The nuclear receptor CAR modulates alcohol-induced liver injury

The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily and a sensor and detoxifier of both xenobiotics and endobiotics. Recent studies also show that CAR participates in metabolism of glucose and lipid, and has an important role in fatty liver disease and diabetes. In this study, we investigate the roles of CAR in chronic and acute alcohol-induced liver injuries. The results showed that absence of CAR in rodents led to significantly increased susceptibility to chronic alcohol-induced liver injury, which was accompanied with elevated hepatocyte apoptosis and fat accumulation. However, pre-activation of CAR by a CAR agonist, TCPOBOP, strongly enhanced the hepatic toxicity by both chronic and acute alcohol infusion in wild-type, but not in CAR(-/-) mice. Gene expression analyses indicated that CAR pre-activation and alcohol infusion synergistically decreased the expression of enzymes that metabolize the alcohol in liver. These results support a role of CAR in modulating alcoholic liver injury and imply a risk of synergistic liver toxicity induced by alcohol and CAR activation.     

Metallothionein-independent zinc protection from alcoholic liver injury

Previous studies using metallothionein (MT)-overexpressing transgenic mice have demonstrated that MT protects the liver from oxidative injury induced by alcohol. The mechanism of action of MT is unknown. Because MT primarily binds to zinc under physiological conditions and releases zinc under oxidative stress and zinc is an antioxidant element, it is likely that zinc mediates the protective action of MT. The present study was undertaken to determine the distinct role of zinc in hepatic protection from alcoholic injury. MT I/II-knockout (MT-KO) mice along with their wild-type controls were treated with three gastric doses of ethanol at 5 g/kg at 12-hour intervals. Zinc sulfate was injected intraperitoneally in a dosage of 5 mg/kg/day for 3 days before ethanol treatment. MT concentrations in MT-KO mice were very low and zinc concentrations in MT-KO mice were lower than in wild-type mice. Zinc treatment significantly elevated hepatic MT concentrations only in wild-type mice and increased zinc concentrations in both MT-KO and wild-type mice. Ethanol treatment caused degenerative morphological changes and necrotic appearance in the livers of MT-KO mice. Microvesicular steatosis was the only ethanol-induced change in the liver of wild-type mice. Ethanol treatment decreased hepatic glutathione concentrations and increased hepatic lipid peroxidation, and the concentrations of lipid peroxide products in the wild-type mice were lower than in the MT-KO mice. All of these alcohol-induced toxic responses were significantly suppressed by zinc treatment in both MT-KO and wild-type mouse livers. These results demonstrate that zinc, independent of MT, plays an important role in protection from alcoholic liver injury. However, MT is required to maintain high levels of zinc in the liver, suggesting that the protective action of MT in the liver is likely mediated by zinc.     

Lactobacillus rhamnosus GG treatment potentiates intestinal hypoxia-inducible factor, promotes intestinal integrity and ameliorates alcohol-induced liver injury

Gut-derived endotoxin is a critical factor in the development and progression of alcoholic liver disease (ALD). Probiotics can treat alcohol-induced liver injury associated with gut leakiness and endotoxemia in animal models, as well as in human ALD; however, the mechanism or mechanisms of their beneficial action are not well defined. We hypothesized that alcohol impairs the adaptive response-induced hypoxia-inducible factor (HIF) and that probiotic supplementation could attenuate this impairment, restoring barrier function in a mouse model of ALD by increasing HIF-responsive proteins (eg, intestinal trefoil factor) and reversing established ALD. C57BJ/6N mice were fed the Lieber DeCarli diet containing 5% alcohol for 8 weeks. Animals received Lactobacillus rhamnosus GG (LGG) supplementation in the last 2 weeks. LGG supplementation significantly reduced alcohol-induced endotoxemia and hepatic steatosis and improved liver function. LGG restored alcohol-induced reduction of HIF-2α and intestinal trefoil factor levels. In vitro studies using the Caco-2 cell culture model showed that the addition of LGG supernatant prevented alcohol-induced epithelial monolayer barrier dysfunction. Furthermore, gene silencing of HIF-1α/2α abolished the LGG effects, indicating that the protective effect of LGG is HIF-dependent. The present study provides a mechanistic insight for utilization of probiotics for the treatment of ALD, and suggests a critical role for intestinal hypoxia and decreased trefoil factor in the development of ALD.     

Deficient liver regeneration after carbon tetrachloride injury in mice lacking type 1 but not type 2 tumor necrosis factor receptor.

Signaling by tumor necrosis factor type 1 receptor (TNFR-1) is required for the initiation of liver regeneration after partial hepatectomy. Using knockout mice that lack either TNFR-1 or TNFR-2, we determined whether signaling through TNF receptors is important for liver injury and hepatocyte proliferation induced by carbon tetrachloride (CCl4). Lack of TNFR-1 inhibited hepatocyte DNA synthesis after CCl4 injection. At 44 hours after the injection, replication of hepatocytes in TNFR-1 was 50% to 90% lower than in wild-type (WT) animals, depending on the dose injected. In WT animals, hepatocyte replication was essentially completed by 4 days after CCl4 injection, but replication at a low level persisted in TNFR-1 mice for at least 2 weeks. TNFR-1 knockout mice had little detectable NF-kappa B and STAT3 binding during the first 5 hours after CCl4, high plasma TNF, and reduced levels of plasma interleukin (IL)-6 and liver IL-6 mRNA. Injection of IL-6 30 minutes before CCl4 administration corrected the deficiency of hepatocyte replication at 44 hours and restored STAT3 binding to normal levels. In contrast, mice lacking TNFR-2 did not differ significantly from WT mice in NF-kappa B and STAT3 binding, IL-6 and TNF levels, or hepatocyte replication. Although AP-1 binding was induced in WT TNFR-1 and TNFR-2 knockout mice, binding in TNFR-2 knockouts was lower than in WT mice. C/EBP binding was much lower in TNFR-1 and TNFR-2 knockout mice than in WT mice. As assessed by morphological analysis and alanine aminotransferase levels, the acute injury caused by CCl4 appeared to be similar in the three groups of animals, but subsequent regeneration was impaired in mice lacking TNFR-1. We conclude that a TNFR-1 signaling pathway involving NF-kappa B, IL-6, and STAT3 is an important component of the hepatocyte mitogenic response induced by CCl4 injury in mouse liver.     

Hepatocyte proliferation is the possible mechanism for the transient decrease in liver injury during steatosis stage of alcoholic liver disease

Steatosis is a frequent pathologic stage in alcoholic liver disease (ALD). Although the mechanisms for increased susceptibility of steatotic liver to injury have been postulated, the ability of these hepatocytes to proliferate and withstand injury is unknown. There are conflicting reports on the status of hepatocyte regeneration following chronic alcohol ingestion. Hence, the objective of this study was to investigate the temporal dynamics between the pattern of liver injury and hepatocyte proliferation during the steatosis stage of ALD. Alcoholic steatosis was induced in male Sprague-Dawley rats by feeding an ethanol (EtOH)-containing Lieber-DeCarli liquid diet for a period of 5 weeks. Microvesicular steatosis was evident in H&E sections by three weeks in the EtOH-treated rats, which further developed into panlobular macrovesicular steatosis by 5 weeks. Plasma transaminase activities indicated progressive increase in liver injury peaking at 3 weeks with significant but mild decrease at 4 and 5 weeks. CYP2E1 protein and activity was significantly increased in EtOH-fed rats as measured by Western blot and pNP hydroxylation assay. PCNA analysis of liver sections indicated that EtOH-treated rats had a significantly higher number of cells in S phase of cell division at weeks 1 (3.20 +/- 0.19), 2 (7.03 +/- 0.92), and 3 (4.23 +/- 1.41) when compared to controls (1.5 +/- 0.22). NF-kappaB DNA binding and Cyclin D1 proteins increased significantly in the EtOH-treated rats corresponding with enhanced hepatic proliferation. These data suggest the transient decline in liver injury during alcoholic steatosis is due to enhanced NF-kappaB-dependent hepatocyte proliferation.     

茶多酚对酒精诱导的小鼠肝脂质过氧化和血清ALT活性变化的影响

目的: 研究茶多酚(TP)对酒精性肝损伤的防治作用。方法: 通过离体和整体实验,采用分光光度法和血清生化检测法,检测肝组织脂质过氧化产物丙二醛(MDA)水平和血清谷丙转氨酶(ALT)的活性。结果: 离体实验中,TP+酒精组肝MDA水平明显低于生理盐水+酒精组(P＜0.01)。整体实验中,TP+酒精组小鼠肝脏MDA水平和血清中ALT活性均显著低于生理盐水+酒精组(P＜0.05)。此外,灌酒精1h后再给TP组,小鼠肝脏MDA含量也明显低于酒精+生理盐水组(P＜0.01)。结论: 茶多酚能抑制酒精引起的小鼠肝组织MDA水平和血清ALT活性的升高,可能具有防治酒精性肝损伤的作用。     

Systematical analysis of impacts of heat stress on the proliferation, apoptosis and metabolism of mouse hepatocyte

Heat stress will stimulate cells of living organisms to generate heat shock proteins (Hsps). In the mouse liver, impacts of heat stress on hepatocyte proliferation, apoptosis and metabolism have not been studied systematically at different temperatures. In this research, the test mice were heated to 40, 42, 44 and 46°C, respectively, for 20?min and recovered at room temperature for 8?h in normal feeding conditions; the control animals were kept at room temperature without heat stress. The expression levels of Hsp70, Pcna, Bax, Bcl2, cytochrome P450 1A2 (CYP1A2), CYP2E1 and analog of CYP3A4 (not reported in mouse before), the parameters reflecting stress strength, cell proliferation, apoptosis and metabolism, were detected by western blotting, immunohistochemistry and semi-quantitative RT-PCR in test and control mice. Haematoxylin-eosin (H&E) staining and TUNEL analysis were further used to study the impacts of heat stress at different temperatures on hepatocellular necrosis and apoptosis. Serum AST and ALT levels, the markers of liver injury, were measured after heat stress at different temperatures. The data show that Hsp70 expression was significantly increased when temperature increased (P??0.05). At higher temperatures (44 or 46°C), expression of Pcna was decreased while hepatocyte necrosis and apoptosis were induced (P?相似文献   

Preservation of intestinal structural integrity by zinc is independent of metallothionein in alcohol-intoxicated mice

Intestinal-derived endotoxins are importantly involved in alcohol-induced liver injury. Disruption of intestinal barrier function and endotoxemia are common features associated with liver inflammation and injury due to acute ethanol exposure. Zinc has been shown to inhibit acute alcohol-induced liver injury. This study was designed to determine the inhibitory effect of zinc on alcohol-induced endotoxemia and whether the inhibition is mediated by metallothionein (MT) or is independent of MT. MT knockout (MT-KO) mice were administered three oral doses of zinc sulfate (2.5 mg zinc ion/kg body weight) every 12 hours before being administered a single dose of ethanol (6 g/kg body weight) by gavage. Ethanol administration caused liver injury as determined by increased serum transaminases, parenchymal fat accumulation, necrotic foci, and an elevation of tumor necrosis factor (TNF-alpha). Increased plasma endotoxin levels were detected in ethanol-treated animals whose small intestinal structural integrity was compromised as determined by microscopic examination. Zinc supplementation significantly inhibited acute ethanol-induced liver injury and suppressed hepatic TNF-alpha production in association with decreased circulating endotoxin levels and a significant protection of small intestine structure. As expected, MT levels remained undetectable in the MT-KO mice under the zinc treatment. These results thus demonstrate that zinc preservation of intestinal structural integrity is associated with suppression of endotoxemia and liver injury induced by acute exposure to ethanol and the zinc protection is independent of MT.     

Changes in the pathogenesis of alcohol-induced liver disease — Preclinical studies

Excessive alcohol consumption presents considerable health risks in humans. A variety of morphologic and functional changes contribute to hepatic injury produced by heavy drinking. The present review summarizes the current knowledge of alcohol-induced liver disease and describes preclinical experimental approaches used to understand alcoholic liver disease (ALD), with a particular emphasis on impaired protein and lipid trafficking, disruption of proteolysis and autophagy, alterations in methionine metabolism and perturbations in metabolic signaling that cause dysfunctional gene expression and the eventual formation of aggresomal Mallory-Denk bodies (MDB) in liver cells. These changes eventually lead to some of the more severe hepatic impairments, including alcoholic hepatitis and fibrosis. Moreover the misuse of alcohol contributes to immune dysfunction and inadequate immune response to viral infections.     

A critical involvement of oxidative stress in acute alcohol-induced hepatic TNF-alpha production

Tumor necrosis factor-alpha (TNF-alpha) production is a critical factor in the pathogenesis of alcoholic liver injury. Both oxidative stress and endotoxin have been implicated in the process of alcohol-induced TNF-alpha production. However, a cause-and-effect relationship between these factors has not been fully defined. The present study was undertaken to determine the mediators of acute alcohol-induced TNF-alpha production using a mouse model of acute alcohol hepatotoxicity. Alcohol administration via gavage at a dose of 6 g/kg to 129/Sv mice induced hepatic TNF-alpha production in Kupffer cells as demonstrated by measuring protein levels, immunohistochemical localization, and mRNA expression. Alcohol intoxication caused liver injury in association with increases in plasma endotoxin and hepatic lipid peroxidation. Treatment with an endotoxin neutralizing protein significantly suppressed alcohol-induced elevation of plasma endotoxin, hepatic lipid peroxidation, and inhibited TNF-alpha production. Treatment with antioxidants, N-ACETYL-L-CYSTEINE, or dimethylsulfoxide, failed to attenuate plasma endotoxin elevation, but significantly inhibited alcohol-induced hepatic lipid peroxidation, TNF-alpha production and steatosis. All treatments prevented alcohol-induced necrotic cell death in the liver. This study thus systemically dissected the relationship among plasma endotoxin elevation, hepatic oxidative stress, and TNF-alpha production following acute alcohol administration, and the results demonstrate that oxidative stress mediates endotoxin-induced hepatic TNF-alpha production in acute alcohol intoxication.     

Attenuated liver fibrosis and depressed serum albumin levels in carbon tetrachloride-treated IL-6-deficient mice.

Chronic intermittent injection of carbon tetrachloride (CCl4) for more than 10 weeks induced liver fibrosis in mice, as evidenced by positive Azan staining and increased intrahepatic collagen content. Preceding the onset of liver fibrosis, interleukin-6 (IL-6) gene expression was enhanced in liver and immunoreactive IL-6 was detected in infiltrating inflammatory cells. To delineate the role of IL-6 in this process, we treated IL-6-deficient mice with CCl4 in a similar manner for 12 weeks, after which fibrotic changes were less evident and serum albumin levels were lower in IL-6-deficient than wild-type mice. Moreover, CCl4-induced expression of transforming growth factor beta1 and hepatocyte growth factor genes in liver was significantly reduced in IL-6-deficient mice. Thus, IL-6 may be vitally involved in fibrotic changes and maintenance of serum albumin levels, partly by modulating intrahepatic expression of these cytokines.     

Alcohol,microbiome, life style influence alcohol and non-alcoholic organ damage

This paper is based upon the “8th Charles Lieber's Satellite Symposium” organized by Manuela G. Neuman at the Research Society on Alcoholism Annual Meeting, on June 25, 2016 at New Orleans, Louisiana, USA.The integrative symposium investigated different aspects of alcohol-induced liver disease (ALD) as well as non-alcohol-induced liver disease (NAFLD) and possible repair. We revealed the basic aspects of alcohol metabolism that may be responsible for the development of liver disease as well as the factors that determine the amount, frequency and which type of alcohol misuse leads to liver and gastrointestinal diseases. We aimed to (1) describe the immuno-pathology of ALD, (2) examine the role of genetics in the development of alcoholic hepatitis (ASH) and NAFLD, (3) propose diagnostic markers of ASH and non-alcoholic steatohepatitis (NASH), (4) examine age and ethnic differences as well as analyze the validity of some models, (5) develop common research tools and biomarkers to study alcohol-induced effects, 6) examine the role of alcohol in oral health and colon and gastrointestinal cancer and (7) focus on factors that aggravate the severity of organ-damage.The present review includes pre-clinical, translational and clinical research that characterizes ALD and NAFLD. Strong clinical and experimental evidence lead to recognition of the key toxic role of alcohol in the pathogenesis of ALD with simple fatty infiltrations and chronic alcoholic hepatitis with hepatic fibrosis or cirrhosis. These latter stages may also be associated with a number of cellular and histological changes, including the presence of Mallory's hyaline, megamitochondria, or perivenular and perisinusoidal fibrosis. Genetic polymorphisms of ethanol metabolizing enzymes and cytochrome p450 (CYP) 2E1 activation may change the severity of ASH and NASH. Other risk factors such as its co-morbidities with chronic viral hepatitis in the presence or absence of human deficiency virus were discussed. Dysregulation of metabolism, as a result of ethanol exposure, in the intestine leads to colon carcinogenesis. The hepatotoxic effects of ethanol undermine the contribution of malnutrition to the liver injury. Dietary interventions such as micro and macronutrients, as well as changes to the microbiota have been suggested. The clinical aspects of NASH, as part of the metabolic syndrome in the aging population, have been presented. The symposium addressed mechanisms and biomarkers of alcohol induced damage to different organs, as well as the role of the microbiome in this dialog. The microbiota regulates and acts as a key element in harmonizing immune responses at intestinal mucosal surfaces. It is known that microbiota is an inducer of proinflammatory T helper 17 cells and regulatory T cells in the intestine. The signals at the sites of inflammation mediate recruitment and differentiation in order to remove inflammatory inducers and promote tissue homeostasis restoration. The change in the intestinal microbiota also influences the change in obesity and regresses the liver steatosis.Evidence on the positive role of moderate alcohol consumption on heart and metabolic diseases as well on reducing steatosis have been looked up. Moreover nutrition as a therapeutic intervention in alcoholic liver disease has been discussed. In addition to the original data, we searched the literature (2008–2016) for the latest publication on the described subjects. In order to obtain the updated data we used the usual engines (Pub Med and Google Scholar).The intention of the eighth symposia was to advance the international profile of the biological research on alcoholism. We also wish to further our mission of leading the forum to progress the science and practice of translational research in alcoholism.     

Myeloid Mixed Lineage Kinase 3 Contributes to Chronic Ethanol-Induced Inflammation and Hepatocyte Injury in Mice

Proinflammatory activity of hepatic macrophages plays a key role during progression of alcoholic liver disease (ALD). Since mixed lineage kinase 3 (MLK3)-dependent phosphorylation of JNK is involved in the activation of macrophages, we tested the hypothesis that myeloid MLK3 contributes to chronic ethanol-induced inflammatory responses in liver, leading to hepatocyte injury and cell death. Primary cultures of Kupffer cells, as well in vivo chronic ethanol feeding, were used to interrogate the role of MLK3 in the progression of liver injury. Phosphorylation of MLK3 was increased in primary cultures of Kupffer cells isolated from ethanol-fed rats compared to cells from pair-fed rats. Kupffer cells from ethanol-fed rats were more sensitive to LPS-stimulated cytokine production; this sensitization was normalized by pharmacological inhibition of MLK3. Chronic ethanol feeding to mice increased MLK3 phosphorylation robustly in F4/80⁺ Kupffer cells, as well as in isolated nonparenchymal cells. MLK3^−/− mice were protected from chronic ethanol-induced phosphorylation of MLK3 and JNK, as well as multiple indicators of liver injury, including increased ALT/AST, inflammatory cytokines, and induction of RIP3. However, ethanol-induced steatosis and hepatocyte apoptosis were not affected by MLK3. Finally, chimeric mice lacking MLK3 only in myeloid cells were also protected from chronic ethanol-induced phosphorylation of JNK, expression of inflammatory cytokines, and increased ALT/AST. MLK3 expression in myeloid cells contributes to phosphorylation of JNK, increased cytokine production, and hepatocyte injury in response to chronic ethanol. Our data suggest that myeloid MLK3 could be targeted for developing potential therapeutic strategies to suppress liver injury in ALD patients.Key words: Alcoholic liver disease (ALD), Kupffer cells, Necroptosis, Toll-like receptor 4 (TLR4), Cytokines     

Adipose tissue,liver and pancreas structural alterations in C57BL/6 mice fed high-fat-high-sucrose diet supplemented with fish oil (n-3 fatty acid rich oil)

Fish oil treatment was used in reversing the morphological and metabolic changes of C57BL/6 mice fed high-fat-high-sucrose (HFHS) diet. Two-month-old male C57BL/6 mice were fed HFHS chow or standard chow (SC). At 3 months of age, HFHS mice were separated into an untreated group (HFHS) and a group treated with fish oil (HFHS-Fo, 1.5 g/kg/day). At 4 months of age, HFHS fed mice had an increase in body mass (BM) and total body fat, when the animals were sacrificed. Both parameters were lower in HFHS-Fo than in HFHS mice. Plasma glucose and insulin levels were not affected among the groups, but HFHS and HFHS-Fo animals had higher homeostasis model assessment for insulin resistance HOMA-IR ratio. HFHS and HFHS-FO mice had increased plasma total cholesterol and LDL-C, HFHS-Fo increased plasma HDL-C and decreased triglycerides levels. The liver mass (LM) and the adipocytes’ size were larger in HFHS mice, while HFHS-Fo mice had a lower LM and smaller adipocytes. The liver steatosis and hepatocyte binucleation were increased in HFHS mice, while HFHS-Fo mice had reduced liver steatosis and hepatocyte binucleation. HFHS-Fo mice had a lower pancreas mass, while HFHS animals had higher islet pancreatic diameter. The SC group showed strong expression for insulin, glucagon and a glucose transporter type 2 GLUT-2 in all pancreatic islets, while in HFHS mice there was less expression for GLUT-2. However, HFHS-Fo mice showed an increase of GLUT-2 expression. In conclusion, dietary fish oil treatment reduces body mass and fat pad adiposity, and also by reducing plasma TG and pancreatic islet hypertrophy in mice fed high-fat-high-sucrose diet. Furthermore, fish oil improves glucagon and GLUT-2 expressions when it is decreased in insulin, but in hepatocyte binucleation and hepatic steatosis where the effect is reduced.     

Tumor necrosis factor receptor p55 is essential for intrahepatic granuloma formation and hepatocellular apoptosis in a murine model of bacterium-induced fulminant hepatitis.

Accumulating evidence implicates tumor necrosis factor (TNF) and Fas systems in liver injury, although the interaction between these two systems remains to be investigated. In this study, we examined Propionibacterium acnes-primed TNF receptor p55-deficient (TNFRp55-/-) or Fas-deficient MRL/MpJ Lpr/Lpr mice challenged with lipopolysaccharide (LPS). Priming with P. acnes caused mononuclear cell infiltration into the hepatic lobules and granuloma formation in the livers of TNFRp55 wild-type mice. Subsequent LPS challenge caused massive liver injury and a marked increase in transaminase levels, leading to acute lethality in control wild-type mice. In contrast, the same treatment caused few pathological changes in livers of TNFRp55-/- mice, and all animals survived. P. acnes and subsequent LPS challenge induced granuloma formation and apoptotic changes, respectively, in livers of MRL/MpJ Lpr/Lpr mice. However, liver injury was 50% of that in control MRL/MpJ +/+ mice, suggesting some role of the Fas-Fas ligand system in this liver injury model. On the other hand, an agonistic anti-Fas antibody caused massive apoptosis and hemorrhagic changes of the liver without any priming with P. acnes, leading to death in both TNFRp55-/- and control wild-type mice. These results suggest that TNFRp55 but not Fas was involved in P. acnes-induced granuloma formation as well as subsequent LPS-induced liver injury and that TNFRp55 and Fas independently induced apoptosis of hepatocytes in vivo.     

Blockade of Notch signaling promotes acetaminophen-induced liver injury

Liver injury after experimental acetaminophen treatment is mediated both by direct hepatocyte injury through a P450-generated toxic metabolite and indirectly by activated liver Kupffer cells and neutrophils. This study was designed to investigate the role of Notch signaling in the regulation of innate immune responses in acetaminophen (APAP)-induced liver injury. Using a mouse model of APAP-induced liver injury, wild-type (WT) and toll-like receptor 4 knockout (TLR4 KO) mice were injected intraperitoneally with APAP or PBS. Some animals were injected with γ-secretase inhibitor DAPT or DMSO vehicle. For the in vitro study, bone marrow-derived macrophages (BMMs) were transfected with Notch1 siRNA, TLR4 siRNA, and non-specific (NS) siRNA and stimulated with LPS. Indeed, paracetamol/acetaminophen-induced liver damage was worse after Notch blockade with DAPT in wild-type mice, which was accompanied by significantly increased ALT levels, diminished hairy and enhancer of split-1 (Hes1), and phosphorylated Stat3 and Akt but enhanced high mobility group box 1 (HMGB1), TLR4, NF-κB, and NLRP3 activation after APAP challenge. Mice receiving DAPT increased macrophage and neutrophil accumulation and hepatocellular apoptosis. However, TLR4 KO mice that received DAPT reduced APAP-induced liver damage and NF-κB, NLRP3, and cleaved caspase-1 activation. BMMs transfected with Notch1 siRNA reduced Hes1 and phosphorylated Stat3 and Akt but augmented HMGB1, TLR4, NF-κB, and NLRP3. Furthermore, TLR4 siRNA knockdown resulted in decreased NF-κB and NLRP3 and cleaved caspase-1 and IL-1β levels following LPS stimulation. These results demonstrate that Notch signaling regulates innate NLRP3 inflammasome activation through regulation of HMGB1/TLR4/NF-κB activation in APAP-induced liver injury. Our novel findings underscore the critical role of the Notch1-Hes1 signaling cascade in the regulation of innate immunity in APAP-triggered liver inflammation. This might imply a novel therapeutic potential for the drug-induced damage-associated lethal hepatitis.     

Increased carbon tetrachloride-induced liver injury and fibrosis in FGFR4-deficient mice

Carbon tetrachloride (CCl(4)) intoxification in rodents is a commonly used model of both acute and chronic liver injury. Recently, we showed that mice in which FGFR4 was ablated from the germline exhibited elevated cholesterol metabolism and bile acid synthesis coincident with unrepressed levels of cytochrome P450 7A (CYP7A), the rate-limiting enzyme in cholesterol disposal. Of the four fibroblast growth factor (FGF) receptor genes expressed in adult liver, FGFR4 is expressed specifically in mature hepatocytes. To determine whether FGFR4 plays a broader role in liver-specific metabolic functions, we examined the impact of both acute and chronic exposure to CCl(4) in FGFR4-deficient mice. Following acute CCl(4) exposure, the FGFR4-deficient mice exhibited accelerated liver injury, a significant increase in liver mass and delayed hepatolobular repair. Chronic CCl(4) exposure resulted in severe fibrosis in livers of FGFR4-deficient mice compared to normal mice. Analysis at both mRNA and protein levels indicated an 8-hour delay in FGFR4-deficient mice in the down-regulation of cytochrome P450 2E1 (CYP2E1) protein, the major enzyme whose products underlie CCl(4)-induced injury. These results show that hepatocyte FGFR4 protects against acute and chronic insult to the liver and prevents accompanying fibrosis. The results show that FGFR4 acts by promotion of processes that restore hepatolobular architecture rather than cellularity while limiting damage due to prolonged CYP2E1 activity.     

对氧磷酶3基因过表达对急性中毒小鼠肝损伤的保护作用及机制

目的 探讨对氧磷酶3（PON3）对急性中毒肝损伤的保护作用及作用机制。 方法 将40只Balb/c小鼠随机分为4组,正常对照（NC）组、敌敌畏（DDVT）对照组、绿色荧光蛋白慢病毒（Lv-GFP）组和重组PON3慢病毒（Lv-PON3）组,每组10只。LV-GFP组和Lv-PON3组小鼠经尾静脉分别注射2×10⁷ TU的Lv-GFP、Lv-PON3慢病毒,3 d后腹腔注射敌敌畏（DDVT）溶液9 mg/kg,DDVT组只需腹腔注射同等剂量DDVT,NC组注射同等剂量生理盐水。各组于DDVT染毒后12 h麻醉处死10只小鼠取肝组织,ELISA法检测各组血清乙酰胆碱酯酶（AChE）、肝功能指标谷丙转氨酶（ALT）和谷草转氨酶（AST）和肝组织氧化应激指标丙二醛（MDA）、过氧化氢酶（CAT）、超氧化物歧化酶（SOD）和谷胱甘肽（GSH）,HE染色法观察肝组织形态学改变,Real-time PCR检测肝组织Kelch样环氧氯丙烷相关蛋白-1（Keap1）、血红蛋白加氧酶（HO-1）和转录因子NF-E2相关因子2（Nrf2） mRNA的表达。 结果 与NC组比较,DDVT组和Lv-GFP组肝组织MDA、ALT、AST水平,HO-1和Nrf2 mRNA表达量明显升高,AChE、CAT、SOD、GSH水平和Keap1 mRNA表达量均明显降低（P<0.05）。与DDVT组比较,Lv-PON3组肝组织MDA、ALT、AST水平,Keap1 mRNA表达量降低,AChE、CAT、SOD和GSH水平均明显著升高,HO-1和Nrf2 mRNA表达量显著上升（P<0.05）。正常对照组肝细胞结构清晰,未出现肝细胞坏死和脂肪病变的情况,DDVT组和Lv-GFP组出现肝细胞坏死和脂肪变性等病理改变,Lv-PON3组肝脏病变组织肝细胞坏死和脂肪变性细胞数有一定的减少。 结论 PON3可通过Keap1-Nrf2/HO-1途径缓解氧化应激反应,改善肝功能,对敌敌畏急性中毒小鼠肝损伤有一定的保护作用。     

Central role of interleukin-6 in burn induced stimulation of hepatic amino acid transport

Burn injury produces a marked and prolonged hypermetabolic state, which is characterized by accelerated hepatic amino acid metabolism and negative nitrogen balance. The effect of burn injury on hepatocyte transport of glutamine, a key substrate in gluconeogenesis and ureagenesis, was examined following the administration of a full thickness 20% total body surface scald injury. The burn injury was induced in wild-type mice, as well as two types of knockout mice: a CD-14 knockout and an interleukin-6 (IL-6) knockout. In both the wild-type and CD-14 knockout mice a latent and profound 2-fold increase in hepatocyte glutamine transport was seen. In contrast, hepatocytes isolated from IL-6 knockout mice failed to show an induction of glutamine transport after burn injury. Hepatocytes isolated from burned wild-type and CD-14 knockout mice produced significantly higher levels of IL-6 as compared to hepatocytes isolated from sham burned mice. Histologic analysis of skin isolated from wild-type mice showed a robust inflammatory response whereas skin from IL-6 deficient mice showed minimal inflammation and decreased granulation tissue. The results of this study suggest that IL-6 may play a key role in the stimulation of hepatic glutamine transport following burn injury.