1 1228 158 CRITICAL ROLE OF VGLL4 IN THE REGULATION OF CHRONIC NORMOBARIC HYPOXIA-INDUCED PULMONARY HYPERTENSION IN MICE. PULMONARY HYPERTENSION (PH), A RARE BUT DEADLY CARDIOPULMONARY DISORDER, IS CHARACTERIZED BY EXTENSIVE REMODELING OF PULMONARY ARTERIES RESULTING FROM ENHANCEMENT OF PULMONARY ARTERY SMOOTH MUSCLE CELL PROLIFERATION AND SUPPRESSED APOPTOSIS; HOWEVER, THE UNDERLYING PATHOPHYSIOLOGICAL MECHANISMS REMAIN LARGELY UNKNOWN. RECENTLY, EPIGENETICS HAS GAINED INCREASING PROMINENCE IN THE DEVELOPMENT OF PH. WE AIMED TO INVESTIGATE THE ROLE OF VESTIGIAL-LIKE FAMILY MEMBER 4 (VGLL4) IN CHRONIC NORMOBARIC HYPOXIA (CNH)-INDUCED PH AND TO ADDRESS WHETHER IT IS ASSOCIATED WITH EPIGENETIC REGULATION. THE RODENT MODEL OF PH WAS ESTABLISHED BY CNH TREATMENT (10% O(2) , 23 HOURS/DAY). WESTERN BLOT, QUANTITATIVE REVERSE TRANSCRIPTION POLYMERASE CHAIN REACTION, IMMUNOFLUORESCENCE, IMMUNOPRECIPITATION, AND ADENO-ASSOCIATED VIRUS TESTS WERE PERFORMED TO EXPLORE THE POTENTIAL MECHANISMS INVOLVED IN CNH-INDUCED PH IN MICE. VGLL4 EXPRESSION WAS UPREGULATED AND CORRELATED WITH CNH IN PH MOUSE LUNG TISSUES IN A TIME-DEPENDENT MANNER. VGLL4 COLOCALIZED WITH ALPHA-SMOOTH MUSCLE ACTIN IN CULTURED PULMONARY ARTERIAL SMOOTH MUSCLE CELLS (PASMCS), AND VGLL4 IMMUNOACTIVITY WAS INCREASED IN PASMCS FOLLOWING HYPOXIA EXPOSURE IN VITRO. VGLL4 KNOCKDOWN ATTENUATED CNH-INDUCED PH AND PULMONARY ARTERY REMODELING BY BLUNTING SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION 3 (STAT3) SIGNALING; CONVERSELY, VGLL4 OVEREXPRESSION EXACERBATED THE DEVELOPMENT OF PH. CNH ENHANCED THE ACETYLATION OF VGLL4 AND INCREASED THE INTERACTION OF AC-H3K9/VGLL4 AND AC-H3K9/STAT3 IN THE LUNG TISSUES, AND LEVELS OF AC-H3K9, P-STAT3/STAT3, AND PROLIFERATION-ASSOCIATED PROTEIN LEVELS WERE MARKEDLY UP-REGULATED, WHEREAS APOPTOSIS-RELATED PROTEIN LEVELS WERE SIGNIFICANTLY DOWNREGULATED, IN THE LUNG TISSUES OF MICE WITH CNH-INDUCED PH. NOTABLY, ABROGATION OF VGLL4 ACETYLATION REVERSED CNH-INDUCED PH AND PULMONARY ARTERY REMODELING AND SUPPRESSED STAT3 SIGNALING. FINALLY, STAT3 KNOCKDOWN ALLEVIATED CNH-INDUCED PH. IN CONCLUSION, VGLL4 ACETYLATION UPREGULATION COULD CONTRIBUTE TO CNH-INDUCED PH AND PULMONARY ARTERY REMODELING VIA STAT3 SIGNALING, AND ABROGATION OF VGLL4 ACETYLATION REVERSED CNH-INDUCED PH. PHARMACOLOGICAL OR GENETIC DELETION OF VGLL4 MIGHT BE A POTENTIAL TARGET FOR THERAPEUTIC INTERVENTIONS IN CNH-INDUCED PH. 2021 2 3128 50 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 3468 44 HYPOXIA-INDUCED DNA HYPERMETHYLATION IN HUMAN PULMONARY FIBROBLASTS IS ASSOCIATED WITH THY-1 PROMOTER METHYLATION AND THE DEVELOPMENT OF A PRO-FIBROTIC PHENOTYPE. BACKGROUND: PULMONARY FIBROSIS IS A DEBILITATING AND LETHAL DISEASE WITH NO EFFECTIVE TREATMENT OPTIONS. UNDERSTANDING THE PATHOLOGICAL PROCESSES AT PLAY WILL DIRECT THE APPLICATION OF NOVEL THERAPEUTIC AVENUES. HYPOXIA HAS BEEN IMPLICATED IN THE PATHOGENESIS OF PULMONARY FIBROSIS YET THE PRECISE MECHANISM BY WHICH IT CONTRIBUTES TO DISEASE PROGRESSION REMAINS TO BE FULLY ELUCIDATED. IT HAS BEEN SHOWN THAT CHRONIC HYPOXIA CAN ALTER DNA METHYLATION PATTERNS IN TUMOUR-DERIVED CELL LINES. THIS EPIGENETIC ALTERATION CAN INDUCE CHANGES IN CELLULAR PHENOTYPE WITH PROMOTER METHYLATION BEING ASSOCIATED WITH GENE SILENCING. OF PARTICULAR RELEVANCE TO IDIOPATHIC PULMONARY FIBROSIS (IPF) IS THE OBSERVATION THAT THY-1 PROMOTER METHYLATION IS ASSOCIATED WITH A MYOFIBROBLAST PHENOTYPE WHERE LOSS OF THY-1 OCCURS ALONGSIDE INCREASED ALPHA SMOOTH MUSCLE ACTIN (ALPHA-SMA) EXPRESSION. THE INITIAL AIM OF THIS STUDY WAS TO DETERMINE WHETHER HYPOXIA REGULATES DNA METHYLATION IN NORMAL HUMAN LUNG FIBROBLASTS (CCD19LU). AS IT HAS BEEN REPORTED THAT HYPOXIA SUPPRESSES THY-1 EXPRESSION DURING LUNG DEVELOPMENT WE ALSO STUDIED THE EFFECT OF HYPOXIA ON THY-1 PROMOTER METHYLATION AND GENE EXPRESSION. METHODS: CCD19LU WERE GROWN FOR UP TO 8 DAYS IN HYPOXIA AND ASSESSED FOR GLOBAL CHANGES IN DNA METHYLATION USING FLOW CYTOMETRY. REAL-TIME PCR WAS USED TO QUANTIFY EXPRESSION OF THY-1, ALPHA-SMA, COLLAGEN I AND III. GENOMIC DNA WAS BISULPHITE TREATED AND METHYLATION SPECIFIC PCR (MSPCR) WAS USED TO EXAMINE THE METHYLATION STATUS OF THE THY-1 PROMOTER. RESULTS: SIGNIFICANT GLOBAL HYPERMETHYLATION WAS DETECTED IN HYPOXIC FIBROBLASTS RELATIVE TO NORMOXIC CONTROLS AND WAS ACCOMPANIED BY INCREASED EXPRESSION OF MYOFIBROBLAST MARKERS. THY-1 MRNA EXPRESSION WAS SUPPRESSED IN HYPOXIC CELLS, WHICH WAS RESTORED WITH THE DEMETHYLATING AGENT 5-AZA-2'-DEOXYCYTIDINE. MSPCR REVEALED THAT THY-1 BECAME METHYLATED FOLLOWING FIBROBLAST EXPOSURE TO 1% O2. CONCLUSION: THESE DATA SUGGEST THAT GLOBAL AND GENE-SPECIFIC CHANGES IN DNA METHYLATION MAY PLAY AN IMPORTANT ROLE IN FIBROBLAST FUNCTION IN HYPOXIA. 2012 4 1210 39 COX2 ENHANCES NEOVASCULARIZATION OF INFLAMMATORY TENOCYTES THROUGH THE HIF-1ALPHA/VEGFA/PDGFB PATHWAY. TENDON INJURIES ARE AMONG THE MOST CHALLENGING IN ORTHOPEDICS. DURING THE EARLY TENDON REPAIR, NEW BLOOD VESSEL FORMATION IS NECESSARY. HOWEVER, EXCESSIVE ANGIOGENESIS ALSO EXACERBATES SCAR FORMATION, LEADING TO PAIN AND DYSFUNCTION. A SIGNIFICANTLY WORSE OUTCOME WAS ASSOCIATED WITH HIGHER EXPRESSION LEVELS OF HYPOXIA-INDUCIBLE FACTOR-1 ALPHA (HIF-1ALPHA), AND ITS TRANSCRIPTIONAL TARGETS VASCULAR ENDOTHELIAL GROWTH FACTOR A (VEGFA) AND PLATELET-DERIVED GROWTH FACTOR B (PDGFB), BUT THE UNDERLYING MOLECULAR MECHANISMS REMAIN UNCLEAR. IN THIS STUDY, LIPOPOLYSACCHARIDE (LPS) WAS USED TO INDUCE AN INFLAMMATORY RESPONSE IN TENOCYTES. LPS INCREASED THE TENOCYTES' INFLAMMATORY FACTOR COX2 EXPRESSION AND ACTIVATED THE HIF-1ALPHA/VEGFA/PDGFB PATHWAY. MOREOVER, THE CONDITIONED MEDIUM FROM THE TENOCYTES BOOSTED RAT AORTIC VASCULAR ENDOTHELIAL CELL (RAOEC) ANGIOGENESIS. FURTHERMORE, TRICHOSTATIN A (TSA), AN INHIBITOR OF HISTONE DEACETYLASE, WAS USED TO TREAT INFLAMMATORY TENOCYTES. THE EXPRESSION LEVELS OF HIF-1ALPHA AND ITS TRANSCRIPTIONAL TARGETS VEGFA AND PDGFB DECREASED, RESULTING IN RAOEC ANGIOGENESIS INHIBITION. FINALLY, THE DUAL-LUCIFERASE REPORTER GENE ASSAY AND CHROMATIN IMMUNOPRECIPITATION (CHIP) ASSAY PROVED THAT THE HIF-1ALPHA/PDGFB PATHWAY PLAYED A MORE CRITICAL ROLE IN TENOCYTE ANGIOGENESIS THAN THE HIF-1ALPHA/VEGFA PATHWAY. TSA COULD ALLEVIATE ANGIOGENESIS MAINLY THROUGH EPIGENETIC REGULATION OF THE HIF-1ALPHA/PDGFB PATHWAY. TAKEN TOGETHER, TSA MIGHT BE A PROMISING ANTI-ANGIOGENESIS DRUG FOR ABNORMAL ANGIOGENESIS, WHICH IS INDUCED BY TENDON INJURIES. 2021 5 1966 34 EPIGENETIC ALTERATION OF PRKCDBP IN COLORECTAL CANCERS AND ITS IMPLICATION IN TUMOR CELL RESISTANCE TO TNFALPHA-INDUCED APOPTOSIS. PURPOSE: PRKCDBP IS A PUTATIVE TUMOR SUPPRESSOR IN WHICH ALTERATION HAS BEEN OBSERVED IN SEVERAL HUMAN CANCERS. WE INVESTIGATED EXPRESSION AND FUNCTION OF PRKCDBP IN COLORECTAL CELLS AND TISSUES TO EXPLORE ITS CANDIDACY AS A SUPPRESSOR IN COLORECTAL TUMORIGENESIS. EXPERIMENTAL DESIGN: EXPRESSION AND METHYLATION STATUS OF PRKCDBP AND ITS EFFECT ON TUMOR GROWTH WERE EVALUATED. TRANSCRIPTIONAL REGULATION BY NF-KAPPAB SIGNALING WAS DEFINED BY LUCIFERASE REPORTER AND CHROMATIN IMMUNOPRECIPITATION ASSAYS. RESULTS: PRKCDBP EXPRESSION WAS HARDLY DETECTABLE IN 29 OF 80 (36%) PRIMARY TUMORS AND 11 OF 19 (58%) CELL LINES, AND ITS ALTERATION CORRELATED WITH TUMOR STAGE AND GRADE. PROMOTER HYPERMETHYLATION WAS COMMONLY FOUND IN CANCERS. PRKCDBP EXPRESSION INDUCED THE G(1) CELL-CYCLE ARREST AND INCREASED CELLULAR SENSITIVITY TO VARIOUS APOPTOTIC STRESSES. PRKCDBP WAS INDUCED BY TNFALPHA, AND ITS LEVEL CORRELATED WITH TUMOR CELL SENSITIVITY TO TNFALPHA-INDUCED APOPTOSIS. PRKCDBP INDUCTION BY TNFALPHA WAS DISRUPTED BY BLOCKING NF-KAPPAB SIGNALING WHILE IT WAS ENHANCED BY RELA TRANSFECTION. THE PRKCDBP PROMOTER ACTIVITY WAS INCREASED IN RESPONSE TO TNFALPHA, AND THIS RESPONSE WAS ABOLISHED BY DISRUPTION OF A KAPPAB SITE IN THE PROMOTER. PRKCDBP DELAYED THE FORMATION AND GROWTH OF XENOGRAFT TUMORS AND IMPROVED TUMOR RESPONSE TO TNFALPHA-INDUCED APOPTOSIS. CONCLUSIONS: PRKCDBP IS A PROAPOPTOTIC TUMOR SUPPRESSOR WHICH IS COMMONLY ALTERED IN COLORECTAL CANCER BY PROMOTER HYPERMETHYLATION, AND ITS GENE TRANSCRIPTION IS DIRECTLY ACTIVATED BY NF-KAPPAB IN RESPONSE TO TNFALPHA. THIS SUGGESTS THAT PRKCDBP INACTIVATION MAY CONTRIBUTE TO TUMOR PROGRESSION BY REDUCING CELLULAR SENSITIVITY TO TNFALPHA AND OTHER STRESSES, PARTICULARLY UNDER CHRONIC INFLAMMATORY MICROENVIRONMENT. 2011 6 3049 35 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 7 984 41 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 8 4001 54 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 9 6294 30 THE PROINFLAMMATORY CYTOKINE TNFALPHA INDUCES DNA DEMETHYLATION-DEPENDENT AND -INDEPENDENT ACTIVATION OF INTERLEUKIN-32 EXPRESSION. IL-32 IS A CYTOKINE INVOLVED IN PROINFLAMMATORY IMMUNE RESPONSES TO BACTERIAL AND VIRAL INFECTIONS. HOWEVER, THE ROLE OF EPIGENETIC EVENTS IN THE REGULATION OF IL-32 GENE EXPRESSION IS UNDERSTUDIED. HERE WE SHOW THAT IL-32 IS REPRESSED BY DNA METHYLATION IN HEK293 CELLS. USING CHIP SEQUENCING, LOCUS-SPECIFIC METHYLATION ANALYSIS, CRISPR/CAS9-MEDIATED GENOME EDITING, AND RT-QPCR (QUANTITATIVE RT-PCR) AND IMMUNOBLOT ASSAYS, WE FOUND THAT SHORT-TERM TREATMENT (A FEW HOURS) WITH THE PROINFLAMMATORY CYTOKINE TUMOR NECROSIS FACTOR ALPHA (TNFALPHA) ACTIVATES IL-32 IN A DNA DEMETHYLATION-INDEPENDENT MANNER. IN CONTRAST, PROLONGED TNFALPHA TREATMENT (SEVERAL DAYS) INDUCED DNA DEMETHYLATION AT THE PROMOTER AND A CPG ISLAND IN THE IL-32 GENE IN A TET (TEN-ELEVEN TRANSLOCATION) FAMILY ENZYME- AND NF-KAPPAB-DEPENDENT MANNER. NOTABLY, THE HYPOMETHYLATION STATUS OF TRANSCRIPTIONAL REGULATORY ELEMENTS IN IL-32 WAS MAINTAINED FOR A LONG TIME (SEVERAL WEEKS), CAUSING ELEVATED IL-32 EXPRESSION EVEN IN THE ABSENCE OF TNFALPHA. CONSIDERING THAT IL-32 CAN, IN TURN, INDUCE TNFALPHA EXPRESSION, WE SPECULATE THAT SUCH FEEDFORWARD EVENTS MAY CONTRIBUTE TO THE TRANSITION FROM AN ACUTE INFLAMMATORY RESPONSE TO CHRONIC INFLAMMATION. 2019 10 26 35 A 6-ALKYLSALICYLATE HISTONE ACETYLTRANSFERASE INHIBITOR INHIBITS HISTONE ACETYLATION AND PRO-INFLAMMATORY GENE EXPRESSION IN MURINE PRECISION-CUT LUNG SLICES. LYSINE ACETYLATIONS ARE POST-TRANSLATIONAL MODIFICATIONS OF CELLULAR PROTEINS, THAT ARE CRUCIAL IN THE REGULATION OF MANY CELLULAR PROCESSES. LYSINE ACETYLATIONS ON HISTONE PROTEINS ARE PART OF THE EPIGENETIC CODE REGULATING GENE EXPRESSION AND ARE INSTALLED BY HISTONE ACETYLTRANSFERASES. OBSERVATIONS THAT INFLAMMATORY LUNG DISEASES, SUCH AS ASTHMA AND CHRONIC OBSTRUCTIVE PULMONARY DISEASE, ARE CHARACTERIZED BY INCREASED HISTONE ACETYLTRANSFERASE ACTIVITY INDICATE THAT DEVELOPMENT OF SMALL MOLECULE INHIBITORS FOR THESE ENZYMES MIGHT BE A VALUABLE APPROACH TOWARDS NEW THERAPIES FOR THESE DISEASES. THE 6-ALKYLSALICYLATE MG149 IS A CANDIDATE TO EXPLORE THIS HYPOTHESIS BECAUSE IT HAS BEEN DEMONSTRATED TO INHIBIT THE MYST TYPE HISTONE ACETYLTRANSFERASES. IN THIS STUDY, WE DETERMINED THE K(I) VALUE FOR INHIBITION OF THE MYST TYPE HISTONE ACETYLTRANSFERASE KAT8 BY MG149 TO BE 39 +/- 7.7 MUM. UPON INVESTIGATING WHETHER THE INHIBITION OF HISTONE ACETYLTRANSFERASES BY MG149 CORRELATES WITH INHIBITION OF HISTONE ACETYLATION IN MURINE PRECISION-CUT LUNG SLICES, INHIBITION OF ACETYLATION WAS OBSERVED USING AN LC-MS/MS BASED ASSAY ON HISTONE H4 RES 4-17, WHICH CONTAINS THE TARGET LYSINE OF KAT8. FOLLOWING UP ON THIS, UPON TREATMENT WITH MG149, REDUCED PRO-INFLAMMATORY GENE EXPRESSION WAS OBSERVED IN LIPOPOLYSACCHARIDE AND INTERFERON GAMMA STIMULATED MURINE PRECISION-CUT LUNG SLICES. BASED ON THIS, WE PROPOSE THAT 6-ALKYLSALICYLATES SUCH AS MG149 HAVE POTENTIAL FOR DEVELOPMENT TOWARDS APPLICATIONS IN THE TREATMENT OF INFLAMMATORY LUNG DISEASES. 2017 11 6297 48 THE PROTECTIVE EFFECT OF HBO1 ON CIGARETTE SMOKE EXTRACT-INDUCED APOPTOSIS IN AIRWAY EPITHELIAL CELLS. PURPOSE: EPIGENETIC MODIFICATION IS ONE OF MOST IMPORTANT MECHANISMS UNDERLYING THE PATHOGENESIS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). THE PURPOSE OF THIS STUDY WAS TO DETERMINE WHETHER HISTONE ACETYLTRANSFERASE BINDING TO ORC1 (HBO1) CAN PROTECT AGAINST CIGARETTE SMOKE (CS)-INDUCED CELL APOPTOSIS AND SUSTAIN NORMAL HISTONE ACETYLATION IN COPD. METHODS: HUMAN LUNG TISSUE SAMPLES WERE OBTAINED FROM PATIENTS WHO UNDERWENT LUNG RESECTION. THE EMPHYSEMA MOUSE MODEL AND HBO1 OVEREXPRESSING MICE WERE EACH ESTABLISHED BY INTRAPERITONEAL INJECTION WITH CIGARETTE SMOKE EXTRACT (CSE) OR INTRATRACHEAL LENTIVIRAL VECTORS INSTILLATION. TUNEL (TERMINAL DEOXYNUCLEOTIDYL TRANSFERASE DUTP NICK END LABELING) ASSAYS WERE USED TO ASSESS APOPTOTIC RATIO IN MICE. THE APOPTOSIS OF HUMAN BRONCHIAL EPITHELIAL CELLS (HBECS) WAS ASSAYED BY FLOW CYTOMETRY. HBO1, B-CELL LYMPHOMA-2 (BCL-2), AND H3K14AC PROTEIN EXPRESSION WERE DETECTED BY WESTERN BLOTTING. HBO1 MRNA EXPRESSION WAS MEASURED BY QUANTITATIVE REAL-TIME POLYMERASE CHAIN REACTION. RESULTS: PROTEIN EXPRESSION OF HBO1 WAS DECREASED SIGNIFICANTLY IN LUNG TISSUE FROM COPD PATIENTS AND CSE-TREATED EMPHYSEMA MOUSE MODELS. OVEREXPRESSION OF HBO1 ATTENUATED CSE-INDUCED EMPHYSEMATOUS CHANGES, AS WELL AS APOPTOSIS IN THE LUNGS OF COPD MICE. IN VITRO, THE HBO1 PROTEIN DEGRADED IN A TIME- AND DOSE-DEPENDENT COURSE WITH CSE TREATMENT. WITH FLOW CYTOMETRY, WE PROVED THAT HBO1 COULD REVERSE THE APOPTOSIS OF HBECS INDUCED BY CSE. FURTHERMORE, HBO1 OVEREXPRESSION PROMOTED THE EXPRESSION OF ANTI-APOPTOTIC BCL-2 PROTEIN AND ENHANCED H3K14 ACETYLATION IN AIRWAY EPITHELIAL CELLS. CONCLUSION: THESE FINDINGS DEMONSTRATE THAT THE KEY HISTONE MODULATOR HBO1 PLAYS A PROTECTIVE ROLE IN COPD PATHOGENESIS THAT MAY SHED LIGHT ON POTENTIAL THERAPEUTIC TARGETS TO INHIBIT THE PROGRESS OF COPD. 2020 12 1461 43 DISRUPTION OF RCAN1.4 EXPRESSION MEDIATED BY YY1/HDAC2 MODULATES CHRONIC RENAL ALLOGRAFT INTERSTITIAL FIBROSIS. CHRONIC ALLOGRAFT DYSFUNCTION (CAD) IS A MAJOR FACTOR THAT HINDERS KIDNEY TRANSPLANT SURVIVAL IN THE LONG RUN. EPITHELIAL-MESENCHYMAL TRANSITION (EMT) HAS BEEN CONFIRMED TO SIGNIFICANTLY CONTRIBUTE TO INTERSTITIAL FIBROSIS/TUBULAR ATROPHY (IF/TA), WHICH IS THE MAIN HISTOPATHOLOGICAL FEATURE OF CAD. ABERRANT EXPRESSION OF THE REGULATOR OF CALCINEURIN 1 (RCAN1), RECOGNIZED AS AN ENDOGENOUS INHIBITOR OF THE CALCINEURIN PHOSPHATASE, HAS BEEN SHOWN TO BE EXTENSIVELY INVOLVED IN VARIOUS KIDNEY DISEASES. HOWEVER, IT REMAINS UNCLEAR HOW RCAN1.4 REGULATES IF/TA FORMATION IN CAD PATIENTS. HEREIN, AN IN VIVO MOUSE RENAL TRANSPLANTATION MODEL AND AN IN VITRO MODEL OF HUMAN RENAL TUBULAR EPITHELIAL CELLS (HK-2) TREATED WITH TUMOR NECROSIS FACTOR-ALPHA (TNF-ALPHA) WERE EMPLOYED. OUR RESULTS PROVED THAT RCAN1.4 EXPRESSION WAS DECREASED IN VIVO AND IN VITRO, IN ADDITION TO THE UP-REGULATION OF YIN YANG 1 (YY1), A TRANSCRIPTION FACTOR THAT HAS BEEN REPORTED TO CONVEY MULTIPLE FUNCTIONS IN CHRONIC KIDNEY DISEASE (CKD). KNOCKING IN OF RCAN1.4 EFFICIENTLY ATTENUATED CHRONIC RENAL ALLOGRAFT INTERSTITIAL FIBROSIS IN VIVO AND INHIBITED TNF-ALPHA-INDUCED EMT IN VITRO THROUGH REGULATING ANTI-OXIDATIVE STRESS AND THE CALCINEURIN/NUCLEAR FACTOR OF ACTIVATED T CELLS CYTOPLASMIC 1 (NFATC1) SIGNALING PATHWAY. IN ADDITION, SUPPRESSION OF YY1 MEDIATED BY SHRNA OR SIRNA ALLEVIATED TNF-ALPHA-INDUCED EMT THROUGH ABOLISHING REACTIVE SPECIES PARTLY IN AN RCAN1.4-DEPENDENT MANNER. NOTABLY, WE CONFIRMED THAT YY1 NEGATIVELY REGULATED RCAN1.4 TRANSCRIPTION BY DIRECTLY INTERACTING WITH THE RCAN1.4 PROMOTER. IN ADDITION, HISTONE DEACETYLASE 2 (HDAC2) INTERACTED WITH YY1 TO FORM A MULTI-MOLECULAR COMPLEX, WHICH WAS INVOLVED IN TNF-ALPHA-INDUCED RCAN1.4 TRANSCRIPTIONAL REPRESSION. THEREFORE, RCAN1.4 IS SUGGESTED TO BE MODULATED BY THE YY1/HDAC2 TRANSCRIPTION REPRESSOR COMPLEX IN AN EPIGENETIC MANNER, WHICH IS A MEDIATED NEPHROPROTECTIVE EFFECT PARTLY THROUGH MODULATING O2?- GENERATION AND THE CALCINEURIN/NFATC1 SIGNALING PATHWAY. THUS, THE YY1-RCAN1.4 AXIS CONSTITUTES AN INNOVATIVE TARGET FOR IF/TA TREATMENT IN CAD PATIENTS. 2023 13 4353 39 MIR-20B-5P ATTENUATES HYPOXIA-INDUCED APOPTOSIS IN CARDIOMYOCYTES VIA THE HIF-1ALPHA/NF-KAPPAB PATHWAY. CHRONIC HYPOXIA IS A COMMON INDUCER OF END-STAGE CARDIOVASCULAR DISEASE. IN CELLS UNDER HYPOXIA, THE HYPOXIA-INDUCIBLE FACTOR-1 (HIF-1) PLAYS A VITAL ROLE IN REGULATING DOWNSTREAM TARGET GENES. HOWEVER, THE MECHANISM OF HYPOXIA IN CARDIOMYOCYTES IS STILL UNCLEAR. IN THIS STUDY, WE AIMED TO IDENTIFY NOVEL DOWNSTREAM EPIGENETIC TARGETS OF HIF-1ALPHA IN CARDIOMYOCYTES UNDER HYPOXIA. H9C2 CELLS WERE EXPOSED TO HYPOXIA CONDITION, AND QUANTITATIVE REAL-TIME PCR ANALYSIS WAS PERFORMED TO EVALUATE THE EXPRESSION OF MIR-20B-5P. THE RESULTS INDICATED THAT THE EXPRESSION OF MIR-20B-5P WAS DOWN-REGULATED IN H9C2 CELLS UNDER LOW OXYGEN CONDITION. MEANWHILE, HIF-1ALPHA OVEREXPRESSION FURTHER DOWN-REGULATED THE MIR-20B-5P EXPRESSION IN H9C2 CELLS TRANSFECTED WITH HIF-1ALPHA PLASMIDS. IN ADDITION, ANNEXIN-V-FITC/PI FLOW CYTOMETRY ANALYSIS SUGGESTED THAT OVEREXPRESSION OF MIR-20B-5P ATTENUATED CELL APOPTOSIS UNDER HYPOXIA CONDITION IN H9C2 CELLS. WESTERN BLOT ANALYSIS SHOWED THAT THE HYPOXIA APPARENTLY INCREASED BAX AND CLEAVED-CASPASE-3, BUT DECREASED BCL-2 EXPRESSION IN H9C2 CELLS, INDICATING THAT HYPOXIA-INDUCED NF-KAPPAB SIGNALING PATHWAY ACTIVATION IS MEDIATED BY MIR-20B-5P. HYPOXIA-INDUCED H9C2 CELL APOPTOSIS WAS REDUCED AFTER HIF-1ALPHA KNOCKDOWN AS SHOWN BY THE FLOW CYTOMETRY ANALYSIS. IN CONCLUSION, WE IDENTIFIED THAT MIR-20B-5P PLAYS AN IMPORTANT ROLE IN MEDIATING CARDIOMYOCYTES APOPTOSIS UNDER HYPOXIA, WHICH IS MEDIATED BY THE HIF-1/NF-KAPPAB SIGNALING PATHWAY. 2020 14 3351 56 HISTONE DEMETHYLASE JARID1B REGULATES PROLIFERATION AND MIGRATION OF PULMONARY ARTERIAL SMOOTH MUSCLE CELLS IN MICE WITH CHRONIC HYPOXIA-INDUCED PULMONARY HYPERTENSION VIA NUCLEAR FACTOR-KAPPA B (NFKB). CHRONIC HYPOXIA-INDUCED PULMONARY HYPERTENSION (PH) IS A DISORDER THAT IS CHARACTERIZED BY INCREASED PULMONARY ARTERIAL PRESSURE RESULTING FROM LUNG DISEASES OR SHORTAGE OF OXYGEN IN THE BODY. EXCESS PROLIFERATION OF PULMONARY VASCULAR CELLS SUCH AS PULMONARY ARTERY ENDOTHELIAL CELLS (PAECS) AND PULMONARY ARTERY SMOOTH MUSCLE CELLS (PASMCS) PLAYS A CRITICAL ROLE IN THE PATHOGENESIS OF PH. RECENT EVIDENCE INDICATES THAT, IN ADDITION TO GENETIC PREDISPOSITION AND ENVIRONMENTAL FACTORS, EPIGENETIC MECHANISMS PLAY A PIVOTAL ROLE IN ETIOLOGY OF PH. IN THIS STUDY, WE INVESTIGATED THE POSSIBLE ROLE PLAYED BY JUMONJI AT-RICH INTERACTIVE DOMAIN 1B (JARID1B), A HISTONE DEMETHYLASE, IN REGULATING THE PROLIFERATION OF VASCULAR SMOOTH MUSCLE CELLS IN CHRONIC HYPOXIA-INDUCED PH CONDITION. QUANTITATIVE POLYMERASE CHAIN REACTION ANALYSIS OF SAMPLES FROM RATS WITH PH SHOWED AN ELEVATED EXPRESSION OF JARID1B IN THEIR PASMCS, POSITIVELY CORRELATING WITH INCREASED NUCLEAR FACTOR-KAPPA B (NFKB) EXPRESSION. FURTHER FUNCTIONAL STUDIES IN VITRO INDICATED THAT OVEREXPRESSION OF JARID1B INCREASED THE PROLIFERATION AND MIGRATION OF PASMCS, WHICH WERE INHIBITED BY DEPLETION OF NFKB. GENOMEWIDE TRANSCRIPTIONAL ANALYSIS REVEALED THAT THE JARID1B REGULATED NFKB SIGNALING PATHWAY BY DIRECTLY BINDING TO ITS PROMOTER. WE HAVE ALSO SHOWN THAT JARID1B INDIRECTLY REGULATES THE EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR VIA NFKB SIGNALING AND HENCE MAY ALSO PLAY A CRUCIAL ROLE IN CONTROLLING PAECS, LEADING TO CHANGES IN VASCULAR ARCHITECTURE IN PH. OUR FINDINGS COULD LEAD TO FURTHER STUDIES ON THE ROLE OF JARID1B IN PH ETIOLOGY AND THEREFORE COULD LEAD TO A POTENTIAL THERAPEUTIC TARGET FOR CHRONIC HYPOXIA INDUCED PULMONARY HYPERTENSION. 2018 15 5636 39 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 16 3096 37 GENOMIC CHARACTERIZATION REVEALS NOVEL MECHANISMS UNDERLYING THE VALOSIN-CONTAINING PROTEIN-MEDIATED CARDIAC PROTECTION AGAINST HEART FAILURE. CHRONIC HYPERTENSION IS A KEY RISK FACTOR FOR HEART FAILURE. HOWEVER, THE UNDERLYING MOLECULAR MECHANISMS ARE NOT FULLY UNDERSTOOD. OUR PREVIOUS STUDIES FOUND THAT THE VALOSIN-CONTAINING PROTEIN (VCP), AN ATPASE-ASSOCIATED PROTEIN, WAS SIGNIFICANTLY DECREASED IN THE HYPERTENSIVE HEART TISSUES. IN THIS STUDY, WE TESTED THE HYPOTHESIS THAT RESTORATION OF VCP PROTECTED THE HEART AGAINST PRESSURE OVERLOAD-INDUCED HEART FAILURE. WITH A CARDIAC-SPECIFIC TRANSGENIC (TG) MOUSE MODEL, WE SHOWED THAT A MODERATE INCREASE OF VCP WAS ABLE TO ATTENUATE CHRONIC PRESSURE OVERLOAD-INDUCED MALADAPTIVE CARDIAC HYPERTROPHY AND DYSFUNCTION. RNA SEQUENCING AND A COMPREHENSIVE BIOINFORMATIC ANALYSIS FURTHER DEMONSTRATED THAT OVEREXPRESSION OF VCP IN THE HEART NORMALIZED THE PRESSURE OVERLOAD-STIMULATED HYPERTROPHIC SIGNALS AND REPRESSED THE STRESS-INDUCED INFLAMMATORY RESPONSE. IN ADDITION, VCP OVEREXPRESSION PROMOTED CELL SURVIVAL BY ENHANCING THE MITOCHONDRIA RESISTANCE TO THE OXIDATIVE STRESS VIA ACTIVATING THE RICTOR-MEDIATED-GENE NETWORKS. VCP WAS ALSO FOUND TO BE INVOLVED IN THE REGULATION OF THE ALTERNATIVE SPLICING AND DIFFERENTIAL ISOFORM EXPRESSION FOR SOME GENES THAT ARE RELATED TO ATP PRODUCTION AND PROTEIN SYNTHESIS BY INTERACTING WITH LONG NO-CODING RNAS AND HISTONE DEACETYLASES, INDICATING A NOVEL EPIGENETIC REGULATION OF VCP IN INTEGRATING CODING AND NONCODING GENOMIC NETWORK IN THE STRESSED HEART. IN SUMMARY, OUR STUDY DEMONSTRATED THAT THE RESCUING OF A DEFICIENT VCP IN THE HEART COULD PREVENT PRESSURE OVERLOAD-INDUCED HEART FAILURE BY RECTIFYING CARDIAC HYPERTROPHIC AND INFLAMMATORY SIGNALING AND ENHANCING THE CARDIAC RESISTANCE TO OXIDATIVE STRESS, WHICH BROUGHT IN NOVEL INSIGHTS INTO THE UNDERSTANDING OF THE MECHANISM OF VCP IN PROTECTING PATIENTS FROM HYPERTENSIVE HEART FAILURE. 2020 17 3390 33 HOPX PLAYS A CRITICAL ROLE IN ANTIRETROVIRAL DRUGS INDUCED EPIGENETIC MODIFICATION AND CARDIAC HYPERTROPHY. PEOPLE LIVING WITH HIV (PLWH) HAVE TO TAKE AN ANTIRETROVIRAL THERAPY (ART) FOR LIFE AND SHOW NONCOMMUNICABLE ILLNESSES SUCH AS CHRONIC INFLAMMATION, IMMUNE ACTIVATION, AND MULTIORGAN DYSREGULATION. RECENT STUDIES SUGGEST THAT LONG-TERM USE OF ART INDUCES COMORBID CONDITIONS AND IS ONE OF THE LEADING CAUSES OF HEART FAILURE IN PLWH. HOWEVER, THE MOLECULAR MECHANISM OF ANTIRETROVIRAL DRUGS (ARVS) INDUCED HEART FAILURE IS UNCLEAR. TO DETERMINE THE MECHANISM OF ARVS INDUCED CARDIAC DYSFUNCTION, WE PERFORMED GLOBAL TRANSCRIPTOMIC PROFILING OF ARVS TREATED NEONATAL RAT VENTRICULAR CARDIOMYOCYTES IN CULTURE. DIFFERENTIALLY EXPRESSED GENES WERE IDENTIFIED BY RNA-SEQUENCING. OUR DATA SHOW THAT ARVS TREATMENT CAUSES UPREGULATION OF SEVERAL BIOLOGICAL FUNCTIONS ASSOCIATED WITH CARDIOTOXICITY, HYPERTROPHY, AND HEART FAILURE. GLOBAL GENE EXPRESSION DATA WERE VALIDATED IN CARDIAC TISSUE ISOLATED FROM HIV PATIENTS HAVING A HISTORY OF ART. INTERESTINGLY, WE FOUND THAT HOMEODOMAIN-ONLY PROTEIN HOMEOBOX (HOPX) EXPRESSION WAS SIGNIFICANTLY INCREASED IN CARDIOMYOCYTES TREATED WITH ARVS AND IN THE HEART TISSUE OF HIV PATIENTS. FURTHERMORE, WE FOUND THAT HOPX PLAYS A CRUCIAL ROLE IN ARVS MEDIATED CELLULAR HYPERTROPHY. MECHANISTICALLY, WE FOUND THAT HOPX PLAYS A CRITICAL ROLE IN EPIGENETIC REGULATION, THROUGH DEACETYLATION OF HISTONE, WHILE THE HDAC INHIBITOR, TRICHOSTATIN A, CAN RESTORE THE ACETYLATION LEVEL OF HISTONE 3 IN THE PRESENCE OF ARVS. 2021 18 1334 31 DEREGULATION OF AIOLOS EXPRESSION IN CHRONIC LYMPHOCYTIC LEUKEMIA IS ASSOCIATED WITH EPIGENETIC MODIFICATIONS. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS CHARACTERIZED BY A CLONAL ACCUMULATION OF MATURE NEOPLASTIC B CELLS THAT ARE RESISTANT TO APOPTOSIS. AIOLOS, A MEMBER OF THE IKAROS FAMILY OF ZINC-FINGER TRANSCRIPTION FACTORS, PLAYS AN IMPORTANT ROLE IN THE CONTROL OF MATURE B LYMPHOCYTE DIFFERENTIATION AND MATURATION. IN THIS STUDY, WE SHOWED THAT AIOLOS EXPRESSION IS UP-REGULATED IN B-CLL CELLS. THIS OVEREXPRESSION DOES NOT IMPLICATE ISOFORM IMBALANCE OR DISTURB AIOLOS SUBCELLULAR LOCALIZATION. THE CHROMATIN STATUS AT THE AIOLOS PROMOTER IN CLL IS DEFINED BY THE DEMETHYLATION OF DNA AND AN ENRICHMENT OF EUCHROMATIN ASSOCIATED HISTONE MARKERS, SUCH AS THE DIMETHYLATION OF THE LYSINE 4 ON HISTONE H3. THESE EPIGENETIC MODIFICATIONS SHOULD ALLOW ITS UPSTREAM EFFECTORS, SUCH AS NUCLEAR FACTOR-KAPPAB, CONSTITUTIVELY ACTIVATED IN CLL, TO GAIN ACCESS TO PROMOTER, RESULTING UP-REGULATION OF AIOLOS. TO DETERMINE THE CONSEQUENCES OF AIOLOS DEREGULATION IN CLL, WE ANALYZED THE EFFECTS OF AIOLOS OVEREXPRESSION OR DOWN-REGULATION ON APOPTOSIS. AIOLOS IS INVOLVED IN CELL SURVIVAL BY REGULATING THE EXPRESSION OF SOME BCL-2 FAMILY MEMBERS. OUR RESULTS STRONGLY SUGGEST THAT AIOLOS DEREGULATION BY EPIGENETIC MODIFICATIONS MAY BE A HALLMARK OF CLL. 2011 19 2432 33 EPIGENETIC SILENCING OF MIR-708 ENHANCES NF-KAPPAB SIGNALING IN CHRONIC LYMPHOCYTIC LEUKEMIA. MICRORNAS (MIRNAS) ARE POST-TRANSCRIPTIONAL REGULATORS OF GENE EXPRESSION AND THEIR DEREGULATION IS INVOLVED IN TUMOR DEVELOPMENT. EPIGENETIC GENE SILENCING IN CANCER BY DNA METHYLATION CONTRIBUTES TO THE SILENCING OF TUMOR-SUPPRESSOR GENES, INCLUDING MIRNAS. WE HAVE RECENTLY SHOWN THAT THE PROMOTER OF MIR-708 IS ABERRANTLY METHYLATED IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). TO CHARACTERIZE THE MOLECULAR SIGNALING NETWORKS THAT ARE INFLUENCED BY MIR-708, WE PERFORMED A LUCIFERASE-BASED SCREEN EVALUATING THE EFFECTS OF ECTOPIC MIR-708 EXPRESSION ON LEUKEMIA-RELEVANT SIGNALING PATHWAYS. WE FOUND THAT MIR-708 STRONGLY REPRESSED NF-KAPPAB SIGNALING, A PATHWAY KNOWN TO BE DEREGULATED IN CLL. AMONG THE PREDICTED MIR-708 TARGETS WAS IKKBETA (INHIBITOR OF KAPPA LIGHT POLYPEPTIDE GENE ENHANCER IN B CELLS, KINASE-BETA/IKBKB), A KEY KINASE FACILITATING NF-KAPPAB SIGNALING. WE VALIDATED THE INTERACTION OF MIR-708 WITH THE 3'-UNTRANSLATED REGION OF IKKBETA AND FOUND THAT MIR-708 OVEREXPRESSION REPRESSES ENDOGENOUS IKKBETA. PHOSPHORYLATION OF THE IKKBETA TARGET IKAPPABALPHA AND EXPRESSION OF KNOWN NF-KAPPAB TARGET GENES WERE IMPAIRED BY MIR-708. FURTHERMORE, WE IDENTIFIED AN ENHANCER REGION DOWNSTREAM OF THE MIR-708 PROMOTER THAT DISPLAYS A DISTINCT DNA METHYLATION STATUS IN CLL. HIGH ENHANCER METHYLATION IS SIGNIFICANTLY CORRELATED WITH LOWER MIR-708 EXPRESSION AND IS PREDOMINANTLY FOUND IN PATIENTS WITH POOR PROGNOSIS AND SHORTER TIME TO TREATMENT. THESE RESULTS DEMONSTRATE THAT MIR-708 REGULATES THE NF-KAPPAB PATHWAY BY TARGETING IKKBETA, AND THAT METHYLATION OF A KEY ENHANCER REGION CONTRIBUTES TO ITS SUPPRESSION IN CLL. 2015 20 3306 31 HIGH-PHOSPHATE-INDUCED CALCIFICATION IS RELATED TO SM22ALPHA PROMOTER METHYLATION IN VASCULAR SMOOTH MUSCLE CELLS. HYPERPHOSPHATEMIA IS CLOSELY RELATED TO VASCULAR CALCIFICATION IN PATIENTS WITH CHRONIC KIDNEY DISEASE. VASCULAR SMOOTH MUSCLE CELLS (VSMCS) EXPOSED TO HIGH PHOSPHATE CONCENTRATIONS IN VITRO UNDERGO PHENOTYPIC TRANSITION TO OSTEOBLAST-LIKE CELLS. MECHANISMS UNDERLYING THIS TRANSDIFFERENTIATION ARE NOT CLEAR. IN THIS STUDY WE USED TWO IN VITRO MODELS, HUMAN AORTIC SMOOTH MUSCLE CELLS AND RAT AORTIC RINGS, TO INVESTIGATE THE PHENOTYPIC TRANSITION OF VSMCS INDUCED BY HIGH PHOSPHATE. WE FOUND THAT HIGH PHOSPHATE CONCENTRATION (3.3 MMOL/L) IN THE MEDIUM WAS ASSOCIATED WITH INCREASED DNA METHYLTRANSFERASE ACTIVITY AND METHYLATION OF THE PROMOTER REGION OF SM22ALPHA. THIS WAS ACCOMPANIED BY LOSS OF THE SMOOTH MUSCLE CELL-SPECIFIC PROTEIN SM22ALPHA, GAIN OF THE OSTEOBLAST TRANSCRIPTION FACTOR CBFA1, AND INCREASED ALKALINE PHOSPHATASE ACTIVITY WITH THE SUBSEQUENT IN VITRO CALCIFICATION. THE ADDITION OF A DEMETHYLATING AGENT (PROCAINE) TO THE HIGH-PHOSPHATE MEDIUM REDUCED DNA METHYLTRANSFERASE ACTIVITY AND PREVENTED METHYLATION OF THE SM22ALPHA PROMOTER, WHICH WAS ACCOMPANIED BY AN INCREASE IN SM22ALPHA EXPRESSION AND LESS CALCIFICATION. ADDITIONALLY, DOWNREGULATION OF SM22ALPHA, EITHER BY SIRNA OR BY A METHYL GROUP DONOR (S-ADENOSYL METHIONINE), RESULTED IN OVEREXPRESSION OF CBFA1. IN CONCLUSION, WE DEMONSTRATE THAT METHYLATION OF SM22ALPHA PROMOTER IS AN IMPORTANT EVENT IN VASCULAR SMOOTH MUSCLE CELL CALCIFICATION AND THAT HIGH PHOSPHATE INDUCES THIS EPIGENETIC MODIFICATION. THESE FINDINGS UNCOVER A NEW INSIGHT INTO MECHANISMS BY WHICH HIGH PHOSPHATE CONCENTRATION PROMOTES VASCULAR CALCIFICATION. 2010