1 3143 162 GLOBAL MIRNA/PROTEOMIC ANALYSES IDENTIFY MIRNAS AT 14Q32 AND 3P21, WHICH CONTRIBUTE TO FEATURES OF CHRONIC IRON-EXPOSED FALLOPIAN TUBE EPITHELIAL CELLS. MALIGNANT TRANSFORMATION OF FALLOPIAN TUBE SECRETORY EPITHELIAL CELLS (FTSECS) IS A KEY CONTRIBUTING EVENT TO THE DEVELOPMENT OF HIGH-GRADE SEROUS OVARIAN CARCINOMA (HGSOC). OUR RECENT FINDINGS IMPLICATE ONCOGENIC TRANSFORMATIVE EVENTS IN CHRONIC IRON-EXPOSED FTSECS, INCLUDING INCREASED EXPRESSION OF ONCOGENIC MEDIATORS, INCREASED TELOMERASE TRANSCRIPTS, AND INCREASED GROWTH/MIGRATORY POTENTIAL. HEREIN, WE EXTEND THESE STUDIES BY IMPLEMENTING AN INTEGRATED TRANSCRIPTOMIC AND MASS SPECTROMETRY-BASED PROTEOMICS APPROACH TO IDENTIFY GLOBAL MIRNA AND PROTEIN ALTERATIONS, FOR WHICH WE ALSO INVESTIGATE A SUBSET OF THESE TARGETS TO IRON-INDUCED FUNCTIONAL ALTERATIONS. PROTEOMIC ANALYSIS IDENTIFIED > 4500 PROTEINS, OF WHICH 243 TARGETS WERE DIFFERENTIALLY EXPRESSED. SIXTY-FIVE DIFFERENTIALLY EXPRESSED MIRNAS WERE IDENTIFIED, OF WHICH 35 WERE ASSOCIATED WITH THE "TOP" PROTEOMIC MOLECULES (> FOURFOLD CHANGE) IDENTIFIED BY INGENUITY PATHWAY ANALYSIS. TWENTY OF THESE 35 MIRNAS ARE AT THE 14Q32 LOCUS (ENCODING A CLUSTER OF 54 MIRNAS) WITH POTENTIAL TO BE REGULATED BY DNA METHYLATION AND HISTONE DEACETYLATION. AT 14Q32, MIR-432-5P AND MIR-127-3P WERE ~ 100-FOLD DOWNREGULATED WHEREAS MIR-138-5P WAS 16-FOLD DOWNREGULATED AT 3P21 IN CHRONIC IRON-EXPOSED FTSECS. COMBINATORIAL TREATMENT WITH METHYLTRANSFERASE AND DEACETYLATION INHIBITORS REVERSED EXPRESSION OF THESE MIRNAS, SUGGESTING CHRONIC IRON EXPOSURE ALTERS MIRNA EXPRESSION VIA EPIGENETIC ALTERATIONS. IN ADDITION, PAX8, AN IMPORTANT TARGET IN HGSOC AND A POTENTIAL MIRNA TARGET (FROM IPA) WAS EPIGENETICALLY DEREGULATED IN IRON-EXPOSED FTSECS. HOWEVER, BOTH PAX8 AND ALDH1A2 (ANOTHER IPA-PREDICTED TARGET) WERE EXPERIMENTALLY IDENTIFIED TO BE INDEPENDENTLY REGULATED BY THESE MIRNAS ALTHOUGH TERT RNA WAS PARTIALLY REGULATED BY MIR-138-5P. INTERESTINGLY, OVEREXPRESSION OF MIR-432-5P DIMINISHED CELL NUMBERS INDUCED BY LONG-TERM IRON EXPOSURE IN FTSECS. COLLECTIVELY, OUR GLOBAL PROFILING APPROACHES UNCOVERED PATTERNS OF MIRNA AND PROTEOMIC ALTERATIONS THAT MAY BE REGULATED BY GENOME-WIDE EPIGENETIC ALTERATIONS AND CONTRIBUTE TO FUNCTIONAL ALTERATIONS INDUCED BY CHRONIC IRON EXPOSURE IN FTSECS. THIS STUDY MAY PROVIDE A PLATFORM TO IDENTIFY FUTURE BIOMARKERS FOR EARLY OVARIAN CANCER DETECTION AND NEW TARGETS FOR THERAPY. 2021 2 1892 17 ENDOMETRIOSIS: EPIDEMIOLOGY, CLASSIFICATION, PATHOGENESIS, TREATMENT AND GENETICS (REVIEW OF LITERATURE). ENDOMETRIOSIS IS A "MYSTERIOUS" DISEASE AND ITS EXACT CAUSE HAS NOT YET BEEN ESTABLISHED. AMONG THE ETIOLOGICAL FACTORS, CONGENITAL, ENVIRONMENTAL, EPIGENETIC, AUTOIMMUNE AND ALLERGIC FACTORS ARE LISTED. IT IS BELIEVED THAT THE PRIMARY MECHANISM OF THE FORMATION OF ENDOMETRIOSIS FOCI IS RETROGRADE MENSTRUATION, I.E., THE PASSAGE OF MENSTRUAL BLOOD THROUGH THE FALLOPIAN TUBES INTO THE PERITONEAL CAVITY AND IMPLANTATION OF EXFOLIATED ENDOMETRIAL CELLS. HOWEVER, SINCE THIS MECHANISM IS ALSO OBSERVED IN HEALTHY WOMEN, OTHER FACTORS MUST ALSO BE INVOLVED IN THE FORMATION OF ENDOMETRIOSIS FOCI. ENDOMETRIOSIS IS IN MANY WOMEN THE CAUSE OF INFERTILITY, CHRONIC PAIN AND THE DETERIORATION OF THE QUALITY OF LIFE. IT ALSO REPRESENTS A SIGNIFICANT FINANCIAL BURDEN ON HEALTH SYSTEMS. THE ARTICLE PRESENTS A REVIEW OF THE LITERATURE ON ENDOMETRIOSIS-A DISEASE AFFECTING WOMEN THROUGHOUT THE WORLD. 2021 3 3605 41 IMPROVING PARP INHIBITOR EFFICACY IN HIGH-GRADE SEROUS OVARIAN CARCINOMA: A FOCUS ON THE IMMUNE SYSTEM. HIGH-GRADE SEROUS OVARIAN CARCINOMA (HGSOC) IS A GENOMICALLY UNSTABLE MALIGNANCY RESPONSIBLE FOR OVER 70% OF ALL DEATHS DUE TO OVARIAN CANCER. WITH ROUGHLY 50% OF ALL HGSOC HARBORING DEFECTS IN THE HOMOLOGOUS RECOMBINATION (HR) DNA REPAIR PATHWAY (E.G., BRCA1/2 MUTATIONS), THE INTRODUCTION OF POLY ADP-RIBOSE POLYMERASE INHIBITORS (PARPI) HAS DRAMATICALLY IMPROVED OUTCOMES FOR WOMEN WITH HR DEFECTIVE HGSOC. BY BLOCKING THE REPAIR OF SINGLE-STRANDED DNA DAMAGE IN CANCER CELLS ALREADY LACKING HIGH-FIDELITY HR PATHWAYS, PARPI CAUSES THE ACCUMULATION OF DOUBLE-STRANDED DNA BREAKS, LEADING TO CELL DEATH. THUS, THIS SYNTHETIC LETHALITY RESULTS IN PARPI SELECTIVELY TARGETING CANCER CELLS, RESULTING IN IMPRESSIVE EFFICACY. DESPITE THIS, RESISTANCE TO PARPI COMMONLY DEVELOPS THROUGH DIVERSE MECHANISMS, SUCH AS THE ACQUISITION OF SECONDARY BRCA1/2 MUTATIONS. PERHAPS LESS WELL DOCUMENTED IS THAT PARPI CAN IMPACT BOTH THE TUMOUR MICROENVIRONMENT AND THE IMMUNE RESPONSE, THROUGH UPREGULATION OF THE STIMULATOR OF INTERFERON GENES (STING) PATHWAY, UPREGULATION OF IMMUNE CHECKPOINTS SUCH AS PD-L1, AND BY STIMULATING THE PRODUCTION OF PRO-INFLAMMATORY CYTOKINES. WHILST TARGETED IMMUNOTHERAPIES HAVE NOT YET FOUND THEIR PLACE IN THE CLINIC FOR HGSOC, THE EVIDENCE ABOVE, AS WELL AS ONGOING STUDIES EXPLORING THE SYNERGISTIC EFFECTS OF PARPI WITH IMMUNE AGENTS, INCLUDING IMMUNE CHECKPOINT INHIBITORS, SUGGESTS POTENTIAL FOR TARGETING THE IMMUNE RESPONSE IN HGSOC. ADDITIONALLY, COMBINING PARPI WITH EPIGENETIC-MODULATING DRUGS MAY IMPROVE PARPI EFFICACY, BY INDUCING A BRCA-DEFECTIVE PHENOTYPE TO SENSITISE RESISTANT CANCER CELLS TO PARPI. FINALLY, INVIGORATING AN IMMUNE RESPONSE DURING PARPI THERAPY MAY ENGAGE ANTI-CANCER IMMUNE RESPONSES THAT POTENTIATE EFFICACY AND MITIGATE THE DEVELOPMENT OF PARPI RESISTANCE. HERE, WE WILL REVIEW THE EMERGING PARPI LITERATURE WITH A FOCUS ON PARPI EFFECTS ON THE IMMUNE RESPONSE IN HGSOC, AS WELL AS THE POTENTIAL OF EPIGENETIC COMBINATION THERAPIES. WE HIGHLIGHT THE POTENTIAL OF TRANSFORMING HGSOC FROM A LETHAL TO A CHRONIC DISEASE AND INCREASING THE LIKELIHOOD OF CURE. 2022 4 3999 33 LOSS OF HDAC3 RESULTS IN NONRECEPTIVE ENDOMETRIUM AND FEMALE INFERTILITY. ENDOMETRIOSIS IS A DISEASE IN WHICH TISSUE THAT NORMALLY GROWS INSIDE THE UTERUS GROWS OUTSIDE THE UTERUS AND CAUSES CHRONIC PELVIC PAIN AND INFERTILITY. HOWEVER, THE EXACT MECHANISMS OF THE PATHOGENESIS OF ENDOMETRIOSIS-ASSOCIATED INFERTILITY ARE UNKNOWN. EPIGENETIC DYSREGULATION HAS RECENTLY BEEN IMPLICATED IN INFERTILITY. HERE, WE REPORT A REDUCTION OF HISTONE DEACETYLASE 3 (HDAC3) PROTEIN AMOUNTS IN EUTOPIC ENDOMETRIUM OF INFERTILE WOMEN WITH ENDOMETRIOSIS COMPARED TO A CONTROL GROUP. TO INVESTIGATE THE EFFECT OF HDAC3 LOSS IN THE UTERUS, WE GENERATED MICE WITH CONDITIONAL ABLATION OF HDAC3 IN PROGESTERONE RECEPTOR (PGR)-POSITIVE CELLS (PGR(CRE/+)HDAC3(F/F) ; HDAC3(D/D) ). LOSS OF HDAC3 IN THE UTERUS OF MICE RESULTS IN INFERTILITY DUE TO IMPLANTATION FAILURE AND DECIDUALIZATION DEFECT. EXPRESSION MICROARRAY AND CHIP-SEQ ANALYSES IDENTIFIED COL1A1 AND COL1A2 AS DIRECT TARGETS OF HDAC3 IN BOTH MICE AND HUMANS. REDUCTION OF HDAC3 ABROGATED DECIDUALIZATION IN A PRIMARY CULTURE OF HUMAN ENDOMETRIAL STROMAL CELLS (HESCS) SIMILAR TO THAT OBSERVED IN INFERTILE PATIENTS WITH ENDOMETRIOSIS. WHEREAS ATTENUATION OF HDAC3 RESULTED IN P300 RECRUITMENT TO COL1A1 AND COL1A2 GENES IN THE UTERUS OF MICE AS WELL AS HESCS, INHIBITION OF P300 PERMITTED HESCS TO UNDERGO DECIDUALIZATION. COLLECTIVELY, WE FOUND ATTENUATION OF HDAC3 AND OVEREXPRESSION OF COLLAGEN TYPE I IN THE EUTOPIC ENDOMETRIUM OF INFERTILE PATIENTS WITH ENDOMETRIOSIS. HDAC3 LOSS CAUSED A DEFECT OF DECIDUALIZATION THROUGH THE ABERRANT TRANSCRIPTIONAL ACTIVATION OF COL1A1 AND COL1A2 GENES IN MICE AND COL1A1 AND COL1A2 GENES IN HUMANS. OUR RESULTS SUGGEST THAT HDAC3 IS CRITICAL FOR ENDOMETRIAL RECEPTIVITY AND DECIDUALIZATION. 2019 5 2942 33 GENETIC AND EPIGENETIC ALTERATIONS OF LTF AT 3P21.3 IN NASOPHARYNGEAL CARCINOMA. TO INVESTIGATE THE ROLES OF LACTOTRANSFERRIN GENE (LTF, ALSO REFERRED TO AS THE LACTOFERRIN GENE, LF), LOCATED AT 3P21.3 WITHIN THE COMMON MINIMAL DELETION REGION, IN THE PATHOGENESIS OF NASOPHARYNGEAL CARCINOMA (NPC), WE FIRST DETECTED ITS EXPRESSION LEVEL IN 33 PRIMARY NPC TISSUES AND 15 CHRONIC NASOPHARYNGITIS TISSUES. ABSENT EXPRESSION OR DOWNREGULATION OF LTF WERE OBSERVED IN 76% (25 OF 33) OF PRIMARY NPC TISSUES. WE FURTHER FOUND THAT 25% (5 OF 20) OF NPC SPECIMENS HAD LOSS OF HETEROZYGOSITY (LOH) AT THE LTF LOCUS. LTF MUTATION ASSESSED BY POLYMERASE CHAIN REACTION SINGLE-STRAND CONFORMATION POLYMORPHISM (PCR-SSCP) AND DNA SEQUENCING WAS NOTED IN 30% (6 OF 20) OF PRIMARY NPC TISSUES. IN ADDITION, HYPER-METHYLATION OF LTF PROMOTER REGION WAS FOUND IN 63.6% (21 OF 33) OF PRIMARY NPC SAMPLES BUT NOT IN CHRONIC NASOPHARYNGITIS TISSUES. THE LTF TRANSCRIPTS IN NPC CELL LINES INCREASED UPON TREATMENT WITH THE DEMETHYLATION COMPOUND, 5-AZA-2-DEOXYCYTIDINE. IN CONCLUSION, OUR DATA INDICATE THAT TWO-HIT SILENCING OF LTF THROUGH GENETIC AND EPIGENETIC CHANGES MAY BE A COMMON AND IMPORTANT EVENT IN THE CARCINOGENESIS OF NPC. 2006 6 6816 33 [EXPRESSION, GENETIC AND EPIGENETIC ALTERATIONS OF LTF GENE IN NASOPHARYNGEAL CARCINOMA CELL LINES]. OBJECTIVE: TO INVESTIGATE THE EXPRESSION OF LTF MRNA IN SEVERAL NASOPHARYNGEAL CANCER (NPC) CELL LINES, AND ANALYZE THE RELATIONSHIP BETWEEN THE GENETIC AND EPIGENETIC CHANGES AND EXPRESSION OF LTF GENE. METHODS: THE EXPRESSION LEVEL OF LTF WAS DETECTED IN NPC CELL LINES HNE1, HNE2, HNE3, CNE1, CNE2, 5-8F, 6-10B CELLS AND TISSUES OF 15 CASES OF CHRONIC NASOPHARYNGITIS BY RT-PCR. THE LTF PROTEIN LEVEL WAS ANALYZED BY WESTERN BLOTTING IN 6-10B CELLS. THEN LOH, MUTATION AND METHYLATION STATUS OF LTF WAS EXAMINED BY MICROSATELLITES ANALYSIS, PCR-SSCP, MSP AND BISULFITE GENOMIC SEQUENCING, RESPECTIVELY. RESULTS: 15 CHRONIC NASOPHARYNGITIS TISSUES SHOWED STABLE LTF EXPRESSION, WHILE THERE WERE WEAK EXPRESSION IN 6-10B CELLS AND ABSENT EXPRESSION IN REMAINING DETECTED NPC CELL LINES. THERE WAS A SIGNIFICANTLY LOWER LTF EXPRESSION IN CHRONIC NASOPHARYNGITIS TISSUES (Z = -3.738, P = 0.000). NO LTF PROTEIN EXPRESSION WAS OBSERVED IN 6-10B CELLS. LOH ANALYSIS DEMONSTRATED THAT ALLELE LOSS OF LTF WASN'T FOUND IN NPC CELL LINES. LTF MUTATION WAS NOTED IN 14.3% (1/7) OF NPC CELL LINES. DNA SEQUENCING CONFIRMED THE MUTATION POINT IN THE PROMOTER REGION (-305 BP TO -50 BP) WAS AT -218 BP (DEL T) OF LTF GENE IN THE HNE1 CELL LINE. METHYLATION OF LTF GENE WAS NOT FOUND IN CHRONIC NASOPHARYNGITIS. HOWEVER, METHYLATION OF LTF PROMOTER WAS DETECTED IN ALL NPC CELL LINES. LTF MRNA EXPRESSION WAS INCREASED IN 5-8F AND 6-10B CELL LINES AFTER TREATMENT WITH 5-AZA-2-DEOXYCYTIDINE. CONCLUSION: THERE IS AN INACTIVATION OF EXPRESSION OF LTF GENE IN THE NPC CELL LINES. ITS MOLECULAR MECHANISM MAY BE RELATED WITH METHYLATION OF PROMOTER REGION AND DELETION MUTATION. 2010 7 4260 30 METTL3-MEDIATED M6A MODIFICATION OF SIRT1 MRNA INHIBITS PROGRESSION OF ENDOMETRIOSIS BY CELLULAR SENESCENCE ENHANCING. BACKGROUND: ENDOMETRIOSIS (EMS), THE ECTOPIC PLANTING OF FUNCTIONAL ENDOMETRIUM OUTSIDE OF THE UTERUS, IS A LEADING CAUSE OF INFERTILITY AND PELVIC PAIN. AS A FUNDAMENTAL MRNA MODIFICATION, N6-METHYLADENOSINE (M6A) PARTICIPATES IN VARIOUS PATHOLOGICAL PROCESSES. HOWEVER, THE ROLE OF M6A RNA MODIFICATION IN ENDOMETRIOSIS REMAINS UNCLEAR. THE PRESENT STUDY EXPLORES METTL3-MEDIATED M6A MODIFICATION AND THE MECHANISMS INVOLVED IN ENDOMETRIOSIS. METHODS: THE DOMINANT M6A REGULATORS IN EMS WERE ANALYSED USING RT?PCR. CANDIDATE TARGETS AND POSSIBLE MECHANISMS OF METTL3 WERE ASSESSED BY M6A-MRNA EPITRANSCRIPTOMIC MICROARRAY AND RNA SEQUENCING. A PRIMARY ESCS MODEL WAS EMPLOYED TO VERIFY THE EFFECT OF METTL3 ON M6A MODIFICATION OF SIRT1 MRNA, AND THE MECHANISM WAS ELUCIDATED BY RT?PCR, WESTERN BLOTTING, MERIP, AND RIP ASSAYS. CCK-8 VIABILITY ASSAYS, TRANSWELL INVASION ASSAYS, EDU PROLIFERATION ASSAYS, WOUND HEALING MIGRATION ASSAYS, AND SENESCENCE-ASSOCIATED BETA-GALACTOSIDASE STAINING WERE PERFORMED TO ILLUMINATE THE POTENTIAL BIOLOGICAL MECHANISM OF METTL3 AND SIRT1 IN ESCS IN VITRO. AN IN VIVO PGRCRE/ + METTL3 -/- FEMALE HOMOZYGOUS MOUSE MODEL AND A NUDE MOUSE XENOGRAFT MODEL WERE EMPLOYED TO FURTHER INVESTIGATE THE PHYSIOLOGIC CONSEQUENCES OF METTL3-MEDIATED M6A ALTERATION ON EMS. RESULTS: OUR DATA SHOW THAT DECREASED METTL3 EXPRESSION SIGNIFICANTLY DOWNREGULATES M6A RNA METHYLATION LEVELS IN ESCS. SILENCING M6A MODIFICATIONS MEDIATED BY METTL3 ACCELERATES ESCS VIABILITY, PROLIFERATION, MIGRATION, AND INVASION IN VITRO. THE M6A READER PROTEIN YTHDF2 BINDS TO M6A MODIFICATIONS TO INDUCE THE DEGRADATION OF SIRT1 MRNA. SIRT1/FOXO3A SIGNALLING PATHWAY ACTIVATION IS SUBSEQUENTLY INHIBITED, PROMOTING THE CELLULAR SENESCENCE OF ESCS AND INHIBITING THE ECTOPIC IMPLANTATION OF ESCS IN VITRO AND IN VIVO. CONCLUSIONS: OUR FINDINGS DEMONSTRATE THAT METTL3-MEDIATED M6A METHYLATION EPIGENETICALLY REGULATES THE ECTOPIC IMPLANTATION OF ESCS, RESULTING IN THE PROGRESSION OF ENDOMETRIOSIS. OUR STUDY ESTABLISHES METTL3-YTHDF2-SIRT1/FOXO3A AS A CRITICAL AXIS AND POTENTIAL MECHANISM IN ENDOMETRIOSIS. 2023 8 3627 38 INACTIVATION OF LARS2, LOCATED AT THE COMMONLY DELETED REGION 3P21.3, BY BOTH EPIGENETIC AND GENETIC MECHANISMS IN NASOPHARYNGEAL CARCINOMA. ALLELIC LOSS OF CHROMOSOME 3P, INCLUDING THE 3P21.3 REGION, IS FOUND IN 95-100% OF PRIMARY NASOPHARYNGEAL CARCINOMA (NPC) BIOPSIES, SUGGESTING THAT THIS REGION SHOULD HARBOR SOME TUMOR SUPPRESSOR GENES (TSGS) CLOSELY RELATED TO NPC DEVELOPMENT. SEVERAL TSGS LOCATED AT 3P21.3, SUCH AS RASSF1A, LTF AND BLU, HAVE BEEN DEMONSTRATED TO BE INVOLVED IN NPC DEVELOPMENT. LARS2 (LEUCYL-TRNA SYNTHETASE 2, MITOCHONDRIAL) IS ANOTHER GENE LOCATED IN THE CHROMOSOME 3 COMMON ELIMINATED REGION-1 (C3CER1) AT 3P21.3. IN THIS STUDY, WE FOCUSSED ON THE EPIGENETIC AND GENETIC ALTERATIONS OF LARS2 IN NPC. THE MRNA EXPRESSION OF LARS2 WAS DETECTED IN 36 NPC AND 8 CHRONIC NASOPHARYNGITIS (NP) TISSUES BY SEMI-QUANTITATIVE REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION (RT-PCR) AND REAL-TIME RT-PCR. SUBSEQUENTLY, THE MUTATION, ALLELIC LOSS, AND METHYLATION STATUS OF LARS2 WERE ANALYSED BY POLYMERASE CHAIN REACTION-SINGLE-STRAND CONFORMATION POLYMORPHISM (PCR-SSCP), HOMOZYGOUS DELETION (HD) ANALYSIS AND METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION IN PRIMARY NPC TISSUES. NO EXPRESSION OR DOWNREGULATION OF LARS2 WAS OBSERVED IN 78% OF PRIMARY NPC TISSUES. NO MUTATIONS, ASSESSED BY PCR-SSCP AND DNA SEQUENCING, WERE FOUND IN THE PROMOTER REGION AND EXON 1 OF LARS2 IN NPC TISSUES, WHEREAS HD WAS DETECTED IN 28% OF NPC SPECIMENS AT THE LARS2 LOCUS. IN ADDITION, HYPERMETHYLATION OF LARS2 WAS FOUND IN 64% OF NPC SAMPLES BUT ONLY IN 12.5% OF NP BIOPSIES. OUR DATA INDICATE THAT INACTIVATION OF LARS2 BY BOTH GENETIC AND EPIGENETIC MECHANISMS MAY BE A COMMON AND IMPORTANT EVENT IN THE CARCINOGENESIS OF NPC. 2009 9 3165 37 GRIK1-AS1 DEFICIENCY ACCELERATES ENDOMETRIOSIS PROGRESSION BY BOOSTING DNMT1-DEPENDENT SFRP1 PROMOTER METHYLATION IN ENDOMETRIAL STROMAL CELLS. BACKGROUND: ENDOMETRIOSIS, A GYNECOLOGICAL DISEASE THAT AFFECTS UP TO 10% OF WOMEN, IS A MAJOR CAUSE OF PAIN AND INFERTILITY. DEREGULATION OF THE EPIGENOME IS ACCOUNTABLE FOR THE ONSET AND PROGRESSION OF ENDOMETRIOSIS, ALTHOUGH ITS EXACT MECHANISM IS UNKNOWN. THE PURPOSE OF THE CURRENT STUDY IS TO EXAMINE THE ROLE OF THE LONG NON-CODING RNA (LNCRNA) GRIK1-AS1 IN THE EPIGENETIC REGULATION OF ENDOMETRIAL STROMAL CELL PROLIFERATION AND THE DEVELOPMENT OF ENDOMETRIOSIS. METHODS: ENDOMETRIOSIS DATASETS WERE SCREENED TO IDENTIFY GRIKI-AS1 AS DRAMATICALLY DECLINING IN ENDOMETRIOSIS. GAIN OR LOSS OF FUNCTION ENDOMETRIAL STROMAL CELL (ESC) MODELS WERE ESTABLISHED. THE ANTI-PROLIFERATION PHENOTYPE WAS INVESTIGATED USING IN VITRO AND IN VIVO EXPERIMENTS. EPIGENETIC REGULATORY NETWORK ANALYSES WERE CONDUCTED TO SUGGEST THE INTRINSIC MOLECULAR MECHANISM. RESULTS: WITH BIOINFORMATIC AND CLINICAL DATA, WE OBSERVED THAT GRIK1-AS1 AND SFRP1 WERE EXPRESSED AT LOW LEVELS IN ENDOMETRIOSIS. OVEREXPRESSED GRIK1-AS1 INHIBITED ESC PROLIFERATION, WHILE SFRP1 KNOCKDOWN RESCUED THE ANTIPROLIFERATIVE ABILITY OF GRIK1-AS1. SPECIFICALLY, METHYLATION-DEPENDENT EXPRESSION INHIBITION OF SFRP1 WAS REVEALED IN ESCS. MECHANISTICALLY, GRIK1-AS1 HAMPERS THE OCCUPANCY OF DNMT1 IN SRFP1 PROMOTER, LEADING TO HYPOMETHYLATION OF SFRP1 AND UPREGULATED SFRP1 EXPRESSION, THEREBY POTENTIALLY SUPPRESSING WNT SIGNALING AND ITS ADVERSE PROLIFERATIVE EFFECT. THERAPEUTICALLY, LENTIVIRUS-MEDIATED UPREGULATION OF GRIK1-AS1 INHIBITED ENDOMETRIOSIS DISEASE PROGRESSION IN VIVO. CONCLUSIONS: OUR STUDY IS A PROOF-OF-CONCEPT DEMONSTRATION FOR GRIKI-AS1-ASSOCIATED ENDOMETRIOSIS PATHOGENESIS AND HIGHLIGHTS A POTENTIAL INTERVENTION TARGET. 2023 10 1996 31 EPIGENETIC AND GENETIC ALTERATIONS OF THE EDNRB GENE IN NASOPHARYNGEAL CARCINOMA. BACKGROUND: LOSS OF HETEROZYGOSITY (LOH) AT 13Q22 IS A COMMON EVENT IN NASOPHARYNGEAL CARCINOMA (NPC). EDNRB GENE LOCATED AT 13Q22 HAS BEEN DEMONSTRATED TO BE HYPERMETHYLATED IN SOME KINDS OF TUMORS. IN THE CURRENT STUDY, WE FOCUSED ON THE EPIGENETIC AND GENETIC ALTERATIONS OF EDNRB IN NPC. METHODS: THE MRNA EXPRESSION OF EDNRB WAS DETECTED BY SEMIQUANTITATIVE RT-PCR AND REAL-TIME QUANTITATIVE PCR IN 49 NPC AND 12 CHRONIC NASOPHARYNGITIS BIOPSIES. THE METHYLATION AND LOH STATUS OF EDNRB WERE EXAMINED BY METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION, MICROSATELLITE PCR AND SEQUENCING. WE ALSO EXAMINED THE MRNA EXPRESSION OF EDNRB IN FOUR NPC CELL LINES AFTER 5-AZA-2'-DEOXYCYTIDINE TREATMENT. RESULTS: EDNRB WAS DOWNREGULATED IN PRIMARY NPC TISSUES AND NPC CELL LINES, AND A RELATIVELY HIGHER METHYLATION LEVEL OF EDNRB WAS FOUND IN NPC BIOPSIES (84%) COMPARED TO THAT IN CHRONIC NASOPHARYNGITIS BIOPSIES (42%). TREATMENT OF NPC CELL LINES WITH 5-AZA-2'-DEOXYCYTIDINE ACTIVATED EDNRB EXPRESSION. LOH OF EDNRB GENE WAS ALSO FOUND AT TWO MICROSATELLITE SITES WITH RATIOS OF 6.25 AND 16.67% IN NPC. CONCLUSION: OUR RESULTS SUGGESTED THAT EDNRB EXPRESSION MAY BE AFFECTED BY ABERRANT PROMOTER METHYLATION AND GENE DELETION AND MAY PLAY A ROLE IN THE DEVELOPMENT OF NPC. 2007 11 4698 31 NFATC2-DEPENDENT EPIGENETIC DOWNREGULATION OF THE TSC2/BECLIN-1 PATHWAY IS INVOLVED IN NEUROPATHIC PAIN INDUCED BY OXALIPLATIN. NEUROPATHIC PAIN IS A COMMON DOSE-LIMITING SIDE EFFECT OF OXALIPLATIN, WHICH HAMPERS THE EFFECTIVE TREATMENT OF TUMORS. HERE, WE FOUND THAT UPREGULATION OF TRANSCRIPTION FACTOR NFATC2 DECREASED THE EXPRESSION OF BECLIN-1, A CRITICAL MOLECULE IN AUTOPHAGY, IN THE SPINAL DORSAL HORN, AND CONTRIBUTED TO NEUROPATHIC PAIN FOLLOWING OXALIPLATIN TREATMENT. MEANWHILE, MANIPULATING AUTOPHAGY LEVELS BY INTRATHECAL INJECTION OF RAPAMYCIN (RAPA) OR 3-METHYLADENINE (3-MA) DIFFERENTIALLY ALTERED MECHANICAL ALLODYNIA IN OXALIPLATIN-TREATED OR NAIVE RATS. UTILIZING CHROMATIN IMMUNOPRECIPITATION-SEQUENCING (CHIP-SEQ) ASSAY COMBINED WITH BIOINFORMATICS ANALYSIS, WE FOUND THAT NFATC2 NEGATIVELY REGULATED THE TRANSCRIPTION OF TUBEROUS SCLEROSIS COMPLEX PROTEIN 2 (TSC2), WHICH CONTRIBUTED TO THE OXALIPLATIN-INDUCED BECLIN-1 DOWNREGULATION. FURTHER ASSAYS REVEALED THAT NFATC2 REGULATED HISTONE H4 ACETYLATION AND METHYLATION IN THE TSC2 PROMOTER SITE 1 IN RATS' DORSAL HORNS WITH OXALIPLATIN TREATMENT. THESE RESULTS SUGGESTED THAT NFATC2 MEDIATED THE EPIGENETIC DOWNREGULATION OF THE TSC2/BECLIN-1 AUTOPHAGY PATHWAY AND CONTRIBUTED TO OXALIPLATIN-INDUCED MECHANICAL ALLODYNIA, WHICH PROVIDED A NEW THERAPEUTIC INSIGHT FOR CHEMOTHERAPY-INDUCED NEUROPATHIC PAIN. 2023 12 5388 40 REDOX REGULATION OF MICRORNAS IN ENDOMETRIOSIS-ASSOCIATED PAIN. ENDOMETRIOSIS IS A CHRONIC, PAINFUL CONDITION WITH UNKNOWN ETIOLOGY. A DIFFERENTIAL EXPRESSION OF MICRORNAS IN THE ENDOMETRIOTIC TISSUES FROM WOMEN WITH ENDOMETRIOSIS WITH PAIN COMPARED TO THOSE WITHOUT SUGGESTED A PLAUSIBLE ROLE FOR MIRNA OR EPIGENETIC MECHANISMS IN THE ETIOLOGY OF ENDOMETRIOTIC PAIN. THE PERITONEAL MILIEU IS INVOLVED IN MAINTENANCE OF ENDOMETRIOTIC LESION AND NOCICEPTION. WE RECENTLY SHOWED THE MECHANISTIC ROLE FOR OXIDIZED-LIPOPROTEINS (OX-LDLS) PRESENT IN PERITONEAL FLUID (PF) IN ENDOMETRIOSIS AND PAIN. WE EXPLORED THE POSSIBILITY OF OX-LDLS MODULATING THE EXPRESSION OF MIRNAS IN A MANNER SIMILAR TO PF FROM WOMEN WITH ENDOMETRIOSIS. EXPRESSION LEVELS OF MIRNAS AND THEIR PREDICTED NOCICEPTIVE AND INFLAMMATORY TARGETS WERE DETERMINED IN PF AND OX-LDL TREATED HUMAN ENDOMETRIAL CELL-LINES. SAMPLES FROM IRB-APPROVED AND CONSENTED PATIENTS WITH AND WITHOUT ENDOMETRIOSIS OR PAIN WERE USED. THESE WERE COMPARED TO ENDOMETRIAL CELL-LINES TREATED WITH VARIOUS FORMS OF OXIDIZED-LIPOPROTEINS. RNA (INCLUDING MIRNAS) WERE ISOLATED FROM TREATED ENDOMETRIAL CELLS AND EXPRESSION LEVELS WERE DETERMINED USING COMMERCIAL MIRNOME ARRAYS. CELL LYSATES WERE USED IN IMMUNOBLOTTING FOR INFLAMMATORY PROTEINS USING A PROTEIN ARRAY. TWENTY MIRNAS INCLUDING ISOFORMS OF MIR-29, MIR-181 AND LET-7 WERE MUTUALLY DIFFERENTIALLY EXPRESSED IN CELLS TREATED WITH PF FROM ENDOMETRIOSIS PATIENTS WITH PAIN AND THOSE TREATED WITH OX-LDL COMPONENTS. THE OX-LDLS AND ENDO-PF TREATMENT ALSO PRODUCED SIGNIFICANT OVEREXPRESSION OF MICRORNA PREDICTED TARGET GENES NERVE GROWTH FACTOR, INTERLEUKIN-6 AND PROSTAGLANDIN E SYNTHASE AND OVEREXPRESSION OF THEIR DOWNSTREAM PROTEIN TARGETS MIP1ALPHA AND MCP1. THIS STUDY SHOWED SIMILARITIES BETWEEN MIRNA REGULATION IN PF FROM ENDOMETRIOTIC WOMEN AND OX-LDLS PRESENT IN ABUNDANCE IN THE PF OF THESE WOMEN. KEY MIRNAS RESPONSIBLE FOR TARGETING NOCICEPTIVE AND INFLAMMATORY MOLECULES WERE DOWNREGULATED IN THE PRESENCE OF OX-LDLS AND ENDO-PF, THUS PLAYING A ROLE IN THE ETIOLOGY OF ENDOMETRIOTIC PAIN. THESE REDOX-SENSITIVE MIRNAS CAN BE OF POTENTIAL USE AS TARGETS IN THE TREATMENT OF ENDOMETRIOSIS-ASSOCIATED PAIN. 2017 13 3755 33 INTEGRATED BIOINFORMATICS ANALYSIS UNCOVERS CHARACTERISTIC GENES AND MOLECULAR SUBTYPING SYSTEM FOR ENDOMETRIOSIS. OBJECTIVE: ENDOMETRIOSIS IS A CHRONIC INFLAMMATORY ESTROGEN-DEPENDENT DISEASE WITH THE GROWTH OF ENDOMETRIAL TISSUES OUTSIDE THE UTERINE CAVITY. NEVERTHELESS, THE ETIOLOGY OF ENDOMETRIOSIS IS STILL UNCLEAR. INTEGRATED BIOINFORMATICS ANALYSIS WAS IMPLEMENTED TO REVEAL THE MOLECULAR MECHANISMS UNDERLYING THIS DISEASE. METHODS: A TOTAL OF FOUR GENE EXPRESSION DATASETS (GSE7305, GSE11691, GSE23339, AND GSE25628) WERE RETRIEVED FROM THE GEO, WHICH WERE MERGED INTO A META-DATASET, FOLLOWED BY THE REMOVAL OF BATCH EFFECTS VIA THE SVA PACKAGE. WEIGHTED GENE CO-EXPRESSION NETWORK ANALYSIS (WGCNA) WAS IMPLEMENTED, AND ENDOMETRIOSIS-RELATED GENES WERE SCREENED UNDER NORMAL AND ENDOMETRIOSIS CONDITIONS. THEREAFTER, CHARACTERISTIC GENES WERE DETERMINED VIA LASSO ANALYSIS. THE DIAGNOSTIC PERFORMANCE WAS ESTIMATED VIA RECEIVER OPERATING CHARACTERISTIC CURVES, AND EPIGENETIC AND POST-TRANSCRIPTIONAL MODIFICATIONS WERE ANALYZED. SMALL MOLECULAR COMPOUNDS WERE PREDICTED. UNSUPERVISED CLUSTERING ANALYSIS WAS CONDUCTED VIA NON-NEGATIVE MATRIX FACTORIZATION ALGORITHM. THE ENRICHED PATHWAYS WERE ANALYZED VIA GENE SET ENRICHMENT ANALYSIS OR GSVA. IMMUNE FEATURES WERE EVALUATED ACCORDING TO IMMUNE-CHECKPOINTS, HLA, RECEPTORS, CHEMOKINES, AND IMMUNE CELLS. RESULTS: IN TOTAL, FOUR CHARACTERISTIC GENES (BGN, AQP1, ELMO1, AND DDR2) WERE DETERMINED FOR ENDOMETRIOSIS, ALL OF WHICH EXHIBITED THE FAVORABLE EFFICACY IN DIAGNOSING ENDOMETRIOSIS. THEIR ABERRANT LEVELS WERE MODULATED BY EPIGENETIC AND POST-TRANSCRIPTIONAL MODIFICATIONS. IN TOTAL, 51 POTENTIAL DRUGS WERE PREDICTED AGAINST ENDOMETRIOSIS. THE CHARACTERISTIC GENES EXHIBITED REMARKABLE ASSOCIATIONS WITH IMMUNOLOGICAL FUNCTION. THREE SUBTYPES WERE CLASSIFIED ACROSS ENDOMETRIOSIS, WITH DIFFERENT MECHANISMS AND IMMUNE FEATURES. CONCLUSION: OUR STUDY REVEALS THE CHARACTERISTIC GENES AND NOVEL MOLECULAR SUBTYPING OF ENDOMETRIOSIS, CONTRIBUTING TO THE EARLY DIAGNOSIS AND INTERVENTION IN ENDOMETRIOSIS. 2022 14 3047 34 GENOME-WIDE ANALYSIS OF DNA METHYLATION IN ENDOMETRIOSIS USING ILLUMINA HUMAN METHYLATION 450 K BEADCHIPS. ENDOMETRIOSIS IS A COMMON CHRONIC GYNECOLOGIC DISORDER CHARACTERIZED BY THE PRESENCE AND GROWTH OF ENDOMETRIAL-LIKE TISSUE OUTSIDE OF THE UTERINE CAVITY. ALTHOUGH THE EXACT ETIOLOGY REMAINS UNCLEAR, EPIGENETIC MODIFICATIONS, SUCH AS DNA METHYLATION, ARE THOUGHT TO CONTRIBUTE TO THE PATHOGENESIS OF ENDOMETRIOSIS. HERE, WE USED THE ILLUMINA HUMAN METHYLATION 450 K BEADCHIP ARRAY TO ANALYZE THE GENOME-WIDE DNA METHYLATION PROFILES OF SIX ENDOMETRIOTIC LESIONS AND SIX EUTOPIC ENDOMETRIA FROM PATIENTS WITH OVARIAN ENDOMETRIOSIS AND SIX ENDOMETRIA OF WOMEN WITHOUT ENDOMETRIOSIS. COMPARED WITH THE EUTOPIC ENDOMETRIA OF WOMEN WITH ENDOMETRIOSIS, 12,159 DIFFERENTIALLY METHYLATED CPG SITES AND 375 DIFFERENTIALLY METHYLATED PROMOTER REGIONS WERE IDENTIFIED IN ENDOMETRIOTIC LESIONS. GO ANALYSES SHOWED THAT THESE PUTATIVE DIFFERENTIALLY METHYLATED GENES WERE PRIMARILY ASSOCIATED WITH IMMUNE RESPONSE, INFLAMMATORY RESPONSE, RESPONSE TO STEROID HORMONE STIMULUS, CELL ADHESION, NEGATIVE REGULATION OF APOPTOSIS, AND ACTIVATION OF THE MAPK ACTIVITY. IN ADDITION, THE EXPRESSION LEVELS OF DNMT1, DNMT3A, DNMT3B, AND MBD2 IN ENDOMETRIOTIC LESIONS AND EUTOPIC ENDOMETRIA WERE SIGNIFICANTLY DECREASED COMPARED WITH CONTROL ENDOMETRIA. OUR FINDINGS SUGGEST THAT ABERRANT DNA METHYLATION STATUS IN ENDOMETRIOTIC LESIONS MAY PLAY A SIGNIFICANT ROLE IN THE PATHOGENESIS AND PROGRESSION OF ENDOMETRIOSIS. 2019 15 3478 37 IDENTIFICATION OF ADPKD-RELATED GENES AND PATHWAYS IN CELLS OVEREXPRESSING PKD2. CONSISTENT WITH THE GENE DOSAGE EFFECT HYPOTHESIS, RENAL CYSTS CAN ARISE IN TRANSGENIC MURINE MODELS OVEREXPRESSING EITHER PKD1 OR PKD2, WHICH ARE CAUSAL GENES FOR AUTOSOMAL DOMINANT POLYCYSTIC KIDNEY DISEASE (ADPKD). TO DETERMINE WHETHER PKD GENE OVEREXPRESSION IS A UNIVERSAL MECHANISM DRIVING CYSTOGENESIS OR IS MERELY RESTRICTED TO RODENTS, OTHER ANIMAL MODELS ARE REQUIRED. PREVIOUSLY, WE FAILED TO OBSERVE ANY RENAL CYSTS IN A TRANSGENIC PORCINE MODEL OF PKD2 OVEREXPRESSION PARTIALLY DUE TO EPIGENETIC SILENCING OF THE TRANSGENE. THUS, TO EXPLORE THE FEASIBILITY OF PORCINE MODELS AND IDENTIFY POTENTIAL GENES/PATHWAYS AFFECTED IN ADPKD, LLC-PK1 CELLS WITH HIGH PKD2 EXPRESSION WERE GENERATED. MRNA SEQUENCING (RNA-SEQ) WAS PERFORMED, AND MYC, IER3, AND ADM WERE FOUND TO BE UPREGULATED GENES COMMON TO THE DIFFERENT PKD2 OVEREXPRESSION CELL MODELS. MYC IS A WELL-CHARACTERIZED FACTOR CONTRIBUTING TO CYSTOGENESIS, AND ADM IS A BIOMARKER FOR CHRONIC KIDNEY DISEASE. THUS, THESE GENES MIGHT BE INDICATORS OF DISEASE PROGRESSION. ADDITIONALLY, SOME ADPKD-ASSOCIATED PATHWAYS, E.G., THE MITOGEN-ACTIVATED PROTEIN KINASE (MAPK) PATHWAY, WERE ENRICHED IN THE CELLS. MOREOVER, GENE ONTOLOGY (GO) ANALYSIS DEMONSTRATED THAT PROLIFERATION, APOPTOSIS, AND CELL CYCLE REGULATION, WHICH ARE HALLMARKS OF ADPKD, WERE ALTERED. THEREFORE, OUR EXPERIMENT IDENTIFIED SOME BIOMARKERS OR INDICATORS OF ADPKD, INDICATING THAT HIGH PKD2 EXPRESSION WOULD LIKELY DRIVE CYSTOGENESIS IN FUTURE PORCINE MODELS. 2020 16 2680 29 EVALUATION OF EPIGENETIC (HDAC, DNMT) AND PAIN (GAD65, TGF) FACTORS FOLLOWING PHOTOBIOMODULATION THERAPY IN A NEUROPATHIC PAIN MODEL. PHOTOBIOMODULATION THERAPY (PBMT) IS CONVERTED TO THE MOST COMMON ANALGESIC TREATMENT BEFORE THE WHOLE MECHANISM IS YET TO BE DISCOVERED. THIS STUDY FOR THE FIRST TIME WAS DESIGNED TO INVESTIGATE ALTERNATIONS OF EPIGENETIC FACTORS AFTER PAIN AND PBMT. THE CCI MODEL WAS CHOSEN TO INDUCE PAIN. PAIN EVALUATION TESTS INCLUDING PLANTAR, ACETONE, VON FREY, AND PINCH WERE DONE WEEKLY. THEN SPINAL CORD TISSUE WAS ISOLATED FOR EVALUATING MRNA EXPRESSION OF DNMT3A, HDAC1, AND NRSF USING RT-QPCR METHOD, AND PROTEIN EXPRESSION FACTORS OF HDAC2 AND DNMT3A USING WESTERN BLOTTING. GAD65 AND TGF-BETA PROTEINS WERE ASSESSED BY THE IHC METHOD. PBMT INCREASED THE PAIN THRESHOLD UP TO THE POINT WHERE IT ROUGHLY MET THE PAIN THRESHOLD OF THE CONTROL GROUP. AFTER THREE WEEKS OF TREATMENT, BOTH PBMT PROTOCOLS DEMONSTRATED A REDUCTION IN ALLODYNIA AND HYPERALGESIA. WHILE SOME MOLECULES, SUCH AS TGF-BETA AND GAD65, INCREASED FOLLOWING PBMT, WE OBSERVED NO INHIBITION OF NRSF, HDAC1, AND DNMT3A EXPRESSION DESPITE IMPLEMENTING TWO DIFFERENT PROTOCOLS. 2023 17 2422 32 EPIGENETIC SIGNATURES DISCRIMINATE PATIENTS WITH PRIMARY SCLEROSING CHOLANGITIS AND ULCERATIVE COLITIS FROM PATIENTS WITH ULCERATIVE COLITIS. BACKGROUND: PRIMARY SCLEROSING CHOLANGITIS (PSC) IS A CHRONIC INFLAMMATORY LIVER DISEASE AFFECTING THE INTRA- AND EXTRAHEPATIC BILE DUCTS, AND IS STRONGLY ASSOCIATED WITH ULCERATIVE COLITIS (UC). IN THIS STUDY, WE EXPLORED THE PERIPHERAL BLOOD DNA METHYLOME AND ITS IMMUNE CELL COMPOSITION IN PATIENTS WITH PSC-UC, UC, AND HEALTHY CONTROLS (HC) WITH THE AIM TO DEVELOP A PREDICTIVE ASSAY IN DISTINGUISHING PATIENTS WITH PSC-UC FROM THOSE WITH UC ALONE. METHODS: THE PERIPHERAL BLOOD DNA METHYLOME OF MALE PATIENTS WITH PSC AND CONCOMITANT UC, UC AND HCS WAS PROFILED USING THE ILLUMINA HUMANMETHYLATION INFINIUM EPIC BEADCHIP (850K) ARRAY. DIFFERENTIALLY METHYLATED CPG POSITION (DMP) AND REGION (DMR) ANALYSES WERE PERFORMED ALONGSIDE GRADIENT BOOSTING CLASSIFICATION ANALYSES TO DISCERN PSC-UC FROM UC PATIENTS. AS OBSERVED DIFFERENCES IN THE DNA METHYLOME COULD BE THE RESULT OF DIFFERENCES IN CELLULAR POPULATIONS, WE ADDITIONALLY EMPLOYED MASS CYTOMETRY (CYTOF) TO CHARACTERIZE THE IMMUNE CELL COMPOSITIONS. RESULTS: GENOME WIDE METHYLATION ANALYSIS DID NOT REVEAL LARGE DIFFERENCES BETWEEN PSC-UC AND UC PATIENTS NOR HCS. NONETHELESS, USING GRADIENT BOOSTING WE WERE CAPABLE OF DISCERNING PSC-UC FROM UC WITH AN AREA UNDER THE RECEIVER OPERATOR CURVE (AUROC) OF 0.80. FOUR CPG SITES ANNOTATED TO THE NINJ2 GENE WERE FOUND TO STRONGLY CONTRIBUTE TO THE PREDICTIVE PERFORMANCE. WHILE CYTOF ANALYSES CORROBORATED THE LARGELY SIMILAR BLOOD CELL COMPOSITION AMONG PATIENTS WITH PSC-UC, UC AND HC, A HIGHER ABUNDANCE OF MYELOID CELLS WAS OBSERVED IN UC COMPARED TO PSC-UC PATIENTS. CONCLUSION: DNA METHYLATION ENABLES DISCERNING PSC-UC FROM UC PATIENTS, WITH A POTENTIAL FOR BIOMARKER DEVELOPMENT. 2022 18 1176 38 CONTROL OF APOBEC3B INDUCTION AND CCCDNA DECAY BY NF-KAPPAB AND MIR-138-5P. BACKGROUND & AIMS: IMMUNE-MEDIATED INDUCTION OF CYTIDINE DEAMINASE APOBEC3B (A3B) EXPRESSION LEADS TO HBV COVALENTLY CLOSED CIRCULAR DNA (CCCDNA) DECAY. HERE, WE AIMED TO DECIPHER THE SIGNALLING PATHWAY(S) AND REGULATORY MECHANISM(S) INVOLVED IN A3B INDUCTION AND RELATED HBV CONTROL. METHODS: DIFFERENTIATED HEPARG CELLS (DHEPARG) KNOCKED-DOWN FOR NF-KAPPAB SIGNALLING COMPONENTS, TRANSFECTED WITH SIRNA OR MICRO RNAS (MIRNA), AND PRIMARY HUMAN HEPATOCYTES +/- HBV OR HBVDELTAX OR HBV-RFP, WERE TREATED WITH LYMPHOTOXIN BETA RECEPTOR (LTBETAR)-AGONIST (BS1). THE BIOLOGICAL OUTCOMES WERE ANALYSED BY REVERSE TRANSCRIPTASE-QPCR, IMMUNOBLOTTING, LUCIFERASE ACTIVITY, CHROMATIN IMMUNE PRECIPITATION, ELECTROPHORETIC MOBILITY-SHIFT ASSAY, TARGETED-BISULFITE-, MIRNA-, RNA-, GENOME-SEQUENCING, AND MASS-SPECTROMETRY. RESULTS: WE FOUND THAT CANONICAL AND NON-CANONICAL NF-KAPPAB SIGNALLING PATHWAYS ARE MANDATORY FOR A3B INDUCTION AND ANTI-HBV EFFECTS. THE DEGREE OF IMMUNE-MEDIATED A3B PRODUCTION IS INDEPENDENT OF A3B PROMOTER DEMETHYLATION BUT IS CONTROLLED POST-TRANSCRIPTIONALLY BY THE MIRNA 138-5P EXPRESSION (HSA-MIR-138-5P), PROMOTING A3B MRNA DECAY. HSA-MIR-138-5P OVER-EXPRESSION REDUCED A3B LEVELS AND ITS ANTIVIRAL EFFECTS. OF NOTE, ESTABLISHED INFECTION INHIBITED BS1-INDUCED A3B EXPRESSION THROUGH EPIGENETIC MODULATION OF A3B PROMOTER. TWELVE DAYS OF TREATMENT WITH A LTBETAR-SPECIFIC AGONIST BS1 IS SUFFICIENT TO REDUCE THE CCCDNA POOL BY 80% WITHOUT INDUCING SIGNIFICANT DAMAGES TO A SUBSET OF CANCER-RELATED HOST GENES. INTERESTINGLY, THE A3B-MEDIATED EFFECT ON HBV IS INDEPENDENT OF THE TRANSCRIPTIONAL ACTIVITY OF CCCDNA AS WELL AS ON RCDNA SYNTHESIS. CONCLUSIONS: ALTOGETHER, A3B REPRESENTS THE ONLY DESCRIBED ENZYME TO TARGET BOTH TRANSCRIPTIONALLY ACTIVE AND INACTIVE CCCDNA. THUS, INHIBITING HSA-MIR-138-5P EXPRESSION SHOULD BE CONSIDERED IN THE COMBINATORIAL DESIGN OF NEW THERAPIES AGAINST HBV, ESPECIALLY IN THE CONTEXT OF IMMUNE-MEDIATED A3B INDUCTION. LAY SUMMARY: IMMUNE-MEDIATED INDUCTION OF CYTIDINE DEAMINASE APOBEC3B IS TRANSCRIPTIONALLY REGULATED BY NF-KAPPAB SIGNALLING AND POST-TRANSCRIPTIONALLY DOWNREGULATED BY HSA-MIR-138-5P EXPRESSION, LEADING TO CCCDNA DECAY. TIMELY CONTROLLED APOBEC3B-MEDIATED CCCDNA DECAY OCCURS INDEPENDENTLY OF CCCDNA TRANSCRIPTIONAL ACTIVITY AND WITHOUT DAMAGE TO A SUBSET OF CANCER-RELATED GENES. THUS, APOBEC3B-MEDIATED CCCDNA DECAY COULD OFFER AN EFFICIENT THERAPEUTIC ALTERNATIVE TO TARGET HEPATITIS B VIRUS CHRONIC INFECTION. 2021 19 663 41 BLOOD ORANGE JUICE INTAKE MODULATES PLASMA AND PBMC MICRORNA EXPRESSION IN OVERWEIGHT AND INSULIN-RESISTANT WOMEN: IMPACT ON MAPK AND NFKAPPAB SIGNALING PATHWAYS. BLOOD ORANGE CONSUMPTION PRESENTS POTENTIAL HEALTH BENEFITS AND MAY MODULATE EPIGENETIC MECHANISMS SUCH AS MICRORNAS (MIRNAS) EXPRESSION. MIRNAS ARE NON-CODING RNAS RESPONSIBLE FOR POST-TRANSCRIPTIONAL GENE REGULATION, AND THESE MOLECULES CAN ALSO BE USED AS BIOMARKERS IN BODY FLUIDS. THIS STUDY WAS DESIGNED TO INVESTIGATE THE EFFECT OF CHRONIC BLOOD ORANGE JUICE (BOJ) INTAKE ON THE INFLAMMATORY RESPONSE AND MIRNA EXPRESSION PROFILE IN PLASMA AND BLOOD CELLS IN OVERWEIGHT WOMEN. THE STUDY COHORT WAS COMPRISED OF TWENTY WOMEN AGED 18-40 YEARS OLD, DIAGNOSED AS OVERWEIGHT, WHO CONSUMED 500 ML/D OF BOJ FOR FOUR WEEKS. CLINICAL DATA WERE COLLECTED AT BASELINE AND AFTER 4 WEEKS OF JUICE CONSUMPTION, E.G., ANTHROPOMETRIC AND HEMODYNAMIC PARAMETERS, FOOD INTAKE, BLOOD CELL COUNT, AND METABOLIC AND INFLAMMATORY BIOMARKERS. BOJ SAMPLES WERE ANALYZED AND CHARACTERIZED. ADDITIONALLY, PLASMA AND BLOOD CELLS WERE ALSO COLLECTED FOR MIRNA EXPRESSION PROFILING AND EVALUATION OF THE EXPRESSION OF GENES AND PROTEINS IN THE MAPK AND NFKAPPAB SIGNALING PATHWAYS. BOJ INTAKE INCREASED THE EXPRESSION OF MIR-144-3P IN PLASMA AND THE EXPRESSION OF MIR-424-5P, MIR-144-3P, AND MIR-130B-3P IN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMC). CONVERSELY, THE BEVERAGE INTAKE DECREASED THE EXPRESSION OF LET-7F-5P AND MIR-126-3P IN PBMC. COMPUTATIONAL ANALYSES IDENTIFIED DIFFERENT TARGETS OF THE DYSREGULATED MIRNA ON INFLAMMATORY PATHWAYS. FURTHERMORE, BOJ INTAKE INCREASED VITAMIN C CONSUMPTION AND THE PJNK/JNK RATIO AND DECREASED THE EXPRESSION OF IL6 MRNA AND NFKAPPAB PROTEIN. THESE RESULTS DEMONSTRATE THAT BOJ REGULATES THE EXPRESSION OF GENES INVOLVED IN THE INFLAMMATORY PROCESS AND DECREASES NFSMALL KA, CYRILLICB-PROTEIN EXPRESSION IN PBMC. 2023 20 5779 42 SPINAL MICRORNA-134-5P TARGETS GLUTAMATE RECEPTOR IONOTROPIC KAINATE 3 TO MODULATE OPIOID INDUCED HYPERALGESIA IN MICE. BACKGROUND: FENTANYL AND ITS ANALOGS ARE EXTENSIVELY USED FOR PAIN RELIEF. HOWEVER, THEIR PARADOXICALLY PRONOCICEPTIVE EFFECTS OFTEN LEAD TO INCREASED OPIOIDS CONSUMPTION AND RISK OF CHRONIC PAIN. COMPARED TO OTHER SYNTHETIC OPIOIDS, REMIFENTANIL HAS BEEN STRONGLY LINKED TO ACUTE OPIOID HYPERALGESIA AFTER EXPOSURE [REMIFENTANIL-INDUCED HYPERALGESIA (RIH)]. THE EPIGENETIC REGULATION OF MICRORNAS (MIRNAS) ON TARGETED MRNAS HAS EMERGED AS AN IMPORTANT PATHOGENESIS IN PAIN. THE CURRENT RESEARCH AIMED AT EXPLORING THE SIGNIFICANCE AND CONTRIBUTIONS OF MIR-134-5P TO THE DEVELOPMENT OF RIH. METHODS: BOTH THE ANTINOCICEPTIVE AND PRONOCICEPTIVE EFFECTS OF TWO COMMONLY USED OPIOIDS WERE ASSESSED, AND MIRNA EXPRESSION PROFILES IN THE SPINAL DORSAL HORN (SDH) OF MICE ACUTELY EXPOSED TO REMIFENTANIL AND REMIFENTANIL EQUIANALGESIC DOSE (RED) SUFENTANIL WERE SCREENED. NEXT, THE CANDIDATE MIRNA LEVEL, CELLULAR DISTRIBUTION, AND FUNCTION WERE EXAMINED BY QPCR, FLUORESCENT IN SITU HYBRIDIZATION (FISH) AND ARGONAUTE-2 IMMUNOPRECIPITATION. FURTHERMORE, BIOINFORMATICS ANALYSIS, LUCIFERASE ASSAYS, MIRNA OVEREXPRESSION, BEHAVIORAL TESTS, GOLGI STAINING, ELECTRON MICROSCOPY, WHOLE-CELL PATCH-CLAMP RECORDING, AND IMMUNOBLOTTING WERE EMPLOYED TO INVESTIGATE THE POTENTIAL TARGETS AND MECHANISMS UNDERLYING RIH. RESULTS: REMIFENTANIL INDUCED SIGNIFICANT PRONOCICEPTIVE EFFECTS AND A DISTINCT MIRNA-PROFILE FROM SUFENTANIL WHEN COMPARED TO SALINE CONTROLS. AMONG TOP 30 DIFFERENTIALLY EXPRESSED MIRNAS SPECTRUM, SPINAL MIR-134-5P WAS DRAMATICALLY DOWNREGULATED IN RIH MICE BUT REMAINED COMPARATIVE IN MICE SUBJECTED TO SUFENTANIL. MOREOVER, GLUTAMATE RECEPTOR IONOTROPIC KAINATE 3 (GRIK3) WAS A TARGET OF MIR-134-5P. THE OVEREXPRESSION OF MIR-134-5P ATTENUATED THE HYPERALGESIC PHENOTYPE, EXCESSIVE DENDRITIC SPINE REMODELING, EXCITATORY SYNAPTIC STRUCTURAL PLASTICITY, AND KAINATE RECEPTOR-MEDIATED MINIATURE EXCITATORY POSTSYNAPTIC CURRENTS (MEPSCS) IN SDH RESULTING FROM REMIFENTANIL EXPOSURE. BESIDES, INTRATHECAL INJECTION OF SELECTIVE KA-R ANTAGONIST WAS ABLE TO REVERSE THE GRIK3 MEMBRANE TRAFFICKING AND RELIEVED RIH. CONCLUSION: THE MIR-134-5P CONTRIBUTES TO REMIFENTANIL-INDUCED PRONOCICEPTIVE FEATURES VIA DIRECTLY TARGETING GRIK3 TO MODULATE DENDRITIC SPINE MORPHOLOGY AND SYNAPTIC PLASTICITY IN SPINAL NEURONS. 2023