trichostatin-a and Sepsis

Sepsis is a syndrome of life-threatening organ dysfunction caused by dysregulated host responses to infection. Macrophage polarization is a key process involved in the pathogenesis of sepsis. Recent evidence has demonstrated that autophagy participates in the regulation of macrophage polarization in different phases of inflammation. Here, we investigated whether trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, promotes the macrophage M2 phenotype by enhancing autophagy to counteract excessive inflammation in a cecal ligation and puncture (CLP) mouse model. TSA stimulation increased the proportions of M2 marker (CD206, CD124 and CD23)-labeled RAW264.7 macrophages. Furthermore, with increasing TSA doses, autophagy was enhanced gradually. Interestingly, the autophagy activator rapamycin (Rap), also known as an mTOR inhibitor, unexpectedly decreased the proportions of M2 marker-labeled macrophages. However, TSA treatment reversed the Rap-induced decreases in CD206-labeled macrophages. Next, we stimulated different groups of RAW264.7 cells with the autophagy inhibitors MHY1485 or 3-methyladenine (3-MA). Inhibition of autophagy at any stage in the process suppressed TSA-induced macrophage M2 polarization, but the effect was not associated with mTOR activity. In vivo, TSA administration promoted peritoneal macrophage M2 polarization, increased LC3 II expression, attenuated sepsis-induced organ (lung, liver and kidney) injury, and altered systemic inflammatory cytokine secretion. However, 3-MA abolished the protective effects of TSA in CLP mice and decreased the number of M2 peritoneal macrophages. Therefore, TSA promotes the macrophage M2 phenotype by enhancing autophagy to reduce systemic inflammation and ultimately improves the survival of mice with polymicrobial sepsis.

Topics: Animals; Autophagy; Biomarkers; Cell Line; Cytokines; Disease Models, Animal; Histone Deacetylase Inhibitors; Hydroxamic Acids; Inflammation; Ligation; Lung; Macrophage Activation; Macrophages, Peritoneal; Male; Mice; Mice, Inbred C57BL; Phenotype; Punctures; RAW 264.7 Cells; Sepsis

Histone deacetylase (HDAC) inhibitors have been recognized as promising approaches to the treatment of various human diseases including cancer, inflammation, neurodegenerative diseases, and metabolic disorders. Several pan-HDAC inhibitors are currently approved only as anticancer drugs. Interestingly, SAHA (vorinostat), one of clinically available pan-HDAC inhibitors, shows an anti-inflammatory effect at concentrations lower than those required for inhibition of tumor cell growth. It was also reported that HDAC6 selective inhibitor tubastatin A has anti-inflammatory and anti-rheumatic effect. In our efforts to develop novel HDAC inhibitors, we rationally designed various HDAC inhibitors based on the structures of two hit compounds identified by virtual screening of chemical database. Among them, 9a ((E)-N-hydroxy-4-(2-styrylthiazol-4-yl)butanamide) was identified as a HDAC6 selective inhibitor (IC50 values of 0.199 μM for HDAC6 versus 13.8 μM for HDAC1), and it did not show significant cytotoxicity against HeLa cells. In vivo biological evaluation of 9a was conducted on a lipopolysaccharide (LPS)-induced mouse model of sepsis. The compound 9a significantly improved 40% survival rate (P = 0.0483), and suppressed the LPS-induced increase of TNF-α and IL-6 mRNA expression in the liver of mice. Our study identified novel HDAC6 selective inhibitor 9a, which may serve as a potential lead for the development of anti-inflammatory or anti-sepsis agents.

Topics: Animals; Cell Line, Tumor; Computer-Aided Design; Drug Design; HeLa Cells; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Male; Mice; Molecular Docking Simulation; Protein Conformation; Sepsis

Acute lung injury (ALI) during sepsis is characterized by bilateral alveolar infiltrates, lung edema and respiratory failure. Here, we examined the efficacy the DNA methyl transferase (DNMT) inhibitor 5-Aza 2-deoxycytidine (Aza), the histone deacetylase (HDAC) inhibitor Trichostatin A (TSA), as well as the combination therapy of Aza and TSA (Aza+TSA) provides in the protection of ALI. In LPS-induced mouse ALI, post-treatment with a single dose of Aza+TSA showed substantial attenuation of adverse lung histopathological changes and inflammation. Importantly, these protective effects were due to substantial macrophage phenotypic changes observed in LPS-stimulated macrophages treated with Aza+TSA as compared with untreated LPS-induced macrophages or LPS-stimulated macrophages treated with either drug alone. Further, we observed significantly lower levels of pro-inflammatory molecules and higher levels of anti-inflammatory molecules in LPS-induced macrophages treated with Aza+TSA than in LPS-induced macrophages treated with either drug alone. The protection was ascribed to dual effects by an inhibition of MAPK-HuR-TNF and activation of STAT3-Bcl2 pathways. Combinatorial treatment with Aza+TSA reduces inflammation and promotes an anti-inflammatory M2 macrophage phenotype in ALI, and has a therapeutic potential for patients with sepsis.

Topics: Acute Lung Injury; Animals; Azacitidine; Decitabine; Drug Combinations; Endotoxemia; Epigenesis, Genetic; Histone Deacetylases; Humans; Hydroxamic Acids; Inflammation; Lipopolysaccharides; Macrophages; Methyltransferases; Mice; Sepsis; Signal Transduction

Sepsis-associated encephalopathy (SAE), which associates with neuronal apoptosis and cognitive disorders, is a common complication of systemic sepsis. However, the mechanism involving its modulation remains to be elucidated. Recent studies showed that histone deacetylases (HDACs) were implicated in neurodegeneration and cognitive functions. The current study was designed to investigate whether septic brain is epigenetically modulated by HDACs, using cecal ligation and peroration (CLP) rats and primary hippocampal neuronal cultures. We found that hippocampal acetylated histone 3 (AcH3), acetylated histone 4 (AcH4), cytoplasmic HDAC4 and Bcl-XL were inhibited in septic brain. Hippocampal Bax and nuclear HDAC4 expressions were enhanced in CLP rats. Administration of HDACs inhibitor, trichostatin A (TSA) or suberoylanilide hydroxamic acid (SAHA) rescued the changes of Bcl-XL and Bax in vivo, and decreased apoptotic cells in vitro. In addition, HDAC4 shRNA transfection significantly enhanced AcH3, AcH4 and Bcl-XL, but suppressed Bax. Neuronal apoptosis was also reduced by transfection of HDAC4 shRNA. Furthermore, CLP rats exhibited significant spatial learning and memory deficits, which could be ameliorated by application of TSA or SAHA without influence on locomotive activity. These results reveal that epigenetic modulation is involved in septic brain, and the inhibition of HDACs may serve as a potential therapeutic approach for SAE treatment.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; bcl-X Protein; Brain Diseases; Cells, Cultured; Cognition Disorders; Epigenesis, Genetic; Histone Deacetylase Inhibitors; Histone Deacetylases; Hydroxamic Acids; Male; Memory Disorders; Neurons; Rats; Rats, Sprague-Dawley; Sepsis; Space Perception; Vorinostat

The histone deacetylase (HDAC) inhibitors have emerged as the useful reagents that epigenetically modulate the expression of various genes. In the present study, the effects of HDAC inhibitors on the expression of inflammation-related genes and lung injury during sepsis were investigated.. Mice were pretreated with two structurally unrelated HDAC inhibitors, Trichostatin A (TSA) and sodium butyrate (SB). Thirty minutes later, mice underwent cecal ligation and puncture (CLP)-induced sepsis. Lung injury and the expression of inflammation-related molecules were determined. In addition, survival was assessed post-CLP.. Our results indicated that administration of TSA or SB alleviated sepsis-induced lung injury. This was accompanied by reduced neutrophil infiltration, decreased intercellular adhesion molecule-1 (ICAM-1) and E-selectin expression in lung tissue, and lower interleukin-6 (IL-6) level in plasma. In addition, treatment with HDAC inhibitors significantly prolonged the survival time of CLP mice.. These data indicated that the HDAC inhibitors, based on modulating the key enzymes linked to acetylation modification, effectively attenuate intrapulmonary inflammatory response, thus significantly alleviating lung injury during sepsis.

Topics: Acetylation; Acute Lung Injury; Animals; Butyrates; Cecum; E-Selectin; Histone Deacetylase Inhibitors; Hydroxamic Acids; Intercellular Adhesion Molecule-1; Interleukin-6; Ligation; Lung; Male; Mice; Mice, Inbred C57BL; Punctures; Sepsis

Muscle wasting during sepsis is in part regulated by glucocorticoids. In recent studies, treatment of cultured muscle cells in vitro with dexamethasone upregulated expression and activity of p300, a histone acetyl transferase (HAT), and reduced expression and activity of the histone deacetylases-3 (HDAC3) and -6, changes that favor hyperacetylation. Here, we tested the hypothesis that sepsis and glucocorticoids regulate p300 and HDAC3 and -6 in skeletal muscle in vivo. Because sepsis-induced metabolic changes are particularly pronounced in white, fast-twitch skeletal muscle, most experiments were performed in extensor digitorum longus muscles. Sepsis in rats upregulated p300 mRNA and protein levels, stimulated HAT activity, and reduced HDAC6 expression and HDAC activity. The sepsis-induced changes in p300 and HDAC expression were prevented by the glucocorticoid receptor antagonist RU38486. Treatment of rats with dexamethasone increased expression of p300 and HAT activity, reduced expression of HDAC3 and -6, and inhibited HDAC activity. Finally, treatment with the HDAC inhibitor trichostatin A resulted in increased muscle proteolysis and expression of the ubiquitin ligase atrogin-1. Taken together, our results suggest for the first time that sepsis-induced muscle wasting may be regulated by glucocorticoid-dependent hyperacetylation caused by increased p300 and reduced HDAC expression and activity. The recent development of pharmacological HDAC activators may provide a novel avenue to prevent and treat muscle wasting in sepsis and other catabolic conditions.

Topics: Animals; Dexamethasone; Disease Models, Animal; Down-Regulation; E1A-Associated p300 Protein; Gene Expression Regulation, Enzymologic; Glucocorticoids; Histone Deacetylase 6; Histone Deacetylase Inhibitors; Histone Deacetylases; Hormone Antagonists; Hydroxamic Acids; Male; Mifepristone; Muscle Proteins; Muscle, Skeletal; Muscular Atrophy; Rats; Rats, Sprague-Dawley; Receptors, Glucocorticoid; RNA, Messenger; Sepsis; Sirtuin 1; SKP Cullin F-Box Protein Ligases; Time Factors; Tripartite Motif Proteins; Ubiquitin-Protein Ligases; Up-Regulation

Interaction of pathogens with cells of the immune system results in activation of inflammatory gene expression. This response, although vital for immune defence, is frequently deleterious to the host due to the exaggerated production of inflammatory proteins. The scope of inflammatory responses reflects the activation state of signalling proteins upstream of inflammatory genes as well as signal-induced assembly of nuclear chromatin complexes that support mRNA expression. Recognition of post-translationally modified histones by nuclear proteins that initiate mRNA transcription and support mRNA elongation is a critical step in the regulation of gene expression. Here we present a novel pharmacological approach that targets inflammatory gene expression by interfering with the recognition of acetylated histones by the bromodomain and extra terminal domain (BET) family of proteins. We describe a synthetic compound (I-BET) that by 'mimicking' acetylated histones disrupts chromatin complexes responsible for the expression of key inflammatory genes in activated macrophages, and confers protection against lipopolysaccharide-induced endotoxic shock and bacteria-induced sepsis. Our findings suggest that synthetic compounds specifically targeting proteins that recognize post-translationally modified histones can serve as a new generation of immunomodulatory drugs.

Topics: Acetylation; Animals; Anti-Inflammatory Agents; Benzodiazepines; Cells, Cultured; Epigenomics; Gene Expression Regulation; Genome-Wide Association Study; Heterocyclic Compounds, 4 or More Rings; Histone Deacetylase Inhibitors; Hydroxamic Acids; Inflammation; Kaplan-Meier Estimate; Lipopolysaccharides; Macrophages; Mice; Mice, Inbred C57BL; Models, Molecular; Protein Serine-Threonine Kinases; Protein Structure, Tertiary; Salmonella Infections; Salmonella typhimurium; Sepsis; Shock, Septic