[Retracted] HCV­E2 inhibits hepatocellular carcinoma metastasis by stimulating mast cells to secrete exosomal shuttle microRNAs.

[This retracts the article DOI: 10.3892/ol.2017.6433.].

The acetylation of MDH1 and IDH1 is associated with energy metabolism in acute liver failure.

The liver is the main organ associated with metabolism. In our previous studies, we identified that the metabolic enzymes malate dehydrogenase 1 (MDH1) and isocitrate dehydrogenase 1 (IDH1) were differentially expressed in ALF. The aim of this study was to explore the changes in the acetylation of MDH1 and IDH1 and the therapeutic effect of histone deacetylase (HDAC) inhibitor in acute liver failure (ALF). Decreased levels of many metabolites were observed in ALF patients. MDH1 and IDH1 were decreased in the livers of ALF patients. The HDAC inhibitor ACY1215 improved the expression of MDH1 and IDH1 after treatment with MDH1-siRNA and IDH1-siRNA. Transfection with mutant plasmids and adeno-associated viruses, identified MDH1 K118 acetylation and IDH1 K93 acetylation as two important sites that regulate metabolism in vitro and in vivo.

Sulforaphane, an NRF2 agonist, alleviates ferroptosis in acute liver failure by regulating HDAC6 activity.

OBJECTIVE: Acute liver failure (ALF) is characterized by severe liver dysfunction, rapid progression and high mortality and is difficult to treat. Studies have found that sulforaphane (SFN), a nuclear factor E2-related factor 2 (NRF2) agonist, has anti-inflammatory, antioxidant and anticancer effects, and has certain protective effects on neurodegenerative diseases, cancer and liver fibrosis. This paper aimed to explore the protective effect of SFN in ALF and it possible mechanisms of action. METHODS: Lipopolysaccharide and D-galactosamine were used to induce liver injury in vitro and in vivo. NRF2 agonist SFN and histone deacetylase 6 (HDAC6) inhibitor ACY1215 were used to observe the protective effect and possible mechanisms of SFN in ALF, respectively. Cell viability, lactate dehydrogenase (LDH), Fe2+, glutathione (GSH) and malondialdehyde (MDA) were detected. The expression of HDAC6, NRF2, glutathione peroxidase 4 (GPX4), acyl-CoA synthetase long-chain family member 4 (ACSL4) and solute carrier family 7 member 11 (SLC7A11) were detected by Western blotting and immunofluorescence. RESULTS: Our results show that NRF2 was activated by SFN. LDH, Fe2+, MDA and ACSL4 were downregulated, while GSH, GPX4 and SLC7A11 were upregulated by SFN in vitro and in vivo, indicating the inhibitory effect of SFN on ferroptosis. Additionally, HDAC6 expression was decreased in the SFN group, indicating that SFN could downregulate the expression of HDAC6 in ALF. After using the HDAC6 inhibitor, ACY1215, SFN further reduced HDAC6 expression and inhibited ferroptosis, indicating that SFN may inhibit ferroptosis by regulating HDAC6 activity. CONCLUSION: SFN has a protective effect on ALF, and the mechanism may include reduction of ferroptosis through the regulation of HDAC6. Please cite this article as: Zhang YQ, Shi CX, Zhang DM, Zhang LY, Wang LW, Gong ZJ. Sulforaphane, an NRF2 agonist, alleviates ferroptosis in acute liver failure by regulating HDAC6 activity. J Integr Med. 2023; 21(5): 464-473.

Interventions for non-alcoholic liver disease: a gut microbial metabolites perspective.

Over the past two decades, non-alcoholic fatty liver disease (NAFLD) has become a leading burden of hepatocellular carcinoma and liver transplantation. Although the exact pathogenesis of NAFLD has not been fully elucidated, recent hypotheses placed more emphasis on the crucial role of the gut microbiome and its derivatives. Reportedly, microbial metabolites such as short-chain fatty acids, amino acid metabolites (indole and its derivatives), bile acids (BAs), trimethylamine N-oxide (TMAO), and endogenous ethanol exhibit sophisticated bioactive properties. These molecules regulate host lipid, glucose, and BAs metabolic homeostasis via modulating nutrient absorption, energy expenditure, inflammation, and the neuroendocrine axis. Consequently, a broad range of research has studied the therapeutic effects of microbiota-derived metabolites. In this review, we explore the interaction of microbial products and NAFLD. We also discuss the regulatory role of existing NAFLD therapies on metabolite levels and investigate the potential of targeting those metabolites to relieve NAFLD.

Sirtuin1 attenuates acute liver failure by reducing reactive oxygen species via hypoxia inducible factor 1α.

BACKGROUND: The occurrence and development of acute liver failure (ALF) is closely related to a series of inflammatory reactions, such as the production of reactive oxygen species (ROS). Hypoxia inducible factor 1α (HIF-1α) is a key factor that regulates oxygen homeostasis and redox, and the stability of HIF-1α is related to the ROS level regulated by Sirtuin (Sirt) family. The activation of Sirt1 will lead to a powerful antioxidant defense system and therapeutic effects in liver disease. However, little is known about the relationship between HIF-1α and Sirt1 in the process of ALF and the molecular mechanism. AIM: To investigate whether HIF-1α may be a target of Sirt1 deacetylation and what the effects on ALF are. METHODS: Mice were administrated lipopolysaccharide (LPS)/D-gal and exposed to hypoxic conditions as animal model, and resveratrol was used as an activator of Sirt1. The cellular model was established with L02 cells stimulated by LPS. N-acetyl-L-cysteine was used to remove ROS, and the expression of Sirt1 was inhibited by nicotinamide. Western blotting was used to detect Sirt1 and HIF-1α activity and related protein expression. The possible signaling pathways involved were analyzed by immunofluorescent staining, co-immunoprecipitation, dihydroethidium staining, and Western blotting. RESULTS: Compared with mice stimulated with LPS alone, the expression of Sirt1 decreased, the level of HIF-1α acetylation increased in hypoxic mice, and the levels of carbonic anhydrase 9 and Bcl-2-adenovirus E1B interacting protein 3 increased significantly, which was regulated by HIF-1α, indicating an increase of HIF-1α activity. Under hypoxia, the down-regulation of Sirt1 activated and acetylated HIF-1α in L02 cells. The inhibition of Sirt1 significantly aggravated this effect and the massive production of ROS. The regulation of ROS was partly through peroxisome proliferator-activated receptor alpha or AMP-activated protein kinase. Resveratrol, a Sirt1 activator, effectively relieved ALF aggravated by hypoxia, the production of ROS, and cell apoptosis. It also induced the deacetylation of HIF-1α and inhibited the activity of HIF-1α. CONCLUSION: Sirt1 may have a protective effect on ALF by inducing HIF-1α deacetylation to reduce ROS.

HDAC6 inhibitor ACY1215 inhibits the activation of NLRP3 inflammasome in acute liver failure by regulating the ATM/F-actin signalling pathway.

Acute liver failure (ALF) is a rare and critical medical condition. This study was designed to investigate the protective effects and underlying mechanism of ACY1215 in ALF mice. Our findings suggested that ACY1215 treatment ameliorates the pathological hepatic damage of ALF and decreases the serum levels of ALT and AST. Furthermore, ACY1215 pretreatment increased the level of ATM, γ-H2AX, Chk2, p53, p21, F-actin and vinculin in ALF. Moreover, ACY1215 inhibited the level of NLRP3, ASC, caspase-1, IL-1ß and IL-18 in ALF. The ATM inhibitor KU55933 could decrease the level of ATM, γ-H2AX, Chk2, p53, p21, F-actin and vinculin in ALF with ACY1215 pretreatment. The F-actin inhibitor cytochalasin B decreased the level of F-actin and vinculin in ALF with ACY1215 pretreatment. However, cytochalasin B had no effect on protein levels of ATM, Chk2, p53 and p21 in ALF with ACY1215 pretreatment. Cytochalasin B could dramatically increase the level of NLRP3, ASC, caspase-1, IL-1ß and IL-18 in ALF with ACY1215 pretreatment. These results indicated that ACY1215 exhibited hepatoprotective properties, which was associated with the inhibition of NLRP3 inflammasome, and this effect of ACY1215 was connected with upregulation of the ATM/F-actin mediated signalling pathways.

Betaine inhibits Toll-like receptor 4 responses and restores intestinal microbiota in acute liver failure mice.

Previous research has revealed that the gut microbiome has a marked impact on acute liver failure (ALF). Here, we evaluated the impact of betaine on the gut microbiota composition in an ALF animal model. The potential protective effect of betaine by regulating Toll-like receptor 4 (TLR4) responses was explored as well. Both mouse and cell experiments included normal, model, and betaine groups. The rat small intestinal cell line IEC-18 was used for in vitro experiments. Betaine ameliorated the small intestine tissue and IEC-18 cell damage in the model group by reducing the high expression of TLR4 and MyD88. Furthermore, the intestinal permeability in the model group was improved by enhancing the expression of the (ZO)-1 and occludin tight junction proteins. There were 509 operational taxonomic units (OTUs) that were identified in mouse fecal samples, including 156 core microbiome taxa. Betaine significantly improved the microbial communities, depleted the gut microbiota constituents Coriobacteriaceae, Lachnospiraceae, Enterorhabdus and Coriobacteriales and markedly enriched the taxa Bacteroidaceae, Bacteroides, Parabacteroides and Prevotella in the model group. Betaine effectively improved intestinal injury in ALF by inhibiting the TLR4/MyD88 signaling pathway, improving the intestinal mucosal barrier and maintaining the gut microbiota composition.

The Beneficial Roles of SIRT1 in Neuroinflammation-Related Diseases.

Sirtuins are the class III of histone deacetylases whose deacetylate of histones is dependent on nicotinamide adenine dinucleotide (NAD+). Among seven sirtuins, SIRT1 plays a critical role in modulating a wide range of physiological processes, including apoptosis, DNA repair, inflammatory response, metabolism, cancer, and stress. Neuroinflammation is associated with many neurological diseases, including ischemic stroke, bacterial infections, traumatic brain injury, Alzheimer's disease (AD), and Parkinson's disease (PD). Recently, numerous studies indicate the protective effects of SIRT1 in neuroinflammation-related diseases. Here, we review the latest progress regarding the anti-inflammatory and neuroprotective effects of SIRT1. First, we introduce the structure, catalytic mechanism, and functions of SIRT1. Next, we discuss the molecular mechanisms of SIRT1 in the regulation of neuroinflammation. Finally, we analyze the mechanisms and effects of SIRT1 in several common neuroinflammation-associated diseases, such as cerebral ischemia, traumatic brain injury, spinal cord injury, AD, and PD. Taken together, this information implies that SIRT1 may serve as a promising therapeutic target for the treatment of neuroinflammation-associated disorders.

TNF-α/HMGB1 inflammation signalling pathway regulates pyroptosis during liver failure and acute kidney injury.

OBJECTIVE: Acute kidney injury (AKI) is a common complication of acute liver failure (ALF). Pyroptosis is a necrosis type related to inflammation. This study aimed to investigate the role of TNF-α/HMGB1 pathway in pyroptosis during ALF and AKI. METHODS: An ALF and AKI mouse model was generated using LPS/D-Gal, and a TNF-α inhibitor, CC-5013, was used to treat the mice. THP-1 cells were induced to differentiate into M1 macrophages, then challenged with either CC-5013 or an HMGB1 inhibitor, glycyrrhizin. pLVX-mCMVZsGreen-PGK-Puros plasmids containing TNF-α wild-type (WT), mutation A94T of TNF-α and mutation P84L of TNF-α were transfected into M1 macrophages. RESULTS: Treatment with CC-5013 decreased the activation of TNF-α/HMGB1 pathway and pyroptosis in the treated mice and cells compared with the control mice and cells. CC-5013 also ameliorated liver and kidney pathological changes and improved liver and renal functions in treated mice, and the number of M1 macrophages in the liver and kidney tissues also decreased. The activation of TNF-α/HMGB1 pathway and pyroptosis increased in the M1 macrophage group compared with the normal group. Similarly, the activation of TNF-α/HMGB1 pathway and pyroptosis in the LPS + WT group also increased. By contrast, the activation of the TNF-α/HMGB1 pathway and pyroptosis decreased in the LPS + A94T and LPS + P84L groups. Moreover, glycyrrhizin inhibited pyroptosis. CONCLUSION: The TNF-α/HMGB1 inflammation signalling pathway plays an important role in pyroptosis during ALF and AKI.

RNA-Seq Revealed a Circular RNA-microRNA-mRNA Regulatory Network in Hantaan Virus Infection.

Hantaan virus (HTNV), a Hantavirus serotype that is prevalent in Asia, causes hemorrhagic fever with renal syndrome (HFRS) with high mortality in human race. However, the pathogenesis of HTNV infection remains elusive. Circular RNAs (circRNAs), a new type of non-coding RNAs, play a crucial role in various pathogenic processes. Nevertheless, circRNA expression profiles and their effects on pathogenesis of HTNV infection are still completely unknown. In the present study, RNA sequencing was performed to analyze the circRNA, microRNA (miRNA), and mRNA expression profiles in HTNV-infected and mock-infected human umbilical vein endothelial cells (HUVECs). A total of 70 circRNAs, 66 miRNAs, and 788 mRNAs were differently expressed. Several differentially expressed RNAs were validated by RT-qPCR. Moreover, we verified that some differentially expressed RNAs, such as circ_0000479, miR-149-5p, miR-330-5p, miR-411-3p, RIG-I, CMPK2, PARP10, and GBP1, promoted or inhibited HTNV replication. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis demonstrated that the host genes of differentially expressed circRNAs were principally involved in the innate immune response, the type I interferon (IFN) signaling pathway, and the cytokine-mediated signaling pathway. Additionally, the circRNA-miRNA-mRNA regulatory network was integrally analyzed. The data showed that there were many circRNA-miRNA-mRNA interactions in HTNV infection. By dual-luciferase reporter assay, we confirmed that circ_0000479 indirectly regulated RIG-I expression by sponging miR-149-5p, hampering viral replication. This study for the first time presents a comprehensive overview of circRNAs induced by HTNV and reveals that a network of enriched circRNAs and circRNA-associated competitive endogenous RNAs (ceRNAs) is involved in the regulation of HTNV infection, thus offering new insight into the mechanisms underlying HTNV-host interaction.

Histone deacetylase 6 inhibitor ACY1215 offers a protective effect through the autophagy pathway in acute liver failure.

AIM: The purpose of the present study was to elucidate the protective effect of histone deacetylase 6 inhibitor ACY1215 on autophagy pathway in acute liver failure (ALF). MAIN METHODS: Lipopolysaccharide (LPS) and d-galactosamine (D-Gal) were used to induce ALF model in C57BL/6 mice. D-Gal and tumor necrosis factor alpha (TNF-α) were applied in L02 cell. Autophagy inhibitor 3-MA and ACY1215 were conducted to induce 3-MA group, ACY1215 group and ACY1215+3-MA group. RESULTS: ACY1215 improved liver histological and functional changes in ALF mice model, whereas the autophagy inhibitor 3-MA aggravated liver tissue pathological and functional damage in ALF mice model group. The apoptotic levels (including apoptotic index/rate and apoptotic proteins) in ALF mice and L02 cell were ameliorated with treatment ACY1215. 3-MA accentuated the apoptotic levels in ACY1215 group. D-Gal/TNF-α could reduce L02 cell mitochondrial membrane potential (ΔΨm) in control group. ACY1215 increased the ΔΨm in ALF model. 3-MA also further reduced the ΔΨm in ACY1215 group. ACY1215 could induce autophagy in ALF mice and cell model group accompanied with an increase in expression of LC3-II and beclin-1 proteins and down-regulation of p62 protein. Moreover, the expression of LC3-II and beclin1 proteins were greatly reduced and the expression of p62 protein was ascended after intervention with 3-MA in ACY1215 group. SIGNIFICANCE: Histone deacetylase 6 inhibitor ACY1215 could protect acute liver failure mice and L02 cell by inhibiting apoptosis pathway through enhancing autophagy way.

Mechanism of glycyrrhizin on ferroptosis during acute liver failure by inhibiting oxidative stress.

The present study aimed to investigate the anti­ferroptosis effects of the HMGB1 inhibitor glycyrrhizin (GLY). The present study used a cell and animal model of acute liver failure (ALF), induced using tumor necrosis factor­α, lipopolysaccharide and D­galactosamine, to investigate the effects of GLY. The expression of glutathione peroxidase 4 (GPX4) and high mobility group protein B1 (HMGB1), heme oxygenase­1 (HO­1) and nuclear factor erythroid 2­related factor 2 (Nrf2) were detected were detected by western blotting in L02 hepatocytes and mouse liver. The expression of GPX4 and HMGB1 in L02 hepatocytes and mouse liver was detected by immunofluorescence. The pathological changes to liver tissues were determined by hematoxylin and eosin staining. The levels of lactate dehydrogenase (LDH), Fe2+, reactive oxygen species (ROS) and glutathione (GSH) were tested using kits. Compared with the normal group, the degree of liver damage and liver function in the model animal group was severe. The protein levels of HMGB1 in L02 cells and liver tissues were significantly increased. The expression of NRF2, HO­1 and GPX4 was significantly decreased. The levels of LDH, Fe2+, malondialdehyde (MDA) and ROS were increased, whereas the level of GSH was decreased. Treatment with GLY reduced the degree of liver damage, the expression of HMGB1 was decreased, and the levels of Nrf2, HO­1 and GPX4 were increased. The levels of LDH, Fe2+, MDA, ROS were decreased, while the level of GSH was increased by GLY treatment. The results of the present study indicated that HMGB1 is involved in the process of ferroptosis. The HMGB1 inhibitor GLY significantly reduced the degree of ferroptosis during ALF by inhibiting oxidative stress.

Inhibition of HDAC6 attenuates LPS-induced inflammation in macrophages by regulating oxidative stress and suppressing the TLR4-MAPK/NF-κB pathways.

BACKGROUND: Histone deacetylase 6 (HDAC6) has been considered as an important regulator in the development of inflammatory diseases. However, the mechanism of HDAC6 in regulating inflammatory responses has not been fully determined. In the present study, we aim to investigate the role and mechanisms of HDAC6 in regulating inflammation in lipopolysaccharide (LPS)-activated macrophages. METHODS: Flow cytometry was used to determine a suitable treatment dosage of ACY-1215 on lipopolysaccharide (LPS)-activated macrophages for the present study. The RAW264.7 macrophages were divided into normal, LPS-treated, and ACY-1215 treated groups, respectively. For the ACY-1215 group, ACY-1215 (10 µM) was added to the medium 2 h prior to treatment with LPS (1 µg/ml) for 24 h. In this study, ROS, inflammatory cytokines, the ultrastructure of mitochondria, mitochondrial membrane potential, RNA and protein expression assay were detected respectively. Subsequently, the effect of HDAC6 knockdown on inflammatory response in LPS-activated RAW264.7 macrophages was also detected. RESULTS: Inhibition of HDAC6 inhibited the overproduction of ROS and suppressed the expression of pro-inflammatory cytokines such as TNF-α, IL-1ß, and IL-6 in LPS-activated RAW264.7 cells. Pretreatment with ACY-1215 could normalize the ultrastructure of mitochondria and mitochondrial membrane potential in LPS-activated macrophages. Moreover, the protein expression of TLR4, Nrf2, HO-1 and the activation of MAPK and NF-κB signaling pathways were normalized by the inhibition of HDAC6. CONCLUSIONS: Inhibition of HDAC6 exhibited protective role against LPS-induced inflammation in RAW264.7 cells by regulating oxidative stress and suppressing the activation of TLR4- MAPK/NF-κB signaling pathway.

Modulations of Histone Deacetylase 2 Offer a Protective Effect through the Mitochondrial Apoptosis Pathway in Acute Liver Failure.

The purpose of this study was to investigate the modulation of histone deacetylase 2 (HDAC2) on mitochondrial apoptosis in acute liver failure (ALF). The cellular model was established with LO2 cells stimulated by tumor necrosis factor alpha (TNF-α)/D-galactosamine (D-gal). Rats were administrated by lipopolysaccharide (LPS)/D-gal as animal model. The cell and animal models were then treated by HDAC2 inhibitor CAY10683. HDAC2 was regulated up or down by lentiviral vector transfection in LO2 cells. The mRNA levels of bcl2 and bax were detected by real-time PCR. The protein levels of HDAC2, bcl2, bax, cytochrome c (cyt c) in mitochondrion and cytosol, apoptosis protease activating factor 1 (apaf1), caspase 3, cleaved-caspase 3, caspase 9, cleaved-caspase 9, acetylated histone H3 (AH3), and histone H3 (H3) were assayed by western blot. Apoptosis was detected by flow cytometry. The serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (TBIL) levels were also assayed. The openness degree of the mitochondrial permeability transition pore (MPTP) was detected by ultraviolet spectrophotometry. The apoptosis of hepatocytes in liver tissues was determined by tunnel staining. The liver tissue pathology was detected by hematoxylin eosin (HE) staining. The ultrastructure of liver tissue was observed by electron microscopy. Compared with cell and rat model groups, the bax mRNA level was decreased, and bcl2 mRNA was increased in the CAY10683 treatment group. The protein levels of HDAC2, bax, cyt c in cytosol, apaf1, cleaved-caspase 3, and cleaved-caspase 9 were decreased, and the apoptosis rate was decreased (P < 0.05), whereas the protein level of bcl2 and cyt c in the mitochondrion was elevated (P < 0.05) in the CAY10683 treatment group. In the HDAC2 down- or upregulated LO2 cells, the mitochondrial apoptosis pathway was inhibited or activated, respectively. After being treated with TNF-α/D-gal in HDAC2 down- or upregulated LO2 cells, the mitochondrial apoptosis pathway was further suppressed or activated, respectively. The MPTP value was elevated in CAY10683-treated groups compared with the rat model group (P < 0.05). Liver tissue pathological damage and apoptotic index in the CAY10683-treated group were significantly reduced. In addition, AH3 was elevated in both cell and animal model groups (P < 0.05). Downregulated or overexpressed HDAC2 could accordingly increase or decrease the AH3 level, and TNF-α/D-gal could enhance the acetylation effect. These results suggested that modulations of histone deacetylase 2 offer a protective effect through the mitochondrial apoptosis pathway in acute liver failure.

Protective role of AGK2 on thioacetamide-induced acute liver failure in mice.

AIMS: The aim of the present study was to investigate the protective effects of AGK2 as a selective SIRT2 inhibitor on thioacetamide (TAA)-induced acute liver failure (ALF) in mice and its potential mechanism. MAIN METHODS: All male C57BL/6 mice were separated into control, TAA, AGK2 + TAA, and AGK2 groups. The histological changes were observed by hematoxylin and eosin (HE) staining. The apoptosis cells of liver tissues were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were used to evaluate the damage of liver function. The inflammatory cytokines of iNOS, TNF-α, IL-1ß was detected by Western blotting and RT-PCR assay. The expression of mitogen-activated protein kinase (MAPK), NF-κB, and apoptosis pathways was determined by Western blotting. KEY FINDINGS: AGK2 improved the damage of TAA-induced liver pathology and function. AGK2 pretreatment also reduced the levels of pro-inflammatory cytokines in ALF liver tissues. AGK2 improved the TAA-induced survival rate. Moreover, AGK2 administration suppressed the increase of phosphorylation NF-κB-p65 and the activation of MAPK pathway. In addition, pretreatment alleviated TAA-induced the liver cells apoptosis. SIGNIFICANCE: AGK2 improve TAA-induced survival rate in mice with ALF, suppress the inflammatory responses by inhibition of MAPK and NF-κB signaling pathways, and decrease the hepatocyte necrosis by inhibition of apoptosis. Pharmacologic inhibition of SIRT2 may be a promising approach for the treatment of ALF.

HDAC2 inhibitor CAY10683 reduces intestinal epithelial cell apoptosis by inhibiting mitochondrial apoptosis pathway in acute liver failure.

INTRODUCTION: Histone deacetylases (HDAC) inhibitor has the effect of anti-tumor and inhibiting apoptosis, and could inhibit the release of inflammatory factors, reducing the damage to liver and enterocytes in acute liver failure (ALF). HDAC2 specific inhibitor CAY10683 was used to verify the protective effect on acute liver failure through reducing intestinal epithelial cell apoptosis by inhibiting mitochondrial apoptosis pathway. MATERIALS AND METHODS: Lipopolysaccharide/D-galactosamine (LPS/D-GalN) was used to induce ALF in Sprague-Dawley rats. A total of 18 healthy rats were randomly divided into three groups. Rats in ALF group and CAY10683 group were given the same amount of normal sodium or CAY10683 2 hours before ALF model protocol was conducted. NCM460 cells were given LPS/D-GalN to establish an apoptotic model. Flow cytometry analysis was used to determine the apoptosis of enterocytes, and TUNEL assay was used to observe the apoptosis of NCM460 cells. The expression of bax was observed by immunofluorescence. The expression of histone proteins, HDAC2 and molecules in the apoptotic signaling pathway were determined by Western blotting and real-time PCR. RESULTS: CAY10683 improves histological and functional changes in ALF model. Compared with control group, LPS/D-GalN induced massive apoptosis of rat intestinal tissues and NCM460 cells (P<0.05), and the apoptosis rate was significantly reduced after CAY10683 treatment (P<0.05). The expression of bax was increased significantly in the model groups (P<0.05), and reduced with the treatment of CAY10683 (P<0.05). Compared with the model group, CAY10683 inhibits mitochondrial apoptosis in intestinal tissues and NCM460 cells (P<0.05). CONCLUSION: CAY10683 reduces the damage to liver and intestinal tissue, and plays an important role in inhibiting mitochondrial apoptosis in ALF rats and in NCM460 cells.

Regulation of Acetylation in High Mobility Group Protein B1 Cytosol Translocation.

High mobility group protein B1 (HMGB1) is a nonhistone that mainly binds to nucleus DNA. As an important late inflammatory transmitter, extracellular HMGB1 is involved in the inflammatory immune response, tumor growth, infiltration, and metastasis. HMGB1 is actively released by activated inflammatory cells or passively released by necrotic cells. Then the released extracellular HMGB1 further induces monocytes/macrophages, neutrophils, and dendritic cells to secrete inflammatory cytokines. Therefore, HMGB1 can not only act as a proinflammatory factor to directly involve in tissue damage, but also acts as an inflammatory medium to aggravate the inflammatory cascade reaction. Studies have shown that the post-translational modification (PTM) participated in the process of HMGB1 cytosol translocation and extracellular release. The acetylation modification is the most common PTM for localization sequence of HMGB1, and the affinity of HMGB1 to DNA depends on the degree of acetylation for HMGB1. The acetylation can weaken the binding of HMGB1 to DNA, which means less HMGB1 cytosol translocation and extracellular release. This article reviews the acetylation regulation mechanisms of cytosol translocation and extracellular release of HMGB1 and provides a therapeutic strategy for controlling HMGB1-induced inflammatory responses in the future.

Histone deacetylases and acetylated histone H3 are involved in the process of hepatitis B virus DNA replication.

AIMS: The aim of this study was to investigate the relationship between anti-HBV treatment and the regulation of HDACs during HBV DNA replication. METHODS: HDAC activities and HBV DNA levels in CHB patients' sera were measured and correlation analysis was made. The changes of HDAC2, HDAC6, AH3 and histone H3 levels in normal control and 4 CHB patient liver tissue samples before and after antiviral treatment were examined. The HDAC inhibitor, TSA, anti-HBV agents, ETV and IFN-α were used to stimulate HepG2.2.15 cells. The levels of HBV DNA, pgRNA in supernatants, and cccDNA in the cells were determined by PCR. The HDAC activity, HDAC6, HDAC2, AH3 and H3 protein levels in cells were tested at days 3, 6, and 9 after treatments. KEY FINDINGS: HDAC activity was positively correlated with HBV DNA in the HBV patients' sera. The levels of HDAC2, HDAC6 and AH3 were notably decreased after antiviral treatment. When compared with antiviral treatment group, the normal liver tissue showed obviously decreased HDAC2, HDAC6 and AH3 protein levels. In vitro study, the level of HBV DNA, the HDAC activity, and the HDAC2, HDAC6 and AH3 protein levels decreased in the ETV, IFN-α and TSA groups compared with the control group. The pgRNA level in supernatants was declined in the IFN-α group and increased in the ETV and TSA groups. cccDNA expression was suppressed by IFN-α. SIGNIFICANCE: The changes of HBV replicative products during antiviral treatment are associated with histone deacetylation. Acetylated histone H3 is involved in the process of hepatitis B virus DNA replication.

Histone deacetylase inhibitor suberoylanilide hydroxamic acid alleviates liver fibrosis by suppressing the transforming growth factor-ß1 signal pathway.

BACKGROUND: Histone deacetylases (HDACs) inhibitors are new anti-fibrotic drugs that inhibit the activity of hepatic stellate cells. The present study focused on the anti-fibrotic function of HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) by suppressing transforming growth factor-ß1 (TGF-ß1) signaling. METHODS: Male Sprague-Dawley rats were used to induce liver fibrosis with carbon tetrachloride (CCl4) and LX2 cell (human hepatic stellate cell line) was stimulated by TGF-ß1. Both animals and cells were treated with SAHA. The Smad7 and connective tissue growth factor (CTGF) mRNA levels were detected by real-time polymerase chain reaction (PCR). Western blotting was used to examine the protein levels of CTGF, Histone H3 (H3), Smad7, Smad2/3, Acetyl-Histone H3 (AH3), HDAC2, α-smooth muscle actin (α-SMA), HDAC6, p-Smad2/3 and HDAC8. In addition, the TGF-ß1 and liver enzyme levels from rat serum were detected. Histopathological changes were examined by hematoxylin and eosin (HE), Sirius red and Masson trichrome staining. The α-SMA expression was detected by immumohistochemical staining. RESULTS: Compared with control group, the TGF-ß1 and liver enzyme levels from rat serum, together with the mRNA levels of CTGF and protein levels of CTGF, HDAC2, α-SMA, HDAC6, p-Smad2/3 and HDAC8 were elevated in fibrotic rats (P < 0.01). But the Smad7 mRNA and AH3 protein levels were notably suppressed in the fibrotic rats (P < 0.01). Pathological examination showed the typical changes of liver fibrosis in the fibrotic rats. After the treatment with SAHA, the levels of liver enzymes, TGF-ß1, CTGF, HDAC2, α-SMA, HDAC6, p-Smad2/3 and HDAC8 were reduced (P < 0.01) and Smad7 and AH3 protein contents were elevated in liver fibrotic rats (P < 0.01). Moreover, immumohistochemistry showed that SAHA significantly suppressed the α-SMA protein content in fibrotic liver (P < 0.01). CONCLUSION: The HDAC inhibitor SAHA alleviated liver fibrosis by suppressing the TGF-ß1 signaling.

Long noncoding RNA LNC473 inhibits the ubiquitination of survivin via association with USP9X and enhances cell proliferation and invasion in hepatocellular carcinoma cells.

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death worldwide. Recent studies reported that lncRNA LINC00473 (LNC473) was involved in cancer progression. However, the clinical significance and functional role of LNC473 in HCC progression is still unknown. In the present study, we found that the LNC473 expression was markedly elevated in HCC tissues and correlated with bigger tumor size, higher BCLC stage, vascular invasion and poor prognosis. Gain- and loss-of-function assay showed that LNC473 enhanced HCC cell proliferation and invasion and induced epithelial-mesenchymal transition (EMT) process. Mechanistically, LNC473 associated with oncoprotein survivin and regulates its stability. Moreover, LNC473 could recruit deubiquitinase USP9X to inhibit the ubiquitination level of survivin and then increase survivin expression. Therefore, our results suggest that LNC473 exerts its functions as an oncogene in HCC progression and may be a therapeutic target for HCC treatment.