Phosphorylation of hepatic farnesoid X receptor by FGF19 signaling-activated Src maintains cholesterol levels and protects from atherosclerosis.

The bile acid (BA) nuclear receptor, farnesoid X receptor (FXR/NR1H4), maintains metabolic homeostasis by transcriptional control of numerous genes, including an intestinal hormone, fibroblast growth factor-19 (FGF19; FGF15 in mice). Besides activation by BAs, the gene-regulatory function of FXR is also modulated by hormone or nutrient signaling-induced post-translational modifications. Recently, phosphorylation at Tyr-67 by the FGF15/19 signaling-activated nonreceptor tyrosine kinase Src was shown to be important for FXR function in BA homeostasis. Here, we examined the role of this FXR phosphorylation in cholesterol regulation. In both hepatic FXR-knockout and FXR-knockdown mice, reconstitution of FXR expression up-regulated cholesterol transport genes for its biliary excretion, including scavenger receptor class B member 1 (Scarb1) and ABC subfamily G member 8 (Abcg5/8), decreased hepatic and plasma cholesterol levels, and increased biliary and fecal cholesterol levels. Of note, these sterol-lowering effects were blunted by substitution of Phe for Tyr-67 in FXR. Moreover, consistent with Src's role in phosphorylating FXR, Src knockdown impaired cholesterol regulation in mice. In hypercholesterolemic apolipoprotein E-deficient mice, expression of FXR, but not Y67F-FXR, ameliorated atherosclerosis, whereas Src down-regulation exacerbated it. Feeding or treatment with an FXR agonist induced Abcg5/8 and Scarb1 expression in WT, but not FGF15-knockout, mice. Furthermore, FGF19 treatment increased occupancy of FXR at Abcg5/8 and Scarb1, expression of these genes, and cholesterol efflux from hepatocytes. These FGF19-mediated effects were blunted by the Y67F-FXR substitution or Src down-regulation or inhibition. We conclude that phosphorylation of hepatic FXR by FGF15/19-induced Src maintains cholesterol homeostasis and protects against atherosclerosis.

MiR-455-5p Attenuates Cerebral Ischemic Reperfusion Injury by Targeting FLT3.

Cerebral ischemia-reperfusion (I/R) injury is a terrible disease which results in the dysfunction and structural damage of brain tissues. Growing evidence implies that miR-455-5p is implicated in the regulation of pathogenesis of several diseases. The aim of this study is to reveal the role of miR-455-5p in cerebral I/R injury and the regulatory mechanism. We established a vitro model by inducing SH-SY5Y and PC-12 cells with oxygen-glucose deprivation and reoxygenation. The experimental cerebral I/R rat model was established by middle cerebral artery occlusion operation. The findings indicated that miR-455-5p expression was downregulated in oxygen-glucose deprivation and reoxygenation induced cells and I/R rat model. In addition, miR-455-5p upregulation inhibited SH-SY5Y cell apoptosis and cerebral damage, whereas miR-455-5p silencing promoted SH-SY5Y cell apoptosis and cerebral damage. Mechanistically, luciferase reporter assay corroborated that miR-455-5p could bind with feline mcDonough sarcoma-like tyrosine kinase 3 (FLT3) mRNA. However, the role of FLT3 in cerebral I/R injury was rarely investigated. Real-time polymerase chain reaction revealed that FTL3 expression was negatively regulated by miR-455-5p. FTL3 upregulation reversed the inhibitory effects of miR-455-5p upregulation on PC-12 and SH-SY5Y cell apoptosis. Therefore, our study verified that miR-455-5p improved cerebral I/R injury by targeting FLT3, which suggests a potential new target for the prevention of cerebral I/R injury.

Brd4 modulates the innate immune response through Mnk2-eIF4E pathway-dependent translational control of IκBα.

Bromodomain-containing factor Brd4 has emerged as an important transcriptional regulator of NF-κB-dependent inflammatory gene expression. However, the in vivo physiological function of Brd4 in the inflammatory response remains poorly defined. We now demonstrate that mice deficient for Brd4 in myeloid-lineage cells are resistant to LPS-induced sepsis but are more susceptible to bacterial infection. Gene-expression microarray analysis of bone marrow-derived macrophages (BMDMs) reveals that deletion of Brd4 decreases the expression of a significant amount of LPS-induced inflammatory genes while reversing the expression of a small subset of LPS-suppressed genes, including MAP kinase-interacting serine/threonine-protein kinase 2 (Mknk2). Brd4-deficient BMDMs display enhanced Mnk2 expression and the corresponding eukaryotic translation initiation factor 4E (eIF4E) activation after LPS stimulation, leading to an increased translation of IκBα mRNA in polysomes. The enhanced newly synthesized IκBα reduced the binding of NF-κB to the promoters of inflammatory genes, resulting in reduced inflammatory gene expression and cytokine production. By modulating the translation of IκBα via the Mnk2-eIF4E pathway, Brd4 provides an additional layer of control for NF-κB-dependent inflammatory gene expression and inflammatory response.

Selective killing of Helicobacter pylori with pH-responsive helix-coil conformation transitionable antimicrobial polypeptides.

Current clinical treatment of Helicobacter pylori infection, the main etiological factor in the development of gastritis, gastric ulcers, and gastric carcinoma, requires a combination of at least two antibiotics and one proton pump inhibitor. However, such triple therapy suffers from progressively decreased therapeutic efficacy due to the drug resistance and undesired killing of the commensal bacteria due to poor selectivity. Here, we report the development of antimicrobial polypeptide-based monotherapy, which can specifically kill H. pylori under acidic pH in the stomach while inducing minimal toxicity to commensal bacteria under physiological pH. Specifically, we designed a class of pH-sensitive, helix-coil conformation transitionable antimicrobial polypeptides (HCT-AMPs) (PGA)m-r-(PHLG-MHH)n, bearing randomly distributed negatively charged glutamic acid and positively charged poly(γ-6-N-(methyldihexylammonium)hexyl-l-glutamate) (PHLG-MHH) residues. The HCT-AMPs showed unappreciable toxicity at physiological pH when they adopted random coiled conformation. Under acidic condition in the stomach, they transformed to the helical structure and exhibited potent antibacterial activity against H. pylori, including clinically isolated drug-resistant strains. After oral gavage, the HCT-AMPs afforded comparable H. pylori killing efficacy to the triple-therapy approach while inducing minimal toxicity against normal tissues and commensal bacteria, in comparison with the remarkable killing of commensal bacteria by 65% and 86% in the ileal contents and feces, respectively, following triple therapy. This strategy renders an effective approach to specifically target and kill H. pylori in the stomach while not harming the commensal bacteria/normal tissues.

BET Inhibition Attenuates Helicobacter pylori-Induced Inflammatory Response by Suppressing Inflammatory Gene Transcription and Enhancer Activation.

Helicobacter pylori infection causes chronic gastritis and peptic ulceration. H. pylori-initiated chronic gastritis is characterized by enhanced expression of many NF-κB-regulated inflammatory cytokines. Brd4 has emerged as an important NF-κB regulator and regulates the expression of many NF-κB-dependent inflammatory genes. In this study, we demonstrated that Brd4 was not only actively involved in H. pylori-induced inflammatory gene mRNA transcription but also H. pylori-induced inflammatory gene enhancer RNA (eRNA) synthesis. Suppression of H. pylori-induced eRNA synthesis impaired H. pylori-induced mRNA synthesis. Furthermore, H. pylori stimulated NF-κB-dependent recruitment of Brd4 to the promoters and enhancers of inflammatory genes to facilitate the RNA polymerase II-mediated eRNA and mRNA synthesis. Inhibition of Brd4 by JQ1 attenuated H. pylori-induced eRNA and mRNA synthesis for a subset of NF-κB-dependent inflammatory genes. JQ1 also inhibited H. pylori-induced interaction between Brd4 and RelA and the recruitment of Brd4 and RNA polymerase II to the promoters and enhancers of inflammatory genes. Finally, we demonstrated that JQ1 suppressed inflammatory gene expression, inflammation, and cell proliferation in H. pylori-infected mice. These studies highlight the importance of Brd4 in H. pylori-induced inflammatory gene expression and suggest that Brd4 could be a potential therapeutic target for the treatment of H. pylori-triggered inflammatory diseases and cancer.

Lidamycin inhibits tumor initiating cells of hepatocellular carcinoma Huh7 through GSK3ß/ß-catenin pathway.

Recently, tumor initiating cells are considered as the central role of tumorigenicity in hepatocellular carcinoma. Enediyne anticancer antibiotic lidamycin with great potential antitumor activity is currently evaluated in Phase II clinical trials. In this study, we evaluated the effect of lidamycin on tumor initiating cells of hepatocellular carcinoma Huh7 and identified the potential mechanism. Flow cytometry analysis and sorting assay, surface marker assay, sphere formation assay, and aldefluor assay were used to evaluate the effect of lidamycin on Huh7 tumor initiating cells in vitro. To investigate the potential mechanism, the activity of GSK3ß/ß-catenin pathway was detected by Western blot and T cell factors transcriptional activity assay. Subcutaneous tumor model in nude mice was used to observe in vivo effect of lidamycin on Huh7 cells. Lidamycin decreased the proportion of EpCAM+ cells and the expression of EpCAM protein. Lidamycin inhibited sphere formation of sorted EpCAM+ cells in 7 d, and of parental cells in three serial passages. The population of aldehyde dehydrogenase-positive cells was reduced by lidamycin. In addition, lidamycin restrained tumor volume and incidence in vivo. Lidamycin activated GSK3ß, and degraded the activity of ß-catenin. Consequently, transcriptional activity of ß-catenin/T cell factors was decreased. In brief, these results suggest that lidamycin suppressed Huh7 tumor initiating cells via GSK3ß/ß-catenin pathway. These findings reveal the potential mechanism of lidamycin on tumor initiating cells and the benefit for further clinical evaluation.

Effect of glycosylation modification on structure and properties of soy protein isolate: A review.

Soybean protein isolate (SPI) is a highly functional protein source used in various food applications, such as emulsion, gelatin, and food packaging. However, its commercial application may be limited due to its poor mechanical properties, barrier properties, and high water sensitivity. Studies have shown that modifying SPI through glycosylation can enhance its functional properties and biological activities, resulting in better application performance. This paper reviews the recent studies on glycosylation modification of SPI, including its quantification method, structural improvements, and enhancement of its functional properties, such as solubility, gelation, emulsifying, and foaming. The review also discusses how glycosylation affects the bioactivity of SPI, such as its antioxidant and antibacterial activity. This review aims to provide a reference for further research on glycosylation modification and lay a foundation for applying SPI in various fields.

A Review of Modified Gelatin: Physicochemical Properties, Modification Methods, and Applications in the Food Field.

Gelatin is a significant multifunctional biopolymer that is widely utilized as a component in food, pharmaceuticals, and cosmetics. Numerous functional qualities are displayed by gelatin, such as its exceptional film-forming ability, gelling qualities, foaming and emulsifying qualities, biocompatibility and biodegradable qualities. Due to its unique structural, physicochemical, and biochemical characteristics, which enhance nutritional content and health benefits as well as the stability, consistency, and elasticity of food products, gelatin is utilized extensively in the food business. Additionally, gelatin has demonstrated excellent performance in encapsulating, delivering, and releasing active ingredients. Gelatin's various modifications, such as chemical, enzymatic, and physical processes, were analyzed to assess their impact on gelatin structures and characteristics. Hopefully, gelatin will be more widely used in various applications after modification using suitable methods.

Hammerhead-type FXR agonists induce an eRNA FincoR that ameliorates nonalcoholic steatohepatitis in mice.

The nuclear receptor, Farnesoid X Receptor (FXR/NR1H4), is increasingly recognized as a promising drug target for metabolic diseases, including nonalcoholic steatohepatitis (NASH). Protein coding genes regulated by FXR are well known, but whether FXR also acts through regulation of long non-coding RNAs (lncRNAs), which vastly outnumber protein-coding genes, remains unknown. Utilizing RNA-seq and GRO-seq analyses in mouse liver, we found that FXR activation affects the expression of many RNA transcripts from chromatin regions bearing enhancer features. Among these we discovered a previously unannotated liver-enriched enhancer-derived lncRNA (eRNA), termed FincoR. We show that FincoR is specifically induced by the hammerhead-type FXR agonists, including GW4064 and tropifexor. CRISPR/Cas9-mediated liver-specific knockdown of FincoR in dietary NASH mice reduced the beneficial effects of tropifexor, an FXR agonist currently in clinical trials for NASH and primary biliary cholangitis (PBC), indicating that that amelioration of liver fibrosis and inflammation in NASH treatment by tropifexor is mediated in part by FincoR. Overall, our findings highlight that pharmacological activation of FXR by hammerhead-type agonists induces a novel eRNA, FincoR, contributing to the amelioration of NASH in mice. FincoR may represent a new drug target for addressing metabolic disorders, including NASH.

Hammerhead-type FXR agonists induce an enhancer RNA Fincor that ameliorates nonalcoholic steatohepatitis in mice.

The nuclear receptor, farnesoid X receptor (FXR/NR1H4), is increasingly recognized as a promising drug target for metabolic diseases, including nonalcoholic steatohepatitis (NASH). Protein-coding genes regulated by FXR are well known, but whether FXR also acts through regulation of long non-coding RNAs (lncRNAs), which vastly outnumber protein-coding genes, remains unknown. Utilizing RNA-seq and global run-on sequencing (GRO-seq) analyses in mouse liver, we found that FXR activation affects the expression of many RNA transcripts from chromatin regions bearing enhancer features. Among these we discovered a previously unannotated liver-enriched enhancer-derived lncRNA (eRNA), termed FXR-induced non-coding RNA (Fincor). We show that Fincor is specifically induced by the hammerhead-type FXR agonists, including GW4064 and tropifexor. CRISPR/Cas9-mediated liver-specific knockdown of Fincor in dietary NASH mice reduced the beneficial effects of tropifexor, an FXR agonist currently in clinical trials for NASH and primary biliary cholangitis (PBC), indicating that amelioration of liver fibrosis and inflammation in NASH treatment by tropifexor is mediated in part by Fincor. Overall, our findings highlight that pharmacological activation of FXR by hammerhead-type agonists induces a novel eRNA, Fincor, contributing to the amelioration of NASH in mice. Fincor may represent a new drug target for addressing metabolic disorders, including NASH.

Non-alcoholic steatohepatitis, also known as NASH, is a severe condition whereby fat deposits around the liver lead to inflammation, swelling, scarring and lasting damage to the organ. Despite being one of the leading causes of liver-related deaths worldwide, the disease has no approved treatment. A protein known as Farnesoid X receptor (or FXR) is increasingly being recognized as a promising drug target for non-alcoholic steatohepatitis. Once activated, FXR helps to regulate the activity of DNA regions which are coding for proteins important for liver health. However, less is known about how FXR may act on non-coding regions, the DNA sequences that do not generate proteins but can be transcribed into RNA molecules with important biological roles. In response, Chen et al. investigated whether FXR activation of non-coding RNAs could be linked to the clinical benefits of hammerhead FXR agonists, a type of synthetic compounds that activates this receptor. To do so, genetic analyses of mouse livers were performed to identify non-coding RNAs generated when FXR was activated by the agonist. These experiments revealed that agonist-activated FXR induced a range of non-coding RNAs transcribed from DNA sequences known as enhancers, which help to regulate gene expression. In particular, hammerhead FXR agonists led to the production of a liver-specific enhancer RNA called Fincor. Additional experiments using tropifexor, a hammerhead FXR agonist currently into clinical trials, showed that this investigational new drug had reduced benefits in a mouse model of non-alcoholic steatohepatitis with low Fincor levels. This suggested that this enhancer RNA may play a key role in mediating the clinical benefits of hammerhead FXR agonists, encouraging further research into its role and therapeutic value.

Rapid Measurement of Antioxidant Properties of Dendrobium officinale Using Near-Infrared Spectroscopy and Chemometrics.

Dendrobium officinale (D. officinale), often used as a dual-use plant with herbal medicine and food applications, has attracted considerable attention for health-benefiting components and wide economic value. The antioxidant ability of D. officinale is of great significance to ensure its health care value and safeguard consumers' interests. However, the common analytical methods for evaluating the antioxidant ability of D. officinale are time-consuming, laborious, and costly. In this study, near-infrared (NIR) spectroscopy and chemometrics were employed to establish a rapid and accurate method for the determination of 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) scavenging capacity, 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging capacity, and ferric reducing antioxidant power (FRAP) in D. officinale. The quantitative models were developed based on the partial least squares (PLS) algorithm. Two wavelength selection methods, namely the genetic algorithm (GA) and competitive adaptive reweighted sampling (CARS) method, were used for model optimization. The CARS-PLS models exhibited superior predictive performance compared to other PLS models. The root mean square errors of cross-validation (RMSECVs) for ABTS, FRAP, and DPPH were 0.44%, 2.64 µmol/L, and 2.06%, respectively. The results demonstrated the potential application of NIR spectroscopy combined with the CARS-PLS model for the rapid prediction of antioxidant activity in D. officinale. This method can serve as an alternative to conventional analytical methods for efficiently quantifying the antioxidant properties in D. officinale.

Efficacy and safety of antibiotics for treatment of leptospirosis: a systematic review and network meta-analysis.

BACKGROUND: Leptospirosis, an important zoonotic bacterial disease, commonly affects resource-poor populations and results in significant morbidity and mortality worldwide. The value of antibiotics in leptospirosis remains unclear, as evidenced by the conflicting opinions published. METHODS: We conducted a search in the PubMed, Web of Science, and Cochrane Library databases for studies. These studies included clinical trials and retrospective studies that evaluated the efficacy or safety of antibiotics for leptospirosis treatment. The primary outcomes assessed were defervescence time, mortality rate, and hospital stays. Subgroup analyses were performed based on whether there were cases involving children and whether there were cases of severe jaundice. Safety was defined as the prevalence of adverse events associated with the use of antibiotics. p scores were utilized to rank the efficacy of the antibiotics. RESULTS: There are included 9 randomized controlled trials (RCTs), 1 control trial (CT), and 3 retrospective studies (RS) involving 920 patients and 8 antibiotics. Six antibiotics resulted in significantly shorter defervescence times compared to the control, namely cefotaxime (MD, - 1.88; 95% CI = - 2.60 to - 1.15), azithromycin (MD, - 1.74; 95% CI = - 2.52 to - 0.95), doxycycline (MD, - 1.53; 95% CI = - 2.05 to - 1.00), ceftriaxone (MD, - 1.22; 95% CI = - 1.89 to - 0.55), penicillin (MD, - 1.22; 95% CI = - 1.80 to - 0.64), and penicillin or ampicillin (MD, - 0.08; 95% CI = - 1.01 to - 0.59). The antibiotics were not effective in reducing the mortality and hospital stays. Common adverse reactions to antibiotics included Jarisch-Herxheimer reaction, rash, headache, and digestive reactions (nausea, vomiting, diarrhea, abdominal pain, and others). CONCLUSIONS: Findings recommend that leptospirosis patients be treated with antibiotics, which significantly reduced the leptospirosis defervescence time. Cephalosporins, doxycycline, and penicillin are suggested, and azithromycin may be a suitable alternative for drug-resistant cases. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42022354938.

Helicobacter pylori activates NF-κB by inducing Ubc13-mediated ubiquitination of lysine 158 of TAK1.

The Helicobacter pylori virulence factor CagA targets a variety of host proteins to alter different cellular responses, including the induction of pro-inflammatory cytokines. We have previously shown that CagA-facilitated lysine 63-linked ubiquitination of TAK1 is essential for the H. pylori-induced NF-κB activation and the expression of proinflammatory cytokines. However, the molecular mechanism for TAK1 ubiquitination and activation in H. pylori-mediated NF-κB activation remains elusive. Here, we identify lysine 158 of TAK1 as the key residue undergoing lysine 63-linked ubiquitination in response to H. pylori infection. Mutation of lysine 158 to arginine prevents the ubiquitination of TAK1 and impairs H. pylori-induced TAK1 and NF-κB activation. Moreover, we demonstrate that E2 ubiquitin conjugating enzyme Ubc13 is involved in H. pylori-mediated TAK1 ubiquitination. Suppressing the activity of Ubc13 by a dominant-negative mutant or siRNA abolishes CagA-facilitated and H. pylori-induced TAK1 and NF-κB activation. These findings further underscore the importance of lysine 63-linked ubiquitination of TAK1 in H. pylori-induced NF-κB activation and NF-κB-mediated inflammatory response.

S632A3, a new glutarimide antibiotic, suppresses lipopolysaccharide-induced pro-inflammatory responses via inhibiting the activation of glycogen synthase kinase 3ß.

Inflammatory mediators including inducible nitric oxide (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α) and Interleukin-6 (IL-6) contribute to the course of a variety of inflammatory diseases. S632A3 is a new member of the glutarimide antibiotics isolated from a cultured broth of Streptomyces hygroscopicus S632 with a potent NF-κB inhibitory activity. In the present study, we investigated the anti-inflammatory effects and the underlying molecular mechanism of S632A3 on lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. S632A3 concentration-dependently inhibited LPS-induced NO and prostaglandin E(2) (PGE(2)) production through the suppression of iNOS and COX-2 at gene transcription levels. In addition, S632A3 suppressed NF-κB-dependent inflammatory responses by inhibiting the activation of glycogen synthase kinase 3ß (GSK-3ß), while the activation of IκB kinase (IKK) complex was unaffected. S632A3 suppressed NF-κB activity by differentially affecting the CREB (cAMP response element-binding protein) and NF-κB p65 interacting with the coactivator CBP (CREB binding protein). S632A3 also inhibited GSK-3ß-elicited iNOS and COX-2 expression. Moreover, S632A3 was shown to inhibit the activation of ASK1 (Apoptosis-signal regulating kinase 1) and p38 mitogen-activated protein kinase, therefore attenuated the LPS-induced NF-κB activity in macrophages. Furthermore, S632A3 significantly reduced the pro-inflammatory cytokines TNF-α and IL-6 production while increased the anti-inflammatory cytokine IL-10 production in LPS-stimulated RAW264.7 cells. Our study thus provides a molecular mechanism by which S632A3 inhibited LPS-induced pro-inflammatory response in macrophages through interfering with the activation of GSK-3ß and ASK1-p38 signaling.

The dark side of NRF2 in arsenic carcinogenesis.

Arsenic is an environmental toxicant that significantly enhances the risk of developing disease, including several cancers. While the epidemiological evidence supporting increased cancer risk due to chronic arsenic exposure is strong, therapies tailored to treat exposed populations are lacking. This can be accredited in large part to the chronic nature and pleiotropic pathological effects associated with prolonged arsenic exposure. Despite this fact, several putative mediators of arsenic promotion of cancer have been identified. Among these, the critical transcription factor NRF2 has been shown to be a key mediator of arsenic's pro-carcinogenic effects. Importantly, the dependence of arsenic-transformed cancer cells on NRF2 upregulation exposes a targetable liability that could be utilized to treat arsenic-promoted cancers. In this chapter, we briefly introduce the "light" vs "dark" side of the NRF2 pathway. We then give a brief overview of arsenic metabolism, and discuss the epidemiological and experimental evidence that support arsenic promotion of different cancers, with a specific emphasis on mechanisms mediated by chronic, non-canonical activation of NRF2 (i.e., the "dark" side). Finally, we briefly highlight how the non-canonical NRF2 pathway plays a role in other arsenic-promoted diseases, as well as research directions that warrant further investigation.

Decreased autophagosome biogenesis, reduced NRF2, and enhanced ferroptotic cell death are underlying molecular mechanisms of non-alcoholic fatty liver disease.

BACKGROUND AND AIMS: Caloric excess and sedentary lifestyles have led to an epidemic of obesity, metabolic syndrome, and non-alcoholic fatty liver disease (NAFLD). The objective of this study was to investigate the mechanisms underlying high fat diet (HFD)-induced NAFLD, and to explore NRF2 activation as a strategy to alleviate NAFLD. APPROACH AND RESULTS: Herein, we demonstrated that high fat diet (HFD) induced lipid peroxidation and ferroptosis, both of which could be alleviated by NRF2 upregulation. Mechanistically, HFD suppressed autophagosome biogenesis through AMPK- and AKT-mediated mTOR activation and decreased ATG7, resulting in KEAP1 stabilization and decreased NRF2 levels in mouse liver. Furthermore, ATG7 is required for HFD-induced NRF2 downregulation, as ATG7 deletion in Cre-inducible ATG7 knockout mice decreased NRF2 levels and enhanced ferroptosis, which was not further exacerbated by HFD. This finding was recapitulated in mouse hepatocytes, which showed a similar phenotype upon treatment with saturated fatty acids (SFAs) but not monounsaturated fatty acids (MUFAs). Finally, NRF2 activation blocked fatty acid (FA)-mediated NRF2 downregulation, lipid peroxidation, and ferroptosis. Importantly, the HFD-induced alterations were also observed in human fatty liver tissue samples. CONCLUSIONS: HFD-mediated autophagy inhibition, NRF2 suppression, and ferroptosis promotion are important molecular mechanisms of obesity-driven metabolic diseases. NRF2 activation counteracts HFD-mediated NRF2 suppression and ferroptotic cell death. In addition, SFA vs. MUFA regulation of NRF2 may underlie their harmful vs. beneficial effects. Our study reveals NRF2 as a key player in the development and progression of fatty liver disease and that NRF2 activation could serve as a potential therapeutic strategy.

NRF2 controls iron homeostasis and ferroptosis through HERC2 and VAMP8.

Enhancing the intracellular labile iron pool (LIP) represents a powerful, yet untapped strategy for driving ferroptotic death of cancer cells. Here, we show that NRF2 maintains iron homeostasis by controlling HERC2 (E3 ubiquitin ligase for NCOA4 and FBXL5) and VAMP8 (mediates autophagosome-lysosome fusion). NFE2L2/NRF2 knockout cells have low HERC2 expression, leading to a simultaneous increase in ferritin and NCOA4 and recruitment of apoferritin into the autophagosome. NFE2L2/NRF2 knockout cells also have low VAMP8 expression, which leads to ferritinophagy blockage. Therefore, deletion of NFE2L2/NRF2 results in apoferritin accumulation in the autophagosome, an elevated LIP, and enhanced sensitivity to ferroptosis. Concordantly, NRF2 levels correlate with HERC2 and VAMP8 in human ovarian cancer tissues, as well as ferroptosis resistance in a panel of ovarian cancer cell lines. Last, the feasibility of inhibiting NRF2 to increase the LIP and kill cancer cells via ferroptosis was demonstrated in preclinical models, signifying the impact of NRF2 inhibition in cancer treatment.

NRF2 and Diabetes: The Good, the Bad, and the Complex.

Despite decades of scientific effort, diabetes continues to represent an incredibly complex and difficult disease to treat. This is due in large part to the multifactorial nature of disease onset and progression and the multiple organ systems affected. An increasing body of scientific evidence indicates that a key mediator of diabetes progression is NRF2, a critical transcription factor that regulates redox, protein, and metabolic homeostasis. Importantly, while experimental studies have confirmed the critical nature of proper NRF2 function in preventing the onset of diabetic outcomes, we have only just begun to scratch the surface of understanding the mechanisms by which NRF2 modulates diabetes progression, particularly across different causative contexts. One reason for this is the contradictory nature of the current literature, which can often be accredited to model discrepancies, as well as whether NRF2 is activated in an acute or chronic manner. Furthermore, despite therapeutic promise, there are no current NRF2 activators in clinical trials for the treatment of patients with diabetes. In this review, we briefly introduce the transcriptional programs regulated by NRF2 as well as how NRF2 itself is regulated. We also review the current literature regarding NRF2 modulation of diabetic phenotypes across the different diabetes subtypes, including a brief discussion of contradictory results, as well as what is needed to progress the NRF2 diabetes field forward.

Brd4 modulates diet-induced obesity via PPARγ-dependent Gdf3 expression in adipose tissue macrophages.

Macrophage-mediated inflammatory response has been implicated in the pathogenesis of obesity and insulin resistance. Brd4 has emerged as a key regulator in the innate immune response. However, the role of Brd4 in obesity-associated inflammation and insulin resistance remains uncharacterized. Here, we demonstrated that myeloid lineage-specific Brd4 knockout (Brd4-CKO) mice were protected from high-fat diet-induced (HFD-induced) obesity with less fat accumulation, higher energy expenditure, and increased lipolysis in adipose tissue. Brd4-CKO mice fed a HFD also displayed reduced local and systemic inflammation with improved insulin sensitivity. RNA-Seq of adipose tissue macrophages (ATMs) from HFD-fed WT and Brd4-CKO mice revealed that expression of antilipolytic factor Gdf3 was significantly decreased in ATMs of Brd4-CKO mice. We also found that Brd4 bound to the promoter and enhancers of Gdf3 to facilitate PPARγ-dependent Gdf3 expression in macrophages. Furthermore, Brd4-mediated expression of Gdf3 acted as a paracrine signal targeting adipocytes to suppress the expression of lipases and the associated lipolysis in cultured cells and mice. Controlling the expression of Gdf3 in ATMs could be one of the mechanisms by which Brd4 modulates lipid metabolism and diet-induced obesity. This study suggests that Brd4 could be a potential therapeutic target for obesity and insulin resistance.

BRD4 inhibition and FXR activation, individually beneficial in cholestasis, are antagonistic in combination.

Activation of farnesoid X receptor (FXR) by obeticholic acid (OCA) reduces hepatic inflammation and fibrosis in patients with primary biliary cholangitis (PBC), a life-threatening cholestatic liver failure. Inhibition of bromodomain-containing protein 4 (BRD4) also has antiinflammatory, antifibrotic effects in mice. We determined the role of BRD4 in FXR function in bile acid (BA) regulation and examined whether the known beneficial effects of OCA are enhanced by inhibiting BRD4 in cholestatic mice. Liver-specific downregulation of BRD4 disrupted BA homeostasis in mice, and FXR-mediated regulation of BA-related genes, including small heterodimer partner and cholesterol 7 alpha-hydroxylase, was BRD4 dependent. In cholestatic mice, JQ1 or OCA treatment ameliorated hepatotoxicity, inflammation, and fibrosis, but surprisingly, was antagonistic in combination. Mechanistically, OCA increased binding of FXR, and the corepressor silencing mediator of retinoid and thyroid hormone receptor (SMRT) decreased NF-κB binding at inflammatory genes and repressed the genes in a BRD4-dependent manner. In patients with PBC, hepatic expression of FXR and BRD4 was significantly reduced. In conclusion, BRD4 is a potentially novel cofactor of FXR for maintaining BA homeostasis and hepatoprotection. Although BRD4 promotes hepatic inflammation and fibrosis in cholestasis, paradoxically, BRD4 is required for the antiinflammatory, antifibrotic actions of OCA-activated FXR. Cotreatment with OCA and JQ1, individually beneficial, may be antagonistic in treatment of liver disease patients with inflammation and fibrosis complications.