1 1854 143 ELEVATED SEMINAL PLASMA ESTRADIOL AND EPIGENETIC INACTIVATION OF ESR1 AND ESR2 IS ASSOCIATED WITH CP/CPPS. CHRONIC PROSTATITIS/CHRONIC PELVIC PAIN SYNDROME (CP/CPPS) IS ASSOCIATED WITH URINARY TRACT SYMPTOMS AND HORMONAL IMBALANCES AMONGST OTHERS. THE HETEROGENEOUS CLINICAL PRESENTATION, UNEXPLORED MOLECULAR BACKGROUND AND LACK OF PROSTATE BIOPSIES COMPLICATE THERAPY. HERE, USING LIQUID BIOPSIES, WE PERFORMED A COMPREHENSIVE TRANSLATIONAL STUDY ON MEN DIAGNOSED WITH CP/CPPS TYPE III (N= 50; MEDIAN AGE 39.8, RANGE 23-65) AND AGE-MATCHED CONTROLS (N= 61; MEDIAN AGE 36.8, RANGE 20-69), CONSIDERING BIOCHEMICAL PARAMETERS OF BLOOD AND EJACULATES, AND EPIGENETIC REGULATION OF THE ESTROGEN RECEPTOR GENES (ESR1 AND ESR2) IN LEUKOCYTES ISOLATED FROM BLOOD (SYSTEMIC REGULATION) AND IN SOMATIC CELLS ISOLATED FROM EJACULATES (LOCAL REGULATION). WE FOUND ELEVATED 17BETA-ESTRADIOL (E(2)) LEVELS IN SEMINAL PLASMA, BUT NOT IN BLOOD PLASMA, THAT WAS SIGNIFICANTLY ASSOCIATED WITH CP/CPPS AND IMPAIRED URINARY TRACT SYMPTOMS. IN EJACULATED SOMATIC CELLS OF CP/CPPS PATIENTS WE FOUND THAT ESR1 AND ESR2 WERE BOTH SIGNIFICANTLY HIGHER METHYLATED IN CPG-PROMOTERS AND EXPRESSIONALLY DOWN-REGULATED IN COMPARISON TO CONTROLS. MAST CELLS ARE REPORTED TO CONTRIBUTE TO CP/CPPS AND ARE ESTROGEN RESPONSIVE. CONSISTENT WITH THIS, WE FOUND THAT E(2) -TREATMENT OF HUMAN MAST CELL LINES (HMC-1 AND LAD2) RESULTED IN ALTERED CYTOKINE AND CHEMOKINE EXPRESSION. INTERESTINGLY, IN HMC-1 CELLS, POSSESSING EPIGENETICALLY INACTIVATED ESR1 AND ESR2, E(2) -TREATMENT LED TO A REDUCED TRANSCRIPTION OF A NUMBER OF INFLAMMATORY GENES. OVERALL, THESE DATA SUGGEST THAT ELEVATED LOCAL E(2) LEVELS ASSOCIATE WITH AN EPIGENETIC DOWN-REGULATION OF THE ESTROGEN RECEPTORS AND HAVE A PROMINENT ROLE IN CP/CPPS. INVESTIGATING E(2) LEVELS IN SEMEN COULD THEREFORE SERVE AS A PROMISING BIOMARKER TO SELECT PATIENTS FOR ESTROGEN TARGETED THERAPY. 2018 2 983 49 CHRONIC PROSTATITIS AFFECTS MALE REPRODUCTIVE HEALTH AND IS ASSOCIATED WITH SYSTEMIC AND LOCAL EPIGENETIC INACTIVATION OF C-X-C MOTIF CHEMOKINE 12 RECEPTOR C-X-C CHEMOKINE RECEPTOR TYPE 4. BACKGROUND/AIMS/OBJECTIVES: CHRONIC PROSTATITIS/CHRONIC PELVIC PAIN SYNDROME (CP/CPPS) HAS DETRIMENTAL EFFECTS ON THE QUALITY OF LIFE INCLUDING THE ASPECT OF SEXUAL DYSFUNCTION. THE AIM OF THE STUDY WAS TO IDENTIFY IF THERE WAS AN ADVERSE EFFECT ON THE MALE GENITAL COMPARTMENT AND IF THERE ARE SYSTEMIC OR COMPARTMENT-SPECIFIC LOCAL SIGNALS FOR EPIGENETIC DYSREGULATION OF INFLAMMATORY FACTORS IN CP/CPPS PATIENTS. METHODS: ONE HUNDRED FIVE NIH IIIB CP/CPPS PATIENTS AND 41 HEALTHY MEN WERE RECRUITED AND UNDERWENT INVESTIGATIONS OF URINES, SEMEN AND BLOOD. PROMOTER METHYLATION AND EXPRESSION OF THE CHEMOKINE C-X-C MOTIF CHEMOKINE 12 AND ITS RECEPTOR C-X-C CHEMOKINE RECEPTOR TYPE 4 (CXCR4) (INVOLVED IN THE RECRUITMENT OF MAST CELLS) WERE ANALYZED IN PROSTATE EPITHELIAL CELL LINES AND IN HEALTHY VOLUNTEERS' AND PATIENTS' BLOOD, EJACULATE CELL PELLETS, AND SEPARATED EJACULATE FRACTIONS (SPERM AND SEMINAL SOMATIC CELLS). RESULTS: INDEPENDENTLY FROM AGE, CP/CPPS NIH IIIB WAS ASSOCIATED WITH SIGNIFICANT IMPAIRMENT OF SPERM MOTILITY, MORPHOLOGY AND SEMEN PH (P < 0.001). PATIENTS OLDER THAN 33 YEARS SHOWED SIGNIFICANTLY INCREASED SEMINAL INTERLEUKIN-8 AND SERUM PROSTATE SPECIFIC ANTIGEN VALUES. IN PATIENTS, THE CXCR4 MRNA-EXPRESSION WAS SIGNIFICANTLY DECREASED IN WHOLE BLOOD AND EJACULATE CELL PELLETS DUE TO PROMOTER HYPERMETHYLATION. ANALYSES ON SEPARATED FRACTIONS OF SPERM AND SEMINAL SOMATIC CELLS REVEALED THAT SPERM DNA WAS UNAFFECTED, WHEREAS SOMATIC CELL DNA WAS DIFFERENTIALLY METHYLATED. CONCLUSIONS: NIH IIIB CP/CPPS HAS NEGATIVE EFFECTS ON SURROGATE PARAMETERS OF MALE FERTILITY AND IS ASSOCIATED SIGNIFICANTLY WITH SYSTEMIC AND LOCAL EPIGENETIC INACTIVATION OF CXCR4. 2017 3 2908 33 GENE EXPRESSION PROFILE OF NORMAL BREAST TISSUE AND BODY MASS INDEX. BACKGROUND: IN HUMAN BREAST, ADIPOSE TISSUE REPRESENTS UP TO 80% OF THE TOTAL VOLUME AND PLAYS A CRITICAL ROLE IN MAMMARY GLAND REMODELING. GIVEN THE EMERGING ROLE OF OBESITY IN BREAST CANCER GROWTH AND DEVELOPMENT, WE EXPLORED THE RELATIONSHIP BETWEEN BODY MASS INDEX (BMI), AS A PROXY OF WOMAN'S OBESITY STATUS, AND THE EXPRESSION IN NORMAL BREAST TISSUE FROM HEALTHY WOMEN OF A SELECTED PANEL OF GENES, KNOWN TO BE INVOLVED IN MAMMARY GLAND HOMEOSTASIS. METHODS: TWO INDEPENDENT PUBLICLY AVAILABLE DATASETS, COMPOSED OF 180 SPECIMENS OF NORMAL BREAST TISSUE FROM REDUCTION MAMMOPLASTY WERE INTERROGATED. DIFFERENTIAL GENE EXPRESSION AMONG BMI CLASSES WAS EVALUATED BY ANOVA, AND PARTIAL CORRELATION COEFFICIENT WAS USED TO ASSAY THE CORRELATION BETWEEN GENES CONTROLLING FOR BMI. RESULTS: DESPITE THE DIFFERENCES IN MICROARRAY PLATFORMS AND ANALYTICAL PROCEDURES, THE TWO DATASETS SHARED A CORE OF 9 GENES DIFFERENTIALLY EXPRESSED IN BMI CLASSES AND SIGNIFICANTLY CORRELATED WITH BMI. FOUR (44%) OF THESE GENES BELONG TO THE FUNCTIONAL CLASS OF CYTOKINES AND CYTOKINE RECEPTORS (IL1R1, IL2RA, IL12A, AND IL12RB2). THE OTHERS BELONG TO THE FUNCTIONAL CLASS OF THE EPIGENETIC REGULATION (MEDAG AND SETD7), SIGNAL TRANSDUCTION (STAT1), CELL ADHESION (ITGAV), AND ENZYMATIC ACTIVITY (STS). CONCLUSIONS: ALTHOUGH EXPLORATORY, PRESENT FINDINGS ARE IN AGREEMENT WITH THE ROLE OF INFLAMMATION MODULATORS IN THE HOMEOSTASIS OF NORMAL BREAST TISSUE AND THE BELIEVE THAT AN INCREASE IN BODY ADIPOSE TISSUE MAY HAVE A POTENTIALLY DANGEROUS LOCAL EFFECT, THROUGH THE INCREASED EXPRESSION OF INFLAMMATION-RELATED GENES AND THE ESTABLISHMENT OF A LOW-GRADE CHRONIC INFLAMMATION. 2021 4 4523 61 MULTIDISCIPLINARY APPROACH TO PROSTATITIS. THE MODERN CLINICAL RESEARCH ON PROSTATITIS STARTED WITH THE WORK OF STAMEY AND COWORKERS WHO DEVELOPED THE BASIC PRINCIPLES WE ARE STILL USING. THEY ESTABLISHED THE SEGMENTED CULTURE TECHNIQUE FOR LOCALIZING THE INFECTIONS IN THE MALES TO THE URETHRA, THE BLADDER, OR THE PROSTATE AND TO DIFFERENTIATE THE MAIN CATEGORIES OF PROSTATITIS. SUCH CATEGORIES WITH SLIGHT MODIFICATIONS ARE STILL USED ACCORDING TO THE NIH CLASSIFICATION: ACUTE BACTERIAL PROSTATITIS, CHRONIC BACTERIAL PROSTATITIS, CHRONIC PELVIC PAIN SYNDROME (CPPS) AND ASYMPTOMATIC PROSTATITIS. PROSTATIC INFLAMMATION IS CONSIDERED AN IMPORTANT FACTOR IN INFLUENCING BOTH PROSTATIC GROWTH AND PROGRESSION OF SYMPTOMS OF BENIGN PROSTATIC HYPERPLASIA AND PROSTATITIS. CHRONIC INFLAMMATION/NEUROINFLAMMATION IS A RESULT OF A DEREGULATED ACUTE PHASE RESPONSE OF THE INNATE IMMUNE SYSTEM AFFECTING SURROUNDING NEURAL TISSUE AT MOLECULAR, STRUCTURAL AND FUNCTIONAL LEVELS. CLINICAL OBSERVATIONS SUGGEST THAT CHRONIC INFLAMMATION CORRELATES WITH CHRONIC PROSTATITIS/CHRONIC PELVIC PAIN SYNDROME (CP/CPPS) AND BENIGN PROSTATIC HYPERPLASIA (BPH) AND AN HISTORY OF CLINICAL CHRONIC PROSTATITIS SIGNIFICANTLY INCREASES THE ODDS FOR PROSTATE CANCER. THE NIHNIDDK CLASSIFICATION BASED ON THE USE OF THE MICROBIOLOGICAL 4- GLASSES LOCALIZATION TEST OR SIMPLIFIED 2-GLASSES TEST, IS CURRENTLY ACCEPTED WORLDWIDE. THE UPOINT SYSTEM IDENTIFIES GROUPS OF CLINICIANS WITH HOMOGENEOUS CLINICAL PRESENTATION AND IS USED TO RECOGNIZE PHENOTYPES TO BE SUBMITTED TO SPECIFIC TREATMENTS. THE UPOINTS ALGORITHM IMPLEMENTED THE ORIGINAL UPOINT ADDING TO THE URINARY DOMAINS (U), PSYCHO-SOCIAL (P), ORGANSPECIFIC (O), INFECTION (I), NEUROLOGICAL (N), MUSCLE TENSION AND TENDERNESS (T) A FURTHER DOMAIN RELATED TO SEXUALITY (S). IN FACT SEXUAL DYSFUNCTION (ERECTILE, EJACULATORY, LIBIDO LOSS) HAS BEEN DESCRIBED IN 46-92% OF CASES WITH A HIGH IMPACT ON THE QUALITY OF LIFE OF PATIENTS WITH CP/CPPS. PROSTATIC ULTRASOUND REPRESENTS THE MOST POPULAR IMAGING TEST IN THE WORK-UP OF EITHER ACUTE AND CHRONIC PROSTATITIS ALTHOUGH NO SPECIFIC HYPO-HYPERECHOIC PATTERN HAS BEEN CLEARLY ASSOCIATED WITH CHRONIC BACTERIAL PROSTATITIS AND CPPS. USE OF A DIGITAL-PROCESSING SOFTWARE TO CALCULATE THE EXTENSION OF PROSTATIC CALCIFICATION AREA AT ULTRASOUND DEMONSTRATED A HIGHER PERCENTAGE OF PROSTATIC CALCIFICATION IN PATIENTS WITH CHRONIC BACTERIAL PROSTATITIS. MULTIPARAMETRIC MAGNETIC RESONANCE IMAGING (MPMRI) IS THE CURRENT STATE-OF-THE ART IMAGING MODALITY IN THE ASSESSMENT OF PATIENTS WITH PROSTATE CANCER ALTHOUGH A VARIETY OF BENIGN CONDITIONS, INCLUDING INFLAMMATION, MAY MIMIC PROSTATE CANCER AND ACT AS CONFOUNDING FACTORS IN THE DISCRIMINATION BETWEEN NEOPLASTIC AND NON-NEOPLASTIC LESIONS. BACTERIA CAN INFECT PROSTATE GLAND BY: ASCENDING THE URETHRA, REFLUX OF URINE INTO THE PROSTATIC DUCTS, DIRECT INOCULATION OF BACTERIA THROUGH INSERTED BIOPSY NEEDLES OR HEMATOGENOUS SEEDING. ENTEROBACTERIACEAE ARE THE PREDOMINANT PATHOGENS IN ACUTE AND CHRONIC BACTERIAL PROSTATITIS, BUT AN INCREASING ROLE OF ENTEROCOCCI HAS BEEN REPORTED. MANY STRAINS OF THESE UROPATHOGENS EXHIBIT THE ABILITY TO FORM BIOFILM AND MULTIDRUG- RESISTANCE. SEXUALLY TRANSMITTED INFECTIONS (STI) AGENTS, IN PARTICULAR CHLAMYDIA TRACHOMATIS AND MYCOPLASMA GENITALIUM, HAVE BEEN ALSO CONSIDERED AS CAUSATIVE PATHOGENS OF CHRONIC BACTERIAL PROSTATITIS. ON THE CONTRARY THE EFFECTIVE ROLE IN GENITAL DISEASES OF OTHER "GENITAL MYCOPLASMAS" IS STILL A MUCH DEBATED ISSUE. SEXUALLY TRANSMITTED INFECTIONS AGENTS SHOULD BE INVESTIGATED BY MOLECULAR METHODS IN BOTH PATIENT AND SEXUAL PARTNER. "NEXT GENERATION" INVESTIGATIONS, SUCH AS CYTOKINE ANALYSIS, CYTOLOGICAL TYPING OF IMMUNE CELLS COULD HELP STRATIFYING THE IMMUNE RESPONSE. EPIGENETIC DYSREGULATION OF INFLAMMATORY FACTORS SHOULD BE INVESTIGATED ACCORDING TO SYSTEMIC AND COMPARTMENT-SPECIFIC SIGNALS. THE SEARCH FOR BIOMARKERS SHOULD ALSO INCLUDE EVALUATION OF HORMONAL PATHWAYS, AS MEASUREMENT OF ESTROGEN LEVELS IN SEMEN. ANTIMICROBIALS ARE THE FIRST LINE AGENTS FOR THE TREATMENT OF BACTERIAL PROSTATITIS. THE SUCCESS OF ANTIMICROBIAL TREATMENT DEPENDS ON THE ANTIBACTERIAL ACTIVITY AND THE PHARMACOKINETIC CHARACTERISTICS OF THE DRUG WHICH MUST REACH HIGH CONCENTRATIONS IN PROSTATE SECRETION AND PROSTATE TISSUE. ACUTE BACTERIAL PROSTATITIS CAN BE A SERIOUS INFECTION WITH A POTENTIAL RISK FOR UROSEPSIS FOR IINITIAL TREATMENT OF SEVERELY ILL PATIENTS, INTRAVENOUS ADMINISTRATION OF HIGH DOSES OF BACTERICIDAL ANTIMICROBIALS, SUCH AS BROAD-SPECTRUM PENICILLINS, THIRD-GENERATION CEPHALOSPORINS OR FLUOROQUINOLONES, IS RECOMMENDED IN COMBINATION WITH AN AMINOGLYCOSIDE. USE OF PIPERACILLIN-TAZOBACTAM AND MEROPENEM IS JUSTIFIED IN PRESENCE OF MULTIRESISTANT GRAMNEGATIVE PATHOGENS. THE ANTIBIOTIC TREATMENT OF CHRONIC PROSTATITIS IS CURRENTLY BASED ON THE USE OF FLUOROQUINOLONES THAT, GIVEN FOR 2 TO 4 WEEKS, CURED ABOUT 70% OF MEN WITH CHRONIC BACTERIAL PROSTATITIS. FOR THE TREATMENT OF CHLAMYDIAL PROSTATITIS MACROLIDES WERE SHOWN TO BE MORE EFFECTIVE THAN FLUOROQUINOLONES, WHEREAS NO DIFFERENCES WERE OBSERVED IN MICROBIOLOGICAL AND CLINICAL EFFICACY BETWEEN MACROLIDES AND TETRACYCLINES FOR THE TREATMENT OF INFECTIONS CAUSED BY INTRACELLULAR PATHOGENS. AMINOGLYCOSIDES AND FOSFOMYCIN COULD BE CONSIDERED AS A THERAPEUTIC ALTERNATIVE FOR THE TREATMENT OF QUINOLONE RESISTANT PROSTATITIS. USE OF ALPHA-BLOCKERS IN CP/CPPS PATIENTS WITH URINARY SYMPTOMS AND ANALGESICS +/- NON STEROIDAL ANTI-INFLAMMATORY DRUGS (NSAID), IN PRESENCE OF PAIN DEMONSTRATED A REDUCTION OF SYMPTOMS REDUCTION AND AN IMPROVEMENT OF QUALITY OF LIFE, ALTHOUGH LONG TERM USE OF NSAID IS LIMITED BY SIDE EFFECT PROFILE. HOWEVER, THE MULTIMODAL THERAPEUTIC REGIMEN BY CONTEMPORARY USE OF ALPHABLOCKERS, ANTIBIOTICS AND ANTI-INFLAMMATORY SHOWED A BETTER CONTROL OF PROSTATITIS SYMPTOMS THAN SINGLE DRUG TREATMENT. NOVEL THERAPEUTIC SUBSTANCES FOR THE TREATMENT OF PAIN, SUCH AS THE CANNABINOID ANANDAMIDE WOULD BE HIGHLY INTERESTING TO TEST. AN ALTERNATIVE FOR THE TREATMENT OF CHRONIC PROSTATITIS/CHRONIC PELVIC PAIN SYNDROME IS PHYTOTHERAPY, AS PRIMARY THERAPY OR IN ASSOCIATION WITH OTHER DRUGS. QUERCETIN, POLLEN EXTRACT, EXTRACT OF SERENOA REPENS AND OTHER MIXTURES OF HERBAL EXTRACTS SHOWED A POSITIVE EFFECT ON SYMPTOMS AND QUALITY OF LIFE WITHOUT SIDE EFFECTS. THE ASSOCIATION OF CP/CPPS WITH ALTERATIONS OF INTESTINAL FUNCTION HAS BEEN DESCRIBED. DIET HAS ITS EFFECTS ON INFLAMMATION BY REGULATION OF THE COMPOSITION OF INTESTINAL FLORA AND DIRECT ACTION ON THE INTESTINAL CELLS (STERILE INFLAMMATION). INTESTINAL BACTERIA (MICROBIOTA) INTERACTS WITH FOOD INFLUENCING THE METABOLIC, IMMUNE AND INFLAMMATORY RESPONSE OF THE ORGANISM. THE INTESTINAL MICROBIOTA HAS PROTECTIVE FUNCTION AGAINST PATHOGENIC BACTERIA, METABOLIC FUNCTION BY SYNTHESIS OF VITAMINS, DECOMPOSITION OF BILE ACIDS AND PRODUCTION OF TROPHIC FACTORS (BUTYRATE), AND MODULATION OF THE INTESTINAL IMMUNE SYSTEM. THE ALTERATION OF THE MICROBIOTA IS CALLED "DYSBIOSIS" CAUSING INVASIVE INTESTINAL DISEASES PATHOLOGIES (LEAKY GUT SYNDROME AND FOOD INTOLERANCES, IRRITABLE BOWEL SYNDROME OR CHRONIC INFLAMMATORY BOWEL DISEASES) AND CORRELATING WITH NUMEROUS SYSTEMIC DISEASES INCLUDING ACUTE AND CHRONIC PROSTATITIS. ADMINISTRATION OF LIVE PROBIOTICS BACTERIA CAN BE USED TO REGULATE THE BALANCE IF INTESTINAL FLORA. SESSIONS OF HYDROCOLONTHERAPY CAN REPRESENT AN INTEGRATION TO THIS THERAPEUTIC APPROACH. FINALLY, MICROBIOLOGICAL EXAMINATION OF SEXUAL PARTNERS CAN OFFER SUPPLEMENTARY INFORMATION FOR TREATMENT. 2019 5 978 39 CHRONIC OXIDATIVE STRESS CAUSES ESTROGEN-INDEPENDENT AGGRESSIVE PHENOTYPE, AND EPIGENETIC INACTIVATION OF ESTROGEN RECEPTOR ALPHA IN MCF-7 BREAST CANCER CELLS. THE ROLE OF CHRONIC OXIDATIVE STRESS IN THE DEVELOPMENT AND AGGRESSIVE GROWTH OF ESTROGEN RECEPTOR (ER)-POSITIVE BREAST CANCER IS WELL KNOWN; HOWEVER, THE MECHANISTIC UNDERSTANDING IS NOT CLEAR. ESTROGEN-INDEPENDENT GROWTH IS ONE OF THE FEATURES OF AGGRESSIVE SUBTYPE OF BREAST CANCER. THEREFORE, THE OBJECTIVE OF THIS STUDY WAS TO EVALUATE THE EFFECT OF OXIDATIVE STRESS ON ESTROGEN SENSITIVITY AND EXPRESSION OF NUCLEAR ESTROGEN RECEPTORS IN ER-POSITIVE BREAST CANCER CELLS. MCF-7 CELLS CHRONICALLY EXPOSED TO HYDROGEN PEROXIDE WERE USED AS A CELL MODEL IN THIS STUDY, AND THEIR GROWTH IN RESPONSE TO 17-BETA ESTRADIOL WAS EVALUATED BY CELL VIABILITY, CELL CYCLE, AND CELL MIGRATION ANALYSIS. RESULTS WERE FURTHER CONFIRMED AT MOLECULAR LEVEL BY ANALYSIS OF GENE EXPRESSIONS AT TRANSCRIPT AND PROTEIN LEVELS. HISTONE H3 MODIFICATIONS, EXPRESSION OF EPIGENETIC REGULATORY GENES, AND THE EFFECT OF DNA DEMETHYLATION WERE ALSO ANALYZED. LOSS OF GROWTH IN RESPONSE TO ESTROGEN WITH A DECREASE IN ERALPHA EXPRESSION WAS OBSERVED IN MCF-7 CELLS ADAPTED TO CHRONIC OXIDATIVE STRESS. INCREASES IN MTTFA AND NRF1 IN THESE CELLS FURTHER SUGGESTED THE ROLE OF MITOCHONDRIA-DEPENDENT REDOX-SENSITIVE GROWTH SIGNALING AS AN ALTERNATIVE PATHWAY TO ESTROGEN-DEPENDENT GROWTH. CHANGES IN EXPRESSION OF EPIGENETIC REGULATORY GENES, LEVELS OF HISTONE H3 MODIFICATIONS AS WELL AS SIGNIFICANT RESTORATIONS OF BOTH ERALPHA EXPRESSION AND ESTROGEN RESPONSE BY 5-AZA-2'-DEOXYCYTIDINE FURTHER CONFIRMED THE EPIGENETIC BASIS FOR ESTROGEN-INDEPENDENT GROWTH IN THESE CELLS. IN CONCLUSION, RESULTS OF THIS STUDY SUGGEST THAT CHRONIC OXIDATIVE STRESS CAN CONVERT ESTROGEN-DEPENDENT NONAGGRESSIVE BREAST CANCER CELLS INTO ESTROGEN-INDEPENDENT AGGRESSIVE FORM POTENTIALLY BY EPIGENETIC MECHANISM. 2015 6 2297 31 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 7 5304 33 PROTEOMIC ANALYSIS OF INFLAMMATORY BIOMARKERS ASSOCIATED WITH BREAST CANCER RECURRENCE. INTRODUCTION: BREAST CANCER IS THE MOST FREQUENT CANCER DETECTED FOR WOMEN, AND WHILE OUR ABILITY TO TREAT BREAST CANCER HAS IMPROVED SUBSTANTIALLY OVER THE YEARS, RECURRENCE REMAINS A MAJOR OBSTACLE. STANDARD SCREENING FOR NEW AND RECURRENT BREAST CANCER INVOLVES CLINICAL BREAST IMAGING. HOWEVER, THERE IS NO CLINICALLY APPROVED NONINVASIVE BODY FLUID TEST FOR THE EARLY DETECTION OF RECURRENT BREAST CANCER. MATERIALS AND METHOD: IN THIS STUDY, WE ANALYZED SERUM SAMPLES FROM BOTH RECURRENT AND NONRECURRENT BREAST CANCER PATIENTS BY DIFFERENT PROTEOMICS METHODS TO IDENTIFY BIOMARKERS IN PATIENTS WITH RECURRENCE OF DISEASE. RESULTS: COMPARATIVE DATA ANALYSIS IDENTIFIED SEVERAL HISTONE DEACETYLASE (HDAC) PROTEINS, WHICH WERE FOUND AT SIGNIFICANTLY HIGHER LEVELS IN THE SERUM OF RECURRENT BREAST CANCER PATIENTS: HDAC9 (C-TERM) (P = 0.0035), HDAC5 (C-TERM) (P = 0.013), SMALL UBIQUITIN-LIKE MODIFIER 1 (N-TERM) (P = 0.017), EMBRYONIC STEM CELL-EXPRESSED RAS (INTER) (P = 0.018), AND HDAC7 (C-TERM) (P = 0.020). CHRONIC INFLAMMATION PLAYS A CRITICAL ROLE IN THE DEVELOPMENT OF THE BREAST CANCER RECURRENCE, AND WE IDENTIFIED SEVERAL PROINFLAMMATORY CYTOKINES THAT WERE PRESENT AT ELEVATED LEVELS ONLY IN RECURRENT BREAST CANCER PATIENT SERUM. CONCLUSIONS: OUR DATA INDICATED THAT THE EPIGENETIC REGULATION OF INFLAMMATORY PROCESSES PLAYS A CRITICAL ROLE IN BREAST CANCER RECURRENCE. THE IDENTIFIED PROTEINS COULD LAY THE GROUNDWORK FOR THE DEVELOPMENT OF A SERUM-BASED BREAST CANCER RECURRENCE ASSAY. 2020 8 344 40 ALTERED BDNF METHYLATION IN PATIENTS WITH CHRONIC MUSCULOSKELETAL PAIN AND HIGH BIOPSYCHOSOCIAL COMPLEXITY. PURPOSE: THE INTERMED INSTRUMENT, WHICH WAS DEVELOPED TO MEASURE PATIENT'S BIOPSYCHOSOCIAL (BPS) COMPLEXITY, REPRESENTS A POWERFUL DIAGNOSTIC AND THERAPEUTIC TOOL. EPIGENETIC CHANGES ARE THE INTERFACE BETWEEN SIGNALS FROM THE ENVIRONMENT AND GENETIC MODIFICATIONS, AFFECTING GENE EXPRESSION, IN PARTICULAR, BY DNA METHYLATION OF CPG DINUCLEOTIDES IN PROMOTOR REGIONS OF THE CORRESPONDING GENES. THE BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) GENE PLAYS A CRUCIAL ROLE IN THE CENTRAL SENSITIZATION (CS) OF PAIN. IN THIS STUDY, WE HYPOTHESIZED THAT CHRONIC PAIN MODIFIES THE METHYLATION LEVELS OF THE BDNF GENE IN A MANNER THAT IS INTERCONNECTED WITH THE BPS STATUS. PATIENTS AND METHODS: FIFTY-EIGHT CHRONIC MUSCULOSKELETAL PAIN PATIENTS (CMSP) WERE ENROLLED IN THE STUDY. DNA WAS EXTRACTED FROM BLOOD SAMPLES, THE METHYLATION LEVELS OF 13 CPG SITES IN THE BDNF PROMOTER WERE MEASURED BY PYROSEQUENCING, AND ASSOCIATION STUDIES WITH VARIOUS PATIENT PARAMETERS AND THE INTERMED SCORES WERE PERFORMED. RESULTS: INTERESTINGLY, A NEGATIVE CORRELATION (-0.40) WAS FOUND BETWEEN THE TOTAL INTERMED SCORES AND THE AVERAGE CPG METHYLATION VALUES OF THE BDNF GENE, BUT NO CORRELATION WAS OBSERVED WITH THE SEVERITY OF PAIN, DEGREE OF ANXIETY, DEPRESSION, OR KINESIOPHOBIA AND CATASTROPHISM. MOREOVER, THE ASSOCIATION WAS INDEPENDENT OF AGE, SEX AND LEVEL OF COMORBIDITIES. CONCLUSION: THIS RESULT SHOWS THAT CMSP, IN ASSOCIATION WITH ITS BIOPSYCHOSOCIAL CONTEXT, EPIGENETICALLY DECREASES THE DEGREE OF METHYLATION OF THE BDNF PROMOTER AND SHOULD THEREFORE INCREASE THE LEVEL OF BDNF TRANSCRIPTION. IT ALSO SUGGESTS A ROLE OF THE INTERMED TOOL TO DETECT A RELATIONSHIP BETWEEN THE BPS COMPLEXITY AND THE EPIGENETIC CONTROL OF A TARGET GENE. THE POSSIBLE UPREGULATION OF BDNF EXPRESSION MIGHT BE, AT LEAST IN PART, THE SIGNAL FOR CHRONIC PAIN-INDUCED CENTRAL SENSITIZATION (CS). THIS COULD PARTLY EXPLAIN WHY PATIENTS WITH A HIGHER LEVEL OF COMPLEXITY FEEL MORE PAIN THAN THOSE WITH LOWER COMPLEXITY. 2020 9 2653 36 EPIGENOTYPING IN PERIPHERAL BLOOD CELL DNA AND BREAST CANCER RISK: A PROOF OF PRINCIPLE STUDY. BACKGROUND: EPIGENETIC CHANGES ARE EMERGING AS ONE OF THE MOST IMPORTANT EVENTS IN CARCINOGENESIS. TWO ALTERATIONS IN THE PATTERN OF DNA METHYLATION IN BREAST CANCER (BC) HAVE BEEN PREVIOUSLY REPORTED; ACTIVE ESTROGEN RECEPTOR-ALPHA (ER-ALPHA) IS ASSOCIATED WITH DECREASED METHYLATION OF ER-ALPHA TARGET (ERT) GENES, AND POLYCOMB GROUP TARGET (PCGT) GENES ARE MORE LIKELY THAN OTHER GENES TO HAVE PROMOTER DNA HYPERMETHYLATION IN CANCER. HOWEVER, WHETHER DNA METHYLATION IN NORMAL UNRELATED CELLS IS ASSOCIATED WITH BC RISK AND WHETHER THESE IMPRINTS CAN BE RELATED TO FACTORS WHICH CAN BE MODIFIED BY THE ENVIRONMENT, IS UNCLEAR. METHODOLOGY/PRINCIPAL FINDINGS: USING QUANTITATIVE METHYLATION ANALYSIS IN A CASE-CONTROL STUDY (N = 1,083) WE FOUND THAT DNA METHYLATION OF PERIPHERAL BLOOD CELL DNA PROVIDES GOOD PREDICTION OF BC RISK. WE ALSO REPORT THAT INVASIVE DUCTAL AND INVASIVE LOBULAR BC IS CHARACTERIZED BY TWO DIFFERENT SETS OF GENES, THE LATTER PARTICULAR BY GENES INVOLVED IN THE DIFFERENTIATION OF THE MESENCHYME (PITX2, TITF1, GDNF AND MYOD1). FINALLY WE DEMONSTRATE THAT ONLY ERT GENES PREDICT ER POSITIVE BC; LACK OF PERIPHERAL BLOOD CELL DNA METHYLATION OF ZNF217 PREDICTED BC INDEPENDENT OF AGE AND FAMILY HISTORY (ODDS RATIO 1.49; 95% CONFIDENCE INTERVAL 1.12-1.97; P = 0.006) AND WAS ASSOCIATED WITH ER-ALPHA BIOACTIVITY IN THE CORRESPONDING SERUM. CONCLUSION/SIGNIFICANCE: THIS FIRST LARGE-SCALE EPIGENOTYPING STUDY DEMONSTRATES THAT DNA METHYLATION MAY SERVE AS A LINK BETWEEN THE ENVIRONMENT AND THE GENOME. FACTORS THAT CAN BE MODULATED BY THE ENVIRONMENT (LIKE ESTROGENS) LEAVE AN IMPRINT IN THE DNA OF CELLS THAT ARE UNRELATED TO THE TARGET ORGAN AND INDICATE THE PREDISPOSITION TO DEVELOP A CANCER. FURTHER RESEARCH WILL NEED TO DEMONSTRATE WHETHER DNA METHYLATION PROFILES WILL BE ABLE TO SERVE AS A NEW TOOL TO PREDICT THE RISK OF DEVELOPING CHRONIC DISEASES WITH SUFFICIENT ACCURACY TO GUIDE PREVENTIVE MEASURES. 2008 10 2436 40 EPIGENETIC SILENCING OF SOCS3 IDENTIFIES A SUBSET OF PROSTATE CANCER WITH AN AGGRESSIVE BEHAVIOR. BACKGROUND: CHRONIC INFLAMMATION AND SUBSEQUENT TISSUTAL ALTERATIONS MAY PLAY A KEY ROLE IN PROSTATE CARCINOGENESIS. IN THIS WAY, MOLECULAR ALTERATIONS OF THE SUPPRESSOR OF CYTOKINE SIGNALING 3 (SOCS3), ONE OF THE MOST IMPORTANT INHIBITORY MOLECULE OF INFLAMMATORY SIGNAL TRANSDUCTION CIRCUITRIES, COULD CONTRIBUTE TO EXPLAIN THE PLEIOTROPIC ROLE OF INTERLEUKIN-6 (IL-6) IN THIS TYPE OF CANCER. METHODS: WE ANALYZED THE METHYLATION STATUS AND MRNA EXPRESSION OF SOCS3 IN 20 BENIGN PROSTATE HYPERPLASIAS (BPH) AND IN 51 PROSTATE CANCER SPECIMENS. WE ANALYZED THE SOCS3 METHYLATION STATUS USING METHYLATION-SPECIFIC PCR. HYPERMETHYLATION WAS CONFIRMED BY SEQUENCING AFTER SUBCLONING. EPIGENETIC SILENCING OF THIS GENE WAS ALSO DEMONSTRATED BY REAL-TIME PCR AND BY IMMUNOHISTOCHEMISTRY. RESULTS AND CORRELATION WITH CLINICAL DATA WERE STATISTICALLY ANALYZED. RESULTS: WE FOUND THAT THE PROMOTER OF SOCS3 WAS METHYLATED IN 39.2% OF PROSTATE CANCER. ON THE CONTRARY, ALL BPH AND NORMAL CONTROLS HAD AN UNMETHYLATED PATTERN. REAL-TIME ANALYSIS SHOWED THAT IN METHYLATED CASES SOCS3 MRNA EXPRESSION WAS REDUCED BY THREE AND FOUR FOLDS AS COMPARED TO BPH AND UNMETHYLATED CASES, RESPECTIVELY. INTERESTINGLY, SOCS3 MRNA LEVEL WAS HIGHER IN UNMETHYLATED PROSTATE CANCER THAN IN BPH. THE IMMUNOHISTOCHEMICAL STAINING ANALYSIS FOR SOCS 3 CONFIRMED MRNA RESULTS. MOREOVER, METHYLATION OF SOCS3 PROMOTER SIGNIFICANTLY ASSOCIATED WITH INTERMEDIATE-HIGH GRADE GLEASON SCORE (P = 0.0007) AND WITH AN UNFAVORABLE CLINICAL OUTCOME (P = 0.0019). CONCLUSIONS: OUR DATA SUGGEST THAT SOCS3 HYPERMETHYLATION MAY BE INVOLVED IN THE PATHOGENESIS OF PROSTATE CANCER AND COULD IDENTIFY A TUMOR SUBSET WITH AN AGGRESSIVE BEHAVIOR. 2011 11 519 33 ASSOCIATIONS BETWEEN GENETIC AND EPIGENETIC VARIATIONS IN CYTOKINE GENES AND MILD PERSISTENT BREAST PAIN IN WOMEN FOLLOWING BREAST CANCER SURGERY. PERSISTENT PAIN FOLLOWING BREAST CANCER SURGERY IS A SIGNIFICANT PROBLEM. BOTH INHERITED AND ACQUIRED MECHANISMS OF INFLAMMATION APPEAR TO PLAY A ROLE IN THE DEVELOPMENT AND MAINTENANCE OF PERSISTENT PAIN. IN THIS LONGITUDINAL STUDY, GROWTH MIXTURE MODELING WAS USED TO IDENTIFY PERSISTENT BREAST PAIN PHENOTYPES BASED ON PAIN ASSESSMENTS OBTAINED PRIOR TO AND MONTHLY FOR 6MONTHS FOLLOWING BREAST CANCER SURGERY. ASSOCIATIONS BETWEEN THE "NO PAIN" AND "MILD PAIN" PHENOTYPES AND SINGLE NUCLEOTIDE POLYMORPHISMS (SNPS) SPANNING 15 CYTOKINE GENES WERE EVALUATED. THE METHYLATION STATUS OF THE CPG SITES FOUND IN THE PROMOTERS OF GENES ASSOCIATED WITH PAIN GROUP MEMBERSHIP WAS DETERMINED USING BISULFITE SEQUENCING. IN THE MULTIVARIATE ANALYSIS, THREE SNPS (I.E., INTERLEUKIN 6 (IL6) RS2069840, C-X-C MOTIF CHEMOKINE LIGAND 8 (CXCL8) RS4073, TUMOR NECROSIS FACTOR (TNF) RS1800610) AND TWO TNF CPG SITES (I.E., C.-350C, C.-344C) WERE ASSOCIATED WITH PAIN GROUP MEMBERSHIP. THESE FINDINGS SUGGEST THAT VARIATIONS IN IL6, CXCL8, AND TNF ARE ASSOCIATED WITH THE DEVELOPMENT AND MAINTENANCE OF MILD PERSISTENT BREAST PAIN. CPG METHYLATION WITHIN THE TNF PROMOTER MAY PROVIDE AN ADDITIONAL MECHANISM THROUGH WHICH TNF ALTERS THE RISK FOR MILD PERSISTENT BREAST PAIN AFTER BREAST CANCER SURGERY. THESE GENETIC AND EPIGENETIC VARIATIONS MAY HELP TO IDENTIFY INDIVIDUALS WHO ARE PREDISPOSED TO THE DEVELOPMENT OF MILD LEVELS OF PERSISTENT BREAST PAIN FOLLOWING BREAST CANCER SURGERY. 2017 12 2400 39 EPIGENETIC REPROGRAMMING OF IMMUNE CELLS IN WOMEN WITH PCOS IMPACT GENES CONTROLLING REPRODUCTIVE FUNCTION. CONTEXT: POLYCYSTIC OVARY SYNDROME (PCOS) IS A CHRONIC DISEASE AFFECTING REPRODUCTIVE FUNCTION AND WHOLE-BODY METABOLISM. ALTHOUGH THE ETIOLOGY IS UNCLEAR, EMERGING EVIDENCE INDICATES THAT THE EPIGENETICS MAY BE A CONTRIBUTING FACTOR. OBJECTIVE: TO DETERMINE THE ROLE OF GLOBAL AND GENOME-WIDE EPIGENETIC MODIFICATIONS IN SPECIFIC IMMUNE CELLS IN PCOS COMPARED WITH CONTROLS AND WHETHER THESE COULD BE RELATED TO CLINICAL FEATURES OF PCOS. DESIGN: CROSS-SECTIONAL STUDY. PARTICIPANTS: WOMEN WITH (N = 17) OR WITHOUT PCOS (N = 17). SETTING: RECRUITED FROM THE GENERAL COMMUNITY. MAIN OUTCOME MEASURES: ISOLATED PERIPHERAL BLOOD MONONUCLEAR CELLS WERE ANALYZED USING MULTICOLOR FLOW CYTOMETRY METHODS TO DETERMINE GLOBAL DNA METHYLATION LEVELS IN A CELL-SPECIFIC FASHION. TRANSCRIPTOMIC AND GENOME-WIDE DNA METHYLATION ANALYSES WERE PERFORMED ON T HELPER CELLS USING RNA SEQUENCING AND REDUCED REPRESENTATION BISULFITE SEQUENCING. RESULTS: WOMEN WITH PCOS HAD LOWER GLOBAL DNA METHYLATION IN MONOCYTES (P = 0.006) AND IN T HELPER (P = 0.004), T CYTOTOXIC (P = 0.004), AND B CELLS (P = 0.03). SPECIFIC GENOME-WIDE DNA METHYLATION ANALYSIS OF T HELPER CELLS FROM WOMEN WITH PCOS IDENTIFIED 5581 DIFFERENTIALLY METHYLATED CPG SITES. FUNCTIONAL GENE ONTOLOGY ENRICHMENT ANALYSIS SHOWED THAT GENES LOCATED AT THE PROXIMITY OF DIFFERENTIALLY METHYLATED CPG SITES BELONG TO PATHWAYS RELATED TO REPRODUCTIVE FUNCTION AND IMMUNE CELL FUNCTION. HOWEVER, THESE GENES WERE NOT ALTERED AT THE TRANSCRIPTOMIC LEVEL. CONCLUSIONS: IT WAS SHOWN THAT PCOS IS ASSOCIATED WITH GLOBAL AND GENE-SPECIFIC DNA METHYLATION REMODELING IN A CELL TYPE-SPECIFIC MANNER. FURTHER INVESTIGATION IS WARRANTED TO DETERMINE WHETHER EPIGENETIC REPROGRAMMING OF IMMUNE CELLS IS IMPORTANT IN DETERMINING THE DIFFERENT PHENOTYPES OF PCOS. 2019 13 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 14 839 27 CHEMO-ENZYMATIC FLUORESCENCE LABELING OF GENOMIC DNA FOR SIMULTANEOUS DETECTION OF GLOBAL 5-METHYLCYTOSINE AND 5-HYDROXYMETHYLCYTOSINE. 5-METHYLCYTOSINE AND 5-HYDROXYMETHYLCYTOSINE ARE EPIGENETIC MODIFICATIONS INVOLVED IN GENE REGULATION AND CANCER. WE PRESENT A NEW, SIMPLE, AND HIGH-THROUGHPUT PLATFORM FOR MULTI-COLOR EPIGENETIC ANALYSIS. THE NOVELTY OF OUR APPROACH IS THE ABILITY TO MULTIPLEX METHYLATION AND DE-METHYLATION SIGNALS IN THE SAME ASSAY. WE UTILIZE AN ENGINEERED METHYLTRANSFERASE ENZYME THAT RECOGNIZES AND LABELS ALL UNMODIFIED CPG SITES WITH A FLUORESCENT COFACTOR. IN COMBINATION WITH THE ALREADY ESTABLISHED LABELING OF THE DE-METHYLATION MARK 5-HYDROXYMETHYLCYTOSINE VIA ENZYMATIC GLYCOSYLATION, WE OBTAINED A ROBUST PLATFORM FOR SIMULTANEOUS EPIGENETIC ANALYSIS OF THESE MARKS. WE ASSESSED THE GLOBAL EPIGENETIC LEVELS IN MULTIPLE SAMPLES OF COLORECTAL CANCER AND OBSERVED A 3.5-FOLD REDUCTION IN 5HMC LEVELS BUT NO CHANGE IN DNA METHYLATION LEVELS BETWEEN SICK AND HEALTHY INDIVIDUALS. WE ALSO MEASURED EPIGENETIC MODIFICATIONS IN CHRONIC LYMPHOCYTIC LEUKEMIA AND OBSERVED A DECREASE IN BOTH MODIFICATION LEVELS (5-HYDROXYMETHYLCYTOSINE: WHOLE BLOOD 30 %; PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS) 40 %. 5-METHYLCYTOSINE: WHOLE BLOOD 53 %; PBMCS 48 %). OUR FINDINGS PROPOSE USING A SIMPLE BLOOD TEST AS A VIABLE METHOD FOR ANALYSIS, SIMPLIFYING SAMPLE HANDLING IN DIAGNOSTICS. IMPORTANTLY, OUR RESULTS HIGHLIGHT THE ASSAY'S POTENTIAL FOR EPIGENETIC EVALUATION OF CLINICAL SAMPLES, BENEFITING RESEARCH AND PATIENT MANAGEMENT. 2023 15 3455 46 HYPOMETHYLATION COORDINATES ANTAGONISTICALLY WITH HYPERMETHYLATION IN CANCER DEVELOPMENT: A CASE STUDY OF LEUKEMIA. BACKGROUND: METHYLATION CHANGES ARE FREQUENT IN CANCERS, BUT UNDERSTANDING HOW HYPER- AND HYPOMETHYLATED REGION CHANGES COORDINATE, ASSOCIATE WITH GENOMIC FEATURES, AND AFFECT GENE EXPRESSION IS NEEDED TO BETTER UNDERSTAND THEIR BIOLOGICAL SIGNIFICANCE. THE FUNCTIONAL SIGNIFICANCE OF HYPERMETHYLATION IS WELL STUDIED, BUT THAT OF HYPOMETHYLATION REMAINS LIMITED. HERE, WITH PAIRED EXPRESSION AND METHYLATION SAMPLES GATHERED FROM A PATIENT/CONTROL COHORT, WE ATTEMPT TO BETTER CHARACTERIZE THE GENE EXPRESSION AND METHYLATION CHANGES THAT TAKE PLACE IN CANCER FROM B CELL CHRONIC LYMPHOCYTE LEUKEMIA (B-CLL) SAMPLES. RESULTS: ACROSS THE DATASET, WE FOUND THAT CONSISTENT DIFFERENTIALLY HYPOMETHYLATED REGIONS (C-DMRS) ACROSS SAMPLES WERE RELATIVELY FEW COMPARED TO THE MANY POORLY CONSISTENT HYPO- AND HIGHLY CONSERVED HYPER-DMRS. HOWEVER, GENES IN THE HYPO-C-DMRS TENDED TO BE ASSOCIATED WITH FUNCTIONS ANTAGONISTIC TO THOSE IN THE HYPER-C-DMRS, LIKE DIFFERENTIATION, CELL-CYCLE REGULATION AND PROLIFERATION, SUGGESTING COORDINATED REGULATION OF METHYLATION CHANGES. HYPO-C-DMRS IN B-CLL WERE FOUND ENRICHED IN KEY SIGNALING PATHWAYS LIKE B CELL RECEPTOR AND P53 PATHWAYS AND GENES/MOTIFS ESSENTIAL FOR B LYMPHOPOIESIS. HYPO-C-DMRS TENDED TO BE PROXIMAL TO GENES WITH ELEVATED EXPRESSION IN CONTRAST TO THE TRANSCRIPTION SILENCING-MECHANISM IMPOSED BY HYPERMETHYLATION. HYPO-C-DMRS TENDED TO BE ENRICHED IN THE REGIONS OF ACTIVATING H4K4ME1/2/3, H3K79ME2, AND H3K27AC HISTONE MODIFICATIONS. IN COMPARISON, THE POLYCOMB REPRESSIVE COMPLEX 2 (PRC2) SIGNATURE, MARKED BY EZH2, SUZ12, CTCF BINDING-SITES, REPRESSIVE H3K27ME3 MARKS, AND "REPRESSED/POISED PROMOTER" STATES WERE ASSOCIATED WITH HYPER-C-DMRS. MOST HYPO-C-DMRS WERE FOUND IN INTRONS (36 %), 3' UNTRANSLATED REGIONS (29 %), AND INTERGENIC REGIONS (24 %). MANY OF THESE GENIC REGIONS ALSO OVERLAPPED WITH ENHANCERS. THE METHYLATION OF CPGS FROM 3'UTR EXONS WAS FOUND TO HAVE WEAK BUT POSITIVE CORRELATION WITH GENE EXPRESSION. IN CONTRAST, METHYLATION IN THE 5'UTR WAS NEGATIVELY CORRELATED WITH EXPRESSION. TO BETTER CHARACTERIZE THE OVERLAP BETWEEN METHYLATION AND EXPRESSION CHANGES, WE IDENTIFIED CORRELATION MODULES THAT ASSOCIATE WITH "APOPTOSIS" AND "LEUKOCYTE ACTIVATION". CONCLUSIONS: DESPITE CLINICAL HETEROGENEITY IN DISEASE PRESENTATION, A NUMBER OF METHYLATION CHANGES, BOTH HYPO AND HYPER, APPEAR TO BE COMMON IN B-CLL. HYPOMETHYLATION APPEARS TO PLAY AN ACTIVE, TARGETED, AND COMPLEMENTARY ROLE IN CANCER PROGRESSION, AND IT INTERPLAYS WITH HYPERMETHYLATION IN A COORDINATED FASHION IN THE CANCER PROCESS. 2016 16 3077 32 GENOME-WIDE METHYL-SEQ ANALYSIS OF BLOOD-BRAIN TARGETS OF GLUCOCORTICOID EXPOSURE. CHRONIC EXPOSURE TO GLUCOCORTICOIDS (GCS) CAN LEAD TO PSYCHIATRIC COMPLICATIONS THROUGH EPIGENETIC MECHANISMS SUCH AS DNA METHYLATION (DNAM). WE SOUGHT TO DETERMINE WHETHER EPIGENETIC CHANGES IN A PERIPHERAL TISSUE CAN SERVE AS A SURROGATE FOR THOSE IN A RELATIVELY INACCESSIBLE TISSUE SUCH AS THE BRAIN. DNA EXTRACTED FROM THE HIPPOCAMPUS AND BLOOD OF MICE TREATED WITH GCS OR VEHICLE SOLUTION WAS ASSAYED USING A GENOME-WIDE DNAM PLATFORM (METHYL-SEQ) TO IDENTIFY DIFFERENTIALLY METHYLATED REGIONS (DMRS) INDUCED BY GC TREATMENT. WE OBSERVED THAT APPROXIMATELY 70% OF THE DMRS IN BOTH TISSUES LOST METHYLATION FOLLOWING GC TREATMENT. OF THE 3,095 DMRS THAT MAPPED TO THE SAME GENES IN BOTH TISSUES, 1,853 DMRS UNDERWENT DNAM CHANGES IN THE SAME DIRECTION. INTERESTINGLY, ONLY 209 DMRS (<7%) OVERLAPPED IN GENOMIC COORDINATES BETWEEN THE 2 TISSUES, SUGGESTING TISSUE-SPECIFIC DIFFERENCES IN GC-TARGETED LOCI. PATHWAY ANALYSIS SHOWED THAT THE DMR-ASSOCIATED GENES WERE MEMBERS OF PATHWAYS INVOLVED IN METABOLISM, IMMUNE FUNCTION, AND NEURODEVELOPMENT. ALSO, CHANGES IN CELL TYPE COMPOSITION OF BLOOD AND BRAIN WERE EXAMINED BY FLUORESCENCE-ACTIVATED CELL SORTING. SEPARATION OF THE CORTEX INTO NEURONAL AND NON-NEURONAL FRACTIONS AND THE LEUKOCYTES INTO T-CELLS, B-CELLS, AND NEUTROPHILS SHOWED THAT GC-INDUCED METHYLATION CHANGES PRIMARILY OCCURRED IN NEURONS AND T-CELLS, WITH THE BLOOD TISSUE ALSO UNDERGOING A SHIFT IN THE PROPORTION OF CONSTITUENT CELL TYPES WHILE THE PROPORTION OF NEURONS AND GLIA IN THE BRAIN REMAINED STABLE. FROM THE CURRENT PILOT STUDY, WE FOUND THAT DESPITE TISSUE-SPECIFIC EPIGENETIC CHANGES AND CELLULAR HETEROGENEITY, BLOOD CAN SERVE AS A SURROGATE FOR GC-INDUCED CHANGES IN THE BRAIN. 2017 17 59 36 A GENOME-WIDE SCREEN IDENTIFIES FREQUENTLY METHYLATED GENES IN HAEMATOLOGICAL AND EPITHELIAL CANCERS. BACKGROUND: GENETIC AS WELL AS EPIGENETIC ALTERATIONS ARE A HALLMARK OF BOTH EPITHELIAL AND HAEMATOLOGICAL MALIGNANCIES. HIGH THROUGHPUT SCREENS ARE REQUIRED TO IDENTIFY EPIGENETIC MARKERS THAT CAN BE USEFUL FOR DIAGNOSTIC AND PROGNOSTIC PURPOSES ACROSS MALIGNANCIES. RESULTS: HERE WE REPORT FOR THE FIRST TIME THE USE OF THE MIRA ASSAY (METHYLATED CPG ISLAND RECOVERY ASSAY) IN COMBINATION WITH GENOME-WIDE CPG ISLAND ARRAYS TO IDENTIFY EPIGENETIC MOLECULAR MARKERS IN CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA (ALL) ON A GENOME-WIDE SCALE. WE IDENTIFIED 30 GENES DEMONSTRATING METHYLATION FREQUENCIES OF > OR =25% IN CHILDHOOD ALL, NINE GENES SHOWED SIGNIFICANTLY DIFFERENT METHYLATION FREQUENCIES IN B VS T-ALL. FOR MAJORITY OF THE GENES EXPRESSION COULD BE RESTORED IN METHYLATED LEUKEMIA LINES AFTER TREATMENT WITH 5-AZADC. FORTY-FOUR PERCENT OF THE GENES REPRESENT TARGETS OF THE POLYCOMB COMPLEX. IN CHRONIC MYELOID LEUKEMIA (CML) TWO OF THE GENES, (TFAP2A AND EBF2), DEMONSTRATED INCREASED METHYLATION IN BLAST CRISIS COMPARED TO CHRONIC PHASE (P < 0.05). FURTHERMORE HYPERMETHYLATION OF AN AUTOPHAGY RELATED GENE ATG16L2 WAS ASSOCIATED WITH POORER PROGNOSIS IN TERMS OF MOLECULAR RESPONSE TO IMATINIB TREATMENT. LASTLY WE DEMONSTRATED THAT TEN OF THESE GENES WERE ALSO FREQUENTLY METHYLATED IN COMMON EPITHELIAL CANCERS. CONCLUSION: IN SUMMARY WE HAVE IDENTIFIED A LARGE NUMBER OF GENES SHOWING FREQUENT METHYLATION IN CHILDHOOD ALL, METHYLATION STATUS OF TWO OF THESE GENES IS ASSOCIATED WITH ADVANCED DISEASE IN CML AND METHYLATION STATUS OF ANOTHER GENE IS ASSOCIATED WITH PROGNOSIS. IN ADDITION A SUBSET OF THESE GENES MAY ACT AS EPIGENETIC MARKERS ACROSS HEMATOLOGICAL MALIGNANCIES AS WELL AS COMMON EPITHELIAL CANCERS. 2010 18 1215 39 CPG PROMOTER METHYLATION STATUS IS NOT A PROGNOSTIC INDICATOR OF GENE EXPRESSION IN BERYLLIUM CHALLENGE. INDIVIDUALS EXPOSED TO BERYLLIUM (BE) MAY DEVELOP BE SENSITIZATION (BES) AND PROGRESS TO CHRONIC BERYLLIUM DISEASE (CBD). RECENT STUDIES WITH OTHER METAL ANTIGENS SUGGEST EPIGENETIC MECHANISMS MAY BE INVOLVED IN INFLAMMATORY DISEASE PROCESSES, INCLUDING GRANULOMATOUS LUNG DISORDERS AND THAT A NUMBER OF METAL CATIONS ALTER GENE METHYLATION. THE OBJECTIVE OF THIS STUDY WAS TO DETERMINE IF BE CAN EXERT AN EPIGENETIC EFFECT ON GENE EXPRESSION BY ALTERING METHYLATION IN THE PROMOTER REGION OF SPECIFIC GENES KNOWN TO BE INVOLVED IN BE ANTIGEN-MEDIATED GENE EXPRESSION. TO INVESTIGATE THIS OBJECTIVE, THREE MACROPHAGE TUMOR MOUSE CELL LINES KNOWN TO DIFFERENTIALLY PRODUCE TUMOR NECROSIS FACTOR (TNF)-ALPHA, BUT NOT INTERFERON (IFN)-GAMMA, IN RESPONSE TO BE ANTIGEN WERE CULTURED WITH BE OR CONTROLS. FOLLOWING CHALLENGES, ELISA WERE PERFORMED TO QUANTIFY INDUCED TNFALPHA AND IFNGAMMA EXPRESSION. BISULFATE-CONVERTED DNA WAS EVALUATED BY PYROSEQUENCING TO QUANTIFY CPG METHYLATION WITHIN THE PROMOTERS OF TNFALPHA AND IFNGAMMA. BE-CHALLENGED H36.12J CELLS EXPRESSED HIGHER LEVELS OF TNFALPHA COMPARED TO EITHER H36.12E CELLS OR P388D.1 CELLS. HOWEVER, THERE WERE NO VARIATIONS IN TNFALPHA PROMOTER CPG METHYLATION LEVELS BETWEEN CELL LINES AT THE SIX CPG SITES TESTED. H36.12J CELL TNFALPHA EXPRESSION WAS SHOWN TO BE METAL-SPECIFIC BY THE INDUCTION OF SIGNIFICANTLY MORE TNFALPHA WHEN EXPOSED TO BE THAN WHEN EXPOSED TO ALUMINUM SULFATE, OR NICKEL (II) CHLORIDE, BUT NOT WHEN EXPOSED TO COBALT (II) CHLORIDE. HOWEVER, H36.12J CELL METHYLATION LEVELS AT THE SIX CPG SITES EXAMINED IN THE TNFALPHA PROMOTER DID NOT CORRELATE WITH CYTOKINE EXPRESSION DIFFERENCES. NONETHELESS, ALL THREE CELL LINES HAD SIGNIFICANTLY MORE PROMOTER METHYLATION AT THE SIX CPG SITES INVESTIGATED WITHIN THE IFNGAMMA PROMOTER (A GENE THAT IS NOT EXPRESSED) WHEN COMPARED TO THE SIX CPG SITES INVESTIGATED IN THE TNFALPHA PROMOTER, REGARDLESS OF TREATMENT CONDITION (P < 1.17 X 10(-9)). THESE FINDINGS SUGGEST THAT, IN THIS CELL SYSTEM, PROMOTER HYPO-METHYLATION MAY BE NECESSARY TO ALLOW EXPRESSION OF METAL-INDUCED TNFALPHA AND THAT PROMOTER HYPER-METHYLATION IN THE IFNGAMMA PROMOTER MAY INTERFERE WITH EXPRESSION. ALSO, AT THE DOZEN CPG SITES INVESTIGATED IN THE PROMOTER REGIONS OF BOTH GENES, BERYLLIUM HAD NO IMPACT ON PROMOTER METHYLATION STATUS, DESPITE ITS ABILITY TO INDUCE PRO-INFLAMMATORY CYTOKINE EXPRESSION. 2016 19 1537 29 DNA METHYLATION IN ADOLESCENTS WITH ANXIETY DISORDER: A LONGITUDINAL STUDY. ANXIETY DISORDERS (AD) TYPICALLY MANIFEST IN CHILDREN AND ADOLESCENTS AND MIGHT PERSIST INTO ADULTHOOD. HOWEVER, THERE ARE STILL FEW DATA CONCERNING EPIGENETIC MECHANISMS ASSOCIATED WITH ONSET, PERSISTENCE OR REMISSION OF AD OVER TIME. WE INVESTIGATED A COHORT OF ADOLESCENTS AND YOUNG ADULTS AT BASELINE (AGE; 13.19 +/- 2.38) AND AFTER 5 YEARS AND CLASSIFIED THEM ACCORDING TO THE AD DIAGNOSIS AND THEIR LONGITUDINAL TRAJECTORIES INTO 4 GROUPS: (1) TYPICALLY DEVELOPING COMPARISONS (TDC; CONTROL GROUP, N = 14); (2) INCIDENT (AD IN THE SECOND EVALUATION ONLY, N = 11); (3) PERSISTENT (AD IN BOTH EVALUATIONS, N = 14) AND (4) REMITTENT (AD IN THE FIRST EVALUATION ONLY, N = 8). DNA METHYLATION WAS EVALUATED WITH THE INFINIUM HUMANMETHYLATION450 BEADCHIP FROM SALIVA SAMPLES COLLECTED AT BOTH EVALUATIONS. GENE SET ENRICHMENT ANALYSIS WAS APPLIED TO CONSIDER BIOLOGICAL PATHWAYS. WE FOUND DECREASED DNA METHYLATION IN TDC GROUP WHILE THE CHRONIC CASES OF AD PRESENTED HYPERMETHYLATION IN CENTRAL NERVOUS SYSTEM DEVELOPMENT PATHWAYS. MOREOVER, WE SHOWED THAT THIS PERSISTENT GROUP ALSO PRESENTED HYPERMETHYLATION WHILE THE OTHER THREE GROUPS WERE ASSOCIATED WITH HYPOMETHYLATION IN NERVOUS SYSTEM DEVELOPMENT PATHWAY. INCIDENCE AND REMISSION GROUPS WERE ASSOCIATED WITH INCREASED AND DECREASED METHYLATION IN NEURON DEVELOPMENT PATHWAYS, RESPECTIVELY. LARGER STUDIES ARE LIKELY TO DETECT SPECIFIC GENES RELEVANT TO AD. 2018 20 6083 39 THE EFFECT OF SMOKING ON DNA METHYLATION OF PERIPHERAL BLOOD MONONUCLEAR CELLS FROM AFRICAN AMERICAN WOMEN. BACKGROUND: REGULAR SMOKING IS ASSOCIATED WITH A WIDE VARIETY OF SYNDROMES WITH PROMINENT INFLAMMATORY COMPONENTS SUCH AS CANCER, OBESITY AND TYPE 2 DIABETES. HEAVY REGULAR SMOKING IS ALSO ASSOCIATED WITH CHANGES IN THE DNA METHYLATION OF PERIPHERAL MONONUCLEAR CELLS. HOWEVER, IN YOUNGER SMOKERS, INFLAMMATORY EPIGENETIC FINDINGS ARE LARGELY ABSENT WHICH SUGGESTS THE INFLAMMATORY RESPONSE(S) TO SMOKING MAY BE DOSE DEPENDENT. TO HELP UNDERSTAND WHETHER PERIPHERAL MONONUCLEAR CELLS HAVE A ROLE IN MEDIATING THESE RESPONSES IN OLDER SMOKERS WITH HIGHER CUMULATIVE SMOKE EXPOSURE, WE EXAMINED GENOME-WIDE DNA METHYLATION IN A GROUP OF WELL CHARACTERIZED ADULT AFRICAN AMERICAN SUBJECTS INFORMATIVE FOR SMOKING, AS WELL AS SERUM C-REACTIVE PROTEIN (CRP) AND INTERLEUKIN-6 RECEPTOR (IL6R) LEVELS. IN ADDITION, COMPLEMENTARY BIOINFORMATIC ANALYSES WERE CONDUCTED TO DELINEATE POSSIBLE PATHWAYS AFFECTED BY LONG-TERM SMOKING. RESULTS: GENOME-WIDE DNA METHYLATION ANALYSIS WITH RESPECT TO SMOKING STATUS YIELDED 910 SIGNIFICANT LOCI AFTER BENJAMINI-HOCHBERG CORRECTION. IN PARTICULAR, TWO LOCI FROM THE AHRR GENE (CG05575921 AND CG23576855) AND ONE LOCUS FROM THE GPR15 GENE (CG19859270) WERE IDENTIFIED AS HIGHLY SIGNIFICANTLY DIFFERENTIALLY METHYLATED BETWEEN SMOKERS AND NON-SMOKERS. THE BIOINFORMATIC ANALYSES SHOWED THAT LONG-TERM CHRONIC SMOKING IS ASSOCIATED WITH ALTERED PROMOTER DNA METHYLATION OF GENES CODING FOR PROTEINS MAPPING TO CRITICAL SUB-NETWORKS MODERATING INFLAMMATION, IMMUNE FUNCTION, AND COAGULATION. CONCLUSIONS: WE CONCLUDE THAT CHRONIC REGULAR SMOKING IS ASSOCIATED WITH CHANGES IN PERIPHERAL MONONUCLEAR CELL METHYLATION SIGNATURE WHICH PERTURB INFLAMMATORY AND IMMUNE FUNCTION PATHWAYS AND MAY CONTRIBUTE TO INCREASED VULNERABILITY FOR COMPLEX ILLNESSES WITH INFLAMMATORY COMPONENTS. 2014