1 4351 136 MIR-199A-5P SILENCING REGULATES THE UNFOLDED PROTEIN RESPONSE IN CHRONIC OBSTRUCTIVE PULMONARY DISEASE AND ALPHA1-ANTITRYPSIN DEFICIENCY. RATIONALE: RETENTION OF ABNORMAL ALPHA1-ANTITRYPSIN (AAT) ACTIVATES THE UNFOLDED PROTEIN RESPONSE IN AAT-DEFICIENT MONOCYTES. THE REGULATORY ROLE OF MICRORNAS (MIRNAS) IN UNFOLDED PROTEIN RESPONSES AND CHRONIC OBSTRUCTIVE PULMONARY DISEASE PATHOGENESIS HAS NOT BEEN INVESTIGATED. OBJECTIVES: TO INVESTIGATE MIRNA EXPRESSION AND FUNCTION IN MM AND ZZ MONOCYTES AND IDENTIFY MIRNA(S) REGULATING THE UNFOLDED PROTEIN RESPONSE. METHODS: PERIPHERAL BLOOD MONOCYTES WERE ISOLATED FROM ASYMPTOMATIC AND SYMPTOMATIC MM AND ZZ INDIVIDUALS FOR MIRNA EXPRESSION PROFILING AND PYROSEQUENCING ANALYSIS. MIRNA/GENE AND PROTEIN EXPRESSION WAS MEASURED WITH QUANTITATIVE POLYMERASE CHAIN REACTION AND WESTERN BLOTTING. OVEREXPRESSION AND INHIBITION STUDIES WERE PERFORMED WITH PRE-MIR OR ANTI-MIR, RESPECTIVELY. LUCIFERASE REPORTER GENES WERE USED TO ELUCIDATE DIRECT MIRNA-TARGET INTERACTIONS. INFLAMMATORY CYTOKINES WERE DETECTED USING THE MESO SCALE DISCOVERY PLEX ASSAYS. MEASUREMENTS AND MAIN RESULTS: FORTY-THREE MIRNAS WERE DIFFERENTIALLY EXPRESSED, WITH MIR-199A-5P MOST HIGHLY UP-REGULATED IN ASYMPTOMATIC ZZ VERSUS MM MONOCYTES. MIR-199A-2 PROMOTER HYPERMETHYLATION INHIBITS MIR-199A-5P EXPRESSION AND WAS INCREASED IN SYMPTOMATIC MM AND ZZ MONOCYTES COMPARED WITH ASYMPTOMATIC COUNTERPARTS. GRP78, ACTIVATING TRANSCRIPTION FACTOR 6, P50, AND P65 WERE INCREASED IN SYMPTOMATIC VERSUS ASYMPTOMATIC ZZ MONOCYTES. RECIPROCAL DOWN- OR UP-REGULATION OF THESE MARKERS WAS OBSERVED AFTER MIRNA MODULATION. DIRECT MIR-199A-5P TARGETING OF ACTIVATING TRANSCRIPTION FACTOR 6, P50, AND P65 BY MIR-199A-5P WAS DEMONSTRATED USING LUCIFERASE REPORTER SYSTEMS. OVEREXPRESSION OF MIR-199A-5P ALSO DECREASED OTHER ARMS OF THE UPR AND EXPRESSION OF CYTOKINES THAT ARE NOT PUTATIVE TARGETS. CONCLUSIONS: MIR-199A-5P IS A KEY REGULATOR OF THE UNFOLDED PROTEIN RESPONSE IN AAT-DEFICIENT MONOCYTES, AND EPIGENETIC SILENCING OF ITS EXPRESSION REGULATES THIS PROCESS IN CHRONIC OBSTRUCTIVE PULMONARY DISEASE.	2014	
                                                                                                                                     
2 4302  48 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	
                                             
3 2331  43 EPIGENETIC REGULATION OF INFLAMMATION BY MICRORNAS IN POST-INFECTIOUS BRONCHIOLITIS OBLITERANS. OBJECTIVES: POST-INFECTIOUS BRONCHIOLITIS OBLITERANS (PIBO) IS A RARE, CHRONIC DISEASE INITIATED BY SEVERE INFECTION AND FOLLOWED BY PERPETUATING INFLAMMATION AND OBLITERATION OF THE SMALL AIRWAYS. MICRORNAS (MIRNAS) HAVE BEEN PROPOSED TO PLAY A CENTRAL ROLE AS EPIGENETIC REGULATORS, WHICH CONTROL RESOLUTION AND PREVENT THE UNCONTROLLED PROGRESS OF INFLAMMATION. THE AIM OF THIS STUDY WAS TO DEFINE BIOMARKERS ON THE LEVEL OF POST-TRANSCRIPTIONAL GENE REGULATION IN ORDER TO CHARACTERISE PIBO. METHODS: A TOTAL OF 39 PATIENTS WITH WELL-DEFINED PIBO AND 31 CONTROLS FROM TWO CENTRES, BARCELONA, SPAIN, AND FRANKFURT, GERMANY, WERE ANALYSED BY NEXT-GENERATION SEQUENCING (NGS). THE EVALUATION OF THE BIOLOGICAL TARGETS OF THE MIRNAS WAS PERFORMED BY PATHWAY ENRICHMENT ANALYSIS AND PROTEIN-PROTEIN INTERACTION NETWORK ANALYSIS RESPECTIVELY. RESULTS: PATIENTS WITH PIBO HAD SIGNIFICANTLY LOWER LUNG FUNCTION VALUES AND INCREASED AIRWAY INFLAMMATION IN INDUCED SPUTUM AS INDICATED BY TOTAL CELL COUNTS, NEUTROPHILS, IL-1BETA, IL-6, IL-8 AND TGF-BETA COMPARED TO CONTROLS.NEXT-GENERATION SEQUENCING ANALYSIS REVEALED A TOTAL OF 22 DYSREGULATED MIRNAS, WHICH PASSED SIGNIFICANCE THRESHOLD FOR PADJ </= 0.001 WITH 17 BEING UPREGULATED AND 5 BEING DOWNREGULATED. OF THESE DYSREGULATED MIRNAS, MIR-335-5P, MIR-186-5P, MIR-30B-5P AND MIR-30C-5P WERE FURTHER VALIDATED USING QRT-PCR. INTERESTINGLY, THESE MIRNAS ARE FUNCTIONALLY IMPLICATED IN CYTOKINE-CYTOKINE RECEPTOR INTERACTION, TGF-BETA SIGNALLING AND FOXO SIGNALLING PATHWAY AND SIGNIFICANTLY CORRELATED WITH LUNG FUNCTION VALUES (FEV1). CONCLUSION: OUR RESULTS DEMONSTRATE AN ABERRANT MIRNA EXPRESSION PROFILE IN PIBO, WHICH IMPACTS PATHWAYS RESPONSIBLE FOR THE REGULATION OF INFLAMMATION AND FIBROSIS. THE DEFINED MIRNAS ARE USEFUL BIOMARKERS AND SHOULD BE ASSESSED AS POTENTIAL TARGET IN THE FIELD OF MIRNA THERAPEUTICS.	2022	
                                                                                                                                                                                                                                                                                           
4 4359  41 MIR-422A SUPPRESSES SMAD4 PROTEIN EXPRESSION AND PROMOTES RESISTANCE TO MUSCLE LOSS. BACKGROUND: LOSS OF MUSCLE MASS AND STRENGTH ARE IMPORTANT SEQUELAE OF CHRONIC DISEASE, BUT THE RESPONSE OF INDIVIDUALS IS REMARKABLY VARIABLE, SUGGESTING IMPORTANT GENETIC AND EPIGENETIC MODULATORS OF MUSCLE HOMEOSTASIS. SUCH FACTORS ARE LIKELY TO MODIFY THE ACTIVITY OF PATHWAYS THAT REGULATE WASTING, BUT TO DATE, FEW SUCH FACTORS HAVE BEEN IDENTIFIED. METHODS: THE EFFECT OF MIR-422A ON SMAD4 EXPRESSION AND TRANSFORMING GROWTH FACTOR (TGF)-BETA SIGNALLING WERE DETERMINED BY WESTERN BLOTTING AND LUCIFERASE ASSAY. MIRNA EXPRESSION WAS DETERMINED BY QPCR IN PLASMA AND MUSCLE BIOPSY SAMPLES FROM A CROSS-SECTIONAL STUDY OF PATIENTS WITH CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) AND A LONGITUDINAL STUDY OF PATIENTS UNDERGOING AORTIC SURGERY, WHO WERE SUBSEQUENTLY ADMITTED TO THE INTENSIVE CARE UNIT (ICU). RESULTS: MIR-422A WAS IDENTIFIED, BY A SCREEN, AS A MICRORNA THAT WAS PRESENT IN THE PLASMA OF PATIENTS WITH COPD AND NEGATIVELY ASSOCIATED WITH MUSCLE STRENGTH AS WELL AS BEING READILY DETECTABLE IN THE MUSCLE OF PATIENTS. IN VITRO, MIR-422A SUPPRESSED SMAD4 EXPRESSION AND INHIBITED TGF-BETA AND BONE MORPHOGENETIC PROTEIN-DEPENDENT LUCIFERASE ACTIVITY IN MUSCLE CELLS. IN MALE PATIENTS WITH COPD AND THOSE UNDERGOING AORTIC SURGERY AND ON THE ICU, A MODEL OF ICU-ASSOCIATED MUSCLE WEAKNESS, QUADRICEPS EXPRESSION OF MIR-422A WAS POSITIVELY ASSOCIATED WITH MUSCLE STRENGTH (MAXIMAL VOLUNTARY CONTRACTION R = 0.59, P < 0.001 AND R = 0.51, P = 0.004, FOR COPD AND AORTIC SURGERY, RESPECTIVELY). FURTHERMORE, PRE-SURGERY LEVELS OF MIR-422A WERE INVERSELY ASSOCIATED WITH THE AMOUNT OF MUSCLE THAT WOULD BE LOST IN THE FIRST POST-OPERATIVE WEEK (R = -0.57, P < 0.001). CONCLUSIONS: THESE DATA SUGGEST THAT DIFFERENCES IN MIR-422A EXPRESSION CONTRIBUTE TO THE SUSCEPTIBILITY TO MUSCLE WASTING ASSOCIATED WITH CHRONIC AND ACUTE DISEASE AND THAT AT LEAST PART OF THIS ACTIVITY MAY BE MEDIATED BY REDUCED TGF-BETA SIGNALLING IN SKELETAL MUSCLE.	2018	
                                                                                                                                                                                                                   
5 6794  38 [EFFECT OF BENZO(A)PYRENE ON THE EXPRESSION OF AHR-REGULATED MICRORNA IN FEMALE AND MALE RAT LUNGS]. SMOKING IS THE MAIN RISK FACTOR FOR LUNG CANCER, MAINLY DUE TO PRESENCE OF NITROSAMINES AND POLYCYCLIC AROMATIC HYDROCARBONS, INCLUDING BENZO[A]PYRENE (BP) IN TOBACCO SMOKE COMPOSITION. THE GENOTOXIC EFFECT OF BP IS BASED ON THE HIGH DNA-BINDING ABILITY OF ITS METABOLITES, WHILE THE EPIGENETIC EFFECTS ARE MEDIATED BY A CHANGE IN THE EXPRESSION OF CANCER RELATED GENES OR REGULATORY RNAS. IT HAS BEEN SHOWN THAT WOMEN HAVE A HIGHER RISK TO DEVELOP LUNG CANCER UPON SMOKING RATHER THAN MEN. WE HYPOTHESIZED THAT CROSSTALK BETWEEN SIGNALING PATHWAYS ACTIVATED BY BP AND ESTROGENS COULD UNDERLIE THE SEX-DEPENDENT DIFFERENCES IN MIRNAS EXPRESSION. TO TEST THIS HYPOTHESIS, MALE AND FEMALE RATS WERE SUBJECTED TO SHORT-TERM OR LONG-TERM BP EXPOSURE. USING IN SILICO ANALYSIS, MIRNAS CONTAINING THE ER- AND AHR-BINDING SITES IN THE PROMOTERS OF THE GENES (OR HOST GENES) WERE SELECTED. DURING CHRONIC EXPOSURE OF BP THE EXPRESSION OF MIR-22-3P, -29A-3P, -126A-3P, -193B-5P IN THE LUNGS OF MALE RATS WERE SIGNIFICANTLY INCREASED, WHILE THE LEVEL OF MIRNA-483-3P WERE DECREASED. EXPRESSION OF MIRNA-483-3P WAS UP-REGULATED DURING CHRONIC BP EXPOSURE IN THE LUNGS OF FEMALE RATS AND THE LEVELS OF OTHER STUDIED MIRNAS WERE UNCHANGED. IN TURN, CHANGES IN THE EXPRESSION OF MIRNAS WERE FOLLOWED BY CHANGES IN THE EXPRESSION OF THEIR TARGET GENES, INCLUDING PTEN, EMP2, IGF1, ITGA6, SLC34A2, AND THE OBSERVED CHANGES IN FEMALE AND MALE RAT LUNGS WERE VARIED. THUS, OUR RESULTS SUGGEST THAT SEX-DEPENDENT EPIGENETIC EFFECTS OF BP MAY BE BASED ON DIFFERENT EXPRESSION OF AHR- AND ER- REGULATED MIRNAS.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
6  164  28 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	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                
7 1297  37 DECREASED MRNA EXPRESSION LEVELS OF DNA METHYLTRANSFERASES TYPE 1 AND 3A IN SYSTEMIC LUPUS ERYTHEMATOSUS. OBJECTIVES: SYSTEMIC LUPUS ERYTHEMATOSUS (SLE) IS A CHRONIC RELAPSING AUTOIMMUNE DISEASE CHARACTERIZED BY THE PRESENCE OF AUTOANTIBODIES DIRECTED AGAINST NUCLEAR ANTIGENS AND BY CHRONIC INFLAMMATION. ALTHOUGH THE ETIOLOGY OF SLE REMAINS UNCLEAR, THE INFLUENCE OF ENVIRONMENT FACTORS, WHICH IS LARGELY REFLECTED BY THE EPIGENETIC MECHANISMS, WITH DNA METHYLATION CHANGES IN PARTICULAR, IS GENERALLY CONSIDERED AS MAIN PLAYERS IN THE PATHOGENESIS OF SLE. WE STUDIED DNA METHYLTRANSFERASES' (DNMTS) TYPE 1, 3A AND 3B TRANSCRIPT LEVELS IN PERIPHERAL BLOOD MONONUCLEAR CELLS FROM PATIENTS DIAGNOSED WITH SYSTEMIC LUPUS ERYTHEMATOSUS AND FROM THE HEALTHY CONTROL SUBJECTS. FURTHERMORE, THE ASSOCIATION OF DNMT1, DNMT3A, AND DNMT3B MRNA LEVELS WITH GENDER, AGE, AND MAJOR CLINICAL MANIFESTATIONS WAS ANALYZED. METHODS: PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS) WERE ISOLATED FROM 32 SLE PATIENTS AND 40 HEALTHY CONTROLS. REVERSE TRANSCRIPTION AND REAL-TIME QUANTITATIVE POLYMERASE CHAIN REACTION (RT-QPCR) ANALYSES WERE USED TO DETERMINE DNMT1, DNMT3A, AND DNMT3B MRNA EXPRESSION LEVELS. RESULTS: SIGNIFICANTLY LOWER DNMT1 (P = 0.015543) AND DNMT3A (P = 0.003652) TRANSCRIPT LEVELS IN SLE PATIENTS WERE OBSERVED COMPARED WITH HEALTHY CONTROLS. NEVERTHELESS, THE DNMT3B MRNA EXPRESSION LEVELS WERE MARKEDLY LOWER COMPARED WITH DNMT1 AND DNMT3A, BOTH IN PBMCS FROM AFFECTED PATIENTS AND THOSE FROM CONTROL SUBJECTS. FURTHERMORE, THE DNMT1 TRANSCRIPT LEVELS WERE POSITIVELY CORRELATED WITH SLE DISEASE ACTIVITY INDEX (SLEDAI) (R (S) = 0.4087, P = 0.020224), WHILE THE DNMT3A TRANSCRIPT LEVELS WERE NEGATIVELY CORRELATED WITH PATIENTS AGE (R (S) = -0.3765, P = 0.03369). CONCLUSIONS: OUR ANALYSES CONFIRMED THE IMPORTANCE OF EPIGENETIC ALTERATIONS IN SLE ETIOLOGY. MOREOVER, OUR RESULTS SUGGEST THAT THE PRESENCE OF SOME CLINICAL MANIFESTATIONS, SUCH AS PHOTOTOSENSITIVITY AND ARTHRITIS, MIGHT BE ASSOCIATED WITH THE DYSREGULATION OF DNA METHYLTRANSFERASES' MRNA EXPRESSION LEVELS.	2017	
                                                                                                                                                                            
8 3199  39 HDAC1 INTERACTS WITH THE P50 NF-?B SUBUNIT VIA ITS NUCLEAR LOCALIZATION SEQUENCE TO CONSTRAIN INFLAMMATORY GENE EXPRESSION. THE NF-?B P50 SUBUNIT IS AN IMPORTANT REGULATOR OF INFLAMMATION, WITH RECENT EXPERIMENTAL EVIDENCE TO SUPPORT IT ALSO HAVING A TUMOR SUPPRESSOR ROLE. CLASSICALLY, P50 FUNCTIONS IN HETERODIMERIC FORM WITH THE RELA (P65) NF-?B SUBUNIT TO ACTIVATE INFLAMMATORY GENES. HOWEVER, P50 ALSO FORMS HOMODIMERS WHICH ACTIVELY REPRESS NF-?B-DEPENDENT INFLAMMATORY GENE EXPRESSION AND EXERT AN IMPORTANT BRAKE ON THE INFLAMMATORY PROCESS. THIS REPRESSIVE ACTIVITY OF P50:P50 IS THOUGHT TO BE IN PART MEDIATED BY AN INTERACTION WITH THE EPIGENETIC REPRESSOR PROTEIN HISTONE DEACETYLASE 1 (HDAC1). HOWEVER, NEITHER THE INTERACTION OF P50 WITH HDAC1 NOR THE REQUIREMENT OF HDAC1 FOR THE REPRESSIVE ACTIVITIES OF P50 HAS BEEN WELL DEFINED. HERE WE EMPLOYED IN SILICO PREDICTION WITH IN VITRO ASSAYS TO MAP SITES OF INTERACTION OF HDAC1 ON THE P50 PROTEIN. DIRECTED MUTAGENESIS OF ONE SUCH REGION RESULTED IN ALMOST COMPLETE LOSS OF HDAC1 BINDING TO P50. TRANSFECTED MUTANT P50 PROTEIN LACKING THE PUTATIVE HDAC1 DOCKING MOTIF RESULTED IN ENHANCED CYTOKINE AND CHEMOKINE EXPRESSION WHEN COMPARED WITH CELLS EXPRESSING A TRANSFECTED WILD TYPE P50. IN ADDITION, EXPRESSION OF THIS MUTANT P50 WAS ASSOCIATED WITH ENHANCED CHEMOATTRACTION OF NEUTROPHILS AND ACETYLATION OF KNOWN INFLAMMATORY GENES DEMONSTRATING THE LIKELY IMPORTANCE OF THE P50:HDAC1 INTERACTION FOR CONTROLLING INFLAMMATION. THESE NEW INSIGHTS PROVIDE AN ADVANCE ON CURRENT KNOWLEDGE OF THE MECHANISMS BY WHICH NF-?B-DEPENDENT GENE TRANSCRIPTION ARE REGULATED AND HIGHLIGHT THE POTENTIAL FOR MANIPULATION OF P50:HDAC1 INTERACTIONS TO BRING ABOUT EXPERIMENTAL MODULATION OF CHRONIC INFLAMMATION AND PATHOLOGIES ASSOCIATED WITH DYSREGULATED NEUTROPHIL ACCUMULATION AND ACTIVATION.	2018	
                                                                                                                                                                                                                                                                                                                                                                                                                 
9 2297  25 EPIGENETIC REGULATION OF ACUTE INFLAMMATORY PAIN. ACUTE PAIN IS ASSOCIATED WITH TISSUE DAMAGE, WHICH RESULTS IN THE RELEASE OF INFLAMMATORY MEDIATORS. RECENT STUDIES POINT TO THE INVOLVEMENT OF EPIGENETIC MECHANISMS (DNA METHYLATION) IN THE DEVELOPMENT OF PAIN. WE HAVE FOUND THAT DURING ACUTE INFLAMMATORY PAIN INDUCED BY THE APPLICATION OF 10% MUSTARD OIL ON THE TONGUES OF RATS, LEVELS OF DNMT3A AND 3B WERE ELEVATED MARKEDLY (36 AND 42 % RESPECTIVELY), WHEREAS THE LEVEL OF DNMT1 WAS NOT CHANGED SIGNIFICANTLY. PREVIOUS INJECTION OF XEFOCAM WITH 0,4 MG/KG DOSE DECREASED LEVELS OF DNMT3A AND 3B (25 AND 24% RESPECTIVELY). THE LEVEL OF DNMT1 WAS NOT CHANGED SIGNIFICANTLY COMPARED TO THE CONTROL GROUP. THE FINDINGS SUPPORT THE IDEA THAT INHIBITORS OF DNA-METHYLTRANSFERASES COULD BE USEFUL FOR PAIN MANAGEMENT. OUR DATA SUGGEST THAT NSAIDS (ALONE OR IN COMBINATION WITH DNMT INHIBITORS) MAY BE PROPOSED AS POSSIBLE EPIGENETIC REGULATORY AGENTS, WHICH MAY PLAY A ROLE IN EPIGENETIC MECHANISMS INDIRECTLY THROUGH ALTERING THE ACTIVITY OF INFLAMMATORY MEDIATORS INVOLVED IN PAIN DEVELOPMENT.	2014	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     
10 1584  25 DNA METHYLATION PROFILES OF SELECTED PRO-INFLAMMATORY CYTOKINES IN ALZHEIMER DISEASE. BY MEANS OF FUNCTIONAL GENOMICS ANALYSIS, WE RECENTLY DESCRIBED THE MRNA EXPRESSION PROFILES OF VARIOUS GENES INVOLVED IN THE NEUROINFLAMMATORY RESPONSE IN THE BRAINS OF SUBJECTS WITH LATE-ONSET ALZHEIMER DISEASE (LOAD). SOME OF THESE GENES, NAMELY INTERLEUKIN (IL)-1BETA AND IL-6, SHOWED DISTINCT EXPRESSION PROFILES WITH PEAK EXPRESSION DURING THE FIRST STAGES OF THE DISEASE AND CONTROL-LIKE LEVELS AT LATER STAGES. IL-1BETA AND IL-6 GENES ARE MODULATED BY DNA METHYLATION IN DIFFERENT CHRONIC AND DEGENERATIVE DISEASES; IT IS ALSO WELL KNOWN THAT LOAD MAY HAVE AN EPIGENETIC BASIS. INDEED, WE AND OTHERS HAVE PREVIOUSLY REPORTED GENE-SPECIFIC DNA METHYLATION ALTERATIONS IN LOAD AND IN RELATED ANIMAL MODELS. BASED ON THESE DATA, WE STUDIED THE DNA METHYLATION PROFILES, AT SINGLE CYTOSINE RESOLUTION, OF IL-1BETA AND IL-6 5'-FLANKING REGION BY BISULPHITE MODIFICATION IN THE CORTEX OF HEALTHY CONTROLS AND LOAD PATIENTS AT 2 DIFFERENT DISEASE STAGES: BRAAK I-II/A AND BRAAK V-VI/C. OUR ANALYSIS PROVIDES EVIDENCE THAT NEUROINFLAMMATION IN LOAD IS ASSOCIATED WITH (AND POSSIBLY MEDIATED BY) EPIGENETIC MODIFICATIONS.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
11 1117  32 COMPARATIVE AND EXPERIMENTAL STUDIES ON THE GENES ALTERED BY CHRONIC HYPOXIA IN HUMAN BRAIN MICROENDOTHELIAL CELLS. BACKGROUND : HYPOXIA INDUCIBLE FACTOR 1 ALPHA (HIF1A) IS A MASTER REGULATOR OF ACUTE HYPOXIA; HOWEVER, WITH CHRONIC HYPOXIA, HIF1A LEVELS RETURN TO THE NORMOXIC LEVELS. IMPORTANTLY, THE GENES THAT ARE INVOLVED IN THE CELL SURVIVAL AND VIABILITY UNDER CHRONIC HYPOXIA ARE NOT KNOWN. THEREFORE, WE TESTED THE HYPOTHESIS THAT CHRONIC HYPOXIA LEADS TO THE UPREGULATION OF A CORE GROUP OF GENES WITH ASSOCIATED CHANGES IN THE PROMOTER DNA METHYLATION THAT MEDIATES THE CELL SURVIVAL UNDER HYPOXIA. RESULTS : WE EXAMINED THE EFFECT OF CHRONIC HYPOXIA (3 DAYS; 0.5% OXYGEN) ON HUMAN BRAIN MICRO ENDOTHELIAL CELLS (HBMEC) VIABILITY AND APOPTOSIS. HYPOXIA CAUSED A SIGNIFICANT REDUCTION IN CELL VIABILITY AND AN INCREASE IN APOPTOSIS. NEXT, WE EXAMINED CHRONIC HYPOXIA ASSOCIATED CHANGES IN TRANSCRIPTOME AND GENOME-WIDE PROMOTER METHYLATION. THE DATA OBTAINED WAS COMPARED WITH 16 OTHER MICROARRAY STUDIES ON CHRONIC HYPOXIA. NINE GENES WERE ALTERED IN RESPONSE TO CHRONIC HYPOXIA IN ALL 17 STUDIES. INTERESTINGLY, HIF1A WAS NOT ALTERED WITH CHRONIC HYPOXIA IN ANY OF THE STUDIES. FURTHERMORE, WE COMPARED OUR DATA TO THREE OTHER STUDIES THAT IDENTIFIED HIF-RESPONSIVE GENES BY VARIOUS APPROACHES. ONLY TWO GENES WERE FOUND TO BE HIF DEPENDENT. WE SILENCED EACH OF THESE 9 GENES USING CRISPR/CAS9 SYSTEM. DOWNREGULATION OF EGLN3 SIGNIFICANTLY INCREASED THE CELL DEATH UNDER CHRONIC HYPOXIA, WHEREAS DOWNREGULATION OF ERO1L, ENO2, ADRENOMEDULLIN, AND SPAG4 REDUCED THE CELL DEATH UNDER HYPOXIA. CONCLUSIONS : WE PROVIDE A CORE GROUP OF GENES THAT REGULATES CELLULAR ACCLIMATIZATION UNDER CHRONIC HYPOXIC STRESS, AND MOST OF THEM ARE HIF INDEPENDENT.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            
12 5114  34 PORPHYROMONAS GINGIVALIS LIPOPOLYSACCHARIDE STIMULATION IN HUMAN PERIODONTAL LIGAMENT STEM CELLS: ROLE OF EPIGENETIC MODIFICATIONS TO THE INFLAMMATION. PERIODONTITIS IS A CHRONIC ORAL INFLAMMATORY DISEASE PRODUCED BY BACTERIA. GINGIVAL RETRACTION AND BONE AND CONNECTIVE TISSUES RESORPTION ARE THE HALLMARKS OF THIS DISEASE. CHRONIC PERIODONTITIS MAY CONTRIBUTE TO THE RISK OF ONSET OR PROGRESSION OF NEUROINFLAMMATORY PATHOLOGICAL CONDITIONS, SUCH AS ALZHEIMER'S DISEASE. THE MAIN GOAL OF THE PRESENT STUDY WAS TO INVESTIGATE IF THE ROLE OF EPIGENETIC MODULATIONS IS INVOLVED IN PERIODONTITIS USING HUMAN PERIODONTAL LIGAMENT STEM CELLS (HPDLSCS) AS AN IN VITRO MODEL SYSTEM. HPDLSCS WERE TREATED WITH LIPOPOLYSACCHARIDE OF PORPHYROMONAS GINGIVALIS AND THE EXPRESSION OF PROTEINS ASSOCIATED WITH DNA METHYLATION AND HISTONE ACETYLATION, SUCH AS DNMT1 AND P300, RESPECTIVELY, AND INFLAMMATORY TRANSCRIPTION FACTOR NF-KB, WERE EXAMINED. IMMUNOFLUORESCENCE, WESTERN BLOT AND NEXT GENERATION SEQUENCING RESULTS DEMONSTRATED THAT P. GINGIVALIS LIPOPOLYSACCHARIDE SIGNIFICANTLY REDUCED DNA METHYLASE DNMT1, WHILE IT MARKEDLY UPREGULATED THE LEVEL OF HISTONE ACETYLTRANSFERASE P300 AND NF-KB IN HPDLSCS. OUR RESULTS SHOWED THAT P. GINGIVALIS LIPOPOLYSACCHARIDE MARKEDLY REGULATE THE GENES INVOLVED IN EPIGENETIC MECHANISM, WHICH MAY RESULT IN INFLAMMATION INDUCTION. WE PROPOSE THAT P. GINGIVALIS LIPOPOLYSACCHARIDE-TREATED HPDLSCS COULD BE A POTENTIAL IN VITRO MODEL SYSTEM TO STUDY EPIGENETICS MODULATIONS ASSOCIATED WITH PERIODONTITIS, WHICH MIGHT BE HELPFUL TO IDENTIFY NOVEL BIOMARKERS LINKED TO THIS ORAL INFLAMMATORY DISEASE.	2017	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              
13 1144  40 CONCOMITANT HETEROCHROMATINISATION AND DOWN-REGULATION OF GENE EXPRESSION UNVEILS EPIGENETIC SILENCING OF RELB IN AN AGGRESSIVE SUBSET OF CHRONIC LYMPHOCYTIC LEUKEMIA IN MALES. BACKGROUND: THE SENSITIVITY OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) CELLS TO CURRENT TREATMENTS, BOTH IN VITRO AND IN VIVO, RELIES ON THEIR ABILITY TO ACTIVATE APOPTOTIC DEATH. CLL CELLS RESISTANT TO DNA DAMAGE-INDUCED APOPTOSIS DISPLAY DEREGULATION OF A SPECIFIC SET OF GENES. METHODS: MICROARRAY HYBRIDIZATION (HUMAN GENECHIP, AFFYMETRIX), IMMUNOFLUORESCENT IN SITU LABELING COUPLED WITH VIDEO-MICROSCOPY RECORDING/ANALYSES, CHROMATIN-IMMUNOPRECIPITATION (CHIP), POLYMERASE CHAIN REACTIONS (PCR), REAL-TIME QUANTITATIVE PCR (RT-QPCR) AND BISULFITE GENOME SEQUENCING WERE THE MAIN METHODS APPLIED. STATISTICAL ANALYSES WERE PERFORMED BY APPLYING GCRMA AND SAM ANALYSIS (MICROARRAY DATA) AND STUDENT'S T-TEST OR MANN & WHITNEY'S U-TEST. RESULTS: HEREIN WE SHOW THAT, REMARKABLY, IN A RESISTANT MALE CLL CELLS THE VAST MAJORITY OF GENES WERE DOWN-REGULATED COMPARED WITH SENSITIVE CELLS, WHEREAS THIS WAS NOT THE CASE IN CELLS DERIVED FROM FEMALES. THIS GENE DOWN-REGULATION WAS FOUND TO BE ASSOCIATED WITH AN OVERALL GAIN OF HETEROCHROMATIN AS EVIDENCED BY IMMUNOFLUORESCENT LABELING OF HETEROCHROMATIN PROTEIN 1ALPHA (HP-1), TRIMETHYLATED HISTONE 3 LYSINE 9 (3METH3K9), AND 5-METHYLCYTIDINE (5METC). NOTABLY, 17 GENES WERE FOUND TO BE COMMONLY DEREGULATED IN RESISTANT MALE AND FEMALE CELL SAMPLES. AMONG THESE, RELB WAS IDENTIFIED AS A DISCRIMINATORY CANDIDATE GENE REPRESSED IN THE MALE AND UPREGULATED IN THE FEMALE RESISTANT CELLS. CONCLUSION: THE MOLECULAR DEFECTS IN THE SILENCING OF RELB INVOLVE AN INCREASE IN H3K9- BUT NOT CPG-ISLAND METHYLATION IN THE PROMOTER REGIONS. INCREASE IN ACETYL-H3 IN RESISTANT FEMALE BUT NOT MALE CLL SAMPLES AS WELL AS A DECREASE OF TOTAL CELLULAR LEVEL OF RELB AFTER AN INHIBITION OF HISTONE DEACETYLASE (HDAC) BY TRICHOSTATIN A (TSA), FURTHER EMPHASIZE THE ROLE OF EPIGENETIC MODIFICATIONS WHICH COULD DISCRIMINATE TWO CLL SUBSETS. TOGETHER, THESE RESULTS HIGHLIGHTED THE EPIGENETIC RELB SILENCING AS A NEW MARKER OF THE PROGRESSIVE DISEASE IN MALES.	2010	
                                                                                 
14 6684  30 VALIDATION OF AN LC-MS BASED APPROACH FOR PROFILING HISTONES IN CHRONIC LYMPHOCYTIC LEUKEMIA. THE IN VITRO EVALUATION OF HISTONES AND THEIR PTMS HAS DRAWN SUBSTANTIAL INTEREST IN THE DEVELOPMENT OF EPIGENETIC THERAPIES. THE DIFFERENTIAL EXPRESSION OF HISTONE ISOFORMS MAY SERVE AS A POTENTIAL MARKER IN THE CLASSIFICATION OF DISEASES AFFECTED BY CHROMATIN ABNORMALITIES. IN THIS STUDY, PROTEIN PROFILING BY LC AND MS WAS USED TO EXPLORE DIFFERENCES IN HISTONE COMPOSITION IN PRIMARY CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) CELLS. EXTENSIVE METHOD VALIDATIONS WERE PERFORMED TO DETERMINE THE EXPERIMENTAL VARIANCES THAT WOULD IMPACT HISTONE RELATIVE ABUNDANCE. THE RESULTING DATA DEMONSTRATED THAT THE PROPOSED METHODOLOGY WAS SUITABLE FOR THE ANALYSIS OF HISTONE PROFILES. IN 4 NORMAL INDIVIDUALS AND 40 CLL PATIENTS, A SIGNIFICANT DECREASE IN THE RELATIVE ABUNDANCE OF HISTONE H2A VARIANTS (H2AFL AND H2AFA/M*) WAS OBSERVED IN PRIMARY CLL CELLS AS COMPARED TO NORMAL B CELLS. PROTEIN IDENTITIES WERE DETERMINED USING HIGH MASS ACCURACY MS AND SHOTGUN PROTEOMICS.	2009	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
15 5269  34 PROMOTER DNA METHYLATION CONTRIBUTES TO HUMAN BETA-DEFENSIN-1 DEFICIENCY IN ATOPIC DERMATITIS. ATOPIC DERMATITIS (AD) IS A CHRONIC INFLAMMATORY SKIN DISEASE CAUSED BY EPIDERMAL BARRIER DYSFUNCTION AND DYSREGULATION OF INNATE AND ADAPTIVE IMMUNITY. EPIGENETIC REGULATION OF HUMAN BETA-DEFENSIN-1 (HBD-1) MIGHT BE ASSOCIATED WITH A VARIETY OF DEFECTS IN THE INNATE IMMUNE SYSTEM DURING AD PATHOGENESIS. WE INVESTIGATED THE POSSIBLE MECHANISM OF DECREASED HBD-1 GENE EXPRESSION IN AD AND DEMONSTRATED THE RESTORATION OF HBD-1 TRANSCRIPTION IN UNDIFFERENTIATED NORMAL HUMAN EPIDERMAL KERATINOCYTE CELLS AFTER TREATMENT WITH A DNA METHYLTRANSFERASE INHIBITOR. WE ALSO CONDUCTED AN IN VITRO METHYLATED REPORTER ASSAY USING A REPORTER CONTAINING 14 CPG SITES. METHYLATION OF THE 14 CPG SITES WITHIN THE HBD-1 5' REGION RESULTED IN AN APPROXIMATELY 86% REDUCTION IN PROMOTER ACTIVITY AND AFFECTED HBD-1 TRANSCRIPTIONAL REGULATION. WE THEN COMPARED METHYLATION FREQUENCIES AT CPG 3 AND CPG 4 BETWEEN NON-LESIONAL AND LESIONAL EPIDERMIS SAMPLES OF PATIENTS WITH SEVERE AD AND BETWEEN THESE PAIRED TISSUES AND HEALTHY CONTROL EPIDERMIS FROM NORMAL VOLUNTEERS WITHOUT AD HISTORY. BISULFITE PYROSEQUENCING DATA SHOWED SIGNIFICANTLY HIGHER METHYLATION FREQUENCIES AT THE CPG 3 AND 4 SITES IN AD LESIONAL SAMPLES THAN IN NON-LESIONAL AD SKIN AND NORMAL SKIN SAMPLES (P < 0.05). THESE RESULTS SUGGEST THAT THE DNA METHYLATION SIGNATURE OF HBD-1 IS A NOVEL DIAGNOSTIC/PROGNOSTIC MARKER AND A PROMISING THERAPEUTIC TARGET FOR THE COMPROMISED STRATUM CORNEUM BARRIER ATTRIBUTED TO HBD-1 DEFICIENCY.	2018	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             
16 1739  37 EARLY DNA METHYLATION CHANGES IN CHILDREN DEVELOPING BETA CELL AUTOIMMUNITY AT A YOUNG AGE. AIMS/HYPOTHESIS: TYPE 1 DIABETES IS A CHRONIC AUTOIMMUNE DISEASE OF COMPLEX AETIOLOGY, INCLUDING A POTENTIAL ROLE FOR EPIGENETIC REGULATION. PREVIOUS EPIGENOMIC STUDIES FOCUSED MAINLY ON CLINICALLY DIAGNOSED INDIVIDUALS. THE AIM OF THE STUDY WAS TO ASSESS EARLY DNA METHYLATION CHANGES ASSOCIATED WITH TYPE 1 DIABETES ALREADY BEFORE THE DIAGNOSIS OR EVEN BEFORE THE APPEARANCE OF AUTOANTIBODIES. METHODS: REDUCED REPRESENTATION BISULPHITE SEQUENCING (RRBS) WAS APPLIED TO STUDY DNA METHYLATION IN PURIFIED CD4(+) T CELL, CD8(+) T CELL AND CD4(-)CD8(-) CELL FRACTIONS OF 226 PERIPHERAL BLOOD MONONUCLEAR CELL SAMPLES LONGITUDINALLY COLLECTED FROM SEVEN TYPE 1 DIABETES-SPECIFIC AUTOANTIBODY-POSITIVE INDIVIDUALS AND CONTROL INDIVIDUALS MATCHED FOR AGE, SEX, HLA RISK AND PLACE OF BIRTH. WE ALSO EXPLORED CORRELATIONS BETWEEN DNA METHYLATION AND GENE EXPRESSION USING RNA SEQUENCING DATA FROM THE SAME SAMPLES. TECHNICAL VALIDATION OF RRBS RESULTS WAS PERFORMED USING PYROSEQUENCING. RESULTS: WE IDENTIFIED 79, 56 AND 45 DIFFERENTIALLY METHYLATED REGIONS IN CD4(+) T CELLS, CD8(+) T CELLS AND CD4(-)CD8(-) CELL FRACTIONS, RESPECTIVELY, BETWEEN TYPE 1 DIABETES-SPECIFIC AUTOANTIBODY-POSITIVE INDIVIDUALS AND CONTROL PARTICIPANTS. THE ANALYSIS OF PRE-SEROCONVERSION SAMPLES IDENTIFIED DNA METHYLATION SIGNATURES AT THE VERY EARLY STAGE OF DISEASE, INCLUDING DIFFERENTIAL METHYLATION AT THE PROMOTER OF IRF5 IN CD4(+) T CELLS. FURTHER, WE VALIDATED RRBS RESULTS USING PYROSEQUENCING AT THE FOLLOWING CPG SITES: CHR19:18118304 IN THE PROMOTER OF ARRDC2; CHR21:47307815 IN THE INTRON OF PCBP3; AND CHR14:81128398 IN THE INTERGENIC REGION NEAR TRAF3 IN CD4(+) T CELLS. CONCLUSIONS/INTERPRETATION: THESE PRELIMINARY RESULTS PROVIDE NOVEL INSIGHTS INTO CELL TYPE-SPECIFIC DIFFERENTIAL EPIGENETIC REGULATION OF GENES, WHICH MAY CONTRIBUTE TO TYPE 1 DIABETES PATHOGENESIS AT THE VERY EARLY STAGE OF DISEASE DEVELOPMENT. SHOULD THESE FINDINGS BE VALIDATED, THEY MAY SERVE AS A POTENTIAL SIGNATURE USEFUL FOR DISEASE PREDICTION AND MANAGEMENT.	2022	
                                                                                                                               
17 3468  41 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	

18 2395  33 EPIGENETIC REPROGRAMMING IN MIST1(-/-) MICE PREDICTS THE MOLECULAR RESPONSE TO CERULEIN-INDUCED PANCREATITIS. GENE EXPRESSION IS AFFECTED BY MODIFICATIONS TO HISTONE CORE PROTEINS WITHIN CHROMATIN. CHANGES IN THESE MODIFICATIONS, OR EPIGENETIC REPROGRAMMING, CAN DICTATE CELL FATE AND PROMOTE SUSCEPTIBILITY TO DISEASE. THE GOAL OF THIS STUDY WAS TO DETERMINE THE EXTENT OF EPIGENETIC REPROGRAMMING IN RESPONSE TO CHRONIC STRESS THAT OCCURS FOLLOWING ABLATION OF MIST1 (MIST1(-/-) ), WHICH IS REPRESSED IN PANCREATIC DISEASE. CHROMATIN IMMUNOPRECIPITATION FOR TRIMETHYLATION OF LYSINE RESIDUE 4 ON HISTONE 3 (H3K4ME3) IN PURIFIED ACINAR CELLS FROM WILD TYPE AND MIST1(-/-) MICE WAS FOLLOWED BY NEXT GENERATION SEQUENCING (CHIP-SEQ) OR CHIP-QPCR. H3K4ME3-ENRICHED GENES WERE ASSESSED FOR EXPRESSION BY QRT-PCR IN PANCREATIC TISSUE BEFORE AND AFTER INDUCTION OF CERULEIN-INDUCED PANCREATITIS. WHILE MOST OF H3K4ME3-ENRICHMENT IS RESTRICTED TO TRANSCRIPTIONAL START SITES, >25% OF ENRICHMENT SITES ARE FOUND WITHIN, DOWNSTREAM OR BETWEEN ANNOTATED GENES. LESS THAN 10% OF THESE SITES WERE ALTERED IN MIST1(-/-) ACINI, WITH MOST CHANGES IN H3K4ME3 ENRICHMENT NOT REFLECTING ALTERED GENE EXPRESSION. INGENUITY PATHWAY ANALYSIS OF GENES DIFFERENTIALLY-ENRICHED FOR H3K4ME3 REVEALED AN ASSOCIATION WITH PANCREATITIS AND PANCREATIC DUCTAL ADENOCARCINOMA IN MIST1(-/-) TISSUE. MOST OF THESE GENES WERE NOT DIFFERENTIALLY EXPRESSED BUT SEVERAL WERE READILY INDUCED BY ACUTE EXPERIMENTAL PANCREATITIS, WITH SIGNIFICANTLY INCREASED EXPRESSION IN MIST1(-/-) TISSUE RELATIVE TO WILD TYPE MICE. WE SUGGEST THAT THE CHRONIC CELL STRESS OBSERVED IN THE ABSENCE OF MIST1 RESULTS IN EPIGENETIC REPROGRAMMING OF GENES INVOLVED IN PROMOTING PANCREATITIS TO A POISED STATE, THEREBY INCREASING THE SENSITIVITY TO EVENTS THAT PROMOTE DISEASE.	2014	
                                                                                                                                                                                                                                                                                                                                                                                                                                              
19 6404  33 THE RS1001179 SNP AND CPG METHYLATION REGULATE CATALASE EXPRESSION IN CHRONIC LYMPHOCYTIC LEUKEMIA. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS AN INCURABLE DISEASE CHARACTERIZED BY AN EXTREMELY VARIABLE CLINICAL COURSE. WE HAVE RECENTLY SHOWN THAT HIGH CATALASE (CAT) EXPRESSION IDENTIFIES PATIENTS WITH AN AGGRESSIVE CLINICAL COURSE. ELUCIDATING MECHANISMS REGULATING CAT EXPRESSION IN CLL IS PREEMINENT TO UNDERSTAND DISEASE MECHANISMS AND DEVELOP STRATEGIES FOR IMPROVING ITS CLINICAL MANAGEMENT. IN THIS STUDY, WE INVESTIGATED THE ROLE OF THE CAT PROMOTER RS1001179 SINGLE NUCLEOTIDE POLYMORPHISM (SNP) AND OF THE CPG ISLAND II METHYLATION ENCOMPASSING THIS SNP IN THE REGULATION OF CAT EXPRESSION IN CLL. LEUKEMIC CELLS HARBORING THE RS1001179 SNP T ALLELE EXHIBITED A SIGNIFICANTLY HIGHER CAT EXPRESSION COMPARED WITH CELLS BEARING THE CC GENOTYPE. CAT PROMOTER HARBORING THE T -BUT NOT C- ALLELE WAS ACCESSIBLE TO ETS-1 AND GR-BETA TRANSCRIPTION FACTORS. MOREOVER, CLL CELLS EXHIBITED LOWER METHYLATION LEVELS THAN NORMAL B CELLS, IN LINE WITH THE HIGHER CAT MRNA AND PROTEIN EXPRESSED BY CLL IN COMPARISON WITH NORMAL B CELLS. METHYLATION LEVELS AT SPECIFIC CPG SITES NEGATIVELY CORRELATED WITH CAT LEVELS IN CLL CELLS. INHIBITION OF METHYLTRANSFERASE ACTIVITY INDUCED A SIGNIFICANT INCREASE IN CAT LEVELS, THUS FUNCTIONALLY VALIDATING THE ROLE OF CPG METHYLATION IN REGULATING CAT EXPRESSION IN CLL. FINALLY, THE CT/TT GENOTYPES WERE ASSOCIATED WITH LOWER METHYLATION AND HIGHER CAT LEVELS, SUGGESTING THAT THE RS1001179 T ALLELE AND CPG METHYLATION MAY INTERACT IN REGULATING CAT EXPRESSION IN CLL. THIS STUDY IDENTIFIES GENETIC AND EPIGENETIC MECHANISMS UNDERLYING DIFFERENTIAL EXPRESSION OF CAT, WHICH COULD BE OF CRUCIAL RELEVANCE FOR THE DEVELOPMENT OF THERAPIES TARGETING REDOX REGULATORY PATHWAYS IN CLL.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                 
20 5972  23 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