Hepatic FOXA3 overexpression prevents Western diet-induced obesity and MASH through TGR5.

Forkhead transcription factor 3 (FOXA3) has been shown to regulate metabolism and development. Hepatic FOXA3 is reduced in obesity and fatty liver disease. However, the role of hepatic FOXA3 in regulating obesity or steatohepatitis remains to be investigated. In this work, C57BL/6 mice were i.v. injected with AAV8-ALB-FOXA3 or the control virus. The mice were then fed a chow or Western diet for 16 weeks. The role of hepatic FOXA3 in energy metabolism and steatohepatitis was investigated. Plasma bile acid composition and the role of Takeda G protein-coupled receptor 5 (TGR5) in mediating the metabolic effects of FOXA3 were determined. Overexpression of hepatic FOXA3 reduced hepatic steatosis in chow-fed mice and attenuated Western diet-induced obesity and steatohepatitis. FOXA3 induced lipolysis and inhibited hepatic genes involved in bile acid uptake, resulting in elevated plasma bile acids. The beneficial effects of hepatic FOXA3 overexpression on Western diet-induced obesity and steatohepatitis were abolished in Tgr5-/- mice. Our data demonstrate that overexpression of hepatic FOXA3 prevents Western diet-induced obesity and steatohepatitis via activation of TGR5.

FABP5+ lipid-loaded macrophages process tumor-derived unsaturated fatty acid signal to suppress T-cell antitumor immunity.

BACKGROUND & AIMS: Tumour-associated macrophages (TAMs) contribute to hepatocellular carcinoma (HCC) progression. However, while the pro-tumour and immunosuppressive roles of lipid-loaded macrophages are well established, the mechanisms by which lipid metabolism enhances the tumour-promoting effects in TAMs remain unclear. METHODS: Single-cell RNA sequencing was performed on mouse and human HCC tumour samples to elucidate the landscape of HCC TAMs. Macrophages were stimulated with various long-chain unsaturated fatty acids (UFAs) to assess immunosuppressive molecules expression in vitro. Additionally, in vivo and in vitro studies were conducted using mice with macrophage-specific deficiencies in fatty acid-binding protein 5 (FABP5) or peroxisome proliferator-activated receptor (PPAR). RESULTS: Single-cell RNA sequencing identified a subpopulation of FABP5+ lipid-loaded TAMs characterized by enhanced immune checkpoint blocker ligands and immunosuppressive molecules in an oncogene-mutant HCC mouse model and human HCC tumours. Mechanistically, long-chain UFAs released by tumour cells activate PPARvia FABP5, resulting in TAM immunosuppressive properties. FABP5 deficiency in macrophages decreases immunosuppressive molecules expression, enhances T-cell-dependent antitumor immunity, diminishes HCC growth, and improves immunotherapy efficacy. CONCLUSIONS: This study demonstrates that UFAs promote tumourigenesis by enhancing the immunosuppressive tumour microenvironment via FABP5-PPAR signaling and provides a proof-of-concept for targeting this pathway to improve tumour immunotherapy.

Podocyte-specific deletion of ubiquitin carboxyl-terminal hydrolase L1 causes podocyte injury by inducing endoplasmic reticulum stress.

Ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) is a unique component of the ubiquitin-proteasome system (UPS), which has multiple activities in maintaining intracellular ubiquitin levels. We previously reported the aberrant low expression of UCHL1 in podocytes of non-immune complex-mediated glomerulonephritis, and recent studies indicate that anti-UCHL1 antibody was responsible for the refractory minimal change disease (MCD), but the specific effect of UCHL1 to the podocytopathy has not been determined. Therefore, we generated podocyte-specific UCHL1 gene knockout (UCHL1cre/cre) rats model. Podocyte-specific UCHL1 knockout rats exhibited severe kidney damage, including segmental/global glomerulosclerosis, kidney function damage and severe proteinuria, compared with littermate control. Subsequently, by carrying out mass spectrometry analysis of isolated glomeruli of rats, abnormal protein accumulation of ECM-receptor Interaction was found in UCHL1cre/cre rats. Mechanistic studies in vivo and in vitro revealed that aberrant protein accumulation after UCHL1 deficiency induced endoplasmic reticulum (ER) stress, unfolded protein reaction (UPR) to reduce the protein level of podocyte skeleton proteins, and CHOP mediated apoptosis as well, which related to the dysfunction of the ubiquitin-proteasome system with decreased free monomeric ubiquitin level, thereby affecting protein ubiquitination and degradation. In addition, inhibition of ER stress by 4-PBA could attenuate the degree of ER stress and podocyte dysfunction. Our study indicates that UCHL1 is a potential target for preventing podocytes injury in some non-immune complex-mediated glomerulopathy.

Mechanism of the switch from NO to H2O2 in endothelium-dependent vasodilation in diabetes.

Coronary microvascular dysfunction is prevalent among people with diabetes and is correlated with cardiac mortality. Compromised endothelial-dependent dilation (EDD) is an early event in the progression of diabetes, but its mechanisms remain incompletely understood. Nitric oxide (NO) is the major endothelium-dependent vasodilatory metabolite in the healthy coronary circulation, but this switches to hydrogen peroxide (H2O2) in coronary artery disease (CAD) patients. Because diabetes is a significant risk factor for CAD, we hypothesized that a similar NO-to-H2O2 switch would occur in diabetes. Vasodilation was measured ex vivo in isolated coronary arteries from wild type (WT) and microRNA-21 (miR-21) null mice on a chow or high-fat/high-sugar diet, and B6.BKS(D)-Leprdb/J (db/db) mice using myography. Myocardial blood flow (MBF), blood pressure, and heart rate were measured in vivo using contrast echocardiography and a solid-state pressure sensor catheter. RNA from coronary arteries, endothelial cells, and cardiac tissues was analyzed via quantitative real-time PCR for gene expression, and cardiac protein expression was assessed via western blot analyses. Superoxide was detected via electron paramagnetic resonance. (1) Ex vivo coronary EDD and in vivo MBF were impaired in diabetic mice. (2) Nω-Nitro-L-arginine methyl ester, an NO synthase inhibitor (L-NAME), inhibited ex vivo coronary EDD and in vivo MBF in WT. In contrast, polyethylene glycol-catalase, an H2O2 scavenger (Peg-Cat), inhibited diabetic mouse EDD ex vivo and MBF in vivo. (3) miR-21 was upregulated in diabetic mouse endothelial cells, and the deficiency of miR-21 prevented the NO-to-H2O2 switch and ameliorated diabetic mouse vasodilation impairments. (4) Diabetic mice displayed increased serum NO and H2O2, upregulated mRNA expression of Sod1, Sod2, iNos, and Cav1, and downregulated Pgc-1α in coronary arteries, but the deficiency of miR-21 reversed these changes. (5) miR-21-deficient mice exhibited increased cardiac PGC-1α, PPARα and eNOS protein and reduced endothelial superoxide. (6) Inhibition of PGC-1α changed the mRNA expression of genes regulated by miR-21, and overexpression of PGC-1α decreased the expression of miR-21 in high (25.5 mM) glucose treated coronary endothelial cells. Diabetic mice exhibit a NO-to-H2O2 switch in the mediator of coronary EDD, which contributes to microvascular dysfunction and is mediated by miR-21. This study represents the first mouse model recapitulating the NO-to-H2O2 switch seen in CAD patients in diabetes.

Hepatocyte Nuclear Factor 4α Prevents the Steatosis-to-NASH Progression by Regulating p53 and Bile Acid Signaling (in mice).

BACKGROUND AND AIMS: Hepatocyte nuclear factor 4α (HNF4α) is highly enriched in the liver, but its role in the progression of nonalcoholic liver steatosis (NAFL) to NASH has not been elucidated. In this study, we investigated the effect of gain or loss of HNF4α function on the development and progression of NAFLD in mice. APPROACH AND RESULTS: Overexpression of human HNF4α protected against high-fat/cholesterol/fructose (HFCF) diet-induced steatohepatitis, whereas loss of Hnf4α had opposite effects. HNF4α prevented hepatic triglyceride accumulation by promoting hepatic triglyceride lipolysis, fatty acid oxidation, and VLDL secretion. Furthermore, HNF4α suppressed the progression of NAFL to NASH. Overexpression of human HNF4α inhibited HFCF diet-induced steatohepatitis in control mice but not in hepatocyte-specific p53-/- mice. In HFCF diet-fed mice lacking hepatic Hnf4α, recapitulation of hepatic expression of HNF4α targets cholesterol 7α-hydroxylase and sterol 12α-hydroxylase and normalized hepatic triglyceride levels and attenuated steatohepatitis. CONCLUSIONS: The current study indicates that HNF4α protects against diet-induced development and progression of NAFLD by coordinating the regulation of lipolytic, p53, and bile acid signaling pathways. Targeting hepatic HNF4α may be useful for treatment of NASH.

Hepatocyte-specific expression of human carboxylesterase 2 attenuates nonalcoholic steatohepatitis in mice.

Human carboxylesterase 2 (CES2) has triacylglycerol hydrolase (TGH) activities and plays an important role in lipolysis. In this study, we aim to determine the role of human CES2 in the progression or reversal of steatohepatitis in diet-induced or genetically obese mice. High-fat/high-cholesterol/high-fructose (HFCF) diet-fed C57BL/6 mice or db/db mice were intravenously injected with an adeno-associated virus expressing human CES2 under the control of an albumin promoter. Human CES2 protected against HFCF diet-induced nonalcoholic fatty liver disease (NAFLD) in C57BL/6J mice and reversed steatohepatitis in db/db mice. Human CES2 also improved glucose tolerance and insulin sensitivity. Mechanistically, human CES2 reduced hepatic triglyceride (T) and free fatty acid (FFA) levels by inducing lipolysis and fatty acid oxidation and inhibiting lipogenesis via suppression of sterol regulatory element-binding protein 1. Furthermore, human CES2 overexpression improved mitochondrial respiration and glycolytic function, and inhibited gluconeogenesis, lipid peroxidation, apoptosis, and inflammation. Our data suggest that hepatocyte-specific expression of human CES2 prevents and reverses steatohepatitis. Targeting hepatic CES2 may be an attractive strategy for treatment of NAFLD.NEW & NOTEWORTHY Human CES2 attenuates high-fat/cholesterol/fructose diet-induced steatohepatitis and reverses steatohepatitis in db/db mice. Mechanistically, human CES2 induces lipolysis, fatty acid and glucose oxidation, and inhibits hepatic glucose production, inflammation, lipid oxidation, and apoptosis. Our data suggest that human CES2 may be targeted for treatment of non-alcoholic steatohepatitis (NASH).

Methionine sulfoxide reductase A attenuates atherosclerosis via repairing dysfunctional HDL in scavenger receptor class B type I deficient mice.

High-density lipoprotein (HDL), a well-known atheroprotective factor, can be converted to proatherogenic particles in chronic inflammation. HDL-targeted therapeutic strategy for atherosclerotic cardiovascular disease (CVD) is currently under development. This study aims to assess the role of methionine sulfoxide reductase A (MsrA) in abnormal HDL and its related disorders in scavenger receptor class B type I deficient (SR-BI-/- ) mice. First, we demonstrated that MsrA overexpression attenuated ROS level and inflammation in HepG2 cells. For the in vivo study, SR-BI-/- mice were intravenously injected with lentivirus to achieve hepatic MsrA overexpression. High-level hepatic MsrA significantly reduced the plasma free cholesterol contents, improved HDL functional proteins apolipoprotein A-I (apoAI), apoE, paraoxonase1 (PON1), and lecithin:cholesterol acyltransferase (LCAT), while decreased the pro-inflammatory property of dysfunctional HDL, contributing to reduced atherosclerosis and hepatic steatosis in Western diet-fed mice. Furthermore, the study revealed that hepatic MsrA altered the expression of several genes controlling HDL biogenesis, cholesterol esterification, cholesterol uptake mediated by low-density lipoprotein receptor (LDLR) and biliary excretion, as well as suppressed nuclear factor κB (NF-κB) signaling pathway, which largely relied on liver X receptor alpha (LXRα)-upregulation. These results provide original evidence that MsrA may be a promising target for the therapy of dysfunctional HDL-related CVD.

Macrophage miR-34a Is a Key Regulator of Cholesterol Efflux and Atherosclerosis.

Macrophages play a crucial role in the pathogenesis of atherosclerosis, but the molecular mechanisms remain poorly understood. Here we show that microRNA-34a (miR-34a) is a key regulator of macrophage cholesterol efflux and reverse cholesterol transport by modulating ATP-binding cassette transporters ATP-binding cassette subfamily A member 1 (ABCA1) and ATP-binding cassette subfamily G member 1 (ABCG1). miR-34a also regulates M1 and M2 macrophage polarization via liver X receptor α. Furthermore, global loss of miR-34a reduces intestinal cholesterol or fat absorption by inhibiting cytochrome P450 enzymes CYP7A1 and sterol 12α-hydroxylase (CYP8B1). Consistent with these findings, macrophage-selective or global ablation of miR-34a markedly inhibits the development of atherosclerosis. Finally, therapeutic inhibition of miR-34a promotes atherosclerosis regression and reverses diet-induced metabolic disorders. Our studies outline a central role of miR-34a in regulating macrophage cholesterol efflux, inflammation, and atherosclerosis, suggesting that miR-34a is a promising target for treatment of cardiometabolic diseases.

Reaction mechanism and kinetics of Criegee intermediate and hydroperoxymethyl formate.

The reaction mechanism and kinetics of the simplest Criegee intermediate CH2OO reaction with hydroperoxymethyl formate (HPMF) was investigated at high-level quantum chemistry calculations. HPMF has two reactive functional groups, -C(O)OH and -OOH. The calculated results of thermodynamic data and rate constants indicated that the insertion reactions of CH2OO with -OOH group of HPMF were more favorable than the reactions of CH2OO with -C(O)OH group. The calculated overall rate constant was 2.33 × 10-13 cm3/(molecule⋅sec) at 298 K and the rate constants decreased as the temperature increased from 200 to 480 K. In addition, we also proved the polymerization reaction mechanism between CH2OO and -OOH of HPMF. This theoretical study interpreted the previous experimental results, and supplied the structures of the intermediate products that couldn't be detected during the experiment.

Hepatic Forkhead Box Protein A3 Regulates ApoA-I (Apolipoprotein A-I) Expression, Cholesterol Efflux, and Atherogenesis.

OBJECTIVE: To determine the role of hepatic FOXA3 (forkhead box A3) in lipid metabolism and atherosclerosis. Approach and Results: Hepatic FOXA3 expression was reduced in diabetic or high fat diet-fed mice or patients with nonalcoholic steatohepatitis. We then used adenoviruses to overexpress or knock down hepatic FOXA3 expression. Overexpression of FOXA3 in the liver increased hepatic ApoA-I (apolipoprotein A-I) expression, plasma HDL-C (high-density lipoprotein cholesterol) level, macrophage cholesterol efflux, and macrophage reverse cholesterol transport. In contrast, knockdown of hepatic FOXA3 expression had opposite effects. We further showed that FOXA3 directly bound to the promoter of the Apoa1 gene to regulate its transcription. Finally, AAV8 (adeno-associated virus serotype 8)-mediated overexpression of human FOXA3 in the hepatocytes of Apoe-/- (apolipoprotein E-deficient) mice raised plasma HDL-C levels and significantly reduced atherosclerotic lesions. CONCLUSIONS: Hepatocyte FOXA3 protects against atherosclerosis by inducing ApoA-I and macrophage reverse cholesterol transport.

Farnesoid X Receptor Activation by Obeticholic Acid Elevates Liver Low-Density Lipoprotein Receptor Expression by mRNA Stabilization and Reduces Plasma Low-Density Lipoprotein Cholesterol in Mice.

Objective- The objective of this study was to determine whether and how activation of farnesoid X receptor (FXR) by obeticholic acid (OCA), a clinical FXR agonist, modulates liver low-density lipoprotein receptor (LDLR) expression under normolipidemic conditions. Approach and Results- Administration of OCA to chow-fed mice increased mRNA and protein levels of LDLR in the liver without affecting the sterol-regulatory element binding protein pathway. Profiling of known LDLR mRNA-binding proteins demonstrated that OCA treatment did not affect expressions of mRNA degradation factors hnRNPD (heterogeneous nuclear ribonucleoprotein D) or ZFP36L1 but increased the expression of Hu antigen R (HuR) an mRNA-stabilizing factor. Furthermore, inducing effects of OCA on LDLR and HuR expression were ablated in Fxr-/- mice. To confirm the post-transcriptional mechanism, we used transgenic mice (albumin-luciferase-untranslated region) that express a human LDLR mRNA 3' untranslated region luciferase reporter gene in the liver. OCA treatment led to significant rises in hepatic bioluminescence signals, Luc-untranslated region chimeric mRNA levels, and endogenous LDLR protein abundance, which were accompanied by elevations of hepatic HuR mRNA and protein levels in OCA-treated transgenic mice. In vitro studies conducted in human primary hepatocytes and HepG2 cells demonstrated that FXR activation by OCA and other agonists elicited the same inducing effect on LDLR expression as in the liver of normolipidemic mice. Furthermore, depletion of HuR in HepG2 cells by short interfering RNA transfection abolished the inducing effect of OCA on LDLR expression. Conclusions- Our study is the first to demonstrate that FXR activation increases LDLR expression in liver tissue by a post-transcriptional regulatory mechanism involving LDLR mRNA-stabilizing factor HuR.

Protein markers of dysfunctional HDL in scavenger receptor class B type I deficient mice.

BACKGROUND: Scavenger receptor class B type I (SR-BI) plays a key role in high density lipoproteins (HDL) metabolism. SR-BI deficiency in mice results in enhanced susceptibility to atherosclerosis with abnormal large, cholesterol enriched, and functional impaired HDL. This study was to characterize the protein markers of dysfunctional HDL in SR-BI deficient (SR-BI-/-) mice and to test if the defective of HDL might be affected by probucol treatment. METHODS: Shotgun proteomics and 2-D gel electrophoresis were performed to examine the profile of HDL protein and distribution of HDL particles isolated from SR-BI-/- mice. HDL's cell-function, paraoxonase 1 (PON1) and myeloperoxidase activity were assessed. The mice were treated with 1.2 mg/g/day probucol for 6 weeks and the impact on HDL protein markers was analyzed. The differential proteins were quantified by Western blotting. RESULTS: The relative amount of protein in SR-BI-/- HDL was decreased by about 25% compared to that in HDL from wild type (WT) mice. Compared to WT HDL, relative protein abundance of representative apoAI and PON1 in SR-BI-/- HDL were significantly reduced, whereas acute-phase protein serum amyloid A (SAA) and apoAIV, proteinase inhibitor proteins α-1-antitrypsin (A1AT) were increased. The distribution of plasma apoAI-containing HDL particles in SR-BI-/- mice was also dramatically altered, although plasma apoAI level was no difference. The protein alterations were accompanied with dysfunction of SR-BI-/- HDL, evidenced by impaired cholesterol homeostasis in macrophages, and reduced anti-oxidative and anti-inflammatory effects. Probucol treatment of SR-BI-/- mice could restored the relative contents of critical proteins including apoAI, PON1, SAA, apoAIV and A1AT on HDL, and improve HDL dysfunction despite decreased HDL-C level. CONCLUSION: SR-BI deficiency leading to dysfunctional HDL is closely related to alteration of HDL protein, suggesting that identification of apoAI, PON1, SAA, apoAIV, and A1AT may serve as the valuable protein markers for diagnosis and therapeutics of dysfunctional HDL-related metabolic diseases.

Hepatic overexpression of methionine sulfoxide reductase A reduces atherosclerosis in apolipoprotein E-deficient mice.

Methionine sulfoxide reductase A (MsrA), a specific enzyme that converts methionine-S-sulfoxide to methionine, plays an important role in the regulation of protein function and the maintenance of redox homeostasis. In this study, we examined the impact of hepatic MsrA overexpression on lipid metabolism and atherosclerosis in apoE-deficient (apoE(-/-)) mice. In vitro study showed that in HepG2 cells, lentivirus-mediated human MsrA (hMsrA) overexpression upregulated the expression levels of several key lipoprotein-metabolism-related genes such as liver X receptor α, scavenger receptor class B type I, and ABCA1. ApoE(-/-) mice were intravenously injected with lentivirus to achieve high-level hMsrA expression predominantly in the liver. We found that hepatic hMsrA expression significantly reduced plasma VLDL/LDL levels, improved plasma superoxide dismutase, and paraoxonase-1 activities, and decreased plasma serum amyloid A level in apoE(-/-) mice fed a Western diet, by significantly altering the expression of several genes in the liver involving cholesterol selective uptake, conversion and excretion into bile, TG biosynthesis, and inflammation. Moreover, overexpression of hMsrA resulted in reduced hepatic steatosis and aortic atherosclerosis. These results suggest that hepatic MsrA may be an effective therapeutic target for ameliorating dyslipidemia and reducing atherosclerosis-related cardiovascular diseases.

PEP-1-MsrA ameliorates inflammation and reduces atherosclerosis in apolipoprotein E deficient mice.

BACKGROUND: Methionine sulfoxide reductase A (MsrA) is a potent intracellular oxidoreductase and serves as an essential factor that protects cells against oxidative damage. However, therapeutic use of exogenous MsrA in oxidative stress-induced diseases is limited, because it cannot enter the cells. The aim of this study is to investigate whether MsrA with PEP-1, a cell penetrating peptide, fused to its N-terminus can protect against oxidative stress in macrophages and can attenuate atherosclerosis in apolipoprotein E deficient (apoE(-/-)) mice. METHODS: MsrA and the fusion protein PEP-1-MsrA were expressed and purified using a pET28a expression system. Transduction of the fusion protein into macrophages was confirmed by Western blot and immunofluorescence staining. Intracellular reactive oxygen species (ROS) and apoptosis levels were measured by flow cytometry. In in vivo study, MsrA or PEP-1-MsrA proteins were intraperitoneally injected into apoE(-/-) mice fed a Western diet for 12 weeks. Plasma lipids levels, inflammatory gene expression, and paraoxonase-1 (PON1) and superoxide dismutase (SOD) activities were assessed. Atherosclerotic lesions were analyzed by Oil Red O staining and immunohistochemistry. RESULTS: PEP-1-MsrA could penetrate the cells and significantly reduced intracellular ROS levels and apoptosis in H2O2-treated macrophages. It also decreased TNFα and IL-1ß mRNA levels and increased the IL-10 mRNA level in lipopolysaccharide-treated macrophages. In in vivo study, PEP-1-MsrA injection significantly increased plasma PON1 and SOD activities and decreased plasma monocyte chemoattractant protein 1 (MCP-1) level compared to the injection of vehicle control or MsrA. In PEP-1-MsrA injected mice, hepatic PON1 levels were increased, while the expression of TNFα and IL-6 mRNA in the liver was suppressed. Although plasma total cholesterol and triglyceride levels did not change, the aortic atherosclerosis in PEP-1-MsrA treated mice was significantly reduced. This was accompanied by a reduction of total and apoptotic macrophages in the lesions. CONCLUSION: Our study provides evidence that PEP-1-MsrA may be a potential therapeutic agent for atherosclerosis-related cardiovascular diseases.

A Method for Prediction of High-Water-Production Areas in Coalbed Methane Blocks Based on γ Logging Data: A Case Study of the Taiyuan Formation in Liulin Area, Eastern Ordos Basin, China.

The roof aquifer of the Carboniferous Taiyuan Formation coal beds in the Liulin area severely restricts the development and utilization of coalbed methane (CBM). A method for quantitatively predicting high-water-production areas was established by analyzing the relationship between the geophysical logging data and water production. The results showed that the logging profile of the limestone aquifers in high-water-production wells was unique, with high acoustic velocity (AC), high γ-ray values (GR), and low resistivity (Rd). The developed pores and fractures in the roof limestone increase the interval transit time. The formation water in the pores and fractures of the roof limestone decreases the resistivity. The clay filling in the pores and fractures of the roof limestone originated from the dissolution product of limestone and hydrodynamic transportation, which resulted in increased GR values. Furthermore, the representative natural GR log data were used to calculate the clay content in limestone, which indicated that the clay content in limestone had a positive correlation with the water yield of the CBM wells. The water-bearing characteristics of roof limestone showed that the water content was higher in the northern area and decreased gradually toward the south. The method for predicting the high-water-production area was helpful for the CBM exploration and production.

Quantitative Characterization of Inorganic Pores in Sinian Doushantou Dolomitic Shale Based on FIB-SEM in Western Hubei Province, China.

Unlike traditional shale gas reservoirs, where organic matter pores dominate, inorganic pores are the primary reservoir space in the Sinian (Ediacaran) high-maturity Doushantou dolomitic shale in western Hubei Province, China. The inorganic pore characteristics of Doushantuo shale and its influence on shale gas aggregation were investigated by examining the TOC content, thermal maturity, mineralogical composition, and field-emission scanning electron microscopy (SEM) and focused ion beam scanning electron microscopy (FIB-SEM) of drill cores. The results show that the shale mineral composition in the study area is primarily dolomite and plate-shaped interparticle-intercrystalline pores associated with dolomite are widespread inorganic pores in dolomitic shale. Interparticle-intercrystalline pores account for 75% of the total pores, with a pore size distribution mainly between 50 and 300 nm, as extracted from the 3D pore network model (PNM). Compared with organic pores, interparticle-intercrystalline pores provide greater space for gas storage and have a strong coupling relationship with the hydrocarbon generation and evolution of organic matter. Therefore, the inorganic pores in the Doushantuo Formation play a vital role in the enrichment and accumulation of shale gas. This study aims to establish a scientific basis for understanding the enrichment mechanism of shale gas in Doushantuo dolomitic shale and other inorganic pore-dominated shales in southern China.

Adipocyte retinoic acid receptor α prevents obesity and steatohepatitis by regulating energy expenditure and lipogenesis.

OBJECTIVE: The adipose tissue-liver axis is a major regulator of the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Retinoic acid signaling plays an important role in development and metabolism. However, little is known about the role of adipose retinoic acid signaling in the development of obesity-associated NAFLD. In this work, the aim was to investigate whether and how retinoic acid receptor alpha (RARα) regulated the development of obesity and NAFLD. METHODS: RARα expression in adipose tissue of db/db or ob/ob mice was determined. Rarαfl/fl mice and adipocyte-specific Rarα-/- (RarαAdi-/- ) mice were fed a chow diet for 1 year or high-fat diet (HFD) for 20 weeks. Primary adipocytes and primary hepatocytes were co-cultured. Metabolic regulation and inflammatory response were characterized. RESULTS: RARα expression was reduced in adipose tissue of db/db or ob/ob mice. RarαAdi-/- mice had increased obesity and steatohepatitis (NASH) when fed a chow diet or HFD. Loss of adipocyte RARα induced lipogenesis and inflammation in adipose tissue and the liver and reduced thermogenesis. In the co-culture studies, loss of RARα in adipocytes induced inflammatory and lipogenic programs in hepatocytes. CONCLUSIONS: The data demonstrate that RARα in adipocytes prevents obesity and NASH via inhibiting lipogenesis and inflammation and inducing energy expenditure.

Loss of adipose ATF3 promotes adipose tissue lipolysis and the development of MASH.

The crosstalk between adipose tissue and the liver is finely controlled to maintain metabolic health. Yet, how adipose tissue controls toxic free fatty acid overflow into the liver remains incompletely understood. Here, we show that adipocyte activating transcription factor 3 (ATF3) was induced in human or mouse obesity. Adipocyte Atf3-/- (Atf3Adi-/-) mice developed obesity, glucose intolerance, and metabolic dysfunction-associated steatohepatitis (MASH) in chow diet, high-fat diet, or Western diet-fed mice. Blocking fatty acid flux by inhibiting hepatocyte CD36, but not the restoration of hepatic AMPK signaling, prevented the aggravation of MASH in Atf3Adi-/- mice. Further studies show that the loss of adipocyte ATF3 increased lipolysis via inducing adipose triglyceride lipase, which in turn induced lipogenesis and inflammation in hepatocytes. Moreover, Atf3Adi-/- mice had reduced energy expenditure and increased adipose lipogenesis and inflammation. Our data demonstrate that adipocyte ATF3 is a gatekeeper in counteracting MASH development under physiological and pathological conditions.

Atf3-mediated metabolic reprogramming in hepatic macrophage orchestrates metabolic dysfunction-associated steatohepatitis.

Metabolic dysfunction-associated steatohepatitis (MASH) is regulated by complex interplay between the macrophages and surrounding cells in the liver. Here, we show that Atf3 regulates glucose-fatty acid cycle in macrophages attenuates hepatocyte steatosis, and fibrogenesis in hepatic stellate cells (HSCs). Overexpression of Atf3 in macrophages protects against the development of MASH in Western diet-fed mice, whereas Atf3 ablation has the opposite effect. Mechanistically, Atf3 improves the reduction of fatty acid oxidation induced by glucose via forkhead box O1 (FoxO1) and Cd36. Atf3 inhibits FoxO1 activity via blocking Hdac1-mediated FoxO1 deacetylation at K242, K245, and K262 and increases Zdhhc4/5-mediated CD36 palmitoylation at C3, C7, C464, and C466; furthermore, macrophage Atf3 decreases hepatocytes lipogenesis and HSCs activation via retinol binding protein 4 (Rbp4). Anti-Rbp4 can prevent MASH progression that is induced by Atf3 deficiency in macrophages. This study identifies Atf3 as a regulator of glucose-fatty acid cycle. Targeting macrophage Atf3 or Rbp4 may be a plausible therapeutic strategy for MASH.

Krüppel-like factor 10 protects against metabolic dysfunction-associated steatohepatitis by regulating HNF4α-mediated metabolic pathways.

BACKGROUND: Krüppel-like factor 10 (KLF10), a zinc finger transcription factor, plays a pivotal role in modulating TGF-ß-mediated cellular processes such as growth, apoptosis, and differentiation. Recent studies have implicated KLF10 in regulating lipid metabolism and glucose homeostasis. This study aimed to elucidate the precise role of hepatic KLF10 in developing metabolic dysfunction-associated steatohepatitis (MASH) in diet-induced obese mice. METHODS: We investigated hepatic KLF10 expression under metabolic stress and the effects of overexpression or ablation of hepatic KLF10 on MASH development and lipidemia. We also determined whether hepatocyte nuclear factor 4α (HNF4α) mediated the metabolic effects of KLF10. RESULTS: Hepatic KLF10 was downregulated in MASH patients and genetically or diet-induced obese mice. AAV8-mediated overexpression of KLF10 in hepatocytes prevented Western diet-induced hypercholesterolemia and steatohepatitis, whereas inactivation of hepatocyte KLF10 aggravated Western diet-induced steatohepatitis. Mechanistically, KLF10 reduced hepatic triglyceride and free fatty acid levels by inducing lipolysis and fatty acid oxidation and inhibiting lipogenesis, and reducing hepatic cholesterol levels by promoting bile acid synthesis. KLF10 highly induced HNF4α expression by directly binding to its promoter. The beneficial effect of KLF10 on MASH development was abolished in mice lacking hepatocyte HNF4α. In addition, the inactivation of KLF10 in hepatic stellate cells exacerbated Western diet-induced liver fibrosis by activating the TGF-ß/SMAD2/3 pathway. CONCLUSIONS: Our data collectively suggest that the transcription factor KLF10 plays a hepatoprotective role in MASH development by inducing HNF4α. Targeting hepatic KLF10 may offer a promising strategy for treating MASH.