1 4400 136 MODULATION OF HEPATIC STELLATE CELLS BY MUTAFLOR((R)) PROBIOTIC IN NON-ALCOHOLIC FATTY LIVER DISEASE MANAGEMENT. BACKGROUND: NAFLD AND NASH ARE EMERGING AS PRIMARY CAUSES OF CHRONIC LIVER DISEASE, INDICATING A NEED FOR AN EFFECTIVE TREATMENT. MUTAFLOR(R) PROBIOTIC, A MICROBIAL TREATMENT OF INTEREST, WAS EFFECTIVE IN SUSTAINING REMISSION IN ULCERATIVE COLITIS PATIENTS. OBJECTIVE: TO CONSTRUCT A GENETIC-EPIGENETIC NETWORK LINKED TO HSC SIGNALING AS A MODULATOR OF NAFLD/NASH PATHOGENESIS, THEN ASSESS THE EFFECTS OF MUTAFLOR((R)) ON THIS NETWORK. METHODS: FIRST, IN SILICO ANALYSIS WAS USED TO CONSTRUCT A GENETIC-EPIGENETIC NETWORK LINKED TO HSC SIGNALING. SECOND, AN INVESTIGATION USING RATS, INCLUDING HFHSD INDUCED NASH AND MUTAFLOR((R)) TREATED ANIMALS, WAS DESIGNED. EXPERIMENTAL PROCEDURES INCLUDED BIOCHEMICAL AND HISTOPATHOLOGIC ANALYSIS OF RAT BLOOD AND LIVER SAMPLES. AT THE MOLECULAR LEVEL, THE EXPRESSION OF GENETIC (FOXA2, TEAD2, AND LATS2 MRNAS) AND EPIGENETIC (MIR-650, RPARP AS-1 LNCRNA) NETWORK WAS MEASURED BY REAL-TIME PCR. PCR RESULTS WERE VALIDATED WITH IMMUNOHISTOCHEMISTRY (ALPHA-SMA AND LATS2). TARGET EFFECTOR PROTEINS, IL-6 AND TGF-BETA, WERE ESTIMATED BY ELISA. RESULTS: MUTAFLOR((R)) ADMINISTRATION MINIMIZED BIOCHEMICAL AND HISTOPATHOLOGIC ALTERATIONS CAUSED BY NAFLD/NASH. HSC ACTIVATION AND EXPRESSION OF PROFIBROGENIC IL-6 AND TGF-BETA EFFECTOR PROTEINS WERE REDUCED VIA INHIBITION OF HEDGEHOG AND HIPPO PATHWAYS. PATHWAYS MAY HAVE BEEN INHIBITED THROUGH UPREGULATION OF RPARP AS-1 LNCRNA WHICH IN TURN DOWNREGULATED THE EXPRESSION OF MIR-650, FOXA2 MRNA AND TEAD2 MRNA AND UPREGULATED LATS2 MRNA EXPRESSION. CONCLUSION: MUTAFLOR((R)) MAY SLOW THE PROGRESSION OF NAFLD/NASH BY MODULATING A GENETIC-EPIGENETIC NETWORK LINKED TO HSC SIGNALING. THE PROBIOTIC MAY BE A USEFUL MODALITY FOR THE PREVENTION AND TREATMENT OF NAFLD/NASH. 2022 2 3128 39 GIPC-REGULATED IGFBP-3 PROMOTES HSC MIGRATION IN VITRO AND PORTAL HYPERTENSION IN VIVO THROUGH A BETA1-INTEGRIN PATHWAY. BACKGROUND & AIMS: TRANSFORMING GROWTH FACTOR (TGF-BETA)-INDUCED ACTIVATION OF QUIESCENT HEPATIC STELLATE CELLS (HSCS) AND THEIR TRANSFORMATION TO MYOFIBROBLASTS IS A KEY EVENT IN LIVER FIBROSIS AND PORTAL HYPERTENSION. GIPC (ALSO REFERRED TO AS SYNECTIN) IS A DOWNSTREAM SIGNAL ACTIVATION MOLECULE OF TGF-BETA AND OTHER RECEPTORS. IN THIS STUDY, WE SOUGHT TO IDENTIFY NOVEL GENES TARGETED BY TGF-BETA AND GIPC AND ELUCIDATE IF AND HOW THEY MAY CONTRIBUTE TO LIVER FIBROSIS. METHODS: WE PERFORMED SEQUENTIAL MESSENGER RNA SEQUENCING ANALYSIS ON TGF-BETA-STIMULATED HSCS AND THEN ON TGF-BETA-STIMULATED HSCS IN THE PRESENCE AND ABSENCE OF GIPC ALSO REFERRED TO AS SYNECTIN (GIPC) KNOCKDOWN. INSULIN-LIKE GROWTH FACTOR BINDING PROTEIN-3 (IGFBP-3) TRANSPORT PROTEIN EMERGED AS A TOP ACTIVATION TARGET OF BOTH TGF-BETA AND GIPC. QUANTITATIVE POLYMERASE CHAIN REACTION, ENZYME-LINKED IMMUNOSORBENT ASSAY, TARGETED CHROMATIN IMMUNOPRECIPITATION, AND WESTERN BLOT ANALYSIS WERE DONE FOR FURTHER CONFIRMATION. RESULTS: IGFBP-3, AN INSULIN GROWTH FACTOR TRANSPORT PROTEIN, EMERGED AS A TOP ACTIVATION TARGET OF BOTH TGF-BETA AND GIPC, WHICH WAS CONFIRMED BY QUANTITATIVE POLYMERASE CHAIN REACTION, ENZYME-LINKED IMMUNOSORBENT ASSAY, AND WESTERN BLOT ANALYSIS. TARGETED CHROMATIN IMMUNOPRECIPITATION SHOWED THAT GIPC INCREASES THE HISTONE 3 LYSINE 27 (H3K27) ACETYLATION ACTIVATING MARK AND CONCURRENTLY DECREASES THE H3K27 INHIBITORY TRIMETHYLATION (H3K27M3) MARK, PROVIDING AN EPIGENETIC CORRELATE TO THE GENE REGULATION CHANGES. IN VIVO, GLOBAL KNOCKOUT OF IGFBP-3 MICE RESULTED IN ATTENUATION OF HSC ACTIVATION MARKERS AND ATTENUATION OF PORTAL PRESSURE IN RESPONSE TO CHRONIC LIVER INJURY MODELS. ANALYSIS OF SERUM LEVELS FROM CIRRHOTIC PATIENTS ALSO SHOWED AN IGFBP-3 INCREASE OF MORE THAN 2-FOLD COMPARED WITH HEALTHY CONTROLS. FINALLY, IN VITRO MECHANISM STUDIES SHOWED THAT IGFBP-3 PROMOTES HSC MIGRATION THROUGH INTEGRIN-DEPENDENT PHOSPHORYLATION OF PROTEIN KINASE B. CONCLUSIONS: TGF-BETA UP-REGULATES IGFBP-3 THROUGH GIPC, LEADING TO INCREASED HSC MIGRATION IN VITRO AND PROMOTES PORTAL HYPERTENSION IN VIVO. THESE STUDIES SUPPORT THE ROLE OF IGFBP-3 AS A POTENTIAL PATHOPHYSIOLOGIC TARGET OR BIOMARKER IN CHRONIC LIVER DISEASE. 2020 3 5604 41 ROSAVIN AMELIORATES HEPATIC INFLAMMATION AND FIBROSIS IN THE NASH RAT MODEL VIA TARGETING HEPATIC CELL DEATH. BACKGROUND: NON-ALCOHOLIC FATTY LIVER DISEASE (NAFLD) REPRESENTS THE MOST COMMON FORM OF CHRONIC LIVER DISEASE THAT URGENTLY NEEDS EFFECTIVE THERAPY. ROSAVIN, A MAJOR CONSTITUENT OF THE RHODIOLA ROSEA PLANT OF THE FAMILY CRASSULACEAE, IS BELIEVED TO EXHIBIT MULTIPLE PHARMACOLOGICAL EFFECTS ON DIVERSE DISEASES. HOWEVER, ITS EFFECT ON NON-ALCOHOLIC STEATOHEPATITIS (NASH), THE PROGRESSIVE FORM OF NAFLD, AND THE UNDERLYING MECHANISMS ARE NOT FULLY ILLUSTRATED. AIM: INVESTIGATE THE PHARMACOLOGICAL ACTIVITY AND POTENTIAL MECHANISM OF ROSAVIN TREATMENT ON NASH MANAGEMENT VIA TARGETING HEPATIC CELL DEATH-RELATED (HSPD1/TNF/MMP14/ITGB1) MRNAS AND THEIR UPSTREAM NONCODING RNA REGULATORS (MIRNA-6881-5P AND LNC-SPARCL1-1:2) IN NASH RATS. RESULTS: HIGH SUCROSE HIGH FAT (HSHF) DIET-INDUCED NASH RATS WERE TREATED WITH DIFFERENT CONCENTRATIONS OF ROSAVIN (10, 20, AND 30 MG/KG/DAY) FOR THE LAST FOUR WEEKS OF DIETARY MANIPULATION. THE DATA REVEALED THAT ROSAVIN HAD THE ABILITY TO MODULATE THE EXPRESSION OF THE HEPATIC CELL DEATH-RELATED RNA PANEL THROUGH THE UPREGULATION OF BOTH (HSPD1/TNF/MMP14/ITGB1) MRNAS AND THEIR EPIGENETIC REGULATORS (MIRNA-6881-5P AND LNC-SPARCL1-1:2). MOREOVER, ROSAVIN AMELIORATED THE DETERIORATION IN BOTH LIVER FUNCTIONS AND LIPID PROFILE, AND THEREBY IMPROVED THE HEPATIC INFLAMMATION, FIBROSIS, AND APOPTOSIS, AS EVIDENCED BY THE DECREASED PROTEIN LEVELS OF IL6, TNF-ALPHA, AND CASPASE-3 IN LIVER SECTIONS OF TREATED ANIMALS COMPARED TO THE UNTREATED NASH RATS. CONCLUSION: ROSAVIN HAS DEMONSTRATED A POTENTIAL ABILITY TO ATTENUATE DISEASE PROGRESSION AND INHIBIT HEPATIC CELL DEATH IN THE NASH ANIMAL MODEL. THE PRODUCED EFFECT WAS CORRELATED WITH UPREGULATION OF THE HEPATIC CELL DEATH-RELATED (HSPD1, TNF, MMP14, AND ITGB1) MRNAS-(MIRNA-6881-5P-(LNC-SPARCL1-1:2) RNA PANEL. 2022 4 3049 29 GENOME-WIDE ANALYSIS REVEALS ZINC TRANSPORTER ZIP9 REGULATED BY DNA METHYLATION PROMOTES RADIATION-INDUCED SKIN FIBROSIS VIA THE TGF-BETA SIGNALING PATHWAY. RADIATION-INDUCED SKIN FIBROSIS IS A DETRIMENTAL AND CHRONIC DISORDER THAT OCCURS AFTER RADIATION EXPOSURE. DNA METHYLATION HAS BEEN CHARACTERIZED AS AN IMPORTANT REGULATORY MECHANISM OF MULTIPLE PATHOLOGICAL PROCESSES. IN THIS STUDY, WE COMPARED THE GENOME-WIDE DNA METHYLATION STATUS IN RADIATION-INDUCED FIBROTIC SKIN AND ADJACENT NORMAL TISSUES OF RATS BY METHYLATED DNA IMMUNOPRECIPITATION SEQUENCING. RADIATION-INDUCED FIBROTIC SKIN SHOWED DIFFERENTIALLY METHYLATED REGIONS ASSOCIATED WITH 3,650 PROTEIN-CODING GENES, 72 MICRORNAS, 5,836 LONG NONCODING RNAS AND 3 PIWI-INTERACTING RNAS. BY INTEGRATING THE MRNA AND METHYLATION PROFILES, THE ZINC TRANSPORTER SLC39A9/ZIP9 WAS INVESTIGATED IN GREATER DETAIL. THE PROTEIN LEVEL OF ZIP9 WAS INCREASED IN IRRADIATED SKIN TISSUES OF HUMANS, MONKEYS, AND RATS, ESPECIALLY IN RADIOGENIC FIBROTIC SKIN TISSUES. RADIATION INDUCED THE DEMETHYLATION OF A CPG DINUCLEOTIDE IN EXON 1 OF ZIP9 THAT RESULTED IN RECRUITMENT OF THE TRANSCRIPTIONAL FACTOR SP1 AND INCREASED ZIP9 EXPRESSION. OVEREXPRESSION OF ZIP9 RESULTED IN ACTIVATION OF THE PROFIBROTIC TRANSFORMING GROWTH FACTOR-BETA SIGNALING PATHWAY THROUGH PROTEIN KINASE B IN HUMAN FIBROBLASTS. IN ADDITION, RADIATION-INDUCED SKIN FIBROSIS WAS ASSOCIATED WITH INCREASED ZINC ACCUMULATION. THE ZINC CHELATOR N,N,N',N'-TETRAKIS(2-PYRIDYLMETHYL)-1,2-ETHYLENEDIAMINE ABROGATED ZIP9-INDUCED ACTIVATION OF THE TRANSFORMING GROWTH FACTOR-BETA SIGNALING PATHWAY AND ATTENUATED RADIATION-INDUCED SKIN FIBROSIS IN A RAT MODEL. IN SUMMARY, OUR FINDINGS ILLUSTRATE EPIGENETIC REGULATION OF ZIP9 AND ITS CRITICAL ROLE IN PROMOTING RADIATION-INDUCED SKIN FIBROSIS. 2020 5 2427 40 EPIGENETIC SILENCING OF MICRORNA-125B-5P PROMOTES LIVER FIBROSIS IN NONALCOHOLIC FATTY LIVER DISEASE VIA INTEGRIN ALPHA8-MEDIATED ACTIVATION OF RHOA SIGNALING PATHWAY. BACKGROUND: NONALCOHOLIC FATTY LIVER DISEASE (NAFLD) IS ONE OF THE MOST COMMON CHRONIC LIVER DISEASES THAT MAY PROGRESS TO LIVER FIBROSIS OR CANCER. THE PRESENT STUDY AIMED TO INVESTIGATE THE ROLE OF MICRORNA-125B-5P (MIR-125B-5P) IN NAFLD AND TO FURTHER EXPLORE UNDERLYING MOLECULAR MECHANISMS. METHODS: A MOUSE MODEL OF NAFLD WAS CONSTRUCTED BY HIGH CHOLESTEROL DIET FEEDING AND A CELL-MODEL WAS DEVELOPED BY TREATING THE MOUSE LIVER CELL LINE NCTC1469 WITH PALMITIC ACID. GAIN- AND LOSS-OF-FUNCTION EXPERIMENTS WERE PERFORMED TO DETERMINE THE EFFECTS OF MIR-125B-5P, INTEGRIN ALPHA8 (ITGA8), AND THE RHOA SIGNALING PATHWAY ON LIVER FIBROSIS IN NAFLD. AFTER THE EXPRESSION LEVELS OF MIR-125B-5P, ITGA8, AND RHOA WERE DETERMINED, LIVER FIBROSIS WAS EVALUATED IN VIVO AND IN VITRO. THE BINDING RELATIONSHIP OF MIR-125B-5P AND ITGA8 WAS THEN VALIDATED. FINALLY, MIR-125B-5P PROMOTER METHYLATION IN NAFLD LIVER TISSUES AND CELLS WAS DETERMINED. RESULTS: IN NAFLD CLINICAL SAMPLES, MOUSE MODEL, AND CELL-MODEL, MIR-125B-5P EXPRESSION WAS REDUCED, WHILE ITGA8 EXPRESSION WAS INCREASED. MOREOVER, MIR-125B-5P TARGETED AND DOWNREGULATED ITGA8, LEADING TO INHIBITION OF THE RHOA SIGNALING PATHWAY. IN NAFLD LIVER TISSUES AND CELLS, THE CPG ISLAND IN THE MIR-125B-5P PROMOTER WAS METHYLATED, CAUSING EPIGENETIC SILENCING OF MIR-125B-5P. BOTH MIR-125B-5P SILENCING AND ITGA8 OVEREXPRESSION PROMOTED IN VITRO AND IN VIVO LIVER FIBROSIS IN NAFLD VIA ACTIVATION OF THE RHOA SIGNALING PATHWAY. CONCLUSIONS: COLLECTIVELY, EPIGENETIC SILENCING OF MIR-125B-5P UPREGULATES ITGA8 EXPRESSION TO ACTIVATE THE RHOA SIGNALING PATHWAY, LEADING TO LIVER FIBROSIS IN NAFLD. 2020 6 3198 27 HDAC-LINKED "PROLIFERATIVE" MIRNA EXPRESSION PATTERN IN PANCREATIC NEUROENDOCRINE TUMORS. EPIGENETIC FACTORS ARE ESSENTIALLY INVOLVED IN CARCINOGENESIS, TUMOR PROMOTION, AND CHEMORESISTANCE. TWO EPIGENETIC KEY PLAYERS ARE MIRNAS AND HISTONE DEACETYLASES (HDACS). AS PREVIOUSLY SHOWN BY OWN THEORETICAL DATABANK ANALYSIS, THE CROSSTALK BETWEEN MIRNAS AND HDACS IS RELEVANT IN DIFFERENT HUMAN CHRONIC DISEASES AND CANCEROGENIC PATHWAYS. WE AIMED TO INVESTIGATE A POTENTIAL CONNECTION BETWEEN THE EXPRESSION OF A WELL-DEFINED SUBSET OF "PROLIFERATION-ASSOCIATED" MIRNAS AND THE EXPRESSION OF HDACS AS WELL AS CLINICAL PARAMETERS IN PANCREATIC NEUROENDOCRINE TUMORS (PNETS). MATERIALS AND METHODS: EXPRESSION LEVELS OF MIRNA132-3P, MIRNA145-5P, MIRNA183-5P, MIRNA34A-5P, AND MIRNA449A IN 57 PNETS RESECTED BETWEEN 1997 AND 2015 WERE MEASURED AND LINKED TO THE IMMUNOHISTOCHEMICAL EXPRESSION PATTERN OF MEMBERS OF THE FOUR HDAC CLASSES ON HUMAN TISSUE MICROARRAYS. ALL PNET CASES WERE CLINICALLY AND PATHOLOGICALLY CHARACTERIZED ACCORDING TO PUBLISHED GUIDELINES. CORRELATION ANALYSIS REVEALED A SIGNIFICANT ASSOCIATION BETWEEN EXPRESSION OF SPECIFIC MIRNAS AND TWO MEMBERS OF THE HDAC FAMILY (HDAC3 AND HDAC4). ADDITIONALLY, A LINKAGE BETWEEN MIRNA EXPRESSION AND CLINICO-PATHOLOGICAL PARAMETERS LIKE GRADING, TNM-STAGING, AND HORMONE ACTIVITY WAS FOUND. MOREOVER, OVERALL AND DISEASE-FREE SURVIVAL IS STATISTICALLY CORRELATED WITH THE EXPRESSION OF THE INVESTIGATED MIRNAS. OVERALL, WE DEMONSTRATED THAT SPECIFIC MIRNAS COULD BE LINKED TO HDAC EXPRESSION IN PNETS. ESPECIALLY MIRNA449A (ASSOCIATED WITH HDAC3/4) SEEMS TO PLAY AN IMPORTANT ROLE IN PNET PROLIFERATION AND COULD BE A POTENTIAL PROGNOSTIC FACTOR FOR POOR SURVIVAL. THESE FIRST DATA COULD HELP, TO IMPROVE OUR KNOWLEDGE OF THE COMPLEX INTERACTIONS OF THE EPIGENETIC DRIVERS IN PNETS FOR FURTHER THERAPEUTIC APPROACHES. 2018 7 5636 44 SERELAXIN ALLEVIATES CARDIAC FIBROSIS THROUGH INHIBITING ENDOTHELIAL-TO-MESENCHYMAL TRANSITION VIA RXFP1. RATIONALE: CARDIAC FIBROSIS IS AN INTEGRAL CONSTITUENT OF EVERY FORM OF CHRONIC HEART DISEASE, AND PERSISTENCE OF FIBROSIS REDUCES TISSUE COMPLIANCE AND ACCELERATES THE PROGRESSION TO HEART FAILURE. RELAXIN-2 IS A HUMAN HORMONE, WHICH HAS VARIOUS PHYSIOLOGICAL FUNCTIONS SUCH AS MEDIATING RENAL VASODILATION IN PREGNANCY. ITS RECOMBINANT FORM SERELAXIN HAS RECENTLY BEEN TESTED IN CLINICAL TRIALS AS A THERAPY FOR ACUTE HEART FAILURE BUT DID NOT MEET ITS PRIMARY ENDPOINTS. THE AIM OF THIS STUDY IS TO EXAMINE WHETHER SERELAXIN HAS AN ANTI-FIBROTIC EFFECT IN THE HEART AND THEREFORE COULD BE BENEFICIAL IN CHRONIC HEART FAILURE. METHODS: WE UTILIZED TWO DIFFERENT CARDIAC FIBROSIS MOUSE MODELS (ASCENDING AORTIC CONSTRICTION (AAC) AND ANGIOTENSIN II (ATII) ADMINISTRATION VIA OSMOTIC MINIPUMPS) TO ASSESS THE ANTI-FIBROTIC POTENTIAL OF SERELAXIN. HISTOLOGICAL ANALYSIS, IMMUNOFLUORESCENCE STAINING AND MOLECULAR ANALYSIS WERE PERFORMED TO ASSESS THE FIBROSIS LEVEL AND INDICATE ENDOTHELIAL CELLS WHICH ARE UNDERGOING ENDMT. IN VITRO TGFBETA1-INDUCED ENDOTHELIAL-TO-MESENCHYMAL TRANSITION (ENDMT) ASSAYS WERE PERFORMED IN HUMAN CORONARY ARTERY ENDOTHELIAL CELLS AND MOUSE CARDIAC ENDOTHELIAL CELLS (MCECS) AND WERE EXAMINED USING MOLECULAR METHODS. CHROMATIN IMMUNOPRECIPITATION-QPCR ASSAY WAS UTILIZED TO IDENTIFY THE SERELAXIN EFFECT ON CHROMATIN REMODELING IN THE RXFP1 PROMOTER REGION IN MCECS. RESULTS: OUR RESULTS DEMONSTRATE A SIGNIFICANT AND DOSE-DEPENDENT ANTI-FIBROTIC EFFECT OF SERELAXIN IN THE HEART IN BOTH MODELS. WE FURTHER SHOW THAT SERELAXIN MEDIATES THIS EFFECT, AT LEAST IN PART, THROUGH INHIBITION OF ENDMT THROUGH THE ENDOTHELIAL RELAXIN FAMILY PEPTIDE RECEPTOR 1 (RXFP1). WE FURTHER DEMONSTRATE THAT SERELAXIN ADMINISTRATION IS ABLE TO INCREASE ITS OWN RECEPTOR EXPRESSION (RXFP1) THROUGH EPIGENETIC REGULATION IN FORM OF HISTONE MODIFICATIONS BY ATTENUATING TGFBETA-PSMAD2/3 SIGNALING IN ENDOTHELIAL CELLS. CONCLUSIONS: THIS STUDY IS THE FIRST TO IDENTIFY THAT SERELAXIN INCREASES THE EXPRESSION OF ITS OWN RECEPTOR RXFP1 AND THAT THIS MEDIATES THE INHIBITION OF ENDMT AND CARDIAC FIBROSIS, SUGGESTING THAT SERELAXIN MAY HAVE A BENEFICIAL EFFECT AS ANTI-FIBROTIC THERAPY IN CHRONIC HEART FAILURE. 2020 8 3327 30 HISTONE DEACETYLASE 4 PROMOTES CHOLESTATIC LIVER INJURY IN THE ABSENCE OF PROHIBITIN-1. PROHIBITIN-1 (PHB1) IS AN EVOLUTIONARILY CONSERVED PLEIOTROPIC PROTEIN THAT PARTICIPATES IN DIVERSE PROCESSES DEPENDING ON ITS SUBCELLULAR LOCALIZATION AND INTERACTOME. RECENT DATA HAVE INDICATED A DIVERSE ROLE FOR PHB1 IN THE PATHOGENESIS OF OBESITY, CANCER, AND INFLAMMATORY BOWEL DISEASE, AMONG OTHERS. DATA PRESENTED HERE SUGGEST THAT PHB1 IS ALSO LINKED TO CHOLESTATIC LIVER DISEASE. EXPRESSION OF PHB1 IS MARKEDLY REDUCED IN PATIENTS WITH PRIMARY BILIARY CIRRHOSIS AND BILIARY ATRESIA OR WITH ALAGILLE SYNDROME, TWO MAJOR PEDIATRIC CHOLESTATIC CONDITIONS. IN THE EXPERIMENTAL MODEL OF BILE DUCT LIGATION, SILENCING OF PHB1 INDUCED LIVER FIBROSIS, REDUCED ANIMAL SURVIVAL, AND INDUCED BILE DUCT PROLIFERATION. IMPORTANTLY, THE MODULATORY EFFECT OF PHB1 IS NOT DEPENDENT ON ITS KNOWN MITOCHONDRIAL FUNCTION. ALSO, PHB1 INTERACTS WITH HISTONE DEACETYLASE 4 (HDAC4) IN THE PRESENCE OF BILE ACIDS. HENCE, PHB1 DEPLETION LEADS TO INCREASED NUCLEAR HDAC4 CONTENT AND ITS ASSOCIATED EPIGENETIC CHANGES. REMARKABLY, HDAC4 SILENCING AND THE ADMINISTRATION OF THE HDAC INHIBITOR PARTHENOLIDE DURING OBSTRUCTIVE CHOLESTASIS IN VIVO PROMOTE GENOMIC REPROGRAMMING, LEADING TO REGRESSION OF THE FIBROTIC PHENOTYPE IN LIVER-SPECIFIC PHB1 KNOCKOUT MICE. CONCLUSION: PHB1 IS AN IMPORTANT MEDIATOR OF CHOLESTATIC LIVER INJURY THAT REGULATES THE ACTIVITY OF HDAC4, WHICH CONTROLS SPECIFIC EPIGENETIC MARKERS; THESE RESULTS IDENTIFY POTENTIAL NOVEL STRATEGIES TO TREAT LIVER INJURY AND FIBROSIS, PARTICULARLY AS A CONSEQUENCE OF CHRONIC CHOLESTASIS. 2015 9 4233 33 METHYLATION OF SEPTIN9 MEDIATED BY DNMT3A ENHANCES HEPATIC STELLATE CELLS ACTIVATION AND LIVER FIBROGENESIS. LIVER FIBROSIS, RESULTING FROM CHRONIC AND PERSISTENT INJURY TO THE LIVER, IS A WORLDWIDE HEALTH PROBLEM. ADVANCED LIVER FIBROSIS RESULTS IN CIRRHOSIS, LIVER FAILURE AND EVEN HEPATOCELLULAR CANCER (HCC), OFTEN EVENTUALLY REQUIRING LIVER TRANSPLANTATION, POSES A HUGE HEALTH BURDEN ON THE GLOBAL COMMUNITY. HOWEVER, THE SPECIFIC PATHOGENESIS OF LIVER FIBROSIS REMAINS NOT FULLY UNDERSTOOD. NUMEROUS BASIC AND CLINICAL STUDIES HAVE PROVIDED EVIDENCE THAT EPIGENETIC MODIFICATIONS, ESPECIALLY DNA METHYLATION, MIGHT CONTRIBUTE TO THE ACTIVATION OF HEPATIC STELLATE CELLS (HSCS), THE PIVOTAL CELL TYPE RESPONSIBLE FOR THE FIBROUS SCAR IN LIVER. HERE, REDUCED REPRESENTATION BISULFITE SEQUENCING (RRBS) AND BISULFITE PYROSEQUENCING PCR (BSP) ANALYSIS IDENTIFIED HYPERMETHYLATION STATUS OF SEPTIN9 (SEPT9) GENE IN LIVER FIBROGENESIS. SEPT9 PROTEIN WAS DRAMATICALLY DECREASED IN LIVERS OF CCL4-TREATED MICE AND IMMORTALIZED HSC-T6 CELLS EXPOSED TO TGF-BETA1. NEVERTHELESS, THE SUPPRESSION OF SEPT9 COULD BE BLOCKED BY DNMT3A-SIRNA AND DNA METHYLTRANSFERASE INHIBITOR, 5-AZA-2'-DEOXYCYTIDINE (5-AZADC). OVEREXPRESSED SEPT9 ATTENUATED TGF-BETA1-INDUCED EXPRESSION OF MYOFIBROBLAST MARKERS ALPHA-SMA AND COL1A1, ACCOMPANIED BY UP-REGULATION OF CELL APOPTOSIS-RELATED PROTEINS. CONVERSELY, RNAI-MEDIATED SILENCING OF SEPT9 ENHANCED ACCUMULATION OF EXTRACELLULAR MATRIX. THESE OBSERVATIONS SUGGESTED THAT SEPT9 CONTRIBUTED TO ALLEVIATE LIVER FIBROSIS MIGHT PARTIALLY THROUGH PROMOTING ACTIVATED HSCS APOPTOSIS AND THIS ANTI-FIBROGENESIS EFFECT MIGHT BE BLOCKED BY DNMT-3A MEDIATED METHYLATION OF SEPT9. THEREFORE, PHARMACOLOGICAL AGENTS THAT INHIBIT SEPT9 METHYLATION AND INCREASE ITS EXPRESSION COULD BE CONSIDERED AS VALUABLE TREATMENTS FOR LIVER FIBROSIS. 2017 10 5914 36 TARGETED-BISULFITE SEQUENCE ANALYSIS OF THE METHYLATION OF CPG ISLANDS IN GENES ENCODING PNPLA3, SAMM50, AND PARVB OF PATIENTS WITH NON-ALCOHOLIC FATTY LIVER DISEASE. BACKGROUND & AIMS: THE PATHOGENESIS OF NON-ALCOHOLIC FATTY LIVER DISEASE (NAFLD) IS AFFECTED BY EPIGENETIC FACTORS AS WELL AS BY GENETIC VARIATION. METHODS: WE PERFORMED TARGETED-BISULFITE SEQUENCING TO DETERMINE THE LEVELS OF DNA METHYLATION OF 4 CPG ISLANDS (CPG99, CPG71, CPG26, AND CPG101) IN THE REGULATORY REGIONS OF PNPLA3, SAMM50, PARVB VARIANT 1, AND PARVB VARIANT 2, RESPECTIVELY. WE COMPARED THE LEVELS OF METHYLATION OF DNA IN THE LIVERS OF THE FIRST AND SECOND SETS OF PATIENTS WITH MILD (FIBROSIS STAGES 0 AND 1) OR ADVANCED (FIBROSIS STAGES 2 TO 4) NAFLD AND IN THOSE OF PATIENTS WITH MILD (F0 TO F2) OR ADVANCED (F3 AND F4) CHRONIC HEPATITIS C INFECTION. THE HEPATIC MRNA LEVELS OF PNPLA3, SAMM50, AND PARVB WERE MEASURED USING QPCR. RESULTS: CPG26, WHICH RESIDES IN THE REGULATORY REGION OF PARVB VARIANT 1, WAS MARKEDLY HYPOMETHYLATED IN THE LIVERS OF PATIENTS WITH ADVANCED NAFLD. CONVERSELY, CPG99 IN THE REGULATORY REGION OF PNPLA3 WAS SUBSTANTIALLY HYPERMETHYLATED IN THESE PATIENTS. THESE DIFFERENCES IN DNA METHYLATION WERE REPLICATED IN A SECOND SET OF PATIENTS WITH NAFLD OR CHRONIC HEPATITIS C. PNPLA3 MRNA LEVELS IN THE LIVER OF THE SAME SECTION OF A BIOPSY SPECIMEN USED FOR GENOMIC DNA PREPARATION WERE LOWER IN PATIENTS WITH ADVANCED NAFLD COMPARED WITH THOSE WITH MILD NAFLD AND CORRELATED INVERSELY WITH CPG99 METHYLATION IN LIVER DNA. MOREOVER, THE LEVELS OF CPG99 METHYLATION AND PNPLA3 MRNA WERE AFFECTED BY THE RS738409 GENOTYPE. CONCLUSIONS: HYPOMETHYLATION OF CPG26 AND HYPERMETHYLATION OF CPG99 MAY CONTRIBUTE TO THE SEVERITY OF FIBROSIS IN PATIENTS WITH NAFLD OR CHRONIC HEPATITIS C INFECTION. 2015 11 2761 30 EXPRESSION OF TESTIS-SPECIFIC GENES, TEX101 AND ODF4, IN CHRONIC MYELOID LEUKEMIA AND EVALUATION OF TEX101 IMMUNOGENICITY. BACKGROUND AND OBJECTIVES: CANCER-TESTIS (CT) ANTIGENS ARE A GROUP OF ANTIGENS WITH A RESTRICTED EXPRESSION IN NORMAL TISSUES, EXCEPT TESTIS, AND THEY HAVE ABERRANT EXPRESSION IN DIFFERENT TUMORS. THIS PATTERN OF EXPRESSION HAS MADE THEM PROMISING TARGETS FOR IMMUNOTHERAPY AND CANCER DETECTION. OUR AIM WAS TO FIND NEW MEMBERS OF THIS GROUP THAT MIGHT BE USEFUL AS MARKERS IN THE DETECTION OF CANCER AND IMMUNOTHERAPY. DESIGN AND SETTING: A DESCRIPTIVE STUDY CONDUCTED IN REFERRAL CENTERS OF TEHRAN UNIVERSITY OF MEDICAL SCIENCE FROM JANUARY 2008 TO JANUARY 2009. PATIENTS AND METHODS: WE ANALYZED THE EXPRESSION OF TWO TESTIS-SPECIFIC GENES NAMED ODF4 (OUTER DENSE FIBER OF SPERM TAILS 4) AND TEX101 (TESTIS EXPRESSED 101) IN 20 CHRONIC MYELOID LEUKEMIA (CML) AND 20 NORMAL SAMPLES BY REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION AND SEQUENCING. IMMUNOGENICITY OF TEX101 WAS EVALUATED BY MEANS OF ENZYME-LINKED IMMUNOSORBENT ASSAY. RESULTS: THESE TWO GENES WERE EXPRESSED IN 30% OF CML PATIENTS BUT NOT IN ANY OF THE HEALTHY DONORS. HUMORAL RESPONSE AGAINST TEX101 WAS NOT DETECTED IN ANY SAMPLES. CONCLUSIONS: TEX101 AND ODF4 ARE CT GENES USEFUL FOR DETECTION OF CML. UNLIKE MANY CT GENES, OVEREXPRESSION OF TEX101 WAS NOT SHOWN TO INDUCE IMMUNOLOGIC RESPONSES IN THESE SAMPLES. ACCORDING TO THE PREVIOUS STUDIES, OVEREXPRESSION OF TEX101 LEADS TO SUPPRESSION OF CANCER INVASION AND METASTASIS; THUS, THE INDUCTION OF THE EXPRESSION OF TEX101 IN CANCER BY EPIGENETIC MECHANISMS MAY BE A TREATMENT STRATEGY. 2012 12 1067 30 CLINICAL UTILITY OF PDSS2 EXPRESSION TO STRATIFY PATIENTS AT RISK FOR RECURRENCE OF HEPATOCELLULAR CARCINOMA. IDENTIFICATION OF NOVEL GENETIC AND EPIGENETIC ALTERATIONS IS REQUIRED FOR OPTIMAL STRATIFICATION OF PATIENTS WITH HEPATOCELLULAR CARCINOMA (HCC) AT RISK FOR RECURRENCE AND ADVERSE PROGNOSIS. COENZYME Q10 (COQ10), WHICH MEDIATES APOPTOSIS, IS SYNTHESIZED BY PRENYL DIPHOSPHATE SYNTHASE SUBUNIT 2 (PDSS2). IN THE PRESENT STUDY WE EVALUATED THE CLINICAL SIGNIFICANCE AND REGULATORY MECHANISMS OF PDSS2 EXPRESSION IN HCC. PDSS2 EXPRESSION LEVELS AND THOSE OF GENES ENCODING POTENTIALLY INTERACTING PROTEINS AS WELL AS THE METHYLATION STATUS OF THE PDSS2 PROMOTER REGION WERE ANALYZED IN HCC CELL LINES. PDSS2 MRNA LEVELS IN 151 PAIRS OF RESECTED SPECIMENS WERE DETERMINED TO EVALUATE THE ASSOCIATION OF PDSS2 EXPRESSION AND CLINICOPATHOLOGICAL FACTORS. THE EXPRESSION AND DISTRIBUTION OF PDSS2 WERE DETERMINED USING IMMUNOHISTOCHEMISTRY. PDSS2 MRNA EXPRESSION WAS DECREASED IN SIX OF NINE HCC CELL LINES AND SIGNIFICANTLY CORRELATED WITH THOSE OF HEPATOCYTE NUCLEAR FACTOR 4ALPHA. PDSS2 TRANSCRIPTION IN HCC CELLS WITH DECREASED PDSS2 EXPRESSION ACCOMPANYING HYPERMETHYLATION WAS REACTIVATED AFTER TREATING THESE CELLS WITH A METHYLATION INHIBITOR. MEAN EXPRESSION LEVELS OF PDSS2 MRNA RELATIVE TO THAT OF UNINVOLVED LIVER DIMINISHED GRADUALLY IN THE ORDER OF CHRONIC HEPATITIS TO CIRRHOSIS, AND EACH WAS SIGNIFICANTLY HIGHER THAN THOSE OF HCCS. PDSS2 AND PDSS2 MRNA LEVELS WERE CONSISTENT. DECREASED PDSS2 MRNA LEVELS WERE DETECTED IN HCC TISSUES OF 56 PATIENTS, CORRELATED WITH SHORTER DISEASE-SPECIFIC SURVIVAL, AND WAS IDENTIFIED AS AN INDEPENDENT PROGNOSTIC FACTOR. PDSS2 IS A PUTATIVE TUMOR SUPPRESSOR, AND PROMOTER HYPERMETHYLATION IS A KEY REGULATORY MECHANISM IN HCC. DECREASED LEVELS OF PDSS2 MRNA EXPRESSION MAY REPRESENT A NOVEL BIOMARKER OF HCC. 2014 13 4001 40 LOSS OF MEN1 LEADS TO RENAL FIBROSIS AND DECREASES HGF-ADAMTS5 PATHWAY ACTIVITY VIA AN EPIGENETIC MECHANISM. BACKGROUND: RENAL FIBROSIS IS A SERIOUS CONDITION THAT RESULTS IN THE DEVELOPMENT OF CHRONIC KIDNEY DISEASES. THE MEN1 GENE IS AN EPIGENETIC REGULATOR THAT ENCODES THE MENIN PROTEIN AND ITS ROLE IN KIDNEY TISSUE REMAINS UNCLEAR. METHODS: KIDNEY HISTOLOGY WAS EXAMINED ON PARAFFIN SECTIONS STAINED WITH HEMATOXYLIN-EOSIN STAINING. MASSON'S TRICHROME STAINING AND SIRIUS RED STAINING WERE USED TO ANALYZE RENAL FIBROSIS. GENE AND PROTEIN EXPRESSION WERE DETERMINED BY QUANTITATIVE REAL-TIME PCR (QPCR) AND WESTERN BLOT, RESPECTIVELY. IMMUNOHISTOCHEMISTRY STAINING IN THE KIDNEY TISSUES FROM MICE OR PATIENTS WAS USED TO EVALUATE PROTEIN LEVELS. FLOW CYTOMETRY WAS USED TO ANALYZE THE CELL CYCLE DISTRIBUTIONS AND APOPTOSIS. RNA-SEQUENCING WAS PERFORMED FOR DIFFERENTIAL EXPRESSION GENES IN THE KIDNEY TISSUES OF THE MEN1F/F AND MEN1?/? MICE. CHROMATIN IMMUNOPRECIPITATION SEQUENCING (CHIP-SEQ) WAS CARRIED OUT FOR IDENTIFICATION OF MENIN- AND H3K4ME3-ENRICHED REGIONS WITHIN THE WHOLE GENOME IN THE MOUSE KIDNEY TISSUE. CHIP-QPCR ASSAYS WERE PERFORMED FOR OCCUPANCY OF MENIN AND H3K4ME3 AT THE GENE PROMOTER REGIONS. LUCIFERASE REPORTER ASSAY WAS USED TO DETECT THE PROMOTER ACTIVITY. THE EXACERBATED UNILATERAL URETERAL OBSTRUCTION (UUO) MODELS IN THE MEN1F/F AND MEN1?/? MICE WERE USED TO ASSESS THE PHARMACOLOGICAL EFFECTS OF RH-HGF ON RENAL FIBROSIS. RESULTS: THE EXPRESSION OF MEN1 IS REDUCE IN KIDNEY TISSUES OF FIBROTIC MOUSE AND HUMAN DIABETIC PATIENTS AND TREATMENT WITH FIBROTIC FACTOR RESULTS IN THE DOWNREGULATION OF MEN1 EXPRESSION IN RENAL TUBULAR EPITHELIAL CELLS (RTECS). DISRUPTION OF MEN1 IN RTECS LEADS TO HIGH EXPRESSION OF ALPHA-SMA AND COLLAGEN 1, WHEREAS MEN1 OVEREXPRESSION RESTRAINS EPITHELIAL-TO-MESENCHYMAL TRANSITION (EMT) INDUCED BY TGF-BETA TREATMENT. CONDITIONAL KNOCKOUT OF MEN1 RESULTED IN CHRONIC RENAL FIBROSIS AND UUO-INDUCED TUBULOINTERSTITIAL FIBROSIS (TIF), WHICH IS ASSOCIATED WITH AN INCREASED INDUCTION OF EMT, G2/M ARREST AND JNK SIGNALING. MECHANISTICALLY, MENIN RECRUITS AND INCREASES H3K4ME3 AT THE PROMOTER REGIONS OF HEPATOCYTE GROWTH FACTOR (HGF) AND A DISINTEGRIN AND METALLOPROTEINASE WITH THROMBOSPONDIN MOTIFS 5 (ADAMTS5) GENES AND ENHANCES THEIR TRANSCRIPTIONAL ACTIVATION. IN THE UUO MICE MODEL, EXOGENOUS HGF RESTORED THE EXPRESSION OF ADAMTS5 AND AMELIORATED RENAL FIBROSIS INDUCED BY MEN1 DEFICIENCY. CONCLUSIONS: THESE FINDINGS DEMONSTRATE THAT MEN1 IS AN ESSENTIAL ANTIFIBROTIC FACTOR IN RENAL FIBROGENESIS AND COULD BE A POTENTIAL TARGET FOR ANTIFIBROTIC THERAPY. 2022 14 3354 36 HISTONE DEMETHYLATION PROFILES IN NONALCOHOLIC FATTY LIVER DISEASE AND PROGNOSTIC VALUES IN HEPATOCELLULAR CARCINOMA: A BIOINFORMATIC ANALYSIS. NONALCOHOLIC FATTY LIVER DISEASE (NAFLD) IS THE MOST COMMON CHRONIC LIVER DISEASE WITH MULTIFACTORIAL PATHOGENESIS; HISTONE DEMETHYLASES (HDMS) ARE EMERGING AS ATTRACTIVE TARGETS. WE IDENTIFIED HDM GENES (INCLUDING KDM5C, KDM6B, KDM8, KDM4A, AND JMJD7) THAT WERE DIFFERENTIALLY EXPRESSED IN NAFLD AND NORMAL SAMPLES BY EXPLORING GENE EXPRESSION PROFILING DATASETS. THERE WAS NO SIGNIFICANT DIFFERENCE IN THE EXPRESSION OF GENES RELATED TO HISTONE DEMETHYLATION BETWEEN MILD AND ADVANCED NAFLD. IN VITRO AND IN VIVO STUDIES INDICATED THAT KDM6B AND JMJD7 WERE UPREGULATED AT THE MRNA LEVEL IN NAFLD. WE EXPLORED THE EXPRESSION LEVELS AND PROGNOSTIC VALUES OF THE IDENTIFIED HDM GENES IN HEPATOCELLULAR CARCINOMA (HCC). KDM5C AND KDM4A WERE UPREGULATED IN HCC COMPARED TO NORMAL TISSUE, WHILE KDM8 SHOWED DOWNREGULATION. THE ABNORMAL EXPRESSION LEVELS OF THESE HDMS COULD PROVIDE PROGNOSTIC VALUES. FURTHERMORE, KDM5C AND KDM4A WERE ASSOCIATED WITH IMMUNE CELL INFILTRATION IN HCC. HDMS WERE ASSOCIATED WITH CELLULAR AND METABOLIC PROCESSES AND MAY BE INVOLVED IN THE REGULATION OF GENE EXPRESSION. DIFFERENTIALLY EXPRESSED HDM GENES IDENTIFIED IN NAFLD MAY PROVIDE VALUE TO UNDERSTANDING PATHOGENESIS AND IN THE DEVELOPMENT OF EPIGENETIC THERAPEUTIC TARGETS. HOWEVER, ON THE BASIS OF THE INCONSISTENT RESULTS OF IN VITRO STUDIES, FUTURE IN VIVO EXPERIMENTS COMBINED WITH TRANSCRIPTOMIC ANALYSIS ARE NEEDED FOR FURTHER VALIDATION. 2023 15 2758 30 EXPRESSION OF HORMONAL CARCINOGENESIS GENES AND RELATED REGULATORY MICRORNAS IN UTERUS AND OVARIES OF DDT-TREATED FEMALE RATS. THE INSECTICIDE DICHLORODIPHENYLTRICHLOROETHANE (DDT) IS A NONMUTAGENIC XENOBIOTIC COMPOUND ABLE TO EXERT ESTROGEN-LIKE EFFECTS RESULTING IN ACTIVATION OF ESTROGEN RECEPTOR-ALPHA (ERALPHA) FOLLOWED BY CHANGED EXPRESSION OF ITS DOWNSTREAM TARGET GENES. IN ADDITION, STUDIES PERFORMED OVER RECENT YEARS SUGGEST THAT DDT MAY ALSO INFLUENCE EXPRESSION OF MICRORNAS. HOWEVER, AN IMPACT OF DDT ON EXPRESSION OF ER, MICRORNAS, AND RELATED TARGET GENES HAS NOT BEEN FULLY ELUCIDATED. HERE, USING REAL-TIME PCR, WE ASSESSED CHANGES IN EXPRESSION OF KEY GENES INVOLVED IN HORMONAL CARCINOGENESIS AS WELL AS POTENTIALLY RELATED REGULATORY ONCOGENIC/TUMOR SUPPRESSOR MICRORNAS AND THEIR TARGET GENES IN THE UTERUS AND OVARIES OF FEMALE WISTAR RATS DURING SINGLE AND CHRONIC MULTIPLE-DOSE DDT EXPOSURE. WE FOUND THAT APPLYING DDT RESULTS IN ALTERED EXPRESSION OF MICRORNAS-221, -222, -205, -126A, AND -429, THEIR TARGET GENES (PTEN, DICER1), AS WELL AS GENES INVOLVED IN HORMONAL CARCINOGENESIS (ESR1, PGR, CCND1, CYP19A1). NOTABLY, CYP19A1 EXPRESSION SEEMS TO BE ALSO REGULATED BY MICRORNAS-221, -222, AND -205. THE DATA SUGGEST THAT EPIGENETIC EFFECTS INDUCED BY DDT AS A POTENTIAL CARCINOGEN MAY BE BASED ON AT LEAST TWO MECHANISMS: (I) ACTIVATION OF ERALPHA FOLLOWED BY ALTERED EXPRESSION OF THE TARGET GENES ENCODING RECEPTOR PGR AND CCND1 AS WELL AS IMPAIRED EXPRESSION OF CYP19A1, AFFECTING, THEREBY, CELL HORMONE BALANCE; AND (II) CHANGED EXPRESSION OF MICRORNAS RESULTING IN IMPAIRED EXPRESSION OF RELATED TARGET GENES INCLUDING REDUCED LEVEL OF CYP19A1 MRNA. 2017 16 3237 33 HEPATIC COX-2 EXPRESSION PROTECTS MICE FROM AN ALCOHOL-HIGH FAT DIET-INDUCED METABOLIC DISORDER BY INVOLVING PROTEIN ACETYLATION RELATED ENERGY METABOLISM. PURPOSE: A DIET HIGH IN FAT AND ETHANOL OFTEN RESULTS IN CHRONIC METABOLIC DISORDER, HEPATIC STEATOSIS, AND LIVER INFLAMMATION. CONSTITUTIVE HEPATIC CYCLOOXYGENASE-2 (COX-2) EXPRESSION COULD PROTECT FROM HIGH FAT-INDUCED METABOLISM DISTURBANCE IN A MURINE MODEL. IN THIS STUDY, WE EXPLORED THE INFLUENCE OF HCOX-2 TRANSGENIC [TG] TO HIGH FAT WITH ETHANOL-INDUCED METABOLIC DISORDER AND LIVER INJURY USING A MOUSE ANIMAL MODEL. METHODS: 12-WEEK-OLD MALE HEPATIC HCOX-2 TRANSGENIC (TG) OR WILD TYPE MICE (WT) WERE FED EITHER A HIGH FAT AND ETHANOL LIQUID DIET (HF+ETH) OR A REGULAR CONTROL DIET (RCD) FOR 5 WEEKS (FOUR GROUPS: RCD/WT, RCD/TG; HF+ETH/TG, HF+ETH/WT). WE ASSESSED METABOLIC BIOMARKERS, CYTOKINE PROFILES, HISTOMORPHOLOGY, AND GENE EXPRESSION TO STUDY THE IMPACT OF PERSISTENT HEPATIC COX-2 EXPRESSION ON DIET-INDUCED LIVER INJURY. RESULTS: IN THE HF+ETH DIET, CONSTITUTIVELY HEPATIC HUMAN COX-2 EXPRESSION PROTECTS MICE FROM BODY WEIGHT GAIN AND WHITE ADIPOSE TISSUE ACCUMULATION, ACCOMPANIED BY IMPROVED IPGTT RESPONSE, SERUM TRIGLYCERIDE/CHOLESTEROL LEVELS, AND LOWER LEVELS OF SERUM AND LIVER INFLAMMATORY CYTOKINES. HISTOLOGICALLY, HCOX-2 MICE SHOWED DECREASED HEPATIC LIPID DROPLETS ACCUMULATION, DECREASED HEPATOCYTE BALLOONING, AND IMPROVED STEATOSIS SCORES. HEPATIC HCOX-2 OVEREXPRESSION ENHANCED AKT INSULIN SIGNALING AND INCREASED FATTY ACID SYNTHESIS IN BOTH RCD AND HF+ETH DIET GROUPS. THE ANTI-LIPOGENIC EFFECT OF HCOX-2 TG IN THE HF+ETH DIET ANIMALS WAS MEDIATED BY INCREASING LIPID DISPOSAL THROUGH ENHANCED BETA-OXIDATION VIA ELEVATIONS IN THE EXPRESSION OF PPARALPHA AND PPARGAMMA, AND INCREASED HEPATIC AUTOPHAGY AS ASSESSED BY THE RATIO OF AUTOPHAGY MARKERS LC3 II/I IN HEPATIC TISSUE. VARIOUS PROTEIN ACETYLATION PATHWAY COMPONENTS, INCLUDING HAT, HDAC1, SIRT1, AND SNAIL1, WERE MODULATED IN HCOX-2 TG MICE IN EITHER RCD OR HF+ETH DIET. CONCLUSIONS: HEPATIC HUMAN COX-2 EXPRESSION PROTECTED MICE FROM THE METABOLIC DISORDER AND LIVER INJURY INDUCED BY A HIGH FAT AND ETHANOL DIET BY ENHANCING HEPATIC LIPID EXPENDITURE. EPIGENETIC REPROGRAMMING OF DIVERSE METABOLIC GENES MIGHT BE INVOLVED IN THE ANTI-LIPOGENIC EFFECT OF COX-2. 2021 17 1335 24 DERMAL FIBROBLASTS CULTURED FROM DONORS WITH TYPE 2 DIABETES MELLITUS RETAIN AN EPIGENETIC MEMORY ASSOCIATED WITH POOR WOUND HEALING RESPONSES. THE PREVALENCE OF TYPE 2 DIABETES MELLITUS (T2DM) IS ESCALATING GLOBALLY. PATIENTS SUFFER FROM MULTIPLE COMPLICATIONS INCLUDING THE DEVELOPMENT OF CHRONIC WOUNDS THAT CAN LEAD TO AMPUTATION. THESE WOUNDS ARE CHARACTERISED BY AN INFLAMMATORY ENVIRONMENT INCLUDING ELEVATED TUMOUR NECROSIS FACTOR ALPHA (TNF-ALPHA). DERMAL FIBROBLASTS (DF) ARE CRITICAL FOR EFFECTIVE WOUND HEALING, SO WE SOUGHT TO ESTABLISH WHETHER THERE WERE ANY DIFFERENCES IN DF CULTURED FROM T2DM DONORS OR THOSE WITHOUT DIABETES (ND-DF). ND- AND T2DM-DF WHEN CULTURED SIMILARLY IN VITRO SECRETED COMPARABLE CONCENTRATIONS OF TNF-ALPHA. FUNCTIONALLY, PRE-TREATMENT WITH TNF-ALPHA REDUCED THE PROLIFERATION OF ND-DF AND TRANSIENTLY ALTERED ND-DF MORPHOLOGY; HOWEVER, T2DM-DF WERE RESISTANT TO THESE TNF-ALPHA INDUCED CHANGES. IN CONTRAST, TNF-ALPHA INHIBITED ND- AND T2DM-DF MIGRATION AND MATRIX METALLOPROTEASE EXPRESSION TO THE SAME DEGREE, ALTHOUGH T2DM-DF EXPRESSED SIGNIFICANTLY HIGHER LEVELS OF TISSUE INHIBITOR OF METALLOPROTEASES (TIMP)-2. FINALLY, TNF-ALPHA SIGNIFICANTLY INCREASED THE SECRETION OF PRO-INFLAMMATORY CYTOKINES (INCLUDING CCL2, CXCL1 AND SERPINE1) IN ND-DF, WHILST THIS EFFECT IN T2DM-DF WAS BLUNTED, PRESUMABLY DUE TO THE TENDENCY TO HIGHER BASELINE PRO-INFLAMMATORY CYTOKINE EXPRESSION OBSERVED IN THIS CELL TYPE. COLLECTIVELY, THESE DATA DEMONSTRATE THAT T2DM-DF EXHIBIT A SELECTIVE LOSS OF RESPONSIVENESS TO TNF-ALPHA, PARTICULARLY REGARDING PROLIFERATIVE AND SECRETORY FUNCTIONS. THIS HIGHLIGHTS IMPORTANT PHENOTYPIC CHANGES IN T2DM-DF THAT MAY EXPLAIN THE SUSCEPTIBILITY TO CHRONIC WOUNDS IN THESE PATIENTS. 2021 18 3718 36 INHIBITION OF BCL6B PROMOTES GASTRIC CANCER BY AMPLIFYING INFLAMMATION IN MICE. BACKGROUND: CHRONIC GASTRITIS HAS BEEN DEMONSTRATED TO BE A KEY CAUSE OF GASTRIC CANCER (GC), AND CONTROL OF GASTRIC INFLAMMATION IS REGARDED AS AN EFFECTIVE TREATMENT FOR THE CLINICAL PREVENTION OF GASTRIC CARCINOGENESIS. HOWEVER, THERE REMAINS AN UNMET NEED TO IDENTIFY THE DOMINANT REGULATORS OF GASTRIC ONCOGENESIS-ASSOCIATED INFLAMMATION IN VIVO. METHODS: THE MOUSE MODEL FOR THE STUDY OF INFLAMMATION-ASSOCIATED GC WAS INDUCED BY BENZO[A]PYRENE (BAP) INTRAGASTRIC ADMINISTRATION IN BCL6B(-/-) AND WILDTYPE MICE ON A C57BL/6 BACKGROUND. 5-AZA-2'-DEOXYCYTIDINE (5-AZA), THE DEMETHYLATION DRUG, WAS INTRAPERITONEALLY INJECTED TO RESTORE BCL6B EXPRESSION. HUMAN GC TISSUE ARRAY WAS USED TO ANALYSE PATIENT SURVIVAL BASED ON BCL6B AND CD3 PROTEIN EXPRESSION. RESULTS: BCL6B WAS GRADUALLY DOWNREGULATED BY ITS OWN PROMOTER HYPERMETHYLATION IN PARALLEL TO AN INCREASING INFLAMMATORY RESPONSE DURING THE PROGRESSION OF BAP-INDUCED GASTRIC CARCINOGENESIS IN MICE. MOREOVER, KNOCKOUT OF BCL6B DRAMATICALLY WORSENED THE SEVERITY OF GASTRIC CANCER AND AGGRAVATED THE INFLAMMATORY RESPONSE IN THE BAP-INDUCED MICE GC MODEL. RE-ACTIVATION OF BCL6B BY 5-AZA IMPEDED INFLAMMATORY AMPLIFICATION AND BAP-INDUCED GC DEVELOPMENT, PROLONGING SURVIVAL TIME IN WILDTYPE MICE, WHEREAS NO NOTABLE CURATIVE EFFECT OCCURRED IN BCL6B(-/-) MICE WITH 5-AZA TREATMENT. FINALLY, SIGNIFICANT NEGATIVE CORRELATIONS WERE DETECTED BETWEEN THE MRNA LEVELS OF BCL6B AND INFLAMMATORY CYTOKINES IN HUMAN GC TISSUES; PATIENTS HARBOURING BCL6B-NEGETIVE AND SEVERE-INFLAMMATION GC TUMOURS WERE FOUND TO EXHIBIT THE SHORTEST SURVIVAL TIME. CONCLUSIONS: EPIGENETIC INACTIVATION OF BCL6B PROMOTES GASTRIC CANCER THROUGH AMPLIFICATION OF THE GASTRIC INFLAMMATORY RESPONSE IN VIVO AND OFFERS A NEW APPROACH FOR GC TREATMENT AND REGENERATIVE MEDICINE. 2019 19 984 31 CHRONIC PSYCHOLOGICAL STRESS ALTERS GENE EXPRESSION IN RAT COLON EPITHELIAL CELLS PROMOTING CHROMATIN REMODELING, BARRIER DYSFUNCTION AND INFLAMMATION. CHRONIC STRESS IS COMMONLY ASSOCIATED WITH ENHANCED ABDOMINAL PAIN (VISCERAL HYPERSENSITIVITY), BUT THE CELLULAR MECHANISMS UNDERLYING HOW CHRONIC STRESS INDUCES VISCERAL HYPERSENSITIVITY ARE POORLY UNDERSTOOD. IN THIS STUDY, WE EXAMINED CHANGES IN GENE EXPRESSION IN COLON EPITHELIAL CELLS FROM A RAT MODEL USING RNA-SEQUENCING TO EXAMINE STRESS-INDUCED CHANGES TO THE TRANSCRIPTOME. FOLLOWING CHRONIC STRESS, THE MOST SIGNIFICANTLY UP-REGULATED GENES INCLUDED ATG16L1, COQ10B, DCAF13, NAT2, PTBP2, RRAS2, SPINK4 AND DOWN-REGULATED GENES INCLUDING ABAT, CITED2, CNNM2, DAB2IP, PLEKHM1, SCD2, AND TAB2. THE PRIMARY ALTERED BIOLOGICAL PROCESSES REVEALED BY NETWORK ENRICHMENT ANALYSIS WERE INFLAMMATION/IMMUNE RESPONSE, TISSUE MORPHOGENESIS AND DEVELOPMENT, AND NUCLEOSOME/CHROMATIN ASSEMBLY. THE MOST SIGNIFICANTLY DOWN-REGULATED PROCESS WAS THE DIGESTIVE SYSTEM DEVELOPMENT/FUNCTION, WHEREAS THE MOST SIGNIFICANTLY UP-REGULATED PROCESSES WERE INFLAMMATORY RESPONSE, ORGANISMAL INJURY, AND CHROMATIN REMODELING MEDIATED BY H3K9 METHYLATION. FURTHERMORE, A SUBPOPULATION OF STRESSED RATS DEMONSTRATED VERY SIGNIFICANTLY ALTERED GENE EXPRESSION AND TRANSCRIPT ISOFORMS, ENRICHED FOR THE DIFFERENTIAL EXPRESSION OF GENES INVOLVED IN THE INFLAMMATORY RESPONSE, INCLUDING UPREGULATION OF CYTOKINE AND CHEMOKINE RECEPTOR GENE EXPRESSION COUPLED WITH DOWNREGULATION OF EPITHELIAL ADHERENS AND TIGHT JUNCTION MRNAS. IN SUMMARY, THESE FINDINGS SUPPORT THAT CHRONIC STRESS IS ASSOCIATED WITH INCREASED LEVELS OF CYTOKINES AND CHEMOKINES, THEIR DOWNSTREAM SIGNALING PATHWAYS COUPLED TO DYSREGULATION OF INTESTINAL CELL DEVELOPMENT AND FUNCTION. EPIGENETIC REGULATION OF CHROMATIN REMODELING LIKELY PLAYS A PROMINENT ROLE IN THIS PROCESS. RESULTS ALSO SUGGEST THAT SUPER ENHANCERS PLAY A PRIMARY ROLE IN CHRONIC STRESS-ASSOCIATED INTESTINAL BARRIER DYSFUNCTION. 2022 20 2155 35 EPIGENETIC MECHANISMS AND METABOLIC REPROGRAMMING IN FIBROGENESIS: DUAL TARGETING OF G9A AND DNMT1 FOR THE INHIBITION OF LIVER FIBROSIS. OBJECTIVE: HEPATIC STELLATE CELLS (HSC) TRANSDIFFERENTIATION INTO MYOFIBROBLASTS IS CENTRAL TO FIBROGENESIS. EPIGENETIC MECHANISMS, INCLUDING HISTONE AND DNA METHYLATION, PLAY A KEY ROLE IN THIS PROCESS. CONCERTED ACTION BETWEEN HISTONE AND DNA-MEHYLTRANSFERASES LIKE G9A AND DNMT1 IS A COMMON THEME IN GENE EXPRESSION REGULATION. WE AIMED TO STUDY THE EFFICACY OF CM272, A FIRST-IN-CLASS DUAL AND REVERSIBLE G9A/DNMT1 INHIBITOR, IN HALTING FIBROGENESIS. DESIGN: G9A AND DNMT1 WERE ANALYSED IN CIRRHOTIC HUMAN LIVERS, MOUSE MODELS OF LIVER FIBROSIS AND CULTURED MOUSE HSC. G9A AND DNMT1 EXPRESSION WAS KNOCKED DOWN OR INHIBITED WITH CM272 IN HUMAN HSC (HHSC), AND TRANSCRIPTOMIC RESPONSES TO TRANSFORMING GROWTH FACTOR-BETA1 (TGFBETA1) WERE EXAMINED. GLYCOLYTIC METABOLISM AND MITOCHONDRIAL FUNCTION WERE ANALYSED WITH SEAHORSE-XF TECHNOLOGY. GENE EXPRESSION REGULATION WAS ANALYSED BY CHROMATIN IMMUNOPRECIPITATION AND METHYLATION-SPECIFIC PCR. ANTIFIBROGENIC ACTIVITY AND SAFETY OF CM272 WERE STUDIED IN MOUSE CHRONIC CCL(4) ADMINISTRATION AND BILE DUCT LIGATION (BDL), AND IN HUMAN PRECISION-CUT LIVER SLICES (PCLSS) IN A NEW BIOREACTOR TECHNOLOGY. RESULTS: G9A AND DNMT1 WERE DETECTED IN STROMAL CELLS IN AREAS OF ACTIVE FIBROSIS IN HUMAN AND MOUSE LIVERS. G9A AND DNMT1 EXPRESSION WAS INDUCED DURING MOUSE HSC ACTIVATION, AND TGFBETA1 TRIGGERED THEIR CHROMATIN RECRUITMENT IN HHSC. G9A/DNMT1 KNOCKDOWN AND CM272 INHIBITED TGFBETA1 FIBROGENIC RESPONSES IN HHSC. TGFBETA1-MEDIATED PROFIBROGENIC METABOLIC REPROGRAMMING WAS ABROGATED BY CM272, WHICH RESTORED GLUCONEOGENIC GENE EXPRESSION AND MITOCHONDRIAL FUNCTION THROUGH ON-TARGET EPIGENETIC EFFECTS. CM272 INHIBITED FIBROGENESIS IN MICE AND PCLSS WITHOUT TOXICITY. CONCLUSIONS: DUAL G9A/DNMT1 INHIBITION BY COMPOUNDS LIKE CM272 MAY BE A NOVEL THERAPEUTIC STRATEGY FOR TREATING LIVER FIBROSIS. 2021