1 3449 155 HYPERMETHYLATION OF THE NRF2 PROMOTER INDUCES FERROPTOSIS BY INHIBITING THE NRF2-GPX4 AXIS IN COPD. BACKGROUND: NUCLEAR FACTOR E2-RELATED FACTOR 2 (NRF2) IS INVOLVED IN OXIDATIVE STRESS AND LUNG INFLAMMATION AND REGULATES THE ETIOLOGY OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). FERROPTOSIS IS CHARACTERIZED BY THE ACCUMULATION OF LIPID REACTIVE OXYGEN SPECIES (ROS) VIA FERROUS ION-DEPENDENT FENTON REACTIONS AND IS INVOLVED IN COPD. HOWEVER, THE ROLE OF NRF2 IN FERROPTOSIS AND ITS EPIGENETIC REGULATION IN THE PATHOGENESIS OF COPD REMAIN UNCLEAR. METHODS: FERROPTOSIS WAS DETECTED BY 4-HNE, MDA, C11BODIPY, DCFH-DA, PEALS' STAINING AND CCK-8 ASSAYS. QPCR AND WESTERN BLOTTING WERE PERFORMED TO EXAMINE THE NRF2 LEVELS IN PERIPHERAL LUNG TISSUES, PRIMARY EPITHELIAL CELLS COLLECTED FROM PATIENTS WITH COPD AND SUBJECTS WITH NORMAL PULMONARY FUNCTION (NEVER-SMOKER [CONTROL-NS]; SMOKER [CONTROL-S]), AND CIGARETTE SMOKE EXTRACT (CSE)-TREATED HUMAN BRONCHIAL EPITHELIAL (HBE) CELLS. ELISA WAS USED TO QUANTIFY IL-8 AND IL-1BETA LEVELS. METHYLATION OF THE NRF2 PROMOTER WAS ANALYZED BY BISULFITE SEQUENCING AND PYROSEQUENCING. RESULTS: FERROPTOSIS WAS INVOLVED IN COPD AND GLUTATHIONE PEROXIDASE 4 (GPX4) EXPRESSION WAS DOWNREGULATED IN THE COPD GROUP. REACTIVE OXYGEN SPECIES (ROS), LIPID PEROXIDES AND MDA WERE INCREASED, BUT GPX4 AND SOD WERE EXHAUSTED IN CSE-TREATED HBE CELLS. THE PRODUCTION OF IL-1BETA AND IL-8 WAS PROMOTED IN HBE CELLS IN RESPONSE TO CSE BUT COULD BE REVERSED BY THE FERROPTOSIS INHIBITOR FER-1. THE NRF2 LEVEL WAS SIGNIFICANTLY DECREASED IN THE COPD GROUP COMPARED WITH THE CONTROL-S AND CONTROL-NS GROUPS. INCREASED NRF2 EXPRESSION ENHANCED GPX4 AND SOD LEVELS AND INHIBITED FERROPTOSIS AND PROINFLAMMATORY CYTOKINES IN THE SUPERNATANT. INHIBITION OF GPX4 REVERSED THE EFFECT OF NRF2 OVEREXPRESSION AND PROMOTED FERROPTOSIS. TWO SPECIFIC CPG SITES WITHIN THE NRF2 PROMOTER WERE HYPERMETHYLATED IN THE COPD GROUP. SIMILARLY, CSE-TREATED HBE CELLS EXHIBITED HYPERMETHYLATION OF THE NRF2 GENE. CONCLUSION: NRF2 EXPRESSION WAS DOWNREGULATED IN THE LUNGS OF COPD PATIENTS DUE TO HYPERMETHYLATION OF THE NRF2 PROMOTER, INHIBITING NRF2/GPX4 AND FERROPTOSIS, WHICH IS RELATED TO THE INITIATION AND PROGRESSION OF COPD. TARGETING NRF2/GPX4 MAY INHIBIT FERROPTOSIS, WHICH COULD PROVIDE STRATEGIES TO DELAY OR TREAT COPD.	2021	
                                                                                                                                                                                                                                                                                                                         
2 3828  41 INVOLVEMENT OF B-CELL CLL/LYMPHOMA 2 PROMOTER METHYLATION IN CIGARETTE SMOKE EXTRACT-INDUCED EMPHYSEMA. ABNORMAL APOPTOTIC EVENTS PLAY AN IMPORTANT ROLE IN THE PATHOGENESIS OF EMPHYSEMA. THE B-CELL CLL/LYMPHOMA 2 (BCL-2) FAMILY PROTEINS ARE ESSENTIAL AND CRITICAL REGULATORS OF APOPTOSIS. WE DETERMINED WHETHER THE ANTI-APOPTOTIC BCL-2 PLAY A ROLE IN THE CIGARETTE SMOKE EXTRACT (CSE)-INDUCED EMPHYSEMA. FURTHERMORE, GIVEN THE INVOLVEMENT OF EPIGENETICS IN CHRONIC OBSTRUCTIVE PULMONARY DISEASE, WE HYPOTHESIZED THAT THE DEREGULATION OF BCL-2 MIGHT BE CAUSED BY GENE METHYLATION. THE EMPHYSEMA IN BALB/C MICE WAS ESTABLISHED BY INTRAPERITONEALLY INJECTION OF CSE. 5-AZA-2'-DEOXYCYTIDINE (AZA; A DEMETHYLATION REAGENT) AND PHOSPHATE-BUFFERED SALINE WERE ALSO ADMINISTERED INTRAPERITONEALLY AS CSE. TUNEL ASSAY WAS USED TO ASSESS APOPTOTIC INDEX OF PULMONARY CELLS. THE METHYLATION STATUS OF CPG DINUCLEOTIDES WITHIN THE BCL-2 PROMOTER WAS OBSERVED IN ALL GROUPS BY BISULFITE SEQUENCING PCR. PULMONARY EXPRESSION OF BCL-2, BAX, AND CYTOCHROME C WERE MEASURED AFTER FOUR WEEKS OF TREATMENT. THE APOPTOTIC INDEX OF PULMONARY CELLS IN CSE INJECTION GROUP WAS MUCH HIGHER THAN CONTROL ((25.88 +/- 7.55)% VS (6.28 +/- 2.96)%). COMPARED TO CONTROL MICE, DECREASED EXPRESSION OF BCL-2 AND HIGH METHYLATION OF BCL-2 PROMOTER WAS OBSERVED IN CSE INJECTED MICE (0.88 +/- 0.08 VS 0.49 +/- 0.11, (3.82 +/- 1.34)% VS (35.68 +/- 5.99)%, P < 0.01).CSE TREATMENT INDUCED LUNG CELL APOPTOSIS AND DECREASED LUNG FUNCTION. AZA TREATMENT INCREASED BCL-2 EXPRESSION WITH BCL-2 PROMOTER DEMETHYLATION. AZA ALSO ALLEVIATED THE LUNG CELL APOPTOSIS AND FUNCTION FAILURE CAUSED BY CSE TREATMENT. THE DECREASED EXPRESSION OF ANTI-APOPTOTIC BCL-2 MIGHT ACCOUNT FOR THE INCREASED APOPTOSIS IN CSE INDUCED-EMPHYSEMA. APPARENTLY, EPIGENETIC ALTERNATION PLAYED A ROLE IN THIS DEREGULATION OF BCL-2 EXPRESSION, AND IT MIGHT SUPPORT THE INVOLVEMENT OF EPIGENETIC EVENTS IN THE PATHOGENESIS OF EMPHYSEMA.	2016	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
3 4302  44 MICRORNA-223 CONTROLS THE EXPRESSION OF HISTONE DEACETYLASE 2: A NOVEL AXIS IN COPD. REDUCED ACTIVITY OF HISTONE DEACETYLASE 2 (HDAC2) HAS BEEN DESCRIBED IN PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD), BUT THE MECHANISMS RESULTING IN DECREASED EXPRESSION OF THIS IMPORTANT EPIGENETIC MODIFIER REMAIN UNKNOWN. HERE, WE EMPLOYED SEVERAL IN VITRO EXPERIMENTS TO ADDRESS THE ROLE OF MICRORNAS (MIRNAS) ON THE REGULATION OF HDAC2 IN ENDOTHELIAL CELLS. MANIPULATION OF MIRNA LEVELS IN HUMAN PULMONARY ARTERY ENDOTHELIAL CELLS (HPAEC) WAS ACHIEVED BY USING ELECTROPORATION WITH ANTI-MIRNAS AND MIRNA MIMICS. TARGET PREDICTION SOFTWARE IDENTIFIED MIR-223 AS A POTENTIAL REPRESSOR OF HDAC2. IN SUBSEQUENT STIMULATION EXPERIMENTS USING INFLAMMATORY CYTOKINES KNOWN TO BE INCREASED IN PATIENTS WITH COPD, MIR-223 WAS FOUND TO BE SIGNIFICANTLY INDUCED. FUNCTIONAL ANALYSIS DEMONSTRATED THAT OVEREXPRESSION OF MIR-223 DECREASED HDAC2 EXPRESSION AND ACTIVITY IN HPAEC. CONVERSELY, HDAC2 EXPRESSION AND ACTIVITY WAS PRESERVED IN ANTI-MIR-223-TREATED CELLS. DIRECT MIRNA-TARGET INTERACTION WAS CONFIRMED BY REPORTER GENE ASSAY. IN A NEXT STEP, REDUCED EXPRESSION OF HDAC2 WAS FOUND TO INCREASE THE LEVELS OF THE CHEMOKINE FRACTALKINE (CX3CL1). IN VIVO STUDIES CONFIRMED ELEVATED EXPRESSION LEVELS OF MIR-223 IN MICE EXPOSED TO CIGARETTE SMOKE AND IN EMPHYSEMATOUS LUNG TISSUE FROM LPS-TREATED MICE. MOREOVER, A SIGNIFICANT INVERSE CORRELATION OF MIR-223 AND HDAC2 EXPRESSION WAS FOUND IN TWO INDEPENDENT COHORTS OF COPD PATIENTS. THESE DATA EMPHASIZE THAT MIR-223, THE MOST PREVALENT MIRNA IN COPD, CONTROLS EXPRESSION AND ACTIVITY OF HDAC2 IN PULMONARY CELLS, WHICH, IN TURN, MIGHT ALTER THE EXPRESSION PROFILE OF CHEMOKINES. THIS PATHWAY PROVIDES A NOVEL PATHOGENIC LINK BETWEEN DYSREGULATED MIRNA EXPRESSION AND EPIGENETIC ACTIVITY IN COPD. KEY MESSAGES: HISTONE DEACETYLASE 2 IS DIRECTLY TARGETED BY MIR-223. LEVELS OF MIR-223 ARE INDUCED BY INTERLEUKIN-1BETA AND TUMOR NECROSIS FACTOR-ALPHA. MIR-223 CONTROLS THE EXPRESSION OF FRACTALKINE BY TARGETING HISTONE DEACETYLASE 2. MIR-223 LEVELS ARE INCREASED IN COPD MOUSE MODELS. MIR-223 LEVELS INVERSELY CORRELATE WITH HDAC2 EXPRESSION IN COPD PATIENTS.	2016	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
4 6297  53 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	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              
5 5422  43 REGULATION OF LUNG EPITHELIAL CELL SENESCENCE IN SMOKING-INDUCED COPD/EMPHYSEMA BY MICROR-125A-5P VIA SP1 MEDIATION OF SIRT1/HIF-1A. CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) AFFECTS THE HEALTH OF MORE THAN 300 MILLION PEOPLE WORLDWIDE; AT PRESENT, THERE IS NO EFFECTIVE DRUG TO TREAT COPD. SMOKING IS THE MOST IMPORTANT RISK FACTOR, BUT THE MOLECULAR MECHANISM BY WHICH SMOKING CAUSES THE DISEASE IS UNCLEAR. THE SENESCENCE OF LUNG EPITHELIAL CELLS IS RELATED TO DEVELOPMENT OF COPD. REGULATION OF MIRNAS IS THE MAIN EPIGENETIC MECHANISM RELATED TO AGING. BETA-GALACTOSE STAINING SHOWED THAT THE LUNG TISSUES OF SMOKERS HAVE A HIGHER DEGREE OF CELLULAR SENESCENCE, AND THE EXPRESSION OF MIR-125A-5P IS HIGH. THIS EFFECT IS OBVIOUS FOR SMOKERS WITH COPD/EMPHYSEMA, AND THERE IS A NEGATIVE CORRELATION BETWEEN MIR-125A-5P LEVELS AND VALUES FOR FORCED EXPIRATORY VOLUME IN ONE SECOND (FEV1)/FORCED VITAL CAPACITY (FVC). AFTER BALB/C MICE WERE CHRONICALLY EXPOSED TO VARIOUS CONCENTRATIONS OF CIGARETTE SMOKE (CS), PLETHYSMOGRAPHY SHOWED THAT LUNG FUNCTION WAS IMPAIRED, LUNG TISSUE SENESCENCE WAS INCREASED, AND THE SENESCENCE-ASSOCIATED SECRETORY PHENOTYPE (SASP) IN BRONCHOALVEOLAR LAVAGE FLUID WAS INCREASED. FOR MOUSE LUNG EPITHELIAL (MLE)-12 CELLS TREATED WITH CIGARETTE SMOKE EXTRACT (CSE), SP1 AND SIRT1 LEVELS WERE LOW, HIF-1ALPHA ACETYLATION LEVELS WERE HIGH, AND CELL SENESCENCE AND SECRETION OF SASP FACTORS WERE ELEVATED. DOWN-REGULATION OF MIR-125A-5P OR UP-REGULATION OF SP1 REVERSED THESE EFFECTS. IN ADDITION, COMPARED WITH MICE EXPOSED TO CS, KNOCKDOWN OF MIR-125A-5P REDUCED LUNG EPITHELIAL CELL SENESCENCE AND COPD/EMPHYSEMA. THEREFORE, IN SMOKING-INDUCED COPD, ELEVATED MIR-125A-5P PARTICIPATES IN THE SENESCENCE OF LUNG EPITHELIAL CELLS THROUGH SP1/SIRT1/HIF-1ALPHA. THESE FINDINGS PROVIDE EVIDENCE RELATED TO THE PATHOGENESIS OF COPD/EMPHYSEMA CAUSED BY CHRONIC SMOKING.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         
6 2774  35 EXTRACELLULAR SUPEROXIDE DISMUTASE (EC-SOD) REGULATES GENE METHYLATION AND CARDIAC FIBROSIS DURING CHRONIC HYPOXIC STRESS. CHRONIC HYPOXIC STRESS INDUCES EPIGENETIC MODIFICATIONS MAINLY DNA METHYLATION IN CARDIAC FIBROBLASTS, INACTIVATING TUMOR SUPPRESSOR GENES (RASSF1A) AND ACTIVATING KINASES (ERK1/2) LEADING TO FIBROBLAST PROLIFERATION AND CARDIAC FIBROSIS. THE RAS/ERK SIGNALING PATHWAY IS AN INTRACELLULAR SIGNAL TRANSDUCTION CRITICALLY INVOLVED IN FIBROBLAST PROLIFERATION. RASSF1A FUNCTIONS THROUGH ITS EFFECT ON DOWNSTREAM ERK1/2. THE ANTIOXIDANT ENZYME, EXTRACELLULAR SUPEROXIDE DISMUTASE (EC-SOD), DECREASES OXIDATIVE STRESS FROM CHRONIC HYPOXIA, BUT ITS EFFECTS ON THESE EPIGENETIC CHANGES HAVE NOT BEEN FULLY EXPLORED. TO TEST OUR HYPOTHESIS, WE USED AN IN-VITRO MODEL: WILD-TYPE C57B6 MALE MICE (WT) AND TRANSGENIC MALES WITH AN EXTRA COPY OF HUMAN HEC-SOD (TG). THE STUDIED ANIMALS WERE HOUSED IN HYPOXIA (10% O(2)) FOR 21 DAYS. THE RIGHT VENTRICULAR TISSUE WAS STUDIED FOR CARDIAC FIBROSIS MARKERS USING RT-PCR AND WESTERN BLOT ANALYSES. PRIMARY C57BL6 MOUSE CARDIAC FIBROBLAST TISSUE CULTURE WAS USED TO STUDY THE IN-VITRO MODEL, THE DOWNSTREAM EFFECTS OF RASSF-1 EXPRESSION AND METHYLATION, AND ITS RELATION TO ERK1/2. OUR FINDINGS SHOWED A SIGNIFICANT INCREASE IN CARDIAC FIBROSIS MARKERS: COLLAGEN 1, ALPHA SMOOTH MUSCLE ACTIN (ASMA), AND SNAIL, IN THE WT HYPOXIC ANIMALS AS COMPARED TO THE TG HYPOXIC GROUP (P < 0.05). THE EXPRESSION OF DNA METHYLATION ENZYMES (DNMT 1&3B) WAS SIGNIFICANTLY INCREASED IN THE WT HYPOXIC MICE AS COMPARED TO THE HYPOXIC TG MICE (P < 0.001). RASSF1A EXPRESSION WAS SIGNIFICANTLY LOWER AND ERK1/2 WAS SIGNIFICANTLY HIGHER IN HYPOXIA WT COMPARED TO THE HYPOXIC TG GROUP (P < 0.05). USE OF SIRNA TO BLOCK RASSF1A GENE EXPRESSION IN MURINE CARDIAC FIBROBLAST TISSUE CULTURE LED TO INCREASED FIBROBLAST PROLIFERATION (P < 0.05). METHYLATION OF THE RASSF1A PROMOTER REGION WAS SIGNIFICANTLY REDUCED IN THE TG HYPOXIC GROUP COMPARED TO THE WT HYPOXIC GROUP (0.59 VS. 0.75, RESPECTIVELY). BASED ON OUR FINDINGS, WE CAN SPECULATE THAT EC-SOD SIGNIFICANTLY ATTENUATES RASSF1A GENE METHYLATION AND CAN ALLEVIATE CARDIAC FIBROSIS INDUCED BY HYPOXIA.	2021	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
7 5418  51 REGULATION OF DNA METHYLATION SIGNATURES ON NF-KAPPAB AND STAT3 PATHWAY GENES AND TET ACTIVITY IN CIGARETTE SMOKE EXTRACT-CHALLENGED CELLS/COPD EXACERBATION MODEL IN VITRO. BACKGROUND: CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) IS A GLOBAL HEALTH PROBLEM. CURRENTLY, THERE IS A LACK OF KNOWLEDGE ABOUT THE PATHOBIOLOGY OF THIS DISEASE AND AVAILABLE THERAPIES ARE INEFFECTIVE. CIGARETTE SMOKING IS THE LEADING CAUSE OF COPD; HOWEVER, NOT ALL SMOKERS DEVELOP COPD. EXACERBATIONS OF COPD CAUSED BY MICROBES ARE COMMON AND DETRIMENTAL. APPROXIMATELY 20-50% OF PATIENT EXACERBATIONS ARE CAUSED BY BACTERIAL COLONIZATION IN THE LOWER AIRWAYS. IT IS GENERALLY ACCEPTED THAT EPIGENETIC MECHANISMS, ESPECIALLY DNA METHYLATION, PLAY AN IMPORTANT ROLE DURING PROGRESSION OF COPD. THUS, WE HYPOTHESIZED THAT DNA METHYLATION PATTERNS VARY SIGNIFICANTLY FOLLOWING SMOKE EXPOSURE AND DURING EXACERBATIONS CAUSED BY BACTERIAL INFECTIONS. TO TEST OUR HYPOTHESIS, WE USED AN IN VITRO STUDY MODEL THAT MIMICS COPD EXACERBATIONS AND PERFORMED EXTENSIVE STUDIES TO UNDERSTAND THE ROLE OF CPG PROMOTER METHYLATION OF NF-KAPPAB AND STAT3-MEDIATED PATHWAY GENES. BOTH NF-KAPPAB AND STAT3 TRANSCRIPTION FACTORS PLAY CRITICAL ROLES IN ORCHESTRATING INFLAMMATORY RESPONSES DURING CIGARETTE SMOKE EXPOSURE. IN BRIEF, HUMAN LUNG ADENOCARCINOMA CELLS WITH TYPE II ALVEOLAR EPITHELIUM CHARACTERISTICS (A549) WERE CHALLENGED WITH CIGARETTE SMOKE EXTRACT (CSE) OR DMSO (CONTROL) FOLLOWED BY A 3-H CHALLENGE WITH BACTERIAL LIPOPOLYSACCHARIDE (LPS; FROM PSEUDOMONAS AERUGINOSA) PRIOR TO THE TERMINATION OF CSE EXPOSURE (COPD EXACERBATION GROUP). THE PRODUCTION OF CYTOKINES/CHEMOKINES, REGULATION OF TRANSCRIPTION FACTORS, AND DNA METHYLATION OF SPECIFIC GENES WERE THEN ASSESSED. WE ALSO STUDIED CHANGES IN THE EXPRESSION AND ACTIVITY OF TEN-ELEVEN TRANSLOCASES (TETS), THE ENZYMES RESPONSIBLE FOR DNA DEMETHYLATION, AND ASSESSED THEIR ROLE IN REGULATING DNA METHYLATION IN THE CSE-CHALLENGED GROUP. RESULTS: THERE WAS A SIGNIFICANT INCREASE IN THE RELEASE OF CYTOKINES/CHEMOKINES (IL-8, MCP-1, IL-6 AND CCL5) IN THE COPD EXACERBATION GROUP AS COMPARED TO THE CONTROL GROUP. HYPOMETHYLATION OF NF-KAPPAB-MEDIATED PATHWAY GENES CORRELATED WITH THEIR INDUCTION IN OUR COPD EXACERBATION STUDY MODEL. FURTHER, WE OBSERVED AN IMPORTANT ROLE OF TET1/2 IN REGULATING THE DNA METHYLATION OF NF-KAPPAB, STAT3, IKK, AND NIK GENES AND CYTOKINE/CHEMOKINE PRODUCTION BY A549 CELLS DURING CSE CHALLENGE. CONCLUSIONS: STUDIES TO FURTHER DEFINE THE ROLE OF TETS IN CSE-MEDIATED EPIGENETIC REGULATION MAY LEAD TO THE DEVELOPMENT OF BETTER AND MORE EFFECTIVE THERAPEUTIC INTERVENTION STRATEGIES FOR COPD.	2020	

8 3447  55 HYPERMETHYLATION OF MITOCHONDRIAL TRANSCRIPTION FACTOR A INDUCED BY CIGARETTE SMOKE IS ASSOCIATED WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE. PURPOSE OF THE STUDY: CIGARETTE SMOKING IS A LEADING ENVIRONMENTAL CONTRIBUTOR TO CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD), BUT ITS EPIGENETIC REGULATION OF MTTFA GENE REMAINS ELUSIVE. THIS STUDY AIMS TO EXPLORE THE RELATIONSHIP OF DNA METHYLATION OF MTTFA AND CIGARETTE SMOKING IN COPD. MATERIALS AND METHODS: WE ANALYZED DNA METHYLATION ON MTTFA PROMOTERS IN CLINICAL SAMPLES FROM COPD PATIENTS AND SUBJECTS WITH NORMAL PULMONARY FUNCTION. EXPRESSION OF MTTFA MRNA IN THE CLINICAL SAMPLES AND MTTFA MRNA AND PROTEIN IN HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS(HUVECS) TREATED WITH CIGARETTE SMOKE EXTRACT (CSE) WAS EVALUATED. MTTFA MRNA AND PROTEIN LEVELS WERE MEASURED TO DETERMINE EFFECTS OF DEMETHYLATION AGENTS ON CSE-TREATED HUVECS. RESULTS: THE DNA METHYLATION LEVEL OF THE MTTFA PROMOTER WAS SIGNIFICANTLY INCREASED IN COPD GROUP. EXPRESSION OF MTTFA MRNA WAS DOWNREGULATED IN THE LUNGS AS A CONSEQUENCE OF HYPERMETHYLATION OF MTTFA PROMOTER. EXPRESSION OF MTTFA MRNA AND PROTEIN WAS DOWNREGULATED IN CSE-TREATED HUVECS AS A CONSEQUENCE OF HYPERMETHYLATION OF THE MTTFA PROMOTER. MTTFA EXPRESSION IN CSE-TREATED HUVECS WAS RESTORED BY THE METHYLATION INHIBITOR, 5-AZA-2'-DEOXYCYTIDINE(AZA). CONCLUSIONS: CIGARETTE SMOKE-INDUCED HYPERMETHYLATION OF THE MTTFA PROMOTER IS RELATED TO THE INITIATION AND PROGRESSION OF COPD. OUR FINDING MAY PROVIDE A NEW STRATEGY FOR THE INTERVENTION OF COPD BY DEVELOPING DEMETHYLATION AGENTS TARGETING MTTFA HYPERMETHYLATION.	2019	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                    
9 4899  50 OXIDATIVE STRESS MEDIATES THE APOPTOSIS AND EPIGENETIC MODIFICATION OF THE BCL-2 PROMOTER VIA DNMT1 IN A CIGARETTE SMOKE-INDUCED EMPHYSEMA MODEL. BACKGROUND: EMPHYSEMA IS A CRUCIAL PATHOLOGICAL CHARACTERISTIC OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). OXIDATIVE STRESS, APOPTOSIS AND EPIGENETIC MECHANISMS CONTRIBUTE TO THE PATHOGENESIS OF EMPHYSEMA. HOWEVER, AN ATTEMPT TO ACCURATELY IDENTIFY WHETHER THESE MECHANISMS INTERACT WITH EACH OTHER AND HOW THEY ARE TRIGGERED HAS NEVER BEEN CONDUCTED. METHOD: THE TOTAL REACTIVE OXYGEN SPECIES (ROS) LEVEL, PULMONARY APOPTOSIS AND B-CELL LYMPHOMA/LEUKEMIA-2 (BCL-2) EXPRESSION, AN APOPTOSIS REGULATOR, WERE DETECTED IN SAMPLES FROM COPD PATIENTS. BISULFITE SEQUENCING PCR (BSP) WAS CONDUCTED TO OBSERVE THE ALTERATIONS IN THE METHYLATION OF THE BCL-2 PROMOTER IN SPECIMENS. THE DYSREGULATION OF DNA METHYLTRANSFERASE ENZYME 1 (DNMT1), A VITAL DNA METHYLTRANSFERASE ENZYME, IN THE LUNGS OF PATIENTS WAS CONFIRMED THROUGH WESTERN BLOTTING. TO FIND OUT INTERACTIONS BETWEEN OXIDATIVE STRESS AND DNA METHYLATION IN EMPHYSEMA, MOUSE MODELS WERE BUILT WITH ANTIOXIDANT TREATMENT AND DNMT1 SILENCING, AND WERE EXAMINED WITH THE PULMONARY APOPTOSIS, BCL-2 AND DNMT1 LEVELS, AND EPIGENETIC ALTERATIONS OF BCL-2. RESULTS: HIGHER ROS LEVELS AND PULMONARY APOPTOSIS WERE OBSERVED IN COPD PATIENTS THAN IN HEALTHY CONTROLS. DOWNREGULATED BCL-2 EXPRESSION WITH INCREASED PROMOTER METHYLATION AND DNMT1 PROTEIN EXPRESSION WAS FOUND IN COPD PATIENTS. ANTIOXIDANT TREATMENT REDUCED THE LEVEL OF ROS, DNMT1 PROTEIN AND EMPHYSEMATOUS PROGRESSION IN THE SMOKING MODELS. FOLLOWING DNMT1 BLOCKADE, SMOKING MODELS SHOWED IMPROVED LUNG FUNCTION, PULMONARY APOPTOSIS, EMPHYSEMATOUS PROGRESSION, AND INCREASED BCL-2 PROTEIN LEVEL WITH LESS PROMOTER METHYLATION THAN EMPHYSEMA MICE. CONCLUSION: CIGARETTE-INDUCED OXIDATIVE STRESS MEDIATES PULMONARY APOPTOSIS AND HYPERMETHYLATION OF THE BCL-2 PROMOTER IN EMPHYSEMA MODELS THROUGH DNMT1.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
10 1591  42 DNA METHYLATION PROFILING IN PERIPHERAL LUNG TISSUES OF SMOKERS AND PATIENTS WITH COPD. BACKGROUND: EPIGENETICS CHANGES HAVE BEEN SHOWN TO BE AFFECTED BY CIGARETTE SMOKING. CIGARETTE SMOKE (CS)-MEDIATED DNA METHYLATION CAN POTENTIALLY AFFECT SEVERAL CELLULAR AND PATHOPHYSIOLOGICAL PROCESSES, ACUTE EXACERBATIONS, AND COMORBIDITY IN THE LUNGS OF PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). WE SOUGHT TO DETERMINE WHETHER GENOME-WIDE LUNG DNA METHYLATION PROFILES OF SMOKERS AND PATIENTS WITH COPD WERE SIGNIFICANTLY DIFFERENT FROM NON-SMOKERS. WE ISOLATED DNA FROM PARENCHYMAL LUNG TISSUES OF PATIENTS INCLUDING EIGHT LIFELONG NON-SMOKERS, EIGHT CURRENT SMOKERS, AND EIGHT PATIENTS WITH COPD AND ANALYZED THE SAMPLES USING ILLUMINA'S INFINIUM HUMANMETHYLATION450 BEADCHIP. RESULTS: OUR DATA REVEALED THAT THE DIFFERENTIALLY METHYLATED GENES WERE RELATED TO TOP CANONICAL PATHWAYS (E.G., G BETA GAMMA SIGNALING, MECHANISMS OF CANCER, AND NNOS SIGNALING IN NEURONS), DISEASE AND DISORDERS (ORGANISMAL INJURY AND ABNORMALITIES, CANCER, AND RESPIRATORY DISEASE), AND MOLECULAR AND CELLULAR FUNCTIONS (CELL DEATH AND SURVIVAL, CELLULAR ASSEMBLY AND ORGANIZATION, CELLULAR FUNCTION AND MAINTENANCE) IN PATIENTS WITH COPD. THE GENOME-WIDE DNA METHYLATION ANALYSIS IDENTIFIED SUGGESTIVE GENES, SUCH AS NOS1AP, TNFAIP2, BID, GABRB1, ATXN7, AND THOC7 WITH DNA METHYLATION CHANGES IN COPD LUNG TISSUES THAT WERE FURTHER VALIDATED BY PYROSEQUENCING. PYROSEQUENCING VALIDATION CONFIRMED HYPER-METHYLATION IN SMOKERS AND PATIENTS WITH COPD AS COMPARED TO NON-SMOKERS. HOWEVER, WE DID NOT DETECT SIGNIFICANT DIFFERENCES IN DNA METHYLATION FOR TNFAIP2, ATXN7, AND THOC7 GENES IN SMOKERS AND COPD GROUPS DESPITE THE CHANGES OBSERVED IN THE GENOME-WIDE ANALYSIS. CONCLUSIONS: OUR STUDY SUGGESTS THAT DNA METHYLATION IN SUGGESTIVE GENES, SUCH AS NOS1AP, BID, AND GABRB1 MAY BE USED AS EPIGENETIC SIGNATURES IN SMOKERS AND PATIENTS WITH COPD IF THE SAME IS VALIDATED IN A LARGER COHORT. FUTURE STUDIES ARE REQUIRED TO CORRELATE DNA METHYLATION STATUS WITH TRANSCRIPTOMICS OF SELECTIVE GENES IDENTIFIED IN THIS STUDY AND ELUCIDATE THEIR ROLE AND INVOLVEMENT IN THE PROGRESSION OF COPD AND ITS EXACERBATIONS.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                   
11  164  30 ABNORMAL HISTONE METHYLATION IS RESPONSIBLE FOR INCREASED VASCULAR ENDOTHELIAL GROWTH FACTOR 165A SECRETION FROM AIRWAY SMOOTH MUSCLE CELLS IN ASTHMA. VASCULAR ENDOTHELIAL GROWTH FACTOR (VEGF), A KEY ANGIOGENIC MOLECULE, IS ABERRANTLY EXPRESSED IN SEVERAL DISEASES INCLUDING ASTHMA WHERE IT CONTRIBUTES TO BRONCHIAL VASCULAR REMODELING AND CHRONIC INFLAMMATION. ASTHMATIC HUMAN AIRWAY SMOOTH MUSCLE CELLS HYPERSECRETE VEGF, BUT THE MECHANISM IS UNCLEAR. IN THIS STUDY, WE DEFINED THE MECHANISM IN HUMAN AIRWAY SMOOTH MUSCLE CELLS FROM NONASTHMATIC AND ASTHMATIC PATIENTS. WE FOUND THAT ASTHMATIC CELLS LACKED A REPRESSION COMPLEX AT THE VEGF PROMOTER, WHICH WAS PRESENT IN NONASTHMATIC CELLS. RECRUITMENT OF G9A, TRIMETHYLATION OF HISTONE H3 AT LYSINE 9 (H3K9ME3), AND A RESULTANT DECREASE IN RNA POLYMERASE II AT THE VEGF PROMOTER WAS CRITICAL TO REPRESSION OF VEGF SECRETION IN NONASTHMATIC CELLS. AT THE ASTHMATIC PROMOTER, H3K9ME3 WAS ABSENT BECAUSE OF FAILED RECRUITMENT OF G9A; RNA POLYMERASE II BINDING, IN ASSOCIATION WITH TATA-BINDING PROTEIN-ASSOCIATED FACTOR 1, WAS INCREASED; H3K4ME3 WAS PRESENT; AND SP1 BINDING WAS EXAGGERATED AND SUSTAINED. IN CONTRAST, DNA METHYLATION AND HISTONE ACETYLATION WERE SIMILAR IN ASTHMATIC AND NONASTHMATIC CELLS. THIS IS THE FIRST STUDY, TO OUR KNOWLEDGE, TO SHOW THAT AIRWAY CELLS IN ASTHMA HAVE ALTERED EPIGENETIC REGULATION OF REMODELING GENE(S). HISTONE METHYLATION AT GENES SUCH AS VEGF MAY BE AN IMPORTANT NEW THERAPEUTIC TARGET.	2012	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                    
12 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	
                                                                                                                                                                                                                                                                                                                                                                                                                     
13 5394  42 REDUCED DNA METHYLATION OF SPHINGOSINE-1 PHOSPHATE RECEPTOR 5 IN ALVEOLAR MACROPHAGES IN COPD: A POTENTIAL LINK TO FAILED EFFEROCYTOSIS. BACKGROUND AND OBJECTIVE: WE PREVIOUSLY SHOWED THAT ALVEOLAR MACROPHAGES FROM COPD PATIENTS ARE DEFECTIVE IN THEIR ABILITY TO PHAGOCYTOSE APOPTOTIC CELLS ('EFFEROCYTOSIS') AND THAT THIS DEFECT IS POTENTIALLY LINKED TO THE SPHINGOSINE-1 PHOSPHATE (S1P) SYSTEM, IN PARTICULAR THE SPHINGOSINE-1 PHOSPHATE RECEPTOR 5 (S1PR5). IN ALVEOLAR MACROPHAGES FROM COPD PATIENTS, S1PR5 MRNA EXPRESSION LEVELS INCREASED AND WERE CORRELATED WITH BOTH LUNG FUNCTION AND EFFEROCYTOSIS. HOWEVER, IT US UNKNOWN WHETHER THESE CHANGES ARE UNDER EPIGENETIC CONTROL VIA DNA METHYLATION OR WHETHER DNA METHYLATION DIRECTLY MODULATES MACROPHAGE FUNCTION. METHODS: BISULFITE SEQUENCING WAS USED TO ASSESS DNA METHYLATION LEVELS AT CPG ISLANDS ASSOCIATED WITH GENES ENCODING SELECTED S1P SYSTEM COMPONENTS, INCLUDING SPHINGOSINE KINASE 1 (SPHK1), S1PR1 AND S1PR5, IN ALVEOLAR MACROPHAGES FROM 20 COPD PATIENTS, 7 HEALTHY SMOKERS AND 10 HEALTHY NON/EX-SMOKERS) BY METHYL QUANTITATIVE REAL-TIME PCR (METHYL QPCR). THE EFFECT OF THE DNA METHYLTRANSFERASE INHIBITOR, 5-AZACYTIDINE ON THE EFFEROCYTOSIS CAPACITY OF THP-1 MACROPHAGES WAS ASSESSED USING FLOW CYTOMETRY. RESULTS: AMONG THE S1P SYSTEM GENES EXAMINED, S1PR5 WAS THE SINGLE TARGET THAT SHOWED SIGNIFICANT CHANGES IN DNA METHYLATION BETWEEN PATIENT GROUPS. ALVEOLAR MACROPHAGES ISOLATED FROM COPD PATIENTS SHOWED LOWER METHYLATION LEVELS IN THE SAME REGION COMPARED TO MACROPHAGES FROM NON/EX-SMOKERS. IN VITRO STUDIES USING THP-1 MACROPHAGES SHOWED THAT DNA DEMETHYLATION WITH 5-AZACYTIDINE INCREASED THE EFFEROCYTOSIS CAPACITY AND DOSE-DEPENDENTLY RESCUED THE CELLS FROM THE CIGARETTE SMOKE-INDUCED DEFECT IN EFFEROCYTOSIS. CONCLUSION: MACROPHAGE FUNCTION CAN BE MODULATED EPIGENETICALLY. REDUCED METHYLATION MAY UNDERLIE THE INCREASED EXPRESSION OF THE S1PR5 GENE IN ALVEOLAR MACROPHAGES AND ASSOCIATED DEFECTIVE EFFEROCYTOSIS IN COPD.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           
14 1011  27 CIGARETTE SMOKE CONDENSATE INDUCES DIFFERENTIAL EXPRESSION AND PROMOTER METHYLATION PROFILES OF CRITICAL GENES INVOLVED IN LUNG CANCER IN NL-20 LUNG CELLS IN VITRO: SHORT-TERM AND CHRONIC EXPOSURE. ESTABLISHING EARLY DIAGNOSTIC MARKERS OF HARM IS CRITICAL FOR EFFECTIVE PREVENTION PROGRAMS AND REGULATION OF TOBACCO PRODUCTS. THIS STUDY EXAMINED EFFECTS OF CIGARETTE SMOKE CONDENSATE (CSC) ON EXPRESSION AND PROMOTER METHYLATION PROFILE OF CRITICAL GENES (DAPK, ECAD, MGMT, AND RASSF1A) INVOLVED IN LUNG CANCER DEVELOPMENT IN DIFFERENT HUMAN LUNG CELL LINES. NL-20 CELLS WERE TREATED WITH 0.1-100 MUG/ML OF CSC FOR 24 TO 72 HRS FOR SHORT-TERM EXPOSURES. DAPK EXPRESSION OR METHYLATION STATUS WAS NOT SIGNIFICANTLY AFFECTED. HOWEVER, CSC TREATMENT RESULTED IN CHANGES IN EXPRESSION AND PROMOTER METHYLATION PROFILE OF ECAD, MGMT, AND RASSF1A. FOR CHRONIC STUDIES, CELLS WERE EXPOSED TO 1 OR 10 MUG/ML CSC UP TO 28 DAYS. CELLS SHOWED MORPHOLOGICAL CHANGES ASSOCIATED WITH TRANSFORMATION AND CHANGES IN INVASION CAPACITIES AND GLOBAL METHYLATION STATUS. THIS STUDY PROVIDES CRITICAL DATA SUGGESTING THAT EPIGENETIC CHANGES COULD SERVE AS AN EARLY BIOMARKER OF HARM DUE TO EXPOSURE TO CIGARETTE SMOKE.	2013	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                
15 3638  44 INCREASED EXPRESSION OF BETA-DEFENSIN 1 (DEFB1) IN CHRONIC OBSTRUCTIVE PULMONARY DISEASE. ON-GOING AIRWAY INFLAMMATION IS CHARACTERISTIC FOR THE PATHOPHYSIOLOGY OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD). HOWEVER, THE KEY FACTORS DETERMINING THE DECREASE IN LUNG FUNCTION, AN IMPORTANT CLINICAL PARAMETER OF COPD, ARE NOT CLEAR. GENOME-WIDE LINKAGE ANALYSES PROVIDE EVIDENCE FOR SIGNIFICANT LINKAGE TO AIRWAY OBSTRUCTION SUSCEPTIBILITY LOCI ON CHROMOSOME 8P23, THE LOCATION OF THE HUMAN DEFENSIN GENE CLUSTER. MOREOVER, A GENETIC VARIATION IN THE DEFENSIN BETA 1 (DEFB1) GENE WAS FOUND TO BE ASSOCIATED WITH COPD. THEREFORE, WE HYPOTHESIZED THAT DEFB1 IS DIFFERENTLY REGULATED AND EXPRESSED IN HUMAN LUNGS DURING COPD PROGRESSION. GENE EXPRESSION OF DEFB1 WAS ASSESSED IN BRONCHIAL EPITHELIUM AND BAL FLUID CELLS OF HEALTHY CONTROLS AND PATIENTS WITH COPD AND USING BISULFITE SEQUENCING AND CHIP ANALYSIS, THE EPIGENETIC CONTROL OF DEFB1 MRNA EXPRESSION WAS INVESTIGATED. WE CAN DEMONSTRATE THAT DEFB1 MRNA EXPRESSION WAS SIGNIFICANTLY INCREASED IN BRONCHOPULMONARY SPECIMEN OF PATIENTS WITH COPD (N = 34) VS. HEALTHY CONTROLS (N = 10) (P<0.0001). FURTHERMORE, A SIGNIFICANT CORRELATION COULD BE DETECTED BETWEEN DEFB1 AND FUNCTIONAL PARAMETERS SUCH AS FEV(1) (P = 0.0024) AND THE FEV(1)/VC RATIO (P = 0.0005). UPREGULATION OF DEFB1 MRNA WAS PARALLELED BY CHANGES IN HDAC1-3, HDAC5 AND HDAC8 MRNA EXPRESSION. WHEREAS BISULFITE SEQUENCING REVEALED NO DIFFERENCES IN THE METHYLATION STATE OF DEFB1 PROMOTER BETWEEN PATIENTS WITH COPD AND CONTROLS, CHIP ANALYSIS SHOWED THAT ENHANCED DEFB1 MRNA EXPRESSION WAS ASSOCIATED WITH THE ESTABLISHMENT OF AN ACTIVE HISTONE CODE. THUS, EXPRESSION OF HUMAN DEFB1 IS UPREGULATED AND RELATED TO THE DECREASE IN PULMONARY FUNCTION IN PATIENTS WITH COPD.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             
16 3767  36 INTEGRATIVE EPIGENOMIC ANALYSIS IN DIFFERENTIATED HUMAN PRIMARY BRONCHIAL EPITHELIAL CELLS EXPOSED TO CIGARETTE SMOKE. CIGARETTE SMOKE (CS) IS ONE OF THE MAJOR RISK FACTORS FOR MANY PULMONARY DISEASES, INCLUDING CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) AND LUNG CANCER. THE FIRST LINE OF DEFENSE FOR CS EXPOSURE IS THE BRONCHIAL EPITHELIAL CELLS. ELUCIDATION OF THE EPIGENETIC CHANGES DURING CS EXPOSURE IS KEY TO GAINING A MECHANISTIC UNDERSTANDING INTO HOW MATURE AND DIFFERENTIATED BRONCHIAL EPITHELIAL CELLS RESPOND TO CS. THEREFORE, WE PERFORMED EPIGENOMIC PROFILING IN CONJUNCTION WITH TRANSCRIPTIONAL PROFILING IN WELL-DIFFERENTIATED HUMAN BRONCHIAL EPITHELIAL (HBE) CELLS CULTURED IN AIR-LIQUID INTERFACE (ALI) EXPOSED TO THE VAPOR PHASE OF CS. THE GENOME-WIDE ENRICHMENT OF HISTONE 3 LYSINE 27 ACETYLATION WAS DETECTED BY CHROMATIN IMMUNOPRECIPITATION FOLLOWED BY NEXT GENERATION SEQUENCING (CHIP-SEQ) IN HBE CELLS AND SUGGESTED THE PLAUSIBLE BINDING OF SPECIFIC TRANSCRIPTION FACTORS RELATED TO CS EXPOSURE. ADDITIONALLY, INTERROGATION OF CHIP-SEQ DATA WITH GENE EXPRESSION PROFILING OF HBE CELLS AFTER CS EXPOSURE FOR DIFFERENT DURATIONS (3 HOURS, 2 DAYS, 4 DAYS) SUGGESTED THAT EARLIER EPIGENETIC CHANGES (3 HOURS AFTER CS EXPOSURE) MAY BE ASSOCIATED WITH LATER GENE EXPRESSION CHANGES INDUCED BY CS EXPOSURE (4 DAYS). THE INTEGRATION OF EPIGENETICS AND GENE EXPRESSION DATA REVEALED SIGNALING PATHWAYS RELATED TO CS-INDUCED EPIGENETIC CHANGES IN HBE CELLS THAT MAY IDENTIFY NOVEL REGULATORY PATHWAYS RELATED TO CS-INDUCED COPD.	2018	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     
17 3524  42 IL-13 REGULATES HUMAN NASAL EPITHELIAL CELL DIFFERENTIATION VIA H3K4ME3 MODIFICATION. INTRODUCTION: EPIGENETIC REGULATION HAS BEEN SHOWN TO PLAY AN IMPORTANT ROLE IN THE DEVELOPMENT OF INFLAMMATORY DISEASES, INCLUDING CHRONIC RHINOSINUSITIS AND NASAL POLYPS. THE LATTER ARE CHARACTERIZED BY EPITHELIAL MIS-DIFFERENTIATION AND INFILTRATION OF INFLAMMATORY CYTOKINES. H3K4ME3 HAS BEEN SHOWN TO BE INVOLVED IN REGULATING LINEAGE COMMITMENT. HOWEVER, THE UNDERLYING MECHANISMS, ESPECIALLY IN HUMAN NASAL EPITHELIAL CELLS (HNEPC), REMAIN UNDEREXPLORED. THE OBJECTIVE OF THIS STUDY WAS TO INVESTIGATE THE ROLE OF H3K4ME3 IN HNEPC DIFFERENTIATION TREATED WITH THE TH2 CYTOKINE IL-13. PATIENTS AND METHODS: THE EXPRESSION LEVELS OF MRNA AND PROTEINS WERE INVESTIGATED USING REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION (RT-PCR) ASSAYS AND WESTERN BLOT IN NASAL POLYP TISSUES AND HUMAN NASAL EPITHELIAL CELLS RESPECTIVELY. WE MEASURED THESE LEVELS OF H3K4ME3, MLL1 AND TARGETED GENES COMPARED WITH CONTROL SUBJECTS. RESULTS: WE DEMONSTRATE THAT EXPRESSION OF H3K4ME3 AND ITS METHYLTRANSFERASE MLL1 WAS SIGNIFICANTLY UPREGULATED IN IL-13-TREATED HNEPC. THIS ELEVATION WAS ALSO OBSERVED IN NASAL POLYPS. EXPRESSION OF CILIA-RELATED TRANSCRIPTION FACTORS FOXJ1 AND DNAI2 DECREASED, WHILE GOBLET CELL-DERIVED GENES CLCA1 AND MUC5A INCREASED UPON IL-13 TREATMENT. MECHANISTICALLY, KNOCKDOWN OF MLL1 RESTORED EXPRESSION OF THESE FOUR GENES INDUCED BY IL-13. CONCLUSION: THESE FINDINGS SUGGEST THAT H3K4ME3 IS A CRITICAL REGULATOR IN CONTROL OF NASAL EPITHELIAL CELL DIFFERENTIATION. MLL1 MAY BE A POTENTIAL THERAPEUTIC TARGET FOR NASAL INFLAMMATORY DISEASES.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
18 5972  27 TET REPRESSION AND INCREASED DNMT ACTIVITY SYNERGISTICALLY INDUCE ABERRANT DNA METHYLATION. CHRONIC INFLAMMATION IS DEEPLY INVOLVED IN VARIOUS HUMAN DISORDERS, SUCH AS CANCER, NEURODEGENERATIVE DISORDERS, AND METABOLIC DISORDERS. INDUCTION OF EPIGENETIC ALTERATIONS, ESPECIALLY ABERRANT DNA METHYLATION, IS ONE OF THE MAJOR MECHANISMS, BUT HOW IT IS INDUCED IS STILL UNCLEAR. HERE, WE FOUND THAT EXPRESSION OF TET GENES, METHYLATION ERASERS, WAS DOWNREGULATED IN INFLAMED MOUSE AND HUMAN TISSUES, AND THAT THIS WAS CAUSED BY UPREGULATION OF TET-TARGETING MIRNAS SUCH AS MIR20A, MIR26B, AND MIR29C, LIKELY DUE TO ACTIVATION OF NF-KAPPAB SIGNALING DOWNSTREAM OF IL-1BETA AND TNF-ALPHA. HOWEVER, TET KNOCKDOWN INDUCED ONLY MILD ABERRANT METHYLATION. NITRIC OXIDE (NO), PRODUCED BY NOS2, ENHANCED ENZYMATIC ACTIVITY OF DNA METHYLTRANSFERASES (DNMTS), METHYLATION WRITERS, AND NO EXPOSURE INDUCED MINIMAL ABERRANT METHYLATION. IN CONTRAST, A COMBINATION OF TET KNOCKDOWN AND NO EXPOSURE SYNERGISTICALLY INDUCED ABERRANT METHYLATION, INVOLVING GENOMIC REGIONS NOT METHYLATED BY EITHER ALONE. THE RESULTS SHOWED THAT A VICIOUS COMBINATION OF TET REPRESSION, DUE TO NF-KAPPAB ACTIVATION, AND DNMT ACTIVATION, DUE TO NO PRODUCTION, IS RESPONSIBLE FOR ABERRANT METHYLATION INDUCTION IN HUMAN TISSUES.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   
19 4584  36 N6-METHYLADENOSINE-METHYLOMIC LANDSCAPE OF LUNG TISSUES OF MICE WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE. CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD), A COMMON RESPIRATORY DISEASE, CAN BE DIVIDED INTO STABLE PHASE AND ACUTE EXACERBATION PHASE (AECOPD) AND IS CHARACTERIZED BY INFLAMMATION AND HYPER-IMMUNITY. METHYLATION OF N6-METHYLADENOSINE (M6A) IS AN EPIGENETIC MODIFICATION THAT REGULATES THE EXPRESSION AND FUNCTIONS OF GENES BY INFLUENCING POST-TRANSCRIPTIONAL RNA MODIFICATIONS. ITS INFLUENCE ON THE IMMUNE REGULATION MECHANISM HAS ATTRACTED GREAT ATTENTION. HEREIN, WE PRESENT THE M6AMETHYLOMIC LANDSCAPE AND OBSERVE HOW THE METHYLATION OF M6A PARTICIPATES IN THE PATHOLOGICAL PROCESS OF COPD. THE M6A MODIFICATION OF 430 GENES INCREASED AND THAT OF 3995 GENES DECREASED IN THE LUNG TISSUES OF MICE WITH STABLE COPD. THE LUNG TISSUES OF MICE WITH AECOPD EXHIBITED 740 GENES WITH HYPERMETHYLATED M6A PEAK AND 1373 GENES WITH LOW M6A PEAK. THESE DIFFERENTIALLY METHYLATED GENES PARTICIPATED IN SIGNALING PATHWAYS RELATED TO IMMUNE FUNCTIONS. TO FURTHER CLARIFY THE EXPRESSION LEVELS OF DIFFERENTIALLY METHYLATED GENES, RNA IMMUNOPRECIPITATION SEQUENCING (MERIP-SEQ) AND RNA-SEQUENCING DATA WERE JOINTLY ANALYZED. IN THE STABLE COPD GROUP, 119 HYPERMETHYLATED MRNAS (82 UPREGULATED AND 37 DOWNREGULATED MRNAS) AND 867 HYPOMETHYLATED MRNAS (419 UPREGULATED AND 448 DOWNREGULATED MRNAS) WERE DIFFERENTIALLY EXPRESSED. IN THE AECOPD GROUP, 87 HYPERMETHYLATED MRNAS (71 UPREGULATED AND 16 DOWNREGULATED MRNAS) AND 358 HYPOMETHYLATED MRNAS (115 UPREGULATED AND 243 DOWNREGULATED MRNAS) SHOWED DIFFERENTIAL EXPRESSION. MANY MRNAS WERE RELATED TO IMMUNE FUNCTION AND INFLAMMATION. TOGETHER, THIS STUDY PROVIDES IMPORTANT EVIDENCE ON THE ROLE OF RNA METHYLATION OF M6A IN COPD.	2023	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           
20 3764  35 INTEGRATIVE ANALYSIS OF DNA METHYLATION AND GENE EXPRESSION DATA IDENTIFIES EPAS1 AS A KEY REGULATOR OF COPD. CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) IS A COMPLEX DISEASE. GENETIC, EPIGENETIC, AND ENVIRONMENTAL FACTORS ARE KNOWN TO CONTRIBUTE TO COPD RISK AND DISEASE PROGRESSION. THEREFORE WE DEVELOPED A SYSTEMATIC APPROACH TO IDENTIFY KEY REGULATORS OF COPD THAT INTEGRATES GENOME-WIDE DNA METHYLATION, GENE EXPRESSION, AND PHENOTYPE DATA IN LUNG TISSUE FROM COPD AND CONTROL SAMPLES. OUR INTEGRATIVE ANALYSIS IDENTIFIED 126 KEY REGULATORS OF COPD. WE IDENTIFIED EPAS1 AS THE ONLY KEY REGULATOR WHOSE DOWNSTREAM GENES SIGNIFICANTLY OVERLAPPED WITH MULTIPLE GENES SETS ASSOCIATED WITH COPD DISEASE SEVERITY. EPAS1 IS DISTINCT IN COMPARISON WITH OTHER KEY REGULATORS IN TERMS OF METHYLATION PROFILE AND DOWNSTREAM TARGET GENES. GENES PREDICTED TO BE REGULATED BY EPAS1 WERE ENRICHED FOR BIOLOGICAL PROCESSES INCLUDING SIGNALING, CELL COMMUNICATIONS, AND SYSTEM DEVELOPMENT. WE CONFIRMED THAT EPAS1 PROTEIN LEVELS ARE LOWER IN HUMAN COPD LUNG TISSUE COMPARED TO NON-DISEASE CONTROLS AND THAT EPAS1 GENE EXPRESSION IS REDUCED IN MICE CHRONICALLY EXPOSED TO CIGARETTE SMOKE. AS EPAS1 DOWNSTREAM GENES WERE SIGNIFICANTLY ENRICHED FOR HYPOXIA RESPONSIVE GENES IN ENDOTHELIAL CELLS, WE TESTED EPAS1 FUNCTION IN HUMAN ENDOTHELIAL CELLS. EPAS1 KNOCKDOWN BY SIRNA IN ENDOTHELIAL CELLS IMPACTED GENES THAT SIGNIFICANTLY OVERLAPPED WITH EPAS1 DOWNSTREAM GENES IN LUNG TISSUE INCLUDING HYPOXIA RESPONSIVE GENES, AND GENES ASSOCIATED WITH EMPHYSEMA SEVERITY. OUR FIRST INTEGRATIVE ANALYSIS OF GENOME-WIDE DNA METHYLATION AND GENE EXPRESSION PROFILES ILLUSTRATES THAT NOT ONLY DOES DNA METHYLATION PLAY A 'CAUSAL' ROLE IN THE MOLECULAR PATHOPHYSIOLOGY OF COPD, BUT IT CAN BE LEVERAGED TO DIRECTLY IDENTIFY NOVEL KEY MEDIATORS OF THIS PATHOPHYSIOLOGY.	2015