Neutral ceramidase regulates breast cancer progression by metabolic programming of TREM2-associated macrophages.

The tumor microenvironment is reprogrammed by cancer cells and participates in all stages of tumor progression. Neutral ceramidase is a key regulator of ceramide, the central intermediate in sphingolipid metabolism. The contribution of neutral ceramidase to the reprogramming of the tumor microenvironment is not well understood. Here, we find that deletion of neutral ceramidase in multiple breast cancer models in female mice accelerates tumor growth. Our result show that Ly6C+CD39+ tumor-infiltrating CD8 T cells are enriched in the tumor microenvironment and display an exhausted phenotype. Deletion of myeloid neutral ceramidase in vivo and in vitro induces exhaustion in tumor-infiltrating Ly6C+CD39+CD8+ T cells. Mechanistically, myeloid neutral ceramidase is required for the generation of lipid droplets and for the induction of lipolysis, which generate fatty acids for fatty-acid oxidation and orchestrate macrophage metabolism. Metabolite ceramide leads to reprogramming of macrophages toward immune suppressive TREM2+ tumor associated macrophages, which promote CD8 T cells exhaustion.

Hepatic Transcriptome and Its Regulation Following Soluble Epoxide Hydrolase Inhibition in Alcohol-Associated Liver Disease.

Alcohol-associated liver disease (ALD) is a serious public health problem with limited pharmacologic options. The goal of the current study was to investigate the efficacy of pharmacologic inhibition of soluble epoxide hydrolase (sEH), an enzyme involved in lipid metabolism, in experimental ALD, and to examine the underlying mechanisms. C57BL/6J male mice were subjected to acute-on-chronic ethanol (EtOH) feeding with or without the sEH inhibitor 4-[[trans-4-[[[[4-trifluoromethoxy phenyl]amino]carbonyl]-amino]cyclohexyl]oxy]-benzoic acid (TUCB). Liver injury was assessed by multiple end points. Liver epoxy fatty acids and dihydroxy fatty acids were measured by targeted metabolomics. Whole-liver RNA sequencing was performed, and free modified RNA bases were measured by mass spectrometry. EtOH-induced liver injury was ameliorated by TUCB treatment as evidenced by reduced plasma alanine aminotransferase levels and was associated with attenuated alcohol-induced endoplasmic reticulum stress, reduced neutrophil infiltration, and increased numbers of hepatic M2 macrophages. TUCB altered liver epoxy and dihydroxy fatty acids and led to a unique hepatic transcriptional profile characterized by decreased expression of genes involved in apoptosis, inflammation, fibrosis, and carcinogenesis. Several modified RNA bases were robustly changed by TUCB, including N6-methyladenosine and 2-methylthio-N6-threonylcarbamoyladenosine. These findings show the beneficial effects of sEH inhibition by TUCB in experimental EtOH-induced liver injury, warranting further mechanistic studies to explore the underlying mechanisms, and highlighting the translational potential of sEH as a drug target for this disease.

Alcohol-mediated susceptibility to lung fibrosis is associated with group 2 innate lymphoid cells in mice.

Chronic alcohol ingestion promotes acute lung injury and impairs immune function. However, the mechanisms involved are incompletely understood. Here, we show that alcohol feeding enhances bleomycin-induced lung fibrosis and inflammation via the regulation of type 2 innate immune responses, especially by group 2 innate lymphoid cells (ILC2s). Neuroimmune interactions have emerged as critical modulators of lung inflammation. We found alcohol consumption induced the accumulation of ILC2 and reduced the production of the neuropeptide calcitonin gene-related peptide (CGRP), primarily released from sensory nerves and pulmonary neuroendocrine cells (PNECs). CGRP potently suppressed alcohol-driven type 2 cytokine signals in vivo. Vagal ganglia TRPV1+ afferents mediated immunosuppression occurs through the release of CGRP. Inactivation of the TRPV1 receptor enhanced bleomycin-induced fibrosis. In addition, mice lacking the CGRP receptor had the increased lung inflammation and fibrosis and type 2 cytokine production as well as exaggerated responses to alcohol feeding. Together, these data indicate that alcohol consumption regulates the interaction of CGRP and ILC2, which is a critical contributor of lung inflammation and fibrosis.

Probiotic-derived nanoparticles inhibit ALD through intestinal miR194 suppression and subsequent FXR activation.

BACKGROUND AND AIMS: Intestinal farnesoid X receptor (FXR) plays a critical role in alcohol-associated liver disease (ALD). We aimed to investigate whether alcohol-induced dysbiosis increased intestinal microRNA194 (miR194) that suppressed Fxr transcription and whether Lactobacillus rhamnosus GG-derived exosome-like nanoparticles (LDNPs) protected against ALD through regulation of intestinal miR194-FXR signaling in mice. APPROACH AND RESULTS: Binge-on-chronic alcohol exposure mouse model was utilized. In addition to the decreased ligand-mediated FXR activation, alcohol feeding repressed intestinal Fxr transcription and increased miR194 expression. This transcriptional suppression of Fxr by miR194 was confirmed in intestinal epithelial Caco-2 cells and mouse enteriods. The alcohol feeding-reduced intestinal FXR activation was further demonstrated by the reduced FXR reporter activity in fecal samples and by the decreased fibroblast growth factor 15 (Fgf15) messenger RNA (mRNA) in intestine and protein levels in the serum, which caused an increased hepatic bile acid synthesis and lipogeneses. We further demonstrated that alcohol feeding increased-miR194 expression was mediated by taurine-upregulated gene 1 (Tug1) through gut microbiota regulation of taurine metabolism. Importantly, 3-day oral administration of LDNPs increased bile salt hydrolase (BSH)-harboring bacteria that decreased conjugated bile acids and increased gut taurine concentration, which upregulated Tug1, leading to a suppression of intestinal miR194 expression and recovery of FXR activation. Activated FXR upregulated FGF15 signaling and subsequently reduced hepatic bile acid synthesis and lipogenesis and attenuated ALD. These protective effects of LDNPs were eliminated in intestinal FxrΔIEC and Fgf15-/- mice. We further showed that miR194 was upregulated, whereas BSH activity and taurine levels were decreased in fecal samples of patients with ALD. CONCLUSIONS: Our results demonstrated that gut microbiota-mediated miR194 regulation contributes to ALD pathogenesis and to the protective effects of LDNPs through modulating intestinal FXR signaling.

Enteric VIP-producing neurons maintain gut microbiota homeostasis through regulating epithelium fucosylation.

Interactions between the enteric nervous system (ENS) and intestinal epithelium are thought to play a vital role in intestinal homeostasis. How the ENS monitors the frontier with commensal and pathogenic microbes while maintaining epithelial function remains unclear. Here, by combining subdiaphragmatic vagotomy with transcriptomics, chemogenetic strategy, and coculture of enteric neuron-intestinal organoid, we show that enteric neurons expressing VIP shape the α1,2-fucosylation of intestinal epithelial cells (IECs). Mechanistically, neuropeptide VIP activates fut2 expression via the Erk1/2-c-Fos pathway through the VIPR1 receptor on IECs. We further demonstrate that perturbation of enteric neurons leads to gut dysbiosis through α1,2-fucosylation in the steady state and results in increased susceptibility to alcohol-associated liver disease (ALD). This was attributed to an imbalance between beneficial Bifidobacterium and opportunistic pathogenic Enterococcus faecalis in ALD. In addition, Bifidobacterium α1,2-fucosidase may promote Bifidobacterium adhesion to the mucosal surface, which restricts Enterococcus faecalis overgrowth and prevents ALD progression.

Nrf2 transcriptional upregulation of IDH2 to tune mitochondrial dynamics and rescue angiogenic function of diabetic EPCs.

Endothelial progenitor cells (EPCs) are reduced in number and impaired in function in diabetic patients. Whether and how Nrf2 regulates the function of diabetic EPCs remains unclear. In this study, we found that the expression of Nrf2 and its downstream genes were decreased in EPCs from both diabetic patients and db/db mice. Survival ability and angiogenic function of EPCs from diabetic patients and db/db mice also were impaired. Gain- and loss-of-function studies, respectively, showed that knockdown of Nrf2 increased apoptosis and impaired tube formation in EPCs from healthy donors and wild-type mice, while Nrf2 overexpression decreased apoptosis and rescued tube formation in EPCs from diabetic patients and db/db mice. Additionally, proangiogenic function of Nrf2-manipulated mouse EPCs was validated in db/db mice with hind limb ischemia. Mechanistic studies demonstrated that diabetes induced mitochondrial fragmentation and dysfunction of EPCs by dysregulating the abundance of proteins controlling mitochondrial dynamics; upregulating Nrf2 expression attenuated diabetes-induced mitochondrial fragmentation and dysfunction and rectified the abundance of proteins controlling mitochondrial dynamics. Further RNA-sequencing analysis demonstrated that Nrf2 specifically upregulated the transcription of isocitrate dehydrogenase 2 (IDH2), a key enzyme regulating tricarboxylic acid cycle and mitochondrial function. Overexpression of IDH2 rectified Nrf2 knockdown- or diabetes-induced mitochondrial fragmentation and EPC dysfunction. In a therapeutic approach, supplementation of an Nrf2 activator sulforaphane enhanced angiogenesis and blood perfusion recovery in db/db mice with hind limb ischemia. Collectively, these findings indicate that Nrf2 is a potential therapeutic target for improving diabetic EPC function. Thus, elevating Nrf2 expression enhances EPC resistance to diabetes-induced oxidative damage and improves therapeutic efficacy of EPCs in treating diabetic limb ischemia likely via transcriptional upregulating IDH2 expression and improving mitochondrial function of diabetic EPCs.

Alcohol-driven metabolic reprogramming promotes development of RORγt-deficient thymic lymphoma.

RORγt is a master regulator of Th17 cells. Despite evidence linking RORγt deficiency/inhibition with metastatic thymic T cell lymphomas, the role of RORγt in lymphoma metabolism is unknown. Chronic alcohol consumption plays a causal role in many human cancers. The risk of T cell lymphoma remains unclear in humans with alcohol use disorders (AUD) after chronic RORγt inhibition. Here we demonstrated that alcohol consumption accelerates RORγt deficiency-induced lymphomagenesis. Loss of RORγt signaling in the thymus promotes aerobic glycolysis and glutaminolysis and increases allocation of glutamine carbon into lipids. Importantly, alcohol consumption results in a shift from aerobic glycolysis to glutaminolysis. Both RORγt deficiency- and alcohol-induced metabolic alterations are mediated by c-Myc, as silencing of c-Myc decreases the effects of alcohol consumption and RORγt deficiency on glutaminolysis, biosynthesis, and tumor growth in vivo. The ethanol-mediated c-Myc activation coupled with increased glutaminolysis underscore the critical role of RORγt-Myc signaling and translation in lymphoma.

Neutral ceramidase-dependent regulation of macrophage metabolism directs intestinal immune homeostasis and controls enteric infection.

It is not clear how the complex interactions between diet and intestinal immune cells protect the gut from infection. Neutral ceramidase (NcDase) plays a critical role in digesting dietary sphingolipids. We find that NcDase is an essential factor that controls intestinal immune cell dynamics. Mice lacking NcDase have reduced cluster of differentiation (CD) 8αß+ T cells and interferon (IFN)-γ+ T cells and increased macrophages in the intestine and fail to clear bacteria after Citrobacter rodentium infection. Mechanistically, cellular NcDase or extracellular vesicle (EV)-related NcDase generates sphingosine, which promotes macrophage-driven Th1 immunity. Loss of NcDase influences sphingosine-controlled glycolytic metabolism in macrophages, which regulates the bactericidal activity of macrophages. Importantly, administration of dietary sphingomyelin and genetic deletion or pharmacological inhibition of SphK1 can protect against C. rodentium infection. Our findings demonstrate that sphingosine profoundly alters macrophage glycolytic metabolism, leading to intestinal macrophage activation and T cell polarization, which prevent pathogen colonization of the gut.

Exosome-like nanoparticles from Mulberry bark prevent DSS-induced colitis via the AhR/COPS8 pathway.

Bark protects the tree against environmental insults. Here, we analyzed whether this defensive strategy could be utilized to broadly enhance protection against colitis. As a proof of concept, we show that exosome-like nanoparticles (MBELNs) derived from edible mulberry bark confer protection against colitis in a mouse model by promoting heat shock protein family A (Hsp70) member 8 (HSPA8)-mediated activation of the AhR signaling pathway. Activation of this pathway in intestinal epithelial cells leads to the induction of COP9 Constitutive Photomorphogenic Homolog Subunit 8 (COPS8). Utilizing a gut epithelium-specific knockout of COPS8, we demonstrate that COPS8 acts downstream of the AhR pathway and is required for the protective effect of MBELNs by inducing an array of anti-microbial peptides. Our results indicate that MBELNs represent an undescribed mode of inter-kingdom communication in the mammalian intestine through an AhR-COPS8-mediated anti-inflammatory pathway. These data suggest that inflammatory pathways in a microbiota-enriched intestinal environment are regulated by COPS8 and that edible plant-derived ELNs may hold the potential as new agents for the prevention and treatment of gut-related inflammatory disease.

Deficiency of Cathelicidin Attenuates High-Fat Diet Plus Alcohol-Induced Liver Injury through FGF21/Adiponectin Regulation.

Alcohol consumption and obesity are known risk factors of steatohepatitis. Here, we report that the deficiency of CRAMP (cathelicidin-related antimicrobial peptide-gene name: Camp) is protective against a high-fat diet (HFD) plus acute alcohol (HFDE)-induced liver injury. HFDE markedly induced liver injury and steatosis in WT mice, which were attenuated in Camp-/- mice. Neutrophil infiltration was lessened in the liver of Camp-/- mice. HFDE feeding dramatically increased epididymal white adipose tissue (eWAT) mass and induced adipocyte hypertrophy in WT mice, whereas these effects were attenuated by the deletion of Camp. Furthermore, Camp-/- mice had significantly increased eWAT lipolysis, evidenced by up-regulated expression of lipolytic enzymes, adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL). The depletion of Camp also increased uncoupling protein 1 (UCP1)-dependent thermogenesis in the brown adipose tissue (BAT) of mice. HFDE fed Camp-/- mice had elevated protein levels of fibroblast growth factor 21 (FGF21) in the eWAT, with an increased adiponectin production, which had been shown to alleviate hepatic fat deposition and inflammation. Collectively, we have demonstrated that Camp-/- mice are protected against HFD plus alcohol-induced liver injury and steatosis through FGF21/adiponectin regulation. Targeting CRAMP could be an effective approach for prevention/treatment of high-fat diet plus alcohol consumption-induced steatohepatitis.

Fat-1 Transgenic Mice With Augmented n3-Polyunsaturated Fatty Acids Are Protected From Liver Injury Caused by Acute-On-Chronic Ethanol Administration.

Alcohol-associated liver disease (ALD) is the leading cause of liver disease worldwide, and alcohol-associated hepatitis (AH), a severe form of ALD, is a major contributor to the mortality and morbidity due to ALD. Many factors modulate susceptibility to ALD development and progression, including nutritional factors such as dietary fatty acids. Recent work from our group and others showed that modulation of dietary or endogenous levels of n6-and n3-polyunsaturated fatty acids (PUFAs) can exacerbate or attenuate experimental ALD, respectively. In the current study, we interrogated the effects of endogenous n3-PUFA enrichment in a mouse model which recapitulates features of early human AH using transgenic fat-1 mice which endogenously convert n6-PUFAs to n3-PUFAs. Male wild type (WT) and fat-1 littermates were provided an ethanol (EtOH, 5% v/v)-containing liquid diet for 10 days, then administered a binge of EtOH (5 g/kg) by oral gavage on the 11th day, 9 h prior to sacrifice. In WT mice, EtOH treatment resulted in liver injury as determined by significantly elevated plasma ALT levels, whereas in fat-1 mice, EtOH caused no increase in this biomarker. Compared to their pair-fed controls, a significant EtOH-mediated increase in liver neutrophil infiltration was observed also in WT, but not fat-1 mice. The hepatic expression of several cytokines and chemokines, including Pai-1, was significantly lower in fat-1 vs WT EtOH-challenged mice. Cultured bone marrow-derived macrophages isolated from fat-1 mice expressed less Pai-1 and Cxcl2 (a canonical neutrophil chemoattractant) mRNA compared to WT when stimulated with lipopolysaccharide. Further, we observed decreased pro-inflammatory M1 liver tissue-resident macrophages (Kupffer cells, KCs), as well as increased liver T regulatory cells in fat-1 vs WT EtOH-fed mice. Taken together, our data demonstrated protective effects of endogenous n3-PUFA enrichment on liver injury caused by an acute-on-chronic EtOH exposure, a paradigm which recapitulates human AH, suggesting that n3-PUFAs may be a viable nutritional adjuvant therapy for this disease.

Exosome-Like Nanoparticles From Lactobacillus rhamnosusGG Protect Against Alcohol-Associated Liver Disease Through Intestinal Aryl Hydrocarbon Receptor in Mice.

Alcohol-associated liver disease (ALD) is a major cause of mortality. Gut barrier dysfunction-induced bacterial translocation and endotoxin release contribute to the pathogenesis of ALD. Probiotic Lactobacillus rhamnosus GG (LGG) is known to be beneficial on experimental ALD by reinforcing the intestinal barrier function. In this study, we aim to investigate whether the protective effects of LGG on intestinal barrier function is mediated by exosome-like nanoparticles (ELNPs) released by LGG. Intestinal epithelial cells and macrophages were treated with LGG-derived ELNPs (LDNPs) isolated from LGG culture. LDNPs increased tight junction protein expression in epithelial cells and protected from the lipopolysaccharide-induced inflammatory response in macrophages. Three-day oral application of LDNPs protected the intestine from alcohol-induced barrier dysfunction and the liver from steatosis and injury in an animal model of ALD. Co-administration of an aryl hydrocarbon receptor (AhR) inhibitor abolished the protective effects of LDNPs, indicating that the effects are mediated, at least in part, by intestinal AhR signaling. We further demonstrated that LDNP administration increased intestinal interleukin-22-Reg3 and nuclear factor erythroid 2-related factor 2 (Nrf2)-tight junction signaling pathways, leading to the inhibition of bacterial translocation and endotoxin release in ALD mice. This protective effect was associated with LDNP enrichment of bacterial tryptophan metabolites that are AhR agonists. Conclusions: Our results suggest that the beneficial effects of LGG and their supernatant in ALD are likely mediated by bacterial AhR ligand-enriched LDNPs that increase Reg3 and Nrf2 expression, leading to the improved barrier function. These findings provide a strategy for the treatment of ALD and other gut barrier dysfunction-associated diseases.

Analysis of sex differences in dietary copper-fructose interaction-induced alterations of gut microbial activity in relation to hepatic steatosis.

BACKGROUND: Inadequate copper intake and increased fructose consumption represent two important nutritional problems in the USA. Dietary copper-fructose interactions alter gut microbial activity and contribute to the development of nonalcoholic fatty liver disease (NAFLD). The aim of this study is to determine whether dietary copper-fructose interactions alter gut microbial activity in a sex-differential manner and whether sex differences in gut microbial activity are associated with sex differences in hepatic steatosis. METHODS: Male and female weanling Sprague-Dawley (SD) rats were fed ad libitum with an AIN-93G purified rodent diet with defined copper content for 8 weeks. The copper content is 6 mg/kg and 1.5 mg/kg in adequate copper diet (CuA) and marginal copper diet (CuM), respectively. Animals had free access to either deionized water or deionized water containing 10% fructose (F) (w/v) as the only drink during the experiment. Body weight, calorie intake, plasma alanine aminotransferase, aspartate aminotransferase, and liver histology as well as liver triglyceride were evaluated. Fecal microbial contents were analyzed by 16S ribosomal RNA (16S rRNA) sequencing. Fecal and cecal short-chain fatty acids (SCFAs) were determined by gas chromatography-mass spectrometry (GC-MS). RESULTS: Male and female rats exhibit similar trends of changes in the body weight gain and calorie intake in response to dietary copper and fructose, with a generally higher level in male rats. Several female rats in the CuAF group developed mild steatosis, while no obvious steatosis was observed in male rats fed with CuAF or CuMF diets. Fecal 16S rRNA sequencing analysis revealed distinct alterations of the gut microbiome in male and female rats. Linear discriminant analysis (LDA) effect size (LEfSe) identified sex-specific abundant taxa in different groups. Further, total SCFAs, as well as, butyrate were decreased in a more pronounced manner in female CuMF rats than in male rats. Of note, the decreased SCFAs are concomitant with the reduced SCFA producers, but not correlated to hepatic steatosis. CONCLUSIONS: Our data demonstrated sex differences in the alterations of gut microbial abundance, activities, and hepatic steatosis in response to dietary copper-fructose interaction in rats. The correlation between sex differences in metabolic phenotypes and alterations of gut microbial activities remains elusive.

Inhibition of Sphingosine-1-Phosphate-Induced Th17 Cells Ameliorates Alcohol-Associated Steatohepatitis in Mice.

BACKGROUND AND AIMS: Chronic alcohol consumption is accompanied by intestinal inflammation. However, little is known about how alterations to the intestinal immune system and sphingolipids contribute to the pathogenesis of alcohol-associated liver disease (ALD). APPROACH AND RESULTS: We used wild-type mice, retinoid-related orphan receptor gamma t (RORγt)-deficient mice, sphingosine kinase-deficient mice, and local gut anti-inflammatory, 5-aminosalicyclic acid-treated mice in a chronic-binge ethanol feeding model. Targeted lipidomics assessed the sphingolipids in gut and liver samples. Gut immune cell populations, the amounts of sphingolipids, and the level of liver injury were examined. Alcohol intake induces a pro-inflammatory shift in immune cell populations in the gut, including an increase in Th17 cells. Using RORγt-deficient mice, we found that Th17 cells are required for alcohol-associated gut inflammation and the development of ALD. Treatment with 5-aminosalicyclic acid decreases alcohol-induced liver injury and reverses gut inflammation by the suppression of CD4+ /RORγt+ /interleukin-17A+ cells. Increased Th17 cells were due to up-regulation of sphingosine kinase 1 activity and RORγt activation. We found that S1P/S1PR1 signaling is required for the development of Th17 cell-mediated ALD. Importantly, in vivo intervention blocking of S1P/S1PR1 signaling markedly attenuated alcohol-induced liver inflammation, steatosis, and damage. CONCLUSIONS: Gut inflammation is a functional alteration of immune cells in ALD. Reducing gut Th17 cells leads to reduced liver damage. S1P signaling was crucial in the pathogenesis of ALD in a Th17 cell-dependent manner. Furthermore, our findings suggest that compounds that reduce gut inflammation locally may represent a unique targeted approach in the treatment of ALD.

Human Beta Defensin 2 Ameliorated Alcohol-Associated Liver Disease in Mice.

Alcohol-associated liver disease (ALD) is a prevalent liver disorder and significant global healthcare burden with limited effective therapeutic options. The gut-liver axis is a critical factor contributing to susceptibility to liver injury due to alcohol consumption. In the current study, we tested whether human beta defensin-2 (hBD-2), a small anti-microbial peptide, attenuates experimental chronic ALD. Male C57Bl/6J mice were fed an ethanol (EtOH)-containing diet for 6 weeks with daily administration of hBD-2 (1.2 mg/kg) by oral gavage during the final week. Two independent cohorts of mice with distinct baseline gut microbiota were used. Oral hBD-2 administration attenuated liver injury in both cohorts as determined by decreased plasma ALT activity. Notably, the degree of hBD-2-mediated reduction of EtOH-associated liver steatosis, hepatocellular death, and inflammation was different between cohorts, suggesting microbiota-specific mechanisms underlying the beneficial effects of hBD-2. Indeed, we observed differential mechanisms of hBD-2 between cohorts, which included an induction of hepatic and small intestinal IL-17A and IL-22, as well as an increase in T regulatory cell abundance in the gut and mesenteric lymph nodes. Lastly, hBD-2 modulated the gut microbiota composition in EtOH-fed mice in both cohorts, with significant decreases in multiple genera including Barnesiella, Parabacteroides, Akkermansia, and Alistipes, as well as altered abundance of several bacteria within the family Ruminococcaceae. Collectively, our results demonstrated a protective effect of hBD-2 in experimental ALD associated with immunomodulation and microbiota alteration. These data suggest that while the beneficial effects of hBD-2 on liver injury are uniform, the specific mechanisms of action are associated with baseline microbiota.

Neutral Ceramidase Mediates Nonalcoholic Steatohepatitis by Regulating Monounsaturated Fatty Acids and Gut IgA+ B Cells.

BACKGROUND AND AIMS: Nonalcoholic steatohepatitis (NASH) is associated with obesity and an increased risk for liver cirrhosis and cancer. Neutral ceramidase (NcDase), which is highly expressed in the intestinal brush border of the small intestine, plays a critical role in digesting dietary sphingolipids (ceramide) to regulate the balance of sphingosine and free fatty acids. It remains unresolved whether obesity-associated alteration of NcDase contributes to the manifestation of NASH. Here, we revealed that NcDase deficiency in murine models of NASH prevents hepatic inflammation and fibrosis but not steatosis. APPROACH AND RESULTS: NcDase-/- mice display reduced stearoyl-CoA desaturase (SCD) 1 expression with a compositional decrease of monounsaturated fatty acids (MUFAs) under the different dietary conditions. We further found that NcDase is a functional regulator of intestinal B cells and influences the abundance and quality of the secretory IgA response toward commensal bacteria. Analysis of composition of the gut microbiota found that Clostridiales colonization was increased in NcDase-/- mice. The colonization of germ-free mice with gut microbiota from NcDase-/- mice resulted in a greater decrease in the expression of SCD1 and the level of MUFAs in the liver relative to gut microbiota from wild-type littermates, which are associated with the alternation of IgA-bound bacteria, including increase of Ruminococcaceae and reduction of Desulfovibrio. Mechanistically, NcDase is a crucial link that controls the expression of SCD1 and MUFA-mediated activation of the Wnt/ß-catenin. Very importantly, our experiments further demonstrated that Wnt3a stimulation can enhance the activity of NcDase in hepatocytes. CONCLUSIONS: Thus, the NcDase-SCD1-Wnt feedback loop promotes the diet-induced steatohepatitis and fibrosis through the regulation of intestinal IgA+ immune cells.

Cathelicidin-related antimicrobial peptide alleviates alcoholic liver disease through inhibiting inflammasome activation.

Alcoholic liver disease (ALD) is associated with gut dysbiosis and hepatic inflammasome activation. While it is known that antimicrobial peptides (AMPs) play a critical role in the regulation of bacterial homeostasis in ALD, the functional role of AMPs in the alcohol-induced inflammasome activation is unclear. The aim of this study was to determine the effects of cathelicidin-related antimicrobial peptide (CRAMP) on inflammasome activation in ALD. CRAMP knockout (Camp-/-) and wild-type (WT) mice were subjected to binge-on-chronic alcohol feeding and synthetic CRAMP peptide was administered. Serum/plasma and hepatic tissue samples from human subjects with alcohol use disorder and/or alcoholic hepatitis were analyzed. CRAMP deficiency exacerbated ALD with enhanced inflammasome activation as shown by elevated serum interleukin (IL)-1ß levels. Although Camp-/- mice had comparable serum endotoxin levels compared to WT mice after alcohol feeding, hepatic lipopolysaccharide (LPS) binding protein (LBP) and cluster of differentiation (CD) 14 were increased. Serum levels of uric acid (UA), a Signal 2 molecule in inflammasome activation, were positively correlated with serum levels of IL-1ß in alcohol use disorder patients with ALD and were increased in Camp-/- mice fed alcohol. In vitro studies showed that CRAMP peptide inhibited LPS binding to macrophages and inflammasome activation stimulated by a combination of LPS and UA. Synthetic CRAMP peptide administration decreased serum UA and IL-1ß concentrations and rescued the liver from alcohol-induced damage in both WT and Camp-/- mice. In summary, CRAMP exhibited a protective role against binge-on-chronic alcohol-induced liver damage via regulation of inflammasome activation by decreasing LPS binding and UA production. CRAMP administration may represent a novel strategy for treating ALD. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Erratum: B7-H3 promotes the cell cycle-mediated chemoresistance of colorectal cancer cells by regulating CDC25A: Erratum.

[This corrects the article DOI: 10.7150/jca.37255.].

Endothelial Overexpression of Metallothionein Prevents Diabetes-Induced Impairment in Ischemia Angiogenesis Through Preservation of HIF-1α/SDF-1/VEGF Signaling in Endothelial Progenitor Cells.

Diabetes-induced oxidative stress is one of the major contributors to dysfunction of endothelial progenitor cells (EPCs) and impaired endothelial regeneration. Thus, we tested whether increasing antioxidant protein metallothionein (MT) in EPCs promotes angiogenesis in a hind limb ischemia (HLI) model in endothelial MT transgenic (JTMT) mice with high-fat diet- and streptozocin-induced diabetes. Compared with littermate wild-type (WT) diabetic mice, JTMT diabetic mice had improved blood flow recovery and angiogenesis after HLI. Similarly, transplantation of JTMT bone marrow-derived mononuclear cells (BM-MNCs) stimulated greater blood flow recovery in db/db mice with HLI than did WT BM-MNCs. The improved recovery was associated with augmented EPC mobilization and angiogenic function. Further, cultured EPCs from patients with diabetes exhibited decreased MT expression, increased cell apoptosis, and impaired tube formation, while cultured JTMT EPCs had enhanced cell survival, migration, and tube formation in hypoxic/hyperglycemic conditions compared with WT EPCs. Mechanistically, MT overexpression enhanced hypoxia-inducible factor 1α (HIF-1α), stromal cell-derived factor (SDF-1), and vascular endothelial growth factor (VEGF) expression and reduced oxidative stress in ischemic tissues. MT's pro-EPC effects were abrogated by siRNA knockdown of HIF-1α without affecting its antioxidant action. These results indicate that endothelial MT overexpression is sufficient to protect against diabetes-induced impairment of angiogenesis by promoting EPC function, most likely through upregulation of HIF-1α/SDF-1/VEGF signaling and reducing oxidative stress.