammatory signals in these cells. On the other hand, ClpP overexpression or activation improved HF/HFr-induced NASH characteristics such as hepatic steatosis, inflammation, fibrosis, and injury in the C57BL/6J mice, whereas ClpP knockdown further augmented steatohepatitis in mice fed a HF/HFr diet. CONCLUSIONS: Reduced ClpP expression and subsequent mitochondrial dysfunction are key to the development of diet-induced NASH. ClpP supplementation through viral transduction or chemical activation represents a potential therapeutic strategy to prevent diet-induced NASH. LAY SUMMARY: Western diets, containing high fat and high fructose, often induce non-alcoholic steatohepatitis (NASH). Mitochondrial dysfunction is considered pathogenically linked to diet-induced NASH. We observed that the mitochondrial protease ClpP decreased in the livers of mice fed a western diet and supplementation of ClpP ameliorated western diet-induced NASH.

Hepatoprotective effects of gemigliptin and empagliflozin in a murine model of diet-induced non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) includes a broad spectrum of liver diseases characterized by steatosis, inflammation, and fibrosis. This study aimed to investigate the potential of dipeptidyl peptidase-4 inhibitors and sodium-glucose cotransporter 2 inhibitors in alleviating the progression of NAFLD. The NAFLD model was generated by feeding male C57BL/6J mice a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) for 7 weeks. After 2 weeks of CDAHFD feeding, the NAFLD model mice were assigned to four groups, namely (ⅰ) VEHICLE, (ⅱ) gemigliptin (GEMI), (ⅲ) empagliflozin (EMPA), and (ⅳ) GEMI + EMPA. For the next 5 weeks, mice received the vehicle or the drug based upon the group to which they belonged. Thereafter, the triglyceride concentration, extent of fibrosis, and the expression of genes encoding inflammatory cytokines, chemokines, and antioxidant enzymes were analyzed in the livers of mice. The NAFLD activity score and hepatic fibrosis grade were assessed via hematoxylin and eosin and Sirius Red staining of the liver tissue samples. All mice belonging to the GEMI, EMPA, and GEMI + EMPA groups showed improvements in the accumulation of liver triglycerides and the expression of inflammatory cytokines and chemokines. Additionally, the oxidative stress was reduced due to inhibition of the c-Jun N-terminal kinase pathway and upregulation of the antioxidant enzymes. Furthermore, in these three groups, the galectin-3 and interleukin 33-induced activity of tumor necrosis factor-α was inhibited, thereby preventing the progression of liver fibrosis. These findings suggest that the GEMI, EMPA, and GEMI + EMPA treatments ameliorate hepatic steatosis, inflammation, oxidative stress, and fibrosis in CDAHFD-induced NAFLD mouse models.

In Vivo Two-Photon Imaging Analysis of Dynamic Degradation of Hepatic Lipid Droplets in MS-275-Treated Mouse Liver.

The accumulation of hepatic lipid droplets (LDs) is a hallmark of non-alcoholic fatty liver disease (NAFLD). Appropriate degradation of hepatic LDs and oxidation of complete free fatty acids (FFAs) are important for preventing the development of NAFLD. Histone deacetylase (HDAC) is involved in the impaired lipid metabolism seen in high-fat diet (HFD)-induced obese mice. Here, we evaluated the effect of MS-275, an inhibitor of HDAC1/3, on the degradation of hepatic LDs and FFA oxidation in HFD-induced NAFLD mice. To assess the dynamic degradation of hepatic LDs and FFA oxidation in fatty livers of MS-275-treated HFD C57BL/6J mice, an intravital two-photon imaging system was used and biochemical analysis was performed. The MS-275 improved hepatic metabolic alterations in HFD-induced fatty liver by increasing the dynamic degradation of hepatic LDs and the interaction between LDs and lysozyme in the fatty liver. Numerous peri-droplet mitochondria, lipolysis, and lipophagy were observed in the MS-275-treated mouse fatty liver. Biochemical analysis revealed that the lipolysis and autophagy pathways were activated in MS-275 treated mouse liver. In addition, MS-275 reduced the de novo lipogenesis, but increased the mitochondrial oxidation and the expression levels of oxidation-related genes, such as PPARa, MCAD, CPT1b, and FGF21. Taken together, these results suggest that MS-275 stimulates the degradation of hepatic LDs and mitochondrial free fatty acid oxidation, thus protecting against HFD-induced NAFLD.

Visfatin exacerbates hepatic inflammation and fibrosis in a methionine-choline-deficient diet mouse model.

BACKGROUND AND AIM: Non-alcoholic fatty liver disease (NAFLD) ranges from simple steatosis to non-alcoholic steatohepatitis, which is characterized by hepatic inflammation that can progress to fibrosis, cirrhosis, and hepatocellular carcinoma. Visfatin, an adipocytokine, was reported to induce pro-inflammatory cytokines and can be associated with liver fibrosis. We investigated the role of visfatin on hepatic inflammation and fibrosis in a methionine-choline-deficient (MCD)-diet-induced steatohepatitis mouse model. METHODS: Eight-week-old male C57BL/6 J mice were randomly assigned into one of three groups: (1) saline-injected control diet group; (2) saline-injected MCD diet group; and (3) visfatin-injected MCD diet group (n = 8 per group). Mice were administered intravenous saline or 10 µg/kg of recombinant murine visfatin for 2 weeks. Histologic assessment of liver and biochemical and molecular measurements of endoplasmic reticulum (ER) stress, reactive oxidative stress (ROS), inflammation, and fibrosis were performed in livers from these animals. RESULTS: Visfatin injection aggravated hepatic steatosis and increased plasma alanine aminotransferase and aspartate aminotransferase concentrations. Visfatin increased inflammatory cell infiltration (as indicated by F4/80, CD68, ly6G, and CD3 mRNA expression) and expression of chemokines in the liver. Visfatin also increased the expression of pro-inflammatory cytokines (IL-1ß, TNF-α, and IL-6) and activated fibrosis markers (CTGF, TIMP1, collagen 1α2, collagen 3α2, αSMA, fibronectin, and vimentin) in liver. Livers of visfatin-injected mice showed upregulation of ER stress and ROS and activation of JNK signaling. CONCLUSIONS: These results suggest that visfatin aggravates hepatic inflammation together with induction of ER and oxidative stress and exacerbates fibrosis in an MCD-diet-fed mouse model of NAFLD.

AMP-Activated Protein Kinase Regulates Circadian Rhythm by Affecting CLOCK in Drosophila.

The circadian clock organizes the physiology and behavior of organisms to their daily environmental rhythms. The central circadian timekeeping mechanism in eukaryotic cells is the transcriptional-translational feedback loop (TTFL). In the Drosophila TTFL, the transcription factors CLOCK (CLK) and CYCLE (CYC) play crucial roles in activating expression of core clock genes and clock-controlled genes. Many signaling pathways converge on the CLK/CYC complex and regulate its activity to fine-tune the cellular oscillator to environmental time cues. We aimed to identify factors that regulate CLK by performing tandem affinity purification combined with mass spectrometry using Drosophila S2 cells that stably express HA/FLAG-tagged CLK and V5-tagged CYC. We identified SNF4Aγ, a homolog of mammalian AMP-activated protein kinase γ (AMPKγ), as a factor that copurified with HA/FLAG-tagged CLK. The AMPK holoenzyme composed of a catalytic subunit AMPKα and two regulatory subunits, AMPKß and AMPKγ, directly phosphorylated purified CLK in vitro Locomotor behavior analysis in Drosophila revealed that knockdown of each AMPK subunit in pacemaker neurons induced arrhythmicity and long periods. Knockdown of AMPKß reduced CLK levels in pacemaker neurons, and thereby reduced pre-mRNA and protein levels of CLK downstream core clock genes, such as period and vrille Finally, overexpression of CLK reversed the long-period phenotype that resulted from AMPKß knockdown. Thus, we conclude that AMPK, a central regulator of cellular energy metabolism, regulates the Drosophila circadian clock by stabilizing CLK and activating CLK/CYC-dependent transcription.SIGNIFICANCE STATEMENT Regulation of the circadian transcription factors CLK and CYC is fundamental to synchronize the core clock with environmental changes. Here, we show that the AMPKγ subunit of AMPK, a central regulator of cellular metabolism, copurifies with the CLK/CYC complex in Drosophila S2 cells. Furthermore, the AMPK holoenzyme directly phosphorylates CLK in vitro This study demonstrates that AMPK activity regulates the core clock in Drosophila by activating CLK, which enhances circadian transcription. In mammals, AMPK affects the core clock by downregulating circadian repressor proteins. It is intriguing to note that AMPK activity is required for core clock regulation through circadian transcription enhancement, whereas the target of AMPK action is different in Drosophila and mammals (positive vs negative element, respectively).

Iron overload by transferrin receptor protein 1 regulation plays an important role in palmitate-induced insulin resistance in human skeletal muscle cells.

Free fatty acid is considered to be one of the major pathogenic factors of inducing insulin resistance. The association between iron disturbances and insulin resistance has recently begun to receive a lot of attention. Although skeletal muscles are a major tissue for iron utilization and storage, the role of iron in palmitate (PA)-induced insulin resistance is unknown. We investigated the molecular mechanism underlying iron dysregulation in PA-induced insulin resistance. Interestingly, we found that PA simultaneously increased intracellular iron and induced insulin resistance. The iron chelator deferoxamine dramatically inhibited PA-induced insulin resistance, and iron donors impaired insulin sensitivity by activating JNK. PA up-regulated transferrin receptor 1 (tfR1), an iron uptake protein, which was modulated by iron-responsive element-binding proteins 2. Knockdown of tfR1 and iron-responsive element-binding proteins 2 prevented PA-induced iron uptake and insulin resistance. PA also translocated the tfR1 by stimulating calcium influx, but the calcium chelator, BAPTA-AM, dramatically reduced iron overload by inhibiting tfR1 translocation and ultimately increased insulin sensitivity. Iron overload may play a critical role in PA-induced insulin resistance. Blocking iron overload may thus be a useful strategy for preventing insulin resistance and diabetes.-Cui, R., Choi, S.-E., Kim, T. H., Lee, H. J., Lee, S. J., Kang, Y., Jeon, J. Y., Kim, H. J., Lee, K.-W. Iron overload by transferrin receptor protein 1 regulation plays an important role in palmitate-induced insulin resistance in human skeletal muscle cells.

CCL20 induced by visfatin in macrophages via the NF-κB and MKK3/6-p38 signaling pathways contributes to hepatic stellate cell activation.

Chemokines interact with hepatic resident cells during inflammation and fibrosis. CC chemokine ligand (CCL) 20 has been reported to be important in inflammation and fibrosis in the liver. We hypothesized that visfatin, an adipocytokine, could play a role in hepatic fibrosis via CCL20. We investigated the effect of visfatin on CCL20 in THP-1 human promonocytic cells and examined the molecular mechanisms involved. Following treatment of THP-1 cells with visfatin, CCL20 expression and secretion were assessed. We assessed the intracellular signaling molecules IKK/NF-κB, JAK2/STAT3, MAPKs, and MKK3/6 by western blotting. We treated THP-1 cells with visfatin and signaling inhibitors, and examined CCL20 mRNA and protein levels. To investigate the effect of visfatin-induced CCL20 expression in hepatic stellate cells (HSCs), LX-2 cells were co-cultured with the culture supernatant of THP-1 cells with or without anti-CCL20 neutralizing antibodies, and fibrosis markers were examined by RT-PCR and immunoblotting. In THP-1 cells, visfatin increased the CCL20 mRNA and protein levels. visfatin increased the activities of the NF-κB, p38, and MLK3/6 signaling pathways but not those of the JAK2/STAT3 and ERK pathways. Visfatin treatment together with an NF-κB, p38, or MLK3 inhibitor reduced the mRNA and protein levels of CCL20. The visfatin-induced CCL20 increased the expression of fibrosis markers and CCR6 in HSCs. Following neutralization of CCL20, the levels of fibrosis markers and CCR6 were decreased. Visfatin increases the expression of CCL20 via the NF-κB and MKK3/6-p38 signaling pathways in macrophages, and visfatin-induced CCL20 expression promotes the fibrosis markers in HSCs.

Transcriptional regulation of autophagy by an FXR-CREB axis.

Lysosomal degradation of cytoplasmic components by autophagy is essential for cellular survival and homeostasis under nutrient-deprived conditions. Acute regulation of autophagy by nutrient-sensing kinases is well defined, but longer-term transcriptional regulation is relatively unknown. Here we show that the fed-state sensing nuclear receptor farnesoid X receptor (FXR) and the fasting transcriptional activator cAMP response element-binding protein (CREB) coordinately regulate the hepatic autophagy gene network. Pharmacological activation of FXR repressed many autophagy genes and inhibited autophagy even in fasted mice, and feeding-mediated inhibition of macroautophagy was attenuated in FXR-knockout mice. From mouse liver chromatin immunoprecipitation and high-throughput sequencing data, FXR and CREB binding peaks were detected at 178 and 112 genes, respectively, out of 230 autophagy-related genes, and 78 genes showed shared binding, mostly in their promoter regions. CREB promoted autophagic degradation of lipids, or lipophagy, under nutrient-deprived conditions, and FXR inhibited this response. Mechanistically, CREB upregulated autophagy genes, including Atg7, Ulk1 and Tfeb, by recruiting the coactivator CRTC2. After feeding or pharmacological activation, FXR trans-repressed these genes by disrupting the functional CREB-CRTC2 complex. This study identifies the new FXR-CREB axis as a key physiological switch regulating autophagy, resulting in sustained nutrient regulation of autophagy during feeding/fasting cycles.

Aberrantly elevated microRNA-34a in obesity attenuates hepatic responses to FGF19 by targeting a membrane coreceptor ß-Klotho.

MicroRNA-34a (miR-34a) is the most highly elevated hepatic miR in obese mice and is also substantially elevated in patients who have steatosis, but its role in obesity and metabolic dysfunction remains unclear. After a meal, FGF19 is secreted from the ileum; binds to a hepatic membrane receptor complex, FGF19 receptor 4 and coreceptor ß-Klotho (ßKL); and mediates postprandial responses under physiological conditions, but hepatic responses to FGF19 signaling were shown to be impaired in patients with steatosis. Here, we show an unexpected functional link between aberrantly elevated miR-34a and impaired ßKL/FGF19 signaling in obesity. In vitro studies show that miR-34a down-regulates ßKL by binding to the 3' UTR of ßKL mRNA. Adenoviral-mediated overexpression of miR-34a in mice decreased hepatic ßKL levels, impaired FGF19-activated ERK and glycogen synthase kinase signaling, and altered expression of FGF19 metabolic target genes. Consistent with these results, ßKL levels were decreased and hepatic responses to FGF19 were severely impaired in dietary obese mice that have elevated miR-34a. Remarkably, in vivo antisense inhibition of miR-34a in obese mice partially restored ßKL levels and improved FGF19 target gene expression and metabolic outcomes, including decreased liver fat. Further, anti-miR-34a treatment in primary hepatocytes of obese mice restored FGF19-activated ERK and glycogen synthase kinase signaling in a ßKL-dependent manner. These results indicate that aberrantly elevated miR-34a in obesity attenuates hepatic FGF19 signaling by directly targeting ßKL. The miR-34a/ßKL/FGF19 axis may present unique therapeutic targets for FGF19-related human diseases, including metabolic disorders and cancer.

Bile acid signal-induced phosphorylation of small heterodimer partner by protein kinase Cζ is critical for epigenomic regulation of liver metabolic genes.

Bile acids (BAs) are recently recognized key signaling molecules that control integrative metabolism and energy expenditure. BAs activate multiple signaling pathways, including those of nuclear receptors, primarily farnesoid X receptor (FXR), membrane BA receptors, and FXR-induced FGF19 to regulate the fed-state metabolism. Small heterodimer partner (SHP) has been implicated as a key mediator of these BA signaling pathways by recruitment of chromatin modifying proteins, but the key question of how SHP transduces BA signaling into repressive histone modifications at liver metabolic genes remains unknown. Here we show that protein kinase Cζ (PKCζ) is activated by BA or FGF19 and phosphorylates SHP at Thr-55 and that Thr-55 phosphorylation is critical for the epigenomic coordinator functions of SHP. PKCζ is coimmunopreciptitated with SHP and both are recruited to SHP target genes after bile acid or FGF19 treatment. Activated phosphorylated PKCζ and phosphorylated SHP are predominantly located in the nucleus after FGF19 treatment. Phosphorylation at Thr-55 is required for subsequent methylation at Arg-57, a naturally occurring mutation site in metabolic syndrome patients. Thr-55 phosphorylation increases interaction of SHP with chromatin modifiers and their occupancy at selective BA-responsive genes. This molecular cascade leads to repressive modifications of histones at metabolic target genes, and consequently, decreased BA pools and hepatic triglyceride levels. Remarkably, mutation of Thr-55 attenuates these SHP-mediated epigenomic and metabolic effects. This study identifies PKCζ as a novel key upstream regulator of BA-regulated SHP function, revealing the role of Thr-55 phosphorylation in epigenomic regulation of liver metabolism.

HDAC11 Regulates Palmitate-induced NLRP3 Inflammasome Activation by Inducing YAP Expression in THP-1 Cells and PBMCs.

Histone deacetylase 11 (HDAC11) has been implicated in the pathogenesis of metabolic diseases characterized by chronic low-grade inflammation, such as obesity. However, the influence of HDAC11 on inflammation and the specific effect of HDAC11 on the palmitic acid (PA)-induced NLR family pyrin domain containing 3 (NLRP3) inflammasome activation are poorly understood. The effect of PA treatment on HDAC11 activity and the NLRP3 inflammasome was investigated in human peripheral blood mononuclear cells and THP-1 cells. The PA-induced responses of key markers of NLRP3 inflammasome activation, including NLRP3 gene expression, caspase-1 p10 activation, cleaved IL-1ß production, and extracellular IL-1ß release, were assessed as well. The role of HDAC11 was explored using a specific inhibitor of HDAC11 and by knockdown using small interfering (si)HDAC11 RNA. The relationship between HDAC11 and yes-associated protein (YAP) in the PA-induced NLRP3 inflammasome was investigated in THP-1 cells with HDAC11 or YAP knockdown. Following PA treatment, HDAC11 activity and protein levels increased significantly, concomitant with activation of the NLRP3 inflammasome. Notably, PA-induced the upregulation of NLRP3, caspase-1 p10 activation, the production of cleaved IL-1ß, and the release of IL-1ß into the extracellular space, all of which were attenuated by FT895 treatment and by HDAC11 knockdown. In THP-1 cells, PA induced the expression of YAP and its interaction with NLRP3, resulting in NLRP3 inflammasome activation, whereas both were inhibited by FT895 and siHDAC11 RNA. These findings demonstrate a pivotal role for HDAC11 in the PA-induced activation of the NLRP3 inflammasome. HDAC11 inhibition thus represents a promising therapeutic strategy for mitigating NLRP3 inflammasome-related inflammation in the context of obesity.

Protective effect of nicotinamide on high glucose/palmitate-induced glucolipotoxicity to INS-1 beta cells is attributed to its inhibitory activity to sirtuins.

This study was initiated to determine whether the protective effect of nicotinamide (NAM) on high glucose/palmitate (HG/PA)-induced INS-1 beta cell death was due to its role as an anti-oxidant, nicotinamide dinucleotide (NAD+) precursor, or inhibitor of NAD+-consuming enzymes such as poly (ADP-ribose) polymerase (PARP) or sirtuins. All anti-oxidants tested were not protective against HG/PA-induced INS-1 cell death. Direct supplementation of NAD+ or indirect supplementation through NAD+ salvage or de novo pathway did not protect the death. Knockdown of the NAD+ salvage pathway enzymes such as nicotinamide phosphoribosyl transferase (NAMPT) or nicotinamide mononucleotide adenyltransferase (NMNAT) did not augment death. On the other hand, pharmacological inhibition or knockdown of PARP did not affect death. However, sirtinol as an inhibitor of NAD-dependant deacetylase or knockdown of SIRT3 or SIRT4 significantly reduced the HG/PA-induced death. These data suggest that protective effect of NAM on beta cell glucolipotoxicity is attributed to its inhibitory activity on sirtuins.

CD38 Inhibition Protects Fructose-Induced Toxicity in Primary Hepatocytes.

A fructose-enriched diet is thought to contribute to hepatic injury in developing non-alcoholic steatohepatitis (NASH). However, the cellular mechanism of fructose-induced hepatic damage remains poorly understood. This study aimed to determine whether fructose induces cell death in primary hepatocytes, and if so, to establish the underlying cellular mechanisms. Our results revealed that treatment with high fructose concentrations for 48 h induced mitochondria-mediated apoptotic death in mouse primary hepatocytes (MPHs). Endoplasmic reticulum stress responses were involved in fructose-induced death as the levels of phosho-eIF2α, phospho-C-Jun-N-terminal kinase (JNK), and C/EBP homologous protein (CHOP) increased, and a chemical chaperone tauroursodeoxycholic acid (TUDCA) prevented cell death. The impaired oxidation metabolism of fatty acids was also possibly involved in the fructose-induced toxicity as treatment with an AMP-activated kinase (AMPK) activator and a PPAR-α agonist significantly protected against fructose-induced death, while carnitine palmitoyl transferase I inhibitor exacerbated the toxicity. However, uric acid-mediated toxicity was not involved in fructose-induced death as uric acid was not toxic to MPHs, and the inhibition of xanthine oxidase (a key enzyme in uric acid synthesis) did not affect cell death. On the other hand, treatment with inhibitors of the nicotinamide adenine dinucleotide (NAD)+-consuming enzyme CD38 or CD38 gene knockdown significantly protected against fructose-induced toxicity in MPHs, and fructose treatment increased CD38 levels. These data suggest that CD38 upregulation plays a role in hepatic injury in the fructose-enriched diet-mediated NASH. Thus, CD38 inhibition may be a promising therapeutic strategy to prevent fructose-enriched diet-mediated NASH.

Protective effect of empagliflozin against palmitate-induced lipotoxicity through AMPK in H9c2 cells.

Sodium-glucose cotransporter 2 (SGLT2) inhibitors have recently emerged as novel cardioprotective agents. However, their direct impact on cardiomyocyte injury is yet to be studied. In this work, we investigate the underlying molecular mechanisms of empagliflozin (EMPA), an SGLT2 inhibitor, in mitigating palmitate (PA)-induced cardiomyocyte injury in H9c2 cells. We found that EMPA significantly attenuated PA-induced impairments in insulin sensitivity, ER stress, inflammatory cytokine gene expression, and cellular apoptosis. Additionally, EMPA elevated AMP levels, activated the AMPK pathway, and increased carnitine palmitoyl transferase1 (CPT1) gene expression, which collectively enhanced fatty acid oxidation and reduced stress signals. This study reveals a novel mechanism of EMPA's protective effects against PA-induced cardiomyocyte injury, providing new therapeutic insights into EMPA as a cardioprotective agent.

Screening and surveillance for Barrett's esophagus: current issues and future directions.

PURPOSE OF REVIEW: Our article discusses the current understanding of screening and surveillance options for Barrett's esophagus and emerging concepts that have the potential to improve the effectiveness and cost-effectiveness of surveillance. RECENT FINDINGS: Although endoscopic surveillance of patients with Barrett's esophagus is commonly practiced in order to detect high-grade dysplasia and early esophageal adenocarcinoma (EAC), the reported incidence of EAC in Barrett's esophagus patients varies widely. Recent studies found the risk of progression from Barrett's esophagus to EAC to be significantly lower than previously reported, raising concerns regarding the limitations of current surveillance strategies. Advances in imaging techniques and their enhanced diagnostic accuracy may improve the value of endoscopic surveillance. Additionally, various efforts are ongoing to identify biomarkers that identify individuals at higher risk of cancer, possibly allowing for individual risk stratification. SUMMARY: These new data highlight some of the opportunities to revise and improve surveillance in patients with Barrett's esophagus. The incorporation of new advances such as imaging techniques and biomarkers has the potential to improve the effectiveness and cost-effectiveness of new surveillance regimens.

Empagliflozin Reduces the Progression of Hepatic Fibrosis in a Mouse Model and Inhibits the Activation of Hepatic Stellate Cells via the Hippo Signalling Pathway.

Hepatic fibrosis is the excessive production and deposition of the extracellular matrix, resulting in the activation of the fibrogenic phenotype of hepatic stellate cells (HSCs). The Hippo/Yes-associated protein (YAP) signalling pathway is a highly conserved kinase cascade that is critical in regulating cell proliferation, differentiation, and survival, and controls stellate cell activation. Empagliflozin, a sodium-glucose cotransporter type-2 inhibitor, is an antidiabetic drug that may prevent fibrotic progression by reducing hepatic steatosis and inflammation. However, little is known about its mechanism of action in liver fibrosis. In this study, we used male C57 BL/6 J mice fed a choline-deficient, l-amino acid-defined, high-fat diet (CDAHFD) as a model for hepatic fibrosis. For 5 weeks, the mice received either a vehicle or empagliflozin based on their assigned group. Empagliflozin attenuated CDAHFD-induced liver fibrosis. Thereafter, we identified the Hippo pathway, along with its effector, YAP, as a key pathway in the mouse liver. Hippo signalling is inactivated in the fibrotic liver, but empagliflozin treatment activated Hippo signalling and decreased YAP activity. In addition, empagliflozin downregulated the expression of pro-fibrogenic genes and activated Hippo signalling in HSCs. We identified a mechanism by which empagliflozin ameliorates liver fibrosis.

Reduction in the Migration Activity of Microglia Treated with Silica-Coated Magnetic Nanoparticles and their Recovery Using Citrate.

Nanoparticles have garnered significant interest in neurological research in recent years owing to their efficient penetration of the blood-brain barrier (BBB). However, significant concerns are associated with their harmful effects, including those related to the immune response mediated by microglia, the resident immune cells in the brain, which are exposed to nanoparticles. We analysed the cytotoxic effects of silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye [MNPs@SiO2(RITC)] in a BV2 microglial cell line using systems toxicological analysis. We performed the invasion assay and the exocytosis assay and transcriptomics, proteomics, metabolomics, and integrated triple-omics analysis, generating a single network using a machine learning algorithm. The results highlight alteration in the mechanisms of the nanotoxic effects of nanoparticles using integrated omics analysis.

Detection of differential proteomes associated with the development of type 2 diabetes in the Zucker rat model using the iTRAQ technique.

Type 2 diabetes (T2D) is closely associated with obesity, and it arises when pancreatic ß cells fail to achieve ß cell compensation. However, the mechanism linking obesity, insulin resistance, and ß cell failure in T2D is not fully understood. To explore this association, we carried out a differential proteomics study using the disease models of Zucker Fatty (ZF) and Zucker Diabetic Fatty (ZDF) rats as the rat models for obese/prediabetes and obese/diabetes, respectively. Differentially expressed islet proteins were identified among ZDF, ZF, and Zucker Lean (ZL, control rat) rats using three iTRAQ experiments, where three biological replicates and two technical replicates were examined to assess both the technical and biological reproducibilities. A total of 54 and 58 proteins were differentially expressed in ZDF versus ZL rats and in ZF versus ZL rats, respectively. Notably, the novel proteins involved in impaired insulin secretion (Scg2, Anxa2, and Rab10), mitochondrial dysfunction (Atp5b and Atp5l), extracellular matrix proteins (Lgal-1, Vim, and Fbn1), and microvascular ischemia (CPA1, CPA2, CPB, Cela2a, and Cela3b) were observed for the first time. With these novel proteins, our proteomics study could provide valuable clues for better understanding the underlying mechanisms associated with the dynamic transition of obesity to T2D.

Supplement of TCA cycle intermediates protects against high glucose/palmitate-induced INS-1 beta cell death.

The aim of this study is to investigate the effect of mitochondrial metabolism on high glucose/palmitate (HG/PA)-induced INS-1 beta cell death. Long-term treatment of INS-1 cells with HG/PA impaired energy-producing metabolism accompanying with depletion of TCA cycle intermediates. Whereas an inhibitor of carnitine palmitoyl transferase 1 augmented HG/PA-induced INS-1 cell death, stimulators of fatty acid oxidation protected the cells against the HG/PA-induced death. Furthermore, whereas mitochondrial pyruvate carboxylase inhibitor phenylacetic acid augmented HG/PA-induced INS-1 cell death, supplementation of TCA cycle metabolites including leucine/glutamine, methyl succinate/α-ketoisocaproic acid, dimethyl malate, and valeric acid or treatment with a glutamate dehydrogenase activator, aminobicyclo-heptane-2-carboxylic acid (BCH), significantly protected the cells against the HG/PA-induced death. In particular, the mitochondrial tricarboxylate carrier inhibitor, benzene tricarboxylate (BTA), also showed a strong protective effect on the HG/PA-induced INS-1 cell death. Knockdown of glutamate dehydrogenase or tricarboxylate carrier augmented or reduced the HG/PA-induced INS-1 cell death, respectively. Both BCH and BTA restored HG/PA-induced reduction of energy metabolism as well as depletion of TCA intermediates. These data suggest that depletion of the TCA cycle intermediate pool and impaired energy-producing metabolism may play a role in HG/PA-induced cytotoxicity to beta cells and thus, HG/PA-induced beta cell glucolipotoxicity can be protected by nutritional or pharmacological maneuver enhancing anaplerosis or reducing cataplerosis.