1 1826 149 EFFECTS OF HISTONE DEACETYLASE INHIBITOR ON EXTRACELLULAR MATRIX PRODUCTION IN HUMAN NASAL POLYP ORGAN CULTURES. BACKGROUND: NASAL POLYPOSIS IS ASSOCIATED WITH A CHRONIC INFLAMMATORY CONDITION OF THE SINONASAL MUCOSA AND INVOLVES MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX (ECM) ACCUMULATION. EPIGENETIC MODULATION BY HISTONE DEACETYLASE (HDAC) INHIBITORS INCLUDING TRICHOSTATIN A (TSA) HAS BEEN REPORTED TO HAVE INHIBITORY EFFECTS ON MYOFIBROBLAST DIFFERENTIATION IN LUNG AND RENAL FIBROBLASTS. THE PURPOSE OF THIS STUDY WAS TO INVESTIGATE THE INHIBITORY EFFECT OF TSA ON MYOFIBROBLAST DIFFERENTIATION AND ECM PRODUCTION IN NASAL POLYP ORGAN CULTURES. METHODS: NASAL POLYP TISSUES FROM 18 PATIENTS WERE ACQUIRED DURING ENDOSCOPIC SINUS SURGERY. AFTER ORGAN CULTURE, NASAL POLYPS WERE STIMULATED WITH TGF-BETA1 AND THEN TREATED WITH TSA. ALPHA-SMOOTH MUSCLE ACTIN (ALPHA-SMA), FIBRONECTIN, AND COLLAGEN TYPE I EXPRESSION LEVELS WERE EXAMINED BY REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION (PCR), REAL-TIME PCR, WESTERN BLOT, AND IMMUNOFLUORESCENT STAINING. HDAC2, HDAC4, AND ACETYLATED H4 EXPRESSION LEVELS WERE ASSAYED BY WESTERN BLOT. CYTOTOXICITY WAS ANALYZED BY THE TERMINAL DEOXYNUCLEOTIDYL TRANSFERASE BIOTIN-DUTP NICK END LABELING ASSAY. RESULTS: THE EXPRESSION LEVELS OF ALPHA-SMA, FIBRONECTIN, AND COLLAGEN TYPE 1 WERE INCREASED IN NASAL POLYP AFTER TRANSFORMING GROWTH FACTOR (TGF) BETA1 TREATMENT. TSA-INHIBITED TGF-BETA1 INDUCED THESE GENE AND PROTEIN EXPRESSION LEVELS. FURTHERMORE, TSA SUPPRESSED PROTEIN EXPRESSION LEVELS OF HDAC2 AND HDAC4. HOWEVER, TSA INDUCED HYPERACETYLATION OF HISTONES H4. TREATMENT WITH TGF-BETA1 WITH OR WITHOUT TSA DID NOT HAVE CYTOTOXIC EFFECT. CONCLUSION: THESE FINDINGS PROVIDE NOVEL INSIGHTS INTO THE EPIGENETIC REGULATION IN MYOFIBROBLAST DIFFERENTIATION AND ECM PRODUCTION OF NASAL POLYP. TSA COULD BE A CANDIDATE OF A THERAPEUTIC AGENT FOR REVERSING THE TGF-BETA1-INDUCED ECM SYNTHESIS THAT LEADS TO NASAL POLYP DEVELOPMENT. 2013 2 1632 43 DNMTS ARE INVOLVED IN TGF-BETA1-INDUCED EPITHELIAL-MESENCHYMAL TRANSITIONS IN AIRWAY EPITHELIAL CELLS. CHRONIC RHINOSINUSITIS (CRS) PATHOGENESIS IS CLOSELY RELATED TO TISSUE REMODELING, INCLUDING EPITHELIAL-MESENCHYMAL TRANSITION (EMT). EPIGENETIC MECHANISMS PLAY KEY ROLES IN EMT. DNA METHYLATION, MEDIATED BY DNA METHYLTRANSFERASES (DNMTS), IS AN EPIGENETIC MARKER THAT IS CRITICAL TO EMT. THE GOAL OF THIS STUDY WAS TO DETERMINE WHETHER DNMTS WERE INVOLVED IN TGF-BETA1-INDUCED EMT AND ELUCIDATE THE UNDERLYING MECHANISMS IN NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. GLOBAL DNA METHYLATION AND DNMT ACTIVITY WERE QUANTIFIED. DNMT EXPRESSION WAS MEASURED USING REAL-TIME PCR (QRT-PCR) IN HUMAN CRS TISSUES. MRNA AND PROTEIN LEVELS OF DNMTS, E-CADHERIN, VIMENTIN, ALPHA-SMA, AND FIBRONECTIN WERE DETERMINED USING RT-PCR AND WESTERN BLOTTING, RESPECTIVELY. DNMT1, DNMT3A, AND DNMT3B GENE EXPRESSION WERE KNOCKED DOWN USING SIRNA TRANSFECTION. MAPK PHOSPHORYLATION AND EMT-RELATED TRANSCRIPTION FACTOR LEVELS WERE DETERMINED USING WESTERN BLOTTING. SIGNALING PATHWAYS WERE ANALYZED USING SPECIFIC INHIBITORS OF MAPK. WE DEMONSTRATED THESE DATA IN PRIMARY NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. GLOBAL DNA METHYLATION, DNMT ACTIVITY, AND DNMT EXPRESSION INCREASED IN CRS TISSUES. DNMT EXPRESSION WAS POSITIVELY CORRELATED WITH LUND-MCKAY CT SCORES. TGF-BETA1 DOSE-DEPENDENTLY INDUCED DNMT EXPRESSION. FURTHER, 5-AZA INHIBITED TGF-BETA1-INDUCED DNMT, SNAIL, AND SLUG EXPRESSION RELATED TO EMT, AS WELL AS P38 AND JNK PHOSPHORYLATION IN A549 CELLS AND TGF-BETA1-INDUCED DNMT EXPRESSION AND EMT IN PRIMARY NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. TGF-BETA1-INDUCED DNMT EXPRESSION LEADS TO DNA METHYLATION AND EMT VIA P38, JNK, SNAIL, AND SLUG SIGNALING PATHWAYS. INHIBITION OF DNMT SUPPRESSED THE EMT PROCESS AND THEREFORE IS POTENTIALLY A CRS THERAPEUTIC STRATEGY. 2022 3 2326 46 EPIGENETIC REGULATION OF HOTAIR IN ADVANCED CHRONIC MYELOID LEUKEMIA. PURPOSE: CHRONIC MYELOID LEUKEMIA (CML) ACCOUNTS FOR ~10% OF LEUKEMIA CASES, AND ITS PROGRESSION INVOLVES EPIGENETIC GENE REGULATION. THIS STUDY INVESTIGATED EPIGENETIC REGULATION OF HOTAIR AND ITS TARGET MICRORNA, MIR-143, IN ADVANCED CML. PATIENTS AND METHODS: WE FIRST ISOLATED BONE MARROW MONONUCLEAR CELLS FROM 70 PATIENTS WITH DIFFERENT PHASES OF CML AND FROM HEALTHY DONORS AS NORMAL CONTROL; WE ALSO CULTURED K562 AND KCL22 CELLS, TREATED WITH DEMETHYLATION DRUG; MTT ASSAY, FLOW CYTOMETRY, QUANTITATIVE REAL-TIME POLYMERASE CHAIN REACTION (QPCR), METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MSP), WESTERN BLOT, LUCIFERASE ASSAY, RNA PULL-DOWN ASSAY AND RNA-BINDING PROTEIN IMMUNOPRECIPITATION (RIP) ASSAY WERE PERFORMED. RESULT: AS MEASURED BY QPCR, HOTAIR EXPRESSION IN K562 CELLS, KCL22 CELLS, AND SAMPLES FROM CASES OF ADVANCED-STAGE CML INCREASED WITH LEVELS OF SEVERAL DNA METHYLTRANSFERASES AND HISTONE DEACETYLATES, INCLUDING DNMT1, DNMT3A, HDAC1, EZH2, AND LSD1, AND MIR-143 LEVELS WERE DECREASED AND HOTAIR LEVELS WERE INCREASED. TREATMENT WITH 5-AZACYTIDINE, A DNA METHYLATION INHIBITOR, DECREASED DNMT1, DNMT3A, HDAC1, EZH2, LSD1 MRNA, PROTEIN LEVELS, AND HOTAIR MRNA LEVELS BUT INCREASED MIR-143 LEVELS. HOTAIR KNOCKDOWN AND MIR-143 OVEREXPRESSION BOTH INHIBITED PROLIFERATION AND PROMOTED APOPTOSIS IN KCL22 AND K562 CELLS THROUGH THE PI3K/AKT PATHWAY. RNA PULL-DOWN, MASS SPECTROMETRY, AND RIP ASSAYS SHOWED THAT HOTAIR INTERACTED WITH EZH2 AND LSD1. A DUAL-LUCIFERASE ASSAY DEMONSTRATED THAT HOTAIR INTERACTED WITH MIR-143. CONCLUSION: OUR FINDINGS DEMONSTRATE THE KEY EPIGENETIC INTERACTIONS OF HOTAIR RELATED TO CML PROGRESSION AND SUGGEST HOTAIR AS A POTENTIAL THERAPEUTIC TARGET FOR ADVANCED CML. FURTHERMORE, OUR RESULTS SUPPORT THE USE OF DEMETHYLATION DRUGS AS A CML TREATMENT STRATEGY. 2018 4 169 39 ABNORMALITIES OF AMPK ACTIVATION AND GLUCOSE UPTAKE IN CULTURED SKELETAL MUSCLE CELLS FROM INDIVIDUALS WITH CHRONIC FATIGUE SYNDROME. BACKGROUND: POST EXERTIONAL MUSCLE FATIGUE IS A KEY FEATURE IN CHRONIC FATIGUE SYNDROME (CFS). ABNORMALITIES OF SKELETAL MUSCLE FUNCTION HAVE BEEN IDENTIFIED IN SOME BUT NOT ALL PATIENTS WITH CFS. TO TRY TO LIMIT POTENTIAL CONFOUNDERS THAT MIGHT CONTRIBUTE TO THIS CLINICAL HETEROGENEITY, WE DEVELOPED A NOVEL IN VITRO SYSTEM THAT ALLOWS COMPARISON OF AMP KINASE (AMPK) ACTIVATION AND METABOLIC RESPONSES TO EXERCISE IN CULTURED SKELETAL MUSCLE CELLS FROM CFS PATIENTS AND CONTROL SUBJECTS. METHODS: SKELETAL MUSCLE CELL CULTURES WERE ESTABLISHED FROM 10 SUBJECTS WITH CFS AND 7 AGE-MATCHED CONTROLS, SUBJECTED TO ELECTRICAL PULSE STIMULATION (EPS) FOR UP TO 24H AND EXAMINED FOR CHANGES ASSOCIATED WITH EXERCISE. RESULTS: IN THE BASAL STATE, CFS CULTURES SHOWED INCREASED MYOGENIN EXPRESSION BUT DECREASED IL6 SECRETION DURING DIFFERENTIATION COMPARED WITH CONTROL CULTURES. CONTROL CULTURES SUBJECTED TO 16 H EPS SHOWED A SIGNIFICANT INCREASE IN BOTH AMPK PHOSPHORYLATION AND GLUCOSE UPTAKE COMPARED WITH UNSTIMULATED CELLS. IN CONTRAST, CFS CULTURES SHOWED NO INCREASE IN AMPK PHOSPHORYLATION OR GLUCOSE UPTAKE AFTER 16 H EPS. HOWEVER, GLUCOSE UPTAKE REMAINED RESPONSIVE TO INSULIN IN THE CFS CELLS POINTING TO AN EXERCISE-RELATED DEFECT. IL6 SECRETION IN RESPONSE TO EPS WAS SIGNIFICANTLY REDUCED IN CFS COMPARED WITH CONTROL CULTURES AT ALL TIME POINTS MEASURED. CONCLUSION: EPS IS AN EFFECTIVE MODEL FOR ELICITING MUSCLE CONTRACTION AND THE METABOLIC CHANGES ASSOCIATED WITH EXERCISE IN CULTURED SKELETAL MUSCLE CELLS. WE FOUND FOUR MAIN DIFFERENCES IN CULTURED SKELETAL MUSCLE CELLS FROM SUBJECTS WITH CFS; INCREASED MYOGENIN EXPRESSION IN THE BASAL STATE, IMPAIRED ACTIVATION OF AMPK, IMPAIRED STIMULATION OF GLUCOSE UPTAKE AND DIMINISHED RELEASE OF IL6. THE RETENTION OF THESE DIFFERENCES IN CULTURED MUSCLE CELLS FROM CFS SUBJECTS POINTS TO A GENETIC/EPIGENETIC MECHANISM, AND PROVIDES A SYSTEM TO IDENTIFY NOVEL THERAPEUTIC TARGETS. 2015 5 1906 33 ENHANCER OF ZESTE HOMOLOG 2-CATALYSED H3K27 TRIMETHYLATION PLAYS A KEY ROLE IN ACUTE-ON-CHRONIC LIVER FAILURE VIA TNF-MEDIATED PATHWAY. ACUTE-ON-CHRONIC LIVER FAILURE IS MAINLY DUE TO HOST IMMUNITY SELF-DESTRUCTION. THE HISTONE H3 LYSINE 27 (H3K27) TRIMETHYLATING ENZYME, ENHANCER OF ZESTE HOMOLOG 2 (EZH2) MEDIATES EPIGENETIC SILENCING OF GENE EXPRESSION AND REGULATES IMMUNITY, ALSO INVOLVES PATHOGENESIS OF SEVERAL LIVER DISEASES. THE CURRENT STUDY WAS TO DETERMINE THE ROLE OF METHYLTRANSFERASE EZH2 AND ITS CATALYSED H3K27 TRIMETHYLATION (H3K27ME3) IN LIVER FAILURE, AND TO FURTHER INVESTIGATE THE POTENTIAL TARGET FOR LIVER FAILURE TREATMENT. EZH2 AND ITS CATALYSED H3K27ME3 WERE DETERMINED IN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMC) FROM LIVER FAILURE PATIENTS AND KUPFFER CELLS FROM EXPERIMENTAL MICE. FURTHERMORE, GSK126 (AN INHIBITOR FOR EZH2 TRIMETHYLATION FUNCTION) WAS APPLIED IN LIVER FAILURE MICE IN VIVO, AND LIPOPOLYSACCHARIDE-STIMULATED MONONUCLEAR CELLS IN VITRO. EZH2 AND H3K27ME3 WERE SIGNIFICANTLY UPREGULATED IN HUMAN PBMC FROM LIVER FAILURE PATIENTS OR MURINE KUPFFER CELLS FROM THE LIVER FAILURE ANIMALS, RESPECTIVELY. GSK126 AMELIORATED DISEASE SEVERITY IN LIVER FAILURE MICE, WHICH MAYBE ATTRIBUTE TO DOWN-REGULATE CIRCULATING AND HEPATIC PROINFLAMMATORY CYTOKINES, ESPECIALLY TNF VIA REDUCING H3K27ME3. IN-DEPTH CHROMATIN IMMUNOPRECIPITATION ANALYSIS UNRAVELLED THAT DECREASED ENRICHMENT OF H3K27ME3 ON TNF PROMOTOR, RESULTING IN TNF ELEVATION IN KUPFFER CELLS FROM LIVER FAILURE MICE. NUCLEAR FACTOR KAPPA B (NF-KAPPAB) AND PROTEIN KINASE B (AKT) SIGNALLING PATHWAYS WERE ACTIVATED UPON LIPOPOLYSACCHARIDE STIMULATION, BUT ATTENUATED BY USING GSK126, ACCOMPANIED WITH DECREASED TNF IN VITRO. IN CONCLUSION, EZH2 AND H3K27ME3 CONTRIBUTED TO THE PATHOGENESIS OF LIVER FAILURE VIA TRIGGERING TNF AND OTHER INDISPENSABLE PROINFLAMMATORY CYTOKINES. EZH2 WAS TO MODIFY H3K27ME3 ENRICHMENT, AS WELL AS, ACTIVATION OF THE DOWNSTREAM NF-KAPPAB AND AKT SIGNALLING PATHWAYS. 2018 6 2300 37 EPIGENETIC REGULATION OF BDNF EXPRESSION IN THE PRIMARY SENSORY NEURONS AFTER PERIPHERAL NERVE INJURY: IMPLICATIONS IN THE DEVELOPMENT OF NEUROPATHIC PAIN. BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) IS KNOWN TO BE UP-REGULATED IN THE DORSAL ROOT GANGLION (DRG) AFTER PERIPHERAL NERVE INJURY, AND TO CONTRIBUTE TO NEUROPATHIC PAIN. HERE, WE FOUND THAT THERMAL HYPERALGESIA AND MECHANICAL ALLODYNIA AT DAY 7 POST-INJURY WERE INHIBITED ONLY WHEN ANTI-BDNF ANTIBODY WAS INTRATHECALLY ADMINISTRATED AT DAY 2 POST-INJURY. CONSISTENT WITH BEHAVIORAL RESULTS, WESTERN BLOT ANALYSIS SHOWED THAT THE EXPRESSION LEVELS OF BDNF PROTEIN IN THE SPINAL DORSAL HORN WERE MARKEDLY INDUCED DURING EARLY STAGE POST-INJURY. MOREOVER, THE MAXIMAL INCREASE IN BDNF MRNA EXPRESSION IN THE DRG WAS OBSERVED AT DAY 1 POST-INJURY, AND SIGNIFICANTLY ELEVATED LEVELS WERE SUSTAINED FOR AT LEAST 14 DAYS. FOUR OF FIVE BDNF MRNA TRANSCRIPTS WERE UP-REGULATED AFTER NERVE INJURY, AND THE MOST INDUCIBLE TRANSCRIPT WAS EXON I. USING A CHROMATIN IMMUNOPRECIPITATION (CHIP) ASSAY, WE FOUND THAT NERVE INJURY PROMOTES HISTONE H3 AND H4 ACETYLATION, TRANSCRIPTIONALLY ACTIVE MODIFICATIONS, AT BDNF PROMOTER I AT DAY 1 POST-INJURY, AND THE LEVELS OF HISTONE ACETYLATION REMAIN ELEVATED FOR AT LEAST 7 DAYS. TAKEN TOGETHER, OUR FINDINGS SUGGEST THAT AN INITIAL INCREASE IN BDNF EXON I EXPRESSION CONTROLLED BY EPIGENETIC MECHANISMS MIGHT HAVE A CRUCIAL ROLE IN THE DEVELOPMENT OF NEUROPATHIC PAIN. 2013 7 4303 46 MICRORNA-223 INHIBITS TISSUE FACTOR EXPRESSION IN VASCULAR ENDOTHELIAL CELLS. OBJECTIVE: ATHEROSCLEROSIS IS A CHRONIC INFLAMMATORY PROCESS, IN WHICH VASCULAR ENDOTHELIAL CELLS (ECS) BECOME DYSFUNCTIONAL OWING TO THE EFFECTS OF CHEMICAL SUBSTANCES, SUCH AS INFLAMMATORY FACTOR AND GROWTH FACTORS. TISSUE FACTOR (TF) EXPRESSION IS INDUCED BY THE ABOVE CHEMICAL SUBSTANCES IN ACTIVATED ECS. TF INITIATES THROMBOSIS ON DISRUPTED ATHEROSCLEROTIC PLAQUES WHICH PLAYS AN ESSENTIAL ROLE DURING THE ONSET OF ACUTE CORONARY SYNDROMES (ACS). INCREASING EVIDENCES SUGGEST THE IMPORTANT ROLE OF MICRORNAS AS EPIGENETIC REGULATORS OF ATHEROSCLEROTIC DISEASE. THE AIM OF OUR STUDY IS TO IDENTIFY IF MICRORNA-223 (MIR-223) TARGETS TF IN ECS. METHODS AND RESULTS: BIOINFORMATIC ANALYSIS SHOWED THAT TF IS A TARGET CANDIDATE OF MIR-223. WESTERN BLOTTING ANALYSIS REVEALED THAT TUMOR NECROSIS FACTOR ALPHA (TNF-ALPHA) INCREASED TF EXPRESSION IN AORTA OF C57BL/6J MICE AND CULTURED ECS (EA.HY926 CELLS AND HUVEC) AFTER 4 H TREATMENT. IN TNF-ALPHA TREATED ECS, TF MRNA WAS ALSO INCREASED MEASURED BY REAL-TIME PCR. REAL-TIME PCR RESULTS SHOWED THAT MIR-223 LEVELS WERE DOWNREGULATED IN TNF-ALPHA-TREATED AORTA OF C57BL/6J MICE AND CULTURED ECS. TRANSFECTION OF ECS WITH MIR-223 MIMIC OR MIR-223 INHIBITOR MODIFIED TF EXPRESSION BOTH IN MRNA AND PROTEIN LEVELS. LUCIFERASE ASSAYS CONFIRMED THAT MIR-223 SUPPRESSED TF EXPRESSION BY BINDING TO THE SEQUENCE OF TF 3'-UNTRANSLATED REGIONS (3'UTR). TF PROCOAGULANT ACTIVITY WAS INHIBITED BY OVEREXPRESSING MIR-223 WITH OR WITHOUT TNF-ALPHA STIMULATION. CONCLUSIONS: MIR-223-MEDIATED SUPPRESSION OF TF EXPRESSION PROVIDES A NOVEL MOLECULAR MECHANISM FOR THE REGULATION OF COAGULATION CASCADE, AND SUGGESTS A CLUE AGAINST THROMBOGENESIS DURING THE PROCESS OF ATHEROSCLEROTIC PLAQUE RUPTURE. 2014 8 2825 46 FLOW-DEPENDENT EPIGENETIC REGULATION OF IGFBP5 EXPRESSION BY H3K27ME3 CONTRIBUTES TO ENDOTHELIAL ANTI-INFLAMMATORY EFFECTS. RATIONALE: ATHEROSCLEROSIS IS A CHRONIC INFLAMMATORY AND EPIGENETIC DISEASE THAT IS INFLUENCED BY DIFFERENT PATTERNS OF BLOOD FLOW. HOWEVER, THE EPIGENETIC MECHANISM WHEREBY ATHEROPROTECTIVE FLOW CONTROLS ENDOTHELIAL GENE PROGRAMMING REMAINS ELUSIVE. HERE, WE INVESTIGATED THE POSSIBILITY THAT FLOW ALTERS ENDOTHELIAL GENE EXPRESSION THROUGH EPIGENETIC MECHANISMS. METHODS: EN FACE STAINING AND WESTERN BLOT WERE USED TO DETECT PROTEIN EXPRESSION. REAL-TIME PCR WAS USED TO DETERMINE RELATIVE GENE EXPRESSION. RNA-SEQUENCING OF HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS TREATED WITH SIRNA OF ENHANCER OF ZESTE HOMOLOG 2 (EZH2) OR LAMINAR FLOW WAS USED FOR TRANSCRIPTIONAL PROFILING. RESULTS: WE FOUND THAT TRIMETHYLATION OF HISTONE 3 LYSINE 27 (H3K27ME3), A REPRESSIVE EPIGENETIC MARK THAT ORCHESTRATES GENE REPRESSION, WAS REDUCED IN LAMINAR FLOW AREAS OF MOUSE AORTA AND FLOW-TREATED HUMAN ENDOTHELIAL CELLS. THE DECREASE OF H3K27ME3 PARALLELED A REDUCTION IN THE EPIGENETIC "WRITER"-EZH2, THE CATALYTIC SUBUNIT OF THE POLYCOMB REPRESSIVE COMPLEX 2 (PRC2). MOREOVER, LAMINAR FLOW DECREASED EXPRESSION OF EZH2 VIA MECHANOSENSITIVE MIR101. GENOME-WIDE TRANSCRIPTOME PROFILING STUDIES IN ENDOTHELIAL CELLS TREATED WITH EZH2 SIRNA AND FLOW REVEALED THE UPREGULATION OF NOVEL MECHANOSENSITIVE GENE IGFBP5 (INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN 5), WHICH IS EPIGENETICALLY SILENCED BY H3K27ME3. FUNCTIONALLY, INHIBITION OF H3K27ME3 BY EZH2 SIRNA OR GSK126 (A SPECIFIC EZH2 INHIBITOR) REDUCED H3K27ME3 LEVELS AND MONOCYTE ADHESION TO ENDOTHELIAL CELLS. ADENOVIRAL OVEREXPRESSION OF IGFBP5 ALSO RECAPITULATED THE ANTI-INFLAMMATORY EFFECTS OF H3K27ME3 INHIBITION. MORE IMPORTANTLY, WE OBSERVED EZH2 UPREGULATION, AND IGFBP5 DOWNREGULATION, IN ADVANCED ATHEROSCLEROTIC PLAQUES FROM HUMAN PATIENTS. CONCLUSION: TAKEN TOGETHER, OUR FINDINGS REVEAL THAT ATHEROPROTECTIVE FLOW REDUCES H3K27ME3 AS A CHROMATIN-BASED MECHANISM TO AUGMENT THE EXPRESSION OF GENES THAT CONFER AN ANTI-INFLAMMATORY RESPONSE IN THE ENDOTHELIUM. OUR STUDY EXEMPLIFIES FLOW-DEPENDENT EPIGENETIC REGULATION OF ENDOTHELIAL GENE EXPRESSION, AND ALSO SUGGESTS THAT TARGETING THE EZH2/H3K27ME3/IGFBP5 PATHWAY MAY OFFER NOVEL THERAPEUTICS FOR INFLAMMATORY DISORDERS SUCH AS ATHEROSCLEROSIS. 2018 9 5860 31 SULFORAPHANE PREVENTS ANGIOTENSIN II-INDUCED CARDIOMYOPATHY BY ACTIVATION OF NRF2 THROUGH EPIGENETIC MODIFICATION. NUCLEAR FACTOR ERYTHROID 2-RELATED FACTOR (NRF2) IS AN IMPORTANT REGULATOR OF CELLULAR ANTIOXIDANT DEFENCE. WE PREVIOUSLY SHOWED THAT SFN PREVENTED ANG II-INDUCED CARDIAC DAMAGE VIA ACTIVATION OF NRF2. HOWEVER, THE UNDERLYING MECHANISM OF SFN'S PERSISTENT CARDIAC PROTECTION REMAINS UNCLEAR. THIS STUDY AIMED TO EXPLORE THE POTENTIAL OF SFN IN ACTIVATING CARDIAC NRF2 THROUGH EPIGENETIC MECHANISMS. WILD-TYPE MICE WERE INJECTED SUBCUTANEOUSLY WITH ANG II, WITH OR WITHOUT SFN. ADMINISTRATION OF CHRONIC ANG II-INDUCED CARDIAC INFLAMMATORY FACTOR EXPRESSION, OXIDATIVE DAMAGE, FIBROSIS AND CARDIAC REMODELLING AND DYSFUNCTION, ALL OF WHICH WERE EFFECTIVELY IMPROVED BY SFN TREATMENT, COUPLED WITH AN UP-REGULATION OF NRF2 AND DOWNSTREAM GENES. BISULFITE GENOME SEQUENCING AND CHROMATIN IMMUNOPRECIPITATION (CHIP) WERE PERFORMED TO DETECT THE METHYLATION LEVEL OF THE FIRST 15 CPGS AND HISTONE H3 ACETYLATION (AC-H3) STATUS IN THE NRF2 PROMOTER REGION, RESPECTIVELY. THE RESULTS SHOWED THAT SFN REDUCED ANG II-INDUCED CPG HYPERMETHYLATION AND PROMOTED AC-H3 ACCUMULATION IN THE NRF2 PROMOTER REGION, ACCOMPANIED BY THE INHIBITION OF GLOBAL DNMT AND HDAC ACTIVITY, AND A DECREASED PROTEIN EXPRESSION OF KEY DNMT AND HDAC ENZYMES. TAKEN TOGETHER, SFN EXERTS ITS CARDIOPROTECTIVE EFFECT THROUGH EPIGENETIC MODIFICATION OF NRF2, WHICH MAY PARTIALLY CONTRIBUTE TO LONG-TERM ACTIVATION OF CARDIAC NRF2. 2021 10 1905 45 ENHANCER OF ZESTE HOMOLOG 2 CONTRIBUTES TO APOPTOSIS BY INACTIVATING JANUS KINASE 2/ SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION SIGNALING IN INFLAMMATORY BOWEL DISEASE. BACKGROUND: INFLAMMATORY BOWEL DISEASE (IBD) IS A PREVALENT WORLDWIDE HEALTH PROBLEM FEATURED BY RELAPSING, CHRONIC GASTROINTESTINAL INFLAMMATION. ENHANCER OF ZESTE HOMOLOG 2 (EZH2) IS A CRITICAL EPIGENETIC REGULATOR IN DIFFERENT PATHOLOGICAL MODELS, SUCH AS CANCER AND INFLAMMATION. HOWEVER, THE ROLE OF EZH2 IN THE IBD DEVELOPMENT IS STILL OBSCURE. AIM: TO EXPLORE THE EFFECT OF EZH2 ON IBD PROGRESSION AND THE UNDERLYING MECHANISM. METHODS: THE IBD MOUSE MODEL WAS CONDUCTED BY ADDING DEXTRAN SODIUM SULFATE (DSS), AND THE EFFECT OF EZH2 ON DSS-INDUCED COLITIS WAS ASSESSED IN THE MODEL. THE FUNCTION OF EZH2 IN REGULATING APOPTOSIS AND PERMEABILITY WAS EVALUATED BY ANNEXIN V-FITC APOPTOSIS DETECTION KIT, TRANSEPITHELIAL ELECTRICAL RESISTANCE ANALYSIS, AND WESTERN BLOT ANALYSIS OF RELATED MARKERS, INCLUDING ZONA OCCLUDENS 1, CLAUDIN-5, AND OCCLUDIN, IN NCM460 AND FETAL HUMAN COLON (FHC) CELLS. THE MECHANICAL INVESTIGATION WAS PERFORMED BY QUANTITATIVE REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION, WESTERN BLOT ANALYSIS, AND CHROMATIN IMMUNOPRECIPITATION ASSAYS. RESULTS: THE COLON LENGTH WAS INHIBITED IN THE DSS-TREATED MICE AND WAS ENHANCED BY THE EZH2 DEPLETION IN THE SYSTEM. DSS TREATMENT CAUSED A DECREASED HISTOLOGICAL SCORE IN THE MICE, WHICH WAS REVERSED BY EZH2 DEPLETION. THE INFLAMMATORY CYTOKINES, SUCH AS TUMOR NECROSIS FACTOR-ALPHA, INTERLEUKIN-6, AND INTERLEUKIN-1BETA, WERE INDUCED IN THE DSS-TREATED MICE, IN WHICH THE DEPLETION OF EZH2 COULD REVERSE THIS EFFECT. MOREOVER, THE TUMOR NECROSIS FACTOR-ALPHA TREATMENT INDUCED THE APOPTOSIS OF NCM460 AND FHC CELLS, IN WHICH EZH2 DEPLETION COULD REVERSE THIS EFFECT IN THE CELLS. MOREOVER, THE DEPLETION OF EZH2 ATTENUATED PERMEABILITY OF COLONIC EPITHELIAL CELLS. MECHANICALLY, THE DEPLETION OF EZH2 OR EZH2 INHIBITOR GSK343 WAS ABLE TO ENHANCE THE EXPRESSION AND THE PHOSPHORYLATION OF JANUS KINASE 2 (JK2) AND SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION IN THE NCM460 AND FHC CELLS. SPECIFICALLY, EZH2 INACTIVATED JAK2 EXPRESSION BY REGULATING HISTONE H3K27ME3. JAK2 INHIBITOR TG101348 WAS ABLE TO REVERSE EZH2 KNOCKDOWN-MEDIATED COLONIC EPITHELIAL CELL PERMEABILITY AND APOPTOSIS. CONCLUSION: THUS, WE CONCLUDED THAT EZH2 CONTRIBUTED TO APOPTOSIS AND INFLAMMATORY RESPONSE BY INACTIVATING JAK2/ SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION SIGNALING IN IBD. EZH2 MAY BE APPLIED AS A POTENTIAL TARGET FOR IBD THERAPY. 2021 11 6704 32 VHL GENE METHYLATION CONTRIBUTES TO EXCESSIVE ERYTHROCYTOSIS IN CHRONIC MOUNTAIN SICKNESS RAT MODEL BY UPREGULATING THE HIF-2ALPHA/EPO PATHWAY. AIMS: HYPOXIA-INDUCIBLE FACTORS (HIFS) PLAY IMPORTANT ROLES IN THE PATHOGENESIS OF ERYTHROCYTOSIS IN CHRONIC MOUNTAIN SICKNESS (CMS). VON HIPPEL-LINDAU (VHL) IS A KEY REGULATOR OF HYPOXIA THAT CAN DIRECT THE POLY-UBIQUITYLATION AND DEGRADATION OF HIFS. EPIGENETIC MECHANISMS ARE BELIEVED TO CONTRIBUTE TOWARD ADAPTION TO CHRONIC HYPOXIA. HERE, WE INVESTIGATED THE CONTRIBUTION AND MECHANISM OF VHL METHYLATION IN RATS WITH ERYTHROCYTOSIS IN CMS. MAIN METHODS: THE METHYLATION STATUS OF VHL WAS MEASURED VIA BISULFITE SEQUENCING PCR, WHILE VHL, DNMT1, DNMT3ALPHA, AND DNMT3BETA EXPRESSION WERE ASSESSED USING REAL-TIME REVERSE TRANSCRIPTION PCR AND WESTERN BLOTTING. HIF-2ALPHA AND EPO EXPRESSION LEVELS IN BONE MARROW WERE DETERMINED VIA IMMUNOHISTOCHEMICAL STAINING, AND ERYTHROID HYPERPLASIA IN BONE MARROW SECTIONS WERE OBSERVED WITH HEMATOXYLIN AND EOSIN STAINING. KEY FINDINGS: WE FOUND THAT CHRONIC HYPOXIA TRIGGERED ERYTHROID HYPERPLASIA IN THE BONE MARROW AND INCREASED THE QUANTITY OF PERIPHERAL RED BLOOD CELLS IN CMS RATS. CHRONIC HYPOXIA SIGNIFICANTLY INDUCED METHYLATION AT THE CPG SITE IN THE VHL PROMOTER, DECREASED VHL EXPRESSION, AND INCREASED HIF-2ALPHA AND EPO EXPRESSION. CHRONIC HYPOXIA INCREASED DNMT3ALPHA AND DNMT3BETA EXPRESSION, CONSISTENT WITH THE DECREASE IN VHL EXPRESSION. THE DNA METHYLTRANSFERASE INHIBITOR 5-AZACYTIDINE REDUCED CHRONIC HYPOXIA-INDUCED ERYTHROID PROLIFERATION IN THE BONE MARROW OF RATS WITH CMS BY SUPPRESSING VHL METHYLATION AND DNMTS EXPRESSION. SIGNIFICANCE: OUR STUDY SUGGESTS THAT VHL METHYLATION CONTRIBUTES TOWARD EXCESSIVE ERYTHROCYTOSIS IN CMS BY UPREGULATING THE HIF-2ALPHA/EPO PATHWAY IN THE BONE MARROW OF RATS. WE DEMONSTRATED THAT THE DNMT INHIBITOR 5-AZACYTIDINE CAN ATTENUATE ERYTHROID HYPERPLASIA IN THE BONE MARROW BY DEMETHYLATING THE VHL PROMOTER. 2021 12 1236 42 CURCUMIN AMELIORATES NEPHROSCLEROSIS VIA SUPPRESSION OF HISTONE ACETYLATION INDEPENDENT OF HYPERTENSION. BACKGROUND: ALTHOUGH HISTONE ACETYLATION, AN EPIGENETIC MODIFICATION, HAS BEEN REPORTED TO BE RELATED TO THE PROGRESSION OF VARIOUS DISEASES, ITS INVOLVEMENT IN NEPHROSCLEROSIS IS UNCLEAR. METHODS: DAHL SALT-SENSITIVE RATS WERE USED AS A MODEL OF NEPHROSCLEROSIS IN THIS STUDY. THE RATS WERE DIVIDED INTO THREE GROUPS: (I) NORMAL-SALT DIET GROUP, (II) HIGH-SALT DIET GROUP (HS), AND (III) HS ADMINISTERED DAILY WITH CURCUMIN, A HISTONE ACETYLTRANSFERASE INHIBITOR (HS+C). AT 6 WEEKS AFTER THE TREATMENT, THE KIDNEYS WERE DISSECTED. MORPHOLOGIC CHANGES WERE ASSESSED BY MASSON'S TRICHROME STAINING. THE NUMBER OF MACROPHAGES, FIBROBLASTS AND THE CELLS EXPRESSING ACETYLATED HISTONE H3 AT LYS 9 (H3K9) WERE ASSESSED BY IMMUNOHISTOCHEMISTRY. RESULTS: ALTHOUGH BOTH HS AND HS+C RATS REVEALED A MARKED INCREASE IN SYSTOLIC BLOOD PRESSURE, SERUM CREATININE WAS INCREASED ONLY IN HS RATS AT 6 WEEKS. IN THE HS RATS, NEPHROSCLEROSIS WAS INDUCED, ACCOMPANYING A SIGNIFICANT ACCUMULATION OF MACROPHAGES AND FIBROBLASTS. THE INFLAMMATION AND FIBROSIS WAS MARKEDLY SUPPRESSED IN THE HS+C GROUP. THE LEVEL OF HISTONE ACETYLATION AT LYS 9 WAS ENHANCED IN THE HS RATS, WHEREAS CURCUMIN ADMINISTRATION SUPPRESSED THE HISTONE ACETYLATION. MOREOVER, IN THE HS RATS, INTERLEUKIN-6 GENE EXPRESSION WAS ASSOCIATED WITH ACETYLATED H3K9, AS REVEALED BY CHROMATIN IMMUNOPRECIPITATION ASSAY. CONCLUSIONS: OUR RESULTS SUGGESTED THAT CURCUMIN AMELIORATES NEPHROSCLEROSIS VIA SUPPRESSION OF HISTONE ACETYLATION, INDEPENDENTLY OF HYPERTENSION. 2016 13 5479 43 RESVERATROL ATTENUATES CIGARETTE SMOKE EXTRACT INDUCED CELLULAR SENESCENCE IN HUMAN AIRWAY EPITHELIAL CELLS BY REGULATING THE MIR-34A/SIRT1/NF-KAPPAB PATHWAY. CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) IS CHARACTERIZED BY ACCELERATED LUNG AGING. SMOKING IS THE CRITICAL RISK FACTOR FOR COPD. CELLULAR SENESCENCE OF AIRWAY EPITHELIAL CELLS IS THE CYTOLOGICAL BASIS OF ACCELERATED LUNG AGING IN COPD, AND THE REGULATION OF MICRORNAS (MIRNAS) IS THE CENTRAL EPIGENETIC MECHANISM OF CELLULAR SENESCENCE. RESVERATROL (RES) IS A POLYPHENOL WITH ANTI-AGING PROPERTIES. THIS STUDY INVESTIGATED WHETHER RES ATTENUATES CIGARETTE SMOKE EXTRACT (CSE)-INDUCED CELLULAR SENESCENCE IN HUMAN AIRWAY EPITHELIAL CELLS (BEAS-2B) THROUGH THE MIR-34A/SIRT1/NUCLEAR FACTOR-KAPPAB (NF-KAPPAB) PATHWAY. BEAS-2B CELLS WERE TREATED WITH RES, CSE AND TRANSFECTED WITH MIR-34A-5P MIMICS. CELLULAR SENESCENCE WAS EVALUATED BY SENESCENCE -RELATED BETA-GALACTOSIDASE (SA-BETA-GAL) STAINING AND EXPRESSION OF SENESCENCE-RELATED GENES (P16, P21, AND P53). THE EXPRESSIONS OF MIR-34A-5P, SIRT1, AND NF-KAPPAB P65 WERE EXAMINED USING QUANTITATIVE REAL TIME POLYMERASE CHAIN REACTION AND WESTERN BLOTTING. THE SENESCENCE-ASSOCIATED SECRETORY PHENOTYPE (SASP) CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) WERE ASSESSED BY ENZYME-LINKED IMMUNOSORBENT ASSAY. THE BINDING BETWEEN MIR-34A-5P AND SIRT1 WAS CONFIRMED BY DUAL-LUCIFERASE REPORTER ASSAY. THE RESULTS SHOWED THAT CSE DOSE-DEPENDENTLY DECREASED CELL VIABILITY AND ELEVATED CELLULAR SENESCENCE, CHARACTERIZED BY INCREASED SA-BETA-GAL STAINING AND SENESCENCE-RELATED GENE EXPRESSIONS (P16, P21, AND P53). FURTHER, CSE DOSE-DEPENDENTLY INCREASED THE EXPRESSION OF MIR-34A-5P AND SASP CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) IN BEAS-2B CELLS. PRETREATMENT WITH RES INHIBITED CSE-INDUCED CELLULAR SENESCENCE AND SECRETION OF SASP CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) IN A DOSE-DEPENDENT MANNER. MOREOVER, RES REVERSED THE CSE-INDUCED DOWN-REGULATION OF SIRT1 AND UP-REGULATION OF MIR-34A-5P AND NF-KAPPAB P65. SIRT1 IS A TARGET OF MIR-34A-5P. OVEREXPRESSION OF MIR-34A-5P VIA TRANSFECTION WITH MIR-34A-5P MIMIC IN BEAS-2B CELLS ATTENUATED THE INHIBITORY EFFECT OF RES ON CELLULAR SENESCENCE, ACCOMPANIED BY REVERSING THE EXPRESSION OF SIRT1 AND NF-KAPPAB P65. IN CONCLUSION, RES ATTENUATED CSE-INDUCED CELLULAR SENESCENCE IN BEAS-2B CELLS BY REGULATING THE MIR-34A/SIRT1/NF-KAPPAB PATHWAY, WHICH MAY PROVIDE A NEW APPROACH FOR COPD TREATMENT. 2022 14 1667 42 DOWNREGULATION OF PCAF BY MIR-181A/B PROVIDES FEEDBACK REGULATION TO TNF-ALPHA-INDUCED TRANSCRIPTION OF PROINFLAMMATORY GENES IN LIVER EPITHELIAL CELLS. ABERRANT CELLULAR RESPONSES TO PROINFLAMMATORY CYTOKINES, SUCH AS TNF-ALPHA, ARE PATHOGENIC FEATURES IN MOST CHRONIC INFLAMMATORY DISEASES. A VARIETY OF EXTRACELLULAR AND INTRACELLULAR FEEDBACK PATHWAYS HAS EVOLVED TO PREVENT AN INAPPROPRIATE CELLULAR REACTION TO THESE PROINFLAMMATORY CYTOKINES. IN THIS STUDY, WE REPORT THAT TNF-ALPHA TREATMENT OF HUMAN AND MOUSE CHOLANGIOCYTES AND HEPATOCYTES DOWNREGULATED EXPRESSION OF P300/CBP-ASSOCIATED FACTOR (PCAF), A COACTIVATOR AND AN ACETYLTRANSFERASE THAT PROMOTES HISTONE ACETYLATION AND GENE TRANSCRIPTION. OF THESE UPREGULATED MICRORNAS IN TNF-ALPHA-TREATED CELLS, MIR-181A/B (MIR-181A AND MIR-181B) SUPPRESSED TRANSLATION OF PCAF MRNA. FUNCTIONAL MANIPULATION OF MIR-181A/B CAUSED RECIPROCAL ALTERATIONS IN PCAF PROTEIN EXPRESSION IN CULTURED CHOLANGIOCYTES AND HEPATOCYTES. INHIBITION OF MIR-181A/B FUNCTION WITH ANTI-MIRS BLOCKED TNF-ALPHA-INDUCED SUPPRESSION OF PCAF EXPRESSION. PROMOTER RECRUITMENT OF PCAF WAS SHOWN TO BE ASSOCIATED WITH TNF-ALPHA-INDUCED TRANSCRIPTION OF INFLAMMATORY GENES. INTRIGUINGLY, PRETREATMENT OF CELLS WITH TNF-ALPHA INHIBITED TRANSCRIPTION OF INFLAMMATORY GENES IN RESPONSE TO SUBSEQUENT TNF-ALPHA STIMULATION. OVEREXPRESSION OF PCAF OR INHIBITION OF MIR-181A/B FUNCTION WITH ANTI-MIRS ATTENUATED THE INHIBITORY EFFECTS OF TNF-ALPHA PRETREATMENT ON EPITHELIAL INFLAMMATORY RESPONSE TO SUBSEQUENT TNF-ALPHA STIMULATION. DOWNREGULATION OF PCAF AND THE INHIBITORY EFFECTS OF TNF-ALPHA PRETREATMENT ON LIVER EPITHELIAL INFLAMMATORY RESPONSE WERE FURTHER CONFIRMED IN A MOUSE MODEL OF TNF-ALPHA I.P. INJECTION. THESE DATA SUGGEST THAT PCAF IS A TARGET FOR MIR-181A/B, AND DOWNREGULATION OF PCAF BY TNF-ALPHA PROVIDES NEGATIVE FEEDBACK REGULATION TO INFLAMMATORY REACTIONS IN LIVER EPITHELIAL CELLS, A PROCESS THAT MAY BE RELEVANT TO THE EPIGENETIC FINE-TUNING OF EPITHELIAL INFLAMMATORY PROCESSES IN GENERAL. 2012 15 3633 37 INCREASE IN HDAC9 SUPPRESSES MYOBLAST DIFFERENTIATION VIA EPIGENETIC REGULATION OF AUTOPHAGY IN HYPOXIA. EXTREMELY REDUCED OXYGEN (O(2)) LEVELS ARE DETRIMENTAL TO MYOGENIC DIFFERENTIATION AND MULTINUCLEATED MYOTUBE FORMATION, AND CHRONIC EXPOSURE TO HIGH-ALTITUDE HYPOXIA HAS BEEN REPORTED TO BE AN IMPORTANT FACTOR IN SKELETAL MUSCLE ATROPHY. HOWEVER, HOW CHRONIC HYPOXIA CAUSES MUSCLE DYSFUNCTION REMAINS UNKNOWN. IN THE PRESENT STUDY, WE FOUND THAT SEVERE HYPOXIA (1% O(2)) SIGNIFICANTLY INHIBITED THE FUNCTION OF C2C12 CELLS (FROM A MYOBLAST CELL LINE). IMPORTANTLY, THE IMPAIRMENT WAS CONTINUOUSLY MANIFESTED EVEN DURING CULTURE UNDER NORMOXIC CONDITIONS FOR SEVERAL PASSAGES. MECHANISTICALLY, WE REVEALED THAT HISTONE DEACETYLASES 9 (HDAC9), A MEMBER OF THE HISTONE DEACETYLASE FAMILY, WAS SIGNIFICANTLY INCREASED IN C2C12 CELLS UNDER HYPOXIC CONDITIONS, THEREBY INHIBITING INTRACELLULAR AUTOPHAGY LEVELS BY DIRECTLY BINDING TO THE PROMOTER REGIONS OF ATG7, BECLIN1, AND LC3. THIS PHENOMENON RESULTED IN THE SEQUENTIAL DEPHOSPHORYLATION OF GSK3BETA AND INACTIVATION OF THE CANONICAL WNT PATHWAY, IMPAIRING THE FUNCTION OF THE C2C12 CELLS. TAKEN TOGETHER, OUR RESULTS SUGGEST THAT HYPOXIA-INDUCED MYOBLAST DYSFUNCTION IS DUE TO ABERRANT EPIGENETIC REGULATION OF AUTOPHAGY, AND OUR EXPERIMENTAL EVIDENCE REVEALS THE POSSIBLE MOLECULAR PATHOGENESIS RESPONSIBLE FOR SOME MUSCLE DISEASES CAUSED BY CHRONIC HYPOXIA AND SUGGESTS A POTENTIAL THERAPEUTIC OPTION. 2019 16 5850 49 SUBEROYLANILIDE HYDROXAMIC ACID (SAHA) REDUCES FIBROSIS MARKERS AND DEACTIVATES HUMAN STELLATE CELLS VIA THE EPITHELIAL-MESENCHYMAL TRANSITION (EMT). HEPATIC FIBROSIS IS KNOWN AS THE ACCUMULATION OF CONNECTIVE TISSUE SECONDARY TO CHRONIC DAMAGE TO THE LIVER. EPITHELIAL-MESENCHYMAL TRANSITION (EMT) CORRESPONDING INCREASE IN LIVER FIBROGENESIS WAS SHOWN WITH IMMUNOHISTOCHEMISTRY AND PCR-BASED STUDIES. SUBEROYLANILIDE HYDROXAMIC ACID (SAHA), A SYNTHETIC COMPOUND APPROVED AS A HISTONE DEACETYLASE INHIBITOR (HDAC) BY THE FDA TO TREAT CUTANEOUS T-CELL LYMPHOMA IS UNDER INVESTIGATION FOR THE TREATMENT OF LUNG AND RENAL FIBROSIS. EXPERIMENTAL MODELING FOR HEPATIC FIBROSIS CAN BE CONSTRUCTED WITH AN LX2 CELL LINE ISOLATED FROM HUMAN HEPATIC STELLATE CELLS (HSCS). IN THIS STUDY, WE AIMED TO INVESTIGATE THE MODULATION OF SAHA IN THE PATHOGENESIS OF LIVER FIBROSIS BY DETECTING THE LEVELS OF PROTEINS; (E-CADHERIN (E-CAD), N-CADHERIN (N-CAD), VIMENTIN (VIM), AND GENES; E-CAD, N-CAD, VIM, TRANSFORMING GROWTH FACTOR-BETA (TGF-BETA), ALPHA-SMOOTH MUSCLE ACTIN (ALPHA-SMA), TYPE 1 COLLAGEN (COL1A1), TYPE 3 COLLAGEN (COL3A1)) THAT PLAY A SIGNIFICANT ROLE IN EMT WITH THE LX2 CELL LINE. WE ALSO EVALUATED THE ACTION OF SAHA WITH CELL PROLIFERATION, CLONOGENIC, AND MIGRATION ASSAY. CELL PROLIFERATION WAS PERFORMED BY FLOW CYTOMETRY. ALL THE PROTEIN LEVELS WERE DETERMINED BY WESTERN BLOT ANALYSIS, AND GENE EXPRESSION LEVELS WERE MEASURED BY REAL-TIME PCR. OUR STUDY OBSERVED THAT SAHA TREATMENT DECREASED CELL VIABILITY, COLONY FORMATION AND MIGRATION IN LX2 CELLS. WE FOUND THAT SAHA INCREASED E-CAD EXPRESSION LEVEL, WHILE IT DECREASED N-CAD, VIM, COL1A1, COL3A1, ALPHA-SMA TGF-BETA GENES EXPRESSION LEVELS. SAHA DECREASED THE LEVEL OF E-CAD, N-CAD, AND VIM PROTEIN LEVELS. WE THOUGHT THAT SAHA POSSESSES POTENT ANTIFIBROTIC AND ANTI-EMT PROPERTIES IN LX2. 2021 17 2349 85 EPIGENETIC REGULATION OF MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX PRODUCTION IN NASAL POLYP-DERIVED FIBROBLASTS. BACKGROUND: NASAL POLYPOSIS IS A MULTI-FACTORIAL DISEASE ASSOCIATED WITH CHRONIC INFLAMMATORY CONDITION OF THE PARANASAL SINUSES. MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX (ECM) ACCUMULATION ARE INVOLVED IN THE PATHOGENESIS OF NASAL POLYPOSIS. OBJECTIVE: THE AIM OF THIS STUDY WAS TO STUDY THE EFFECT OF TRICHOSTATIN A (TSA), A HISTONE DEACETYLASE (HDAC) INHIBITOR, ON TRANSFORMING GROWTH FACTOR (TGF)-BETA1-INDUCED MYOFIBROBLAST DIFFERENTIATION AND ECM ACCUMULATION IN NASAL POLYP-DERIVED FIBROBLASTS (NPDFS). METHODS: NASAL POLYP-DERIVED FIBROBLASTS WERE ISOLATED FROM NASAL POLYPS OF PATIENTS WHO HAVE CHRONIC RHINOSINUSITIS WITH NASAL POLYP. TSA WAS TREATED IN TGF-BETA1-INDUCED NPDFS. EXPRESSION LEVELS OF HDAC2, ALPHA-SMOOTH MUSCLE ACTIN (SMA), TGF-BETA1, COLLAGEN TYPE I, ACETYLATED HISTONE H3, ACETYLATED HISTONE H4, PHOSPHORYLATED SMAD2/3 AND SMAD7 WERE DETERMINED BY RT-PCR, WESTERN BLOT AND/OR IMMUNOFLUORESCENT STAINING. THE TOTAL COLLAGEN AMOUNT PRODUCTION WAS ANALYSED BY SIRCOL SOLUBLE COLLAGEN ASSAY AND CONTRACTILE ACTIVITY WAS MEASURED BY COLLAGEN GEL CONTRACTION ASSAY. HDAC2 INHIBITION BY TSA OR HDAC2 SILENCING WAS ESTABLISHED BY RT-PCR AND WESTERN BLOT. THE EPIGENETIC EFFECT ON ALPHA-SMA GENE INACTIVATION WAS EXAMINED BY CHROMATIN IMMUNOPRECIPITATION ASSAY. PROLIFERATION WAS DETERMINED BY KI67-POSITIVE CELL STAINING AND CYTOTOXICITY WAS ASSESSED BY 3-(4,5- DIMETHYLTHIAZOL-2YL)-2,5-DIPHENYL-2H-TETRAZOLIUM BROMIDE (MTT) ASSAY. RESULTS: THE EXPRESSION LEVELS OF HDAC2, ALPHA-SMA AND TGF-BETA1 WERE INCREASED IN NASAL POLYP TISSUES COMPARED TO NORMAL INFERIOR TURBINATE TISSUES. TSA AND HDAC2 SILENCING INHIBITED EXPRESSION LEVELS ALPHA-SMA, COLLAGEN AND HDAC2. TSA INDUCED HYPERACETYLATION OF HISTONE AND SUPPRESSED OPENING OF ALPHA-SMA GENE PROMOTER IN TGF-BETA1-INDUCED NPDFS. TSA INHIBITED TGF-BETA1-INDUCED SMAD 2/3 AND RESCUED TGF-BETA1-SUPPRESSED SMAD7 SIGNALLING PATHWAY. FINALLY, TSA BLOCKED PROLIFERATION IN TGF-BETA1-INDUCED NPDFS AND HAS NO CYTOTOXIC EFFECT IN NPDFS. CONCLUSIONS AND CLINICAL RELEVANCE: THESE RESULTS SUGGEST THAT HDAC INHIBITION IS ASSOCIATED WITH MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLUAR MATRIX ACCUMULATION IN NASAL POLYPOSIS. TSA MAY BE USEFUL AS AN INHIBITOR OF NASAL POLYP GROWTH, AND THUS HAS POTENTIAL TO BE USED AS A NOVEL TREATMENT OPTION FOR NASAL POLYPOSIS. 2012 18 1831 44 EFFECTS OF MATERNAL SEPARATION AND ANTIDEPRESSANT DRUG ON EPIGENETIC REGULATION OF THE BRAIN-DERIVED NEUROTROPHIC FACTOR EXON I PROMOTER IN THE ADULT RAT HIPPOCAMPUS. AIM: EARLY LIFE STRESS CAN INDUCE EPIGENETIC CHANGES THROUGH GENETIC AND ENVIRONMENTAL INTERACTIONS AND IS A RISK FACTOR FOR DEPRESSION. BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) HAS BEEN IMPLICATED IN THE PATHOPHYSIOLOGY OF DEPRESSION AND ANTIDEPRESSANT DRUG ACTION. WE INVESTIGATED EPIGENETIC CHANGES AT THE BDNF EXON I PROMOTER IN THE HIPPOCAMPUS OF ADULT RATS SUBJECTED TO MATERNAL SEPARATION (MS) DURING EARLY LIFE AND TREATED WITH AN ANTIDEPRESSANT DRUG AS ADULTS. METHODS: RAT PUPS WERE SUBJECTED TO MS FROM POSTNATAL DAY 1 TO 21 AND RECEIVED CHRONIC ESCITALOPRAM (ESC) AS ADULTS. WE ASSESSED THE EFFECTS OF MS AND ESC ON BDNF EXON I AND DNA METHYLTRANSFERASES (DNMT) MRNA LEVELS (QUANTITATIVE REVERSE-TRANSCRIPTION POLYMERASE CHAIN REACTION), ACETYLATED HISTONE H3, AND MECP2 BINDING TO THE BDNF PROMOTER I (CHROMATIN IMMUNOPRECIPITATION FOLLOWED BY REAL-TIME POLYMERASE CHAIN REACTION), AND BDNF PROTEIN LEVELS (ENZYME-LINKED IMMUNOSORBENT ASSAY). RESULTS: THE LEVELS OF BDNF PROTEIN, EXON I MRNA, HISTONE H3 ACETYLATION, AND DNMT1 AND DNMT3A MRNA WERE ALTERED IN THE MS GROUP COMPARED WITH THE CONTROL GROUP. SIGNIFICANT DECREASES WERE OBSERVED IN THE BDNF PROTEIN, EXON I MRNA, AND HISTONE H3 ACETYLATION LEVELS AND THERE WERE SIGNIFICANT INCREASES IN DNMT1 AND DNMT3A MRNA LEVELS. THE COMPARISON BETWEEN THE MS + ESC AND MS GROUPS REVEALED SIGNIFICANT INCREASES IN BDNF PROTEIN, EXON I MRNA, AND HISTONE H3 ACETYLATION LEVELS AND SIGNIFICANT DECREASES IN MECP2 AND DNMT1 AND DNMT3A MRNA LEVELS. CONCLUSION: THESE FINDINGS INDICATE THAT MS INDUCED EPIGENETIC CHANGES AT THE BDNF EXON I PROMOTER AND THESE CHANGES WERE PREVENTED BY ANTIDEPRESSANT DRUG TREATMENT DURING ADULTHOOD. 2018 19 3983 37 LONG-TERM EXPOSURE TO CIGARETTE SMOKE EXTRACT INDUCES HYPOMETHYLATION AT THE RUNX3 AND IGF2-H19 LOCI IN IMMORTALIZED HUMAN UROTHELIAL CELLS. CIGARETTE SMOKING IS THE SINGLE MOST IMPORTANT EPIDEMIOLOGICAL RISK FACTOR FOR BLADDER CANCER BUT IT IS NOT KNOWN WHETHER EXPOSURE OF UROTHELIAL CELLS TO THE SYSTEMIC SOLUBLE CONTENTS OF CIGARETTE SMOKE IS DIRECTLY CAUSATIVE TO BLADDER CANCER AND THE ASSOCIATED EPIGENETIC CHANGES SUCH AS TUMOR SUPPRESSOR GENE HYPERMETHYLATION. WE UNDERTOOK THIS STUDY TO INVESTIGATE IF LONG-TERM TREATMENT OF HUMAN UROTHELIAL CELLS WITH CIGARETTE SMOKE EXTRACT (CSE) RESULTS IN TUMOR SUPPRESSOR GENE HYPERMETHYLATION, A PHENOTYPE THAT WAS PREVIOUSLY ASSOCIATED WITH LONG-TERM CONSTANT CSE TREATMENT OF AIRWAY EPITHELIAL CELLS. WE CHRONICALLY TREATED AN IMMORTALIZED HUMAN UROTHELIAL CELL LINE UROTSA WITH CSE USING A CYCLIC DAILY REGIMEN BUT THE CELLS WERE CULTURED IN CSE-FREE MEDIUM BETWEEN DAILY TREATMENTS. BISULFITE SEQUENCING AND REAL-TIME PCR ARRAY-BASED METHYLATION PROFILING WERE EMPLOYED TO EVALUATE METHYLATION CHANGES AT TUMOR SUPPRESSOR GENE LOCI IN THE CHRONICALLY CSE-TREATED CELLS VERSUS THE PASSAGE-MATCHED UNTREATED CONTROL CELLS. THE RUNX3 TUMOR SUPPRESSOR GENE PROMOTER WAS HYPOMETHYLATED WITH A SIGNIFICANT INCREASE IN PROPORTION OF THE COMPLETELY UNMETHYLATED HAPLOTYPE AFTER THE LONG-TERM CSE TREATMENT; WHEREAS RUNX3 PROMOTER HYPERMETHYLATION WAS PREVIOUSLY REPORTED FOR BLADDER CANCERS OF SMOKERS. HYPOMETHYLATION INDUCED BY THE LONG-TERM CSE TREATMENT WAS ALSO OBSERVED FOR THE IGF2-H19 LOCUS. THE METHYLATION STATUS AT THE PRSS8/PROSTASIN AND 16 ADDITIONAL LOCI HOWEVER, WAS UNAFFECTED BY THE CHRONIC CSE TREATMENT. TRANSIENT CSE TREATMENT OVER 1 DAILY REGIMEN RESULTED IN TRANSCRIPTIONAL DOWN-REGULATION OF RUNX3 AND H19, BUT ONLY THE H19 TRANSCRIPTION WAS DOWN-REGULATED IN THE CHRONICALLY CSE-TREATED UROTHELIAL CELLS. TRANSCRIPTION OF A KEY ENZYME IN ONE-CARBON METABOLISM, DIHYDROFOLATE REDUCTASE (DHFR) WAS GREATLY REDUCED BY THE LONG-TERM CSE TREATMENT, POTENTIALLY SERVING AS A MECHANISM FOR THE HYPOMETHYLATION PHENOTYPE VIA A REDUCED SUPPLY OF METHYL DONOR. IN CONCLUSION, CHRONIC CYCLIC CSE TREATMENT OF UROTHELIAL CELLS INDUCED HYPOMETHYLATION RATHER THAN HYPERMETHYLATION AT SPECIFIC LOCI. 2013 20 531 45 ASTROCYTE REACTIVITY FOLLOWING BLAST EXPOSURE INVOLVES ABERRANT HISTONE ACETYLATION. BLAST INDUCED NEUROTRAUMA (BINT) IS A PREVALENT INJURY WITHIN MILITARY AND CIVILIAN POPULATIONS. THE INJURY IS CHARACTERIZED BY PERSISTENT INFLAMMATION AT THE CELLULAR LEVEL WHICH MANIFESTS AS A MULTITUDE OF COGNITIVE AND FUNCTIONAL IMPAIRMENTS. EPIGENETIC REGULATION OF TRANSCRIPTION OFFERS AN IMPORTANT CONTROL MECHANISM FOR GENE EXPRESSION AND CELLULAR FUNCTION WHICH MAY UNDERLIE CHRONIC INFLAMMATION AND RESULT IN NEURODEGENERATION. WE HYPOTHESIZE THAT ALTERED HISTONE ACETYLATION PATTERNS MAY BE INVOLVED IN BLAST INDUCED INFLAMMATION AND THE CHRONIC ACTIVATION OF GLIAL CELLS. THIS STUDY AIMED TO ELUCIDATE CHANGES TO HISTONE ACETYLATION OCCURRING FOLLOWING INJURY AND THE ROLES THESE CHANGES MAY HAVE WITHIN THE PATHOLOGY. SPRAGUE DAWLEY RATS WERE SUBJECTED TO EITHER A 10 OR 17 PSI BLAST OVERPRESSURE WITHIN AN ADVANCED BLAST SIMULATOR (ABS). SHAM ANIMALS UNDERWENT THE SAME PROCEDURES WITHOUT BLAST EXPOSURE. MEMORY IMPAIRMENTS WERE MEASURED USING THE NOVEL OBJECT RECOGNITION (NOR) TEST AT 2 AND 7 DAYS POST-INJURY. TISSUES WERE COLLECTED AT 7 DAYS FOR WESTERN BLOT AND IMMUNOHISTOCHEMISTRY (IHC) ANALYSIS. SHAM ANIMALS SHOWED INTACT MEMORY AT EACH TIME POINT. THE NOVEL OBJECT DISCRIMINATION DECREASED SIGNIFICANTLY BETWEEN TWO AND 7 DAYS FOR EACH INJURY GROUP (P < 0.05). THIS IS INDICATIVE OF THE ONSET OF MEMORY IMPAIRMENT. WESTERN BLOT ANALYSIS SHOWED GLIAL FIBRILLARY ACIDIC PROTEIN (GFAP), A KNOWN MARKER OF ACTIVATED ASTROCYTES, WAS ELEVATED IN THE PREFRONTAL CORTEX (PFC) FOLLOWING BLAST EXPOSURE FOR BOTH INJURY GROUPS. ANALYSIS OF HISTONE PROTEIN EXTRACT SHOWED NO CHANGES IN THE LEVEL OF ANY TOTAL HISTONE PROTEINS WITHIN THE PFC. HOWEVER, ACETYLATION LEVELS OF HISTONE H2B, H3, AND H4 WERE DECREASED IN BOTH GROUPS (P < 0.05). CO-LOCALIZATION IMMUNOFLUORESCENCE WAS USED TO FURTHER INVESTIGATE ANY POTENTIAL CORRELATION BETWEEN DECREASED HISTONE ACETYLATION AND ASTROCYTE ACTIVATION. THESE EXPERIMENTS SHOWED A SIMILAR DECREASE IN H3 ACETYLATION IN ASTROCYTES EXPOSED TO A 17 PSI BLAST BUT NOT A 10 PSI BLAST. FURTHER INVESTIGATION OF GENE EXPRESSION BY POLYMERASE CHAIN REACTION (PCR) ARRAY, SHOWED DYSREGULATION OF SEVERAL CYTOKINE AND CYTOKINE RECEPTORS THAT ARE INVOLVED IN NEUROINFLAMMATORY PROCESSES. WE HAVE SHOWN ABERRANT HISTONE ACETYLATION PATTERNS INVOLVED IN BLAST INDUCED ASTROGLIOSIS AND COGNITIVE IMPAIRMENTS. FURTHER UNDERSTANDING OF THEIR ROLE IN THE INJURY PROGRESSION MAY LEAD TO NOVEL THERAPEUTIC TARGETS. 2016