1 4993 177 PENTOXIFYLLINE-INDUCED PROTEIN EXPRESSION CHANGE IN RAW 264.7 CELLS AS DETERMINED BY IMMUNOPRECIPITATION-BASED HIGH PERFORMANCE LIQUID CHROMATOGRAPHY. ALTHOUGH PENTOXIFYLLINE (PTX) WAS IDENTIFIED AS A COMPETITIVE NON-SELECTIVE PHOSPHODIESTERASE INHIBITOR, ITS PHARMACOLOGICAL EFFECT HAS NOT BEEN CLEARLY ELUCIDATED. THE PRESENT STUDY EXPLORED THE EFFECT OF LOW DOSE 10 MUG/ML PTX (THERAPEUTIC DOSE) COMPARED TO HIGH DOSE 300 MUG/ML PTX (EXPERIMENTAL DOSE) IN RAW 264.7 CELLS THROUGH IMMUNOPRECIPITATION-BASED HIGH PERFORMANCE LIQUID CHROMATOGRAPHY (IP-HPLC), IMMUNOHISTOCHEMISTRY, AND WESTERN BLOT. 10 MUG/ML PTX INCREASED THE EXPRESSION OF PROLIFERATION (KI-67, PCNA, CYCLIN D2, CDC25A), EPIGENETIC MODIFICATION (KDM4D, PCAF, HMGB1), PROTEIN TRANSLATION (DOHH, DHPS, EIF5A1), RAS SIGNALING (KRAS, PAKT1/2/3, PI3K), NFKB SIGNALING (NFKB, GADD45, P38), PROTECTION (HSP70, SOD1, GSTO1/2), SURVIVAL (PAKT1/2/3, SP1, SIRTUIN 6), NEUROMUSCULAR DIFFERENTIATION (NSEGAMMA, MYOSIN-1A, DESMIN), OSTEOBLASTIC DIFFERENTIATION (BMP2, RUNX2, OSTERIX), ACUTE INFLAMMATION (TNFALPHA, IL-1, CXCR4), INNATE IMMUNITY (BETA-DEFENSIN 1, LACTOFERRIN, TLR-3, -4), CELL-MEDIATED IMMUNITY (CD4, CD8, CD80), WHILE DECREASED THE EXPRESSION OF ER STRESS (EIF2ALPHA, EIF2AK3, ATF6ALPHA), FIBROSIS (FGF2, CTGF, COLLAGEN 3A1), AND CHRONIC INFLAMMATION (CD68, MMP-2, -3, COX2) VERSUS THE UNTREATED CONTROLS. THE ACTIVATION OF PROLIFERATION BY 10 MUG/ML PTX WAS ALSO SUPPORTED BY THE INCREASE OF CMYC-MAX HETERODIMER AND BETA-CATENIN-TCF1 COMPLEX IN DOUBLE IP-HPLC. 10 MUG/ML PTX ENHANCED FAS-MEDIATED APOPTOSIS BUT DIMINISHED P53-MEDIATED APOPTOSIS, AND DOWNREGULATED MANY ANGIOGENESIS PROTEINS (ANGIOGENIN, VEGF-A, AND FLT4), BUT UPREGULATED HIF1ALPHA, VEGFR2, AND CMG2 REACTIVELY. WHEREAS, 300 MUG/ML PTX CONSISTENTLY DECREASED PROLIFERATION, EPIGENETIC MODIFICATION, RAS AND NFKB SIGNALING, NEUROMUSCULAR AND OSTEOBLASTIC DIFFERENTIATION, BUT INCREASED APOPTOSIS, ER STRESS, AND FIBROSIS COMPARED TO 10 MUG/ML PTX. THESE DATA SUGGEST PTX HAS DIFFERENT BIOLOGICAL EFFECT ON RWA 264.7 CELLS DEPENDING ON THE CONCENTRATION OF 10 MUG/ML AND 300 MUG/ML PTX. THE LOW DOSE 10 MUG/ML PTX ENHANCED RAS/NFKB SIGNALING, PROLIFERATION, DIFFERENTIATION, AND INFLAMMATION, PARTICULARLY, IT STIMULATED NEUROMUSCULAR AND OSTEOBLASTIC DIFFERENTIATION, INNATE IMMUNITY, AND CELL-MEDIATED IMMUNITY, BUT ATTENUATED ER STRESS, FIBROSIS, ANGIOGENESIS, AND CHRONIC INFLAMMATION, WHILE THE HIGH DOSE 300 MUG/ML PTX WAS FOUND TO ALLEVIATE THE 10 MUG/ML PTX-INDUCED BIOLOGICAL EFFECTS, RESULTED IN THE SUPPRESSION OF RAS/NFKB SIGNALING, PROLIFERATION, NEUROMUSCULAR AND OSTEOBLASTIC DIFFERENTIATION, AND INFLAMMATION. 2022 2 3510 24 IDENTIFYING NOVEL B-CELL TARGETS FOR CHRONIC INFLAMMATORY AUTOIMMUNE DISEASE BY SCREENING OF CHEMICAL PROBES IN A PATIENT-DERIVED CELL ASSAY. B-CELL SECRETION OF AUTOANTIBODIES DRIVES AUTOIMMUNE DISEASES, INCLUDING SYSTEMIC LUPUS ERYTHEMATOSUS AND IDIOPATHIC INFLAMMATORY MYOSITIS. FEW THERAPIES ARE PRESENTLY AVAILABLE FOR TREATMENT OF THESE PATIENTS, OFTEN RESULTING IN UNSATISFACTORY EFFECTS AND HELPING ONLY SOME OF THE PATIENTS. WE DEVELOPED A SCREENING ASSAY FOR EVALUATION OF NOVEL TARGETS SUSPENDING B-CELL MATURATION INTO ANTIBODY SECRETING CELLS, WHICH COULD CONTRIBUTE TO FUTURE DRUG DEVELOPMENT. THE ASSAY WAS EMPLOYED FOR TESTING 43 HIGH QUALITY CHEMICAL PROBES AND COMPOUNDS INHIBITING UNDER-EXPLORED PROTEIN TARGETS, USING PRIMARY CELLS FROM PATIENTS WITH AUTOIMMUNE DISEASE. PROBES INHIBITING BROMODOMAIN FAMILY PROTEINS AND HISTONE METHYL TRANSFERASES DEMONSTRATED ABROGATION OF B-CELL FUNCTIONS TO A DEGREE COMPARABLE TO A POSITIVE CONTROL, THE JAK INHIBITOR TOFACITINIB. INHIBITION OF EACH TARGET RENDERED A SPECIFIC FUNCTIONAL CELL AND POTENTIAL DISEASE MODIFYING EFFECT, INDICATING SPECIFIC EPIGENETIC PROTEIN TARGETS AS POTENTIAL NEW INTERVENTION POINTS FOR FUTURE DRUG DISCOVERY AND DEVELOPMENT EFFORTS. 2021 3 1667 31 DOWNREGULATION OF PCAF BY MIR-181A/B PROVIDES FEEDBACK REGULATION TO TNF-ALPHA-INDUCED TRANSCRIPTION OF PROINFLAMMATORY GENES IN LIVER EPITHELIAL CELLS. ABERRANT CELLULAR RESPONSES TO PROINFLAMMATORY CYTOKINES, SUCH AS TNF-ALPHA, ARE PATHOGENIC FEATURES IN MOST CHRONIC INFLAMMATORY DISEASES. A VARIETY OF EXTRACELLULAR AND INTRACELLULAR FEEDBACK PATHWAYS HAS EVOLVED TO PREVENT AN INAPPROPRIATE CELLULAR REACTION TO THESE PROINFLAMMATORY CYTOKINES. IN THIS STUDY, WE REPORT THAT TNF-ALPHA TREATMENT OF HUMAN AND MOUSE CHOLANGIOCYTES AND HEPATOCYTES DOWNREGULATED EXPRESSION OF P300/CBP-ASSOCIATED FACTOR (PCAF), A COACTIVATOR AND AN ACETYLTRANSFERASE THAT PROMOTES HISTONE ACETYLATION AND GENE TRANSCRIPTION. OF THESE UPREGULATED MICRORNAS IN TNF-ALPHA-TREATED CELLS, MIR-181A/B (MIR-181A AND MIR-181B) SUPPRESSED TRANSLATION OF PCAF MRNA. FUNCTIONAL MANIPULATION OF MIR-181A/B CAUSED RECIPROCAL ALTERATIONS IN PCAF PROTEIN EXPRESSION IN CULTURED CHOLANGIOCYTES AND HEPATOCYTES. INHIBITION OF MIR-181A/B FUNCTION WITH ANTI-MIRS BLOCKED TNF-ALPHA-INDUCED SUPPRESSION OF PCAF EXPRESSION. PROMOTER RECRUITMENT OF PCAF WAS SHOWN TO BE ASSOCIATED WITH TNF-ALPHA-INDUCED TRANSCRIPTION OF INFLAMMATORY GENES. INTRIGUINGLY, PRETREATMENT OF CELLS WITH TNF-ALPHA INHIBITED TRANSCRIPTION OF INFLAMMATORY GENES IN RESPONSE TO SUBSEQUENT TNF-ALPHA STIMULATION. OVEREXPRESSION OF PCAF OR INHIBITION OF MIR-181A/B FUNCTION WITH ANTI-MIRS ATTENUATED THE INHIBITORY EFFECTS OF TNF-ALPHA PRETREATMENT ON EPITHELIAL INFLAMMATORY RESPONSE TO SUBSEQUENT TNF-ALPHA STIMULATION. DOWNREGULATION OF PCAF AND THE INHIBITORY EFFECTS OF TNF-ALPHA PRETREATMENT ON LIVER EPITHELIAL INFLAMMATORY RESPONSE WERE FURTHER CONFIRMED IN A MOUSE MODEL OF TNF-ALPHA I.P. INJECTION. THESE DATA SUGGEST THAT PCAF IS A TARGET FOR MIR-181A/B, AND DOWNREGULATION OF PCAF BY TNF-ALPHA PROVIDES NEGATIVE FEEDBACK REGULATION TO INFLAMMATORY REACTIONS IN LIVER EPITHELIAL CELLS, A PROCESS THAT MAY BE RELEVANT TO THE EPIGENETIC FINE-TUNING OF EPITHELIAL INFLAMMATORY PROCESSES IN GENERAL. 2012 4 6772 34 [ADVANCES IN EPIGENETIC MARKERS OF DERMATOMYOSITIS/POLYMYOSITIS]. IDIOPATHIC INFLAMMATORY MYOPATHY (IIM) IS A RARE GROUP OF AUTOIMMUNE DISEASES, CHARACTERIZED BY CHRONIC MUSCLE WEAKNESS, MUSCLE FATIGUE AND INFILTRATION OF SINGLE NUCLEAR CELLS IN SKELETAL MUSCLE. ITS SUBTYPES INCLUDE DERMATOMYOSITIS (DM), POLYMYOSITIS (PM), INCLUSION BODY MYOSITIS (IBM) AND IMMUNE-MEDIATED NECROTIZING MYOSITIS (IMNM), AND THE MOST COMMON SUBTYPES ARE DM AND PM. PM IS AN AUTOIMMUNE DISEASE MAINLY MANIFESTED BY MUSCLE DAMAGE. WHEN THE SKIN IS INVOLVED, IT IS CALLED DM. THE INCIDENCE OF IIM WAS RELATIVELY LOW, WHICH WAS 1.16-19 PER MILLION PEOPLE/YEAR, BUT THE MORTALITY WAS HIGH AND THE PROGNOSIS WAS POOR. THE PATHOGENESIS OF IIM IS STILL UNCLEAR. PREVIOUS STUDIES SUGGEST THAT BOTH IMMUNE AND NON-IMMUNE MECHANISMS ARE INVOLVED IN ITS PATHOGENESIS, ESPECIALLY CELLULAR AND HUMORAL IMMUNITY. IN RECENT YEARS, RESEARCHERS HAVE CONDUCTED A NUMBER OF STUDIES ON THE PATHOGENESIS OF IIM, ESPECIALLY IN THE STUDY OF DM/PM WITH THE APPLICATION OF HIGH-THROUGHPUT BIOMETRICS. EPIGENETICS IS A DISCIPLINE THAT REFERS TO THE GENETIC PHENOMENA OF DNA METHYLATION SPECTRUM, CHROMATIN STRUCTURE STATE AND GENE EXPRESSION SPECTRUM TRANSFERRED BETWEEN CELLS WITHOUT ANY CHANGES IN DNA SEQUENCE, INCLUDING DNA METHYLATION, CHROMATIN MODIFICATION AND NON-CODING RNA CHANGES. A LARGE NUMBER OF STUDIES HAVE SHOWN THAT EPIGENETIC MODIFICATION PLAYS AN IMPORTANT ROLE IN MANY DISEASES, ESPECIALLY IN CANCER. RECENT STUDIES HAVE ALSO FOUND A SERIES OF EPIGENETIC MARKERS RELATED TO THE OCCURRENCE AND DEVELOPMENT OF DM/PM, MAINLY IN THE ASPECT OF NON-CODING RNA CHANGES, SUCH AS MIR-10A, MIR-206, ETC. AND THERE HAS ALSO BEEN SOME RESEARCH ON DNA METHYLATION. HOWEVER, NO STUDIES HAVE BEEN REPORTED ON WHETHER CHROMATIN MODIFICATION IS INVOLVED IN THE PATHOGENESIS OF DM/PM. THE PATHOGENESIS OF DM/PM IS COMPLEX AND DIVERSE. WITH THE DEVELOPMENT OF RESEARCH, CERTAIN MICRORNAS (MIRNAS) AND LONG NON-CODING RNAS (LNCRNAS) MAY BECOME BIOLOGICAL MARKERS FOR THE EARLY DIAGNOSIS OF DM/PM. THEREFORE, THIS PAPER MAINLY EXPOUNDS THE RESEARCH PROGRESS OF THE BIOMARKERS OF DM/PM FROM THE ASPECT OF EPIGENETICS. 2019 5 6217 29 THE JAK INHIBITOR TOFACITINIB INHIBITS STRUCTURAL DAMAGE IN OSTEOARTHRITIS BY MODULATING JAK1/TNF-ALPHA/IL-6 SIGNALING THROUGH MIR-149-5P. BACKGROUND: OSTEOARTHRITIS (OA), A COMMON ARTICULAR BONE DEGENERATIVE DISEASE, IS EXACERBATED BY PROINFLAMMATORY CYTOKINE SIGNALING. MOUNTING EVIDENCE SUGGESTS THAT EPIGENETIC MODIFIERS, NAMELY MICRORNAS (MIRS), ARE DYSREGULATED IN ARTICULAR CHONDROCYTES (ACS) DURING OA. METHODS: AN INITIAL DATABASE SEARCH LED TO THE IDENTIFICATION OF MIR-149-5P, WHICH WAS DOWNREGULATED IN CLINICAL OA SAMPLES AND CONTRIBUTED TO CHRONIC INFLAMMATION, BY INCREASING TNF-ALPHA/IL-6 SIGNALING WITHIN THE SYNOVIUM, AND OA PROGRESSION. RESULTS: WE OVEREXPRESSED MIR-149-5P IN THE HUMAN CHONDROCYTE CELL LINES C20A4 AND C28/I2 TO EXAMINE ITS ROLE IN CHONDROCYTE HYPERTROPHY AND OSTEOCLASTOGENESIS AND FOUND A SIGNIFICANT DECREASE IN IL-6 EXPRESSION, AN INCREASE IN SOX9 EXPRESSION, AND A REDUCTION IN CHONDROCYTE HYPERTROPHY. WE EVALUATED THE THERAPEUTIC EFFECTS OF TOFACITINIB (JAK INHIBITOR) BY SUPPRESSING INFLAMMATION AND RESTORING MIR-149-5P EXPRESSION. TOFACITINIB-TREATED C20A4 AND C28/I2 CELLS HAD A SIGNIFICANTLY LOWER EXPRESSION OF JAK/IL-6/TNF-ALPHA AND AN INCREASED LEVEL OF MIR-149-5P. NOTABLY, TOFACITINIB TREATMENT REDUCED AC HYPERTROPHY AND SECRETION OF RANKL AND IL-6. FINALLY, AN OA MOUSE MODEL WAS USED TO EVALUATE THE THERAPEUTIC POTENTIAL OF TOFACITINIB. INTRA-ARTICULAR INJECTION OF TOFACITINIB SIGNIFICANTLY LOWERED ARTHRITIS SCORES AND BONE DEGRADATION IN TREATED MICE COMPARED WITH THEIR CONTROL COUNTERPARTS. CONCLUSION: WE SHOW FOR THE FIRST TIME THAT TOFACITINIB SUPPRESSES THE EXPRESSION LEVEL OF JAK1/TNF-ALPHA/IL-6 BY UPREGULATING MIR-149-5P LEVEL. OUR FINDINGS REVEALED THE FUNCTIONAL ASSOCIATION BETWEEN PROINFLAMMATORY JAK1/TNF-ALPHA/IL-6 SIGNALING AND ACS DEVELOPMENT AND HIGHLIGHT THE THERAPEUTIC POTENTIAL OF TOFACITINIB IN OA. 2021 6 5716 28 SIRT6 PROTECTS VASCULAR SMOOTH MUSCLE CELLS FROM OSTEOGENIC TRANSDIFFERENTIATION VIA RUNX2 IN CHRONIC KIDNEY DISEASE. VASCULAR CALCIFICATION (VC) IS REGARDED AS AN IMPORTANT PATHOLOGICAL CHANGE LACKING EFFECTIVE TREATMENT AND ASSOCIATED WITH HIGH MORTALITY. SIRTUIN 6 (SIRT6) IS A MEMBER OF THE SIRTUIN FAMILY, A CLASS III HISTONE DEACETYLASE AND A KEY EPIGENETIC REGULATOR. SIRT6 HAS A PROTECTIVE ROLE IN PATIENTS WITH CHRONIC KIDNEY DISEASE (CKD). HOWEVER, THE EXACT ROLE AND MOLECULAR MECHANISM OF SIRT6 IN VC IN PATIENTS WITH CKD REMAIN UNCLEAR. HERE, WE DEMONSTRATED THAT SIRT6 WAS MARKEDLY DOWNREGULATED IN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS) AND IN THE RADIAL ARTERY TISSUE OF PATIENTS WITH CKD WITH VC. SIRT6-TRANSGENIC (SIRT6-TG) MICE SHOWED ALLEVIATED VC, WHILE VASCULAR SMOOTH MUSCLE CELL-SPECIFIC (VSMC-SPECIFIC) SIRT6 KNOCKED-DOWN MICE SHOWED SEVERE VC IN CKD. SIRT6 SUPPRESSED THE OSTEOGENIC TRANSDIFFERENTIATION OF VSMCS VIA REGULATION OF RUNT-RELATED TRANSCRIPTION FACTOR 2 (RUNX2). COIMMUNOPRECIPITATION (CO-IP) AND IMMUNOPRECIPITATION (IP) ASSAYS CONFIRMED THAT SIRT6 BOUND TO RUNX2. MOREOVER, RUNX2 WAS DEACETYLATED BY SIRT6 AND FURTHER PROMOTED NUCLEAR EXPORT VIA EXPORTIN 1 (XPO1), WHICH IN TURN CAUSED DEGRADATION OF RUNX2 THROUGH THE UBIQUITIN-PROTEASOME SYSTEM. THESE RESULTS DEMONSTRATED THAT SIRT6 PREVENTED VC BY SUPPRESSING THE OSTEOGENIC TRANSDIFFERENTIATION OF VSMCS, AND AS SUCH TARGETING SIRT6 MAY BE AN APPEALING THERAPEUTIC TARGET FOR VC IN CKD. 2022 7 2202 31 EPIGENETIC MODIFICATION OF MIR-10A REGULATES RENAL DAMAGE BY TARGETING CREB1 IN TYPE 2 DIABETES MELLITUS. EMERGING EVIDENCE HAS SHOWN THAT MICRORNA-MEDIATED GENE EXPRESSION MODULATION PLAYS A CRUCIAL ROLE IN THE PATHOGENESIS OF TYPE 2 DIABETES MELLITUS, BUT THE NOVEL MIRNAS INVOLVED IN TYPE 2 DIABETES AND ITS FUNCTIONAL REGULATORY MECHANISMS STILL NEED TO BE DETERMINED. IN THIS STUDY, WE ASSESSED THE ROLE OF MIR-10A IN EXTRACELLULAR MATRIX ACCUMULATION IN THE KIDNEY OF DIABETIC MELLITUS INDUCED BY COMBINING ADMINISTRATION OF CHRONIC HIGH FAT DIET (HFD) AND LOW DOSAGE OF STREPTOZOTOCIN (STZ, 35MG/KG). HERE, WE FOUND THAT HFD/STZ ADMINISTRATION DECREASED THE LEVEL OF MICRORNA (MIR-10A) EXPRESSION IN ICR STRAIN MICE. OVEREXPRESSION OF MIR-10A ALLEVIATED THE INCREASED RATIO OF URINE ALBUMIN-TO-CREATININE (ACR) RATIO OF HFD/STZ MICE. IN CONTRAST, KNOCKDOWN OF MIR-10A INCREASED THE RATIO OF KIDNEY ACR IN NAIVE MICE. FURTHERMORE, CAMP RESPONSE ELEMENT BINDING PROTEIN 1 (CREB1) WAS VALIDATED AS A TARGET OF MIR-10A IN VITRO AND IN VIVO. CREB1 AND ITS DOWNSTREAM FIBRONECTIN (FN, EXTRACELLULAR MATRIX) WERE INCREASED IN HFD/STZ-TREATED MICE, WHICH WAS REVERSED BY KIDNEY MIR-10A OVEREXPRESSION. THE CONTENT OF CREB1 AND FN WAS INCREASED BY MIR-10A KNOCKDOWN IN KIDNEY OF NAIVE MICE. FURTHERMORE, HISTONE DEACETYLASE 3 (HDAC3) WAS REVEALED TO BE INCREASED IN KIDNEY OF HFD/STZ MICE, ACCOMPANIED WITH THE AUGMENTATION OF ACR RATIO AND FN LEVEL. KNOCKDOWN OF HDAC3 WITH SIRNA SIGNIFICANTLY CAUSED THE INCREASE OF MIR-10A, RESULTING IN THE DECREASE IN CREB1 AND FN EXPRESSION IN KIDNEY OF HFD/STZ MICE. CONTRARILY, HDAC3 OVEREXPRESSION MEDIATED BY LENTIVIRUS DECREASED MIR-10A CONTENT, AND ENHANCED ACR VALUE, CREB1 AND FN FORMATION IN NAIVE MICE. COLLECTIVELY, THESE RESULTS ELUCIDATE THAT HDAC3/MIR-10A/CREB1 SERVES AS A NEW MECHANISM UNDERLYING KIDNEY INJURY, PROVIDING POTENTIAL THERAPEUTIC TARGETS IN TYPE 2 DIABETES. 2016 8 6149 26 THE EXPRESSION PROFILE OF MIR-23B IS NOT ALTERED IN PERIPHERAL BLOOD MONONUCLEAR CELLS OF PATIENTS WITH IDIOPATHIC INFLAMMATORY MYOPATHIES. IDIOPATHIC INFLAMMATORY MYOPATHIES (IIM) BELONG TO A GROUP OF AUTOIMMUNE DISORDERS, PRIMARILY CHARACTERIZED BY CHRONIC INFLAMMATION OF HUMAN SKELETAL MUSCLE TISSUE. THE ETIOLOGY OF THESE DISEASES IS UNKNOWN, HOWEVER, GENETIC PREDISPOSITION PLAYS A SIGNIFICANT ROLE IN DISEASE ONSET. BESIDE THE KNOWN GENETIC RISK LOCATED IN THE MHC COMPLEX, THE EPIGENETIC MODIFICATIONS INCLUDING CHANGES IN MIRNAS EXPRESSION PROFILES HAVE BEEN RECENTLY IMPLICATED RECENTLY IN MANY AUTOIMMUNE DISEASES. MICRO RNA MOLECULES ARE INVOLVED IN MANY PHYSIOLOGICAL PROCESSES, INCLUDING THE REGULATION OF THE IMMUNE RESPONSE. IN OUR STUDY WE HAVE FOCUSED ON THE MIR-23B, AS IT REPRESENTS A NOVEL PROMISING AUTOIMMUNITY REGULATOR MOLECULE. DOWNREGULATION OF MIR-23B WAS RECENTLY DESCRIBED IN PATIENTS WITH RHEUMATOID ARTHRITIS AND SYSTEMIC LUPUS ERYTHEMATOSUS. WE HAVE MEASURED THE EXPRESSION MIR-23B PERIPHERAL BLOOD MONONUCLEAR CELLS OF PATIENTS WITH DERMATOMYOSITIS AND POLYMYOSITIS. NO MEANINGFUL DIFFERENCE WAS FOUND IN COMPARISON WITH HEALTHY CONTROLS. 2013 9 6702 33 VEGFA PROMOTER GENE HYPERMETHYLATION AT HIF1ALPHA BINDING SITE IS AN EARLY CONTRIBUTOR TO CKD PROGRESSION AFTER RENAL ISCHEMIA. CHRONIC HYPOXIA IS A MAJOR CONTRIBUTOR TO CHRONIC KIDNEY DISEASE (CKD) AFTER ACUTE KIDNEY INJURY (AKI). HOWEVER, THE TEMPORAL RELATION BETWEEN THE ACUTE INSULT AND MALADAPTIVE RENAL RESPONSE TO HYPOXIA REMAINS UNCLEAR. IN THIS STUDY, WE ANALYZED THE TIME-COURSE OF RENAL HEMODYNAMICS, OXIDATIVE STRESS, INFLAMMATION, AND FIBROSIS, AS WELL AS EPIGENETIC MODIFICATIONS, WITH FOCUS ON HIF1ALPHA/VEGF SIGNALING, IN THE AKI TO CKD TRANSITION. SHAM-OPERATED, RIGHT NEPHRECTOMY (UNX), AND UNX PLUS RENAL ISCHEMIA (IR + UNX) GROUPS OF RATS WERE INCLUDED AND STUDIED AT 1, 2, 3, OR 4 MONTHS. THE IR + UNX GROUP DEVELOPED CKD CHARACTERIZED BY PROGRESSIVE PROTEINURIA, RENAL DYSFUNCTION, TUBULAR PROLIFERATION, AND FIBROSIS. AT FIRST MONTH POST-ISCHEMIA, THERE WAS A TWOFOLD SIGNIFICANT INCREASE IN OXIDATIVE STRESS AND REDUCTION IN GLOBAL DNA METHYLATION THAT WAS MAINTAINED THROUGHOUT THE STUDY. HIF1ALPHA AND VEGFA EXPRESSION WERE DEPRESSED IN THE FIRST AND SECOND-MONTHS POST-ISCHEMIA, AND THEN HIF1ALPHA BUT NOT VEGFA EXPRESSION WAS RECOVERED. INTERESTINGLY, HYPERMETHYLATION OF THE VEGFA PROMOTER GENE AT THE HIF1ALPHA BINDING SITE WAS FOUND, SINCE EARLY STAGES OF THE CKD PROGRESSION. OUR FINDINGS SUGGEST THAT RENAL HYPOPERFUSION, INEFFICIENT HYPOXIC RESPONSE, INCREASED OXIDATIVE STRESS, DNA HYPOMETHYLATION, AND, VEGFA PROMOTER GENE HYPERMETHYLATION AT HIF1ALPHA BINDING SITE, ARE EARLY DETERMINANTS OF AKI-TO-CKD TRANSITION. 2021 10 5417 39 REGULATION OF DNA METHYLATION IN RHEUMATOID ARTHRITIS SYNOVIOCYTES. RHEUMATOID ARTHRITIS (RA) IS A CHRONIC INFLAMMATORY DISEASE IN WHICH FIBROBLAST-LIKE SYNOVIOCYTES (FLS) EXHIBIT AN AGGRESSIVE PHENOTYPE. ALTHOUGH THE MECHANISMS RESPONSIBLE ARE NOT WELL DEFINED, EPIGENETIC DETERMINANTS SUCH AS DNA METHYLATION MIGHT CONTRIBUTE. DNA METHYLTRANSFERASES (DNMTS) ARE CRITICAL ENZYMES THAT ESTABLISH AND MAINTAIN DNA METHYLATION. WE EVALUATED WHETHER PROINFLAMMATORY CYTOKINES MIGHT CONTRIBUTE TO DIFFERENTIAL DNA METHYLATION PREVIOUSLY DESCRIBED IN RA FLS THROUGH ALTERED DNMT EXPRESSION. FLS WERE OBTAINED FROM RA AND OSTEOARTHRITIS (OA) SYNOVIUM AT THE TIME OF TOTAL JOINT REPLACEMENT. GENE EXPRESSION WAS DETERMINED BY QUANTITATIVE REAL-TIME PCR AND PROTEIN EXPRESSION BY WESTERN BLOT ANALYSIS. DNMT ACTIVITY WAS MEASURED WITH A FUNCTIONAL ASSAY, AND GLOBAL METHYLATION WAS DETERMINED BY AN IMMUNOASSAY THAT DETECTS METHYLCYTOSINE. RESTING EXPRESSION OF DNMT1, -3A, AND -3B MRNA WERE SIMILAR IN RA AND OA FLS. WESTERN BLOT SHOWED ABUNDANT DNMT1 AND DNMT3A PROTEIN. EXPOSURE TO IL-1 DECREASED DNMT1 AND DNMT3A MRNA EXPRESSION IN FLS. DOSE RESPONSES DEMONSTRATED DECREASED DNMT EXPRESSION AT CONCENTRATIONS AS LOW AS 1 PG/ML OF IL-1. DNMT MRNA LEVELS DECREASED RAPIDLY, WITH SIGNIFICANT SUPPRESSION AFTER 2-8 H OF IL-1 STIMULATION. IL-1 STIMULATION OF OA FLS DID NOT AFFECT METHYLATION OF LINE1 SITES BUT LED TO DEMETHYLATION OF A CHI3L1 LOCUS THAT IS HYPOMETHYLATED IN RA FLS. CHRONIC IL-1 STIMULATION ALSO MIMICKED THE EFFECT OF A DNMT INHIBITOR ON FLS GENE EXPRESSION. EXPOSURE TO PROINFLAMMATORY MEDIATORS REVERSIBLY ALTERS DNA METHYLATION IN FLS BY DECREASING DNMT EXPRESSION AND FUNCTION. THESE DATA SUGGEST THAT IL-1 CAN POTENTIALLY IMPRINT CELLS IN CHRONIC INFLAMMATORY DISEASES. 2013 11 2784 35 EZH2 PROMOTES EXTRACELLULAR MATRIX DEGRADATION VIA NUCLEAR FACTOR-KAPPAB (NF-KAPPAB) AND P38 SIGNALING PATHWAYS IN PULPITIS. PULPITIS IS A COMPLICATED CHRONIC INFLAMMATORY PROCESS WHICH CAN BE IN A DYNAMIC BALANCE BETWEEN DAMAGE AND REPAIR. THE EXTRACELLULAR MATRIX PLAYS AN IMPORTANT REGULATORY ROLE IN WOUND HEALING AND TISSUE REPAIR. THE AIM OF THIS STUDY WAS TO EXPLORE THE ROLE OF THE EPIGENETIC MARK, ENHANCER OF ZESTE HOMOLOG 2 (EZH2) ON THE DEGRADATION OF EXTRACELLULAR MATRIX DURING PULPITIS. QUANTITATIVE POLYMERASE CHAIN REACTION WAS USED TO ASSESS THE EXPRESSION OF MATRIX METALLOPROTEINASES (MMPS) AND TYPE I COLLAGEN IN HUMAN DENTAL PULP CELLS (HDPCS) UPON EZH2 AND EI1 (EZH2 INHIBITOR) STIMULATION. THE MECHANISM OF EZH2 AFFECTING EXTRACELLULAR MATRIX WAS EXPLORED THROUGH QUANTITATIVE POLYMERASE CHAIN REACTION AND WESTERN BLOT. A RAT MODEL OF DENTAL PULP INFLAMMATION WAS ESTABLISHED, AND THE EXPRESSION OF TYPE I COLLAGEN IN DENTAL PULP UNDER EZH2 STIMULATION WAS DETECTED BY IMMUNOHISTOCHEMICAL STAINING. EZH2 UPREGULATED THE EXPRESSION OF MMP-1, MMP-3, MMP-8, AND MMP-10 AND DECREASED THE PRODUCTION OF TYPE I COLLAGEN IN HDPCS, WHILE EI1 HAD THE OPPOSITE EFFECT. EZH2 ACTIVATED THE NUCLEAR FACTOR-KAPPA B (NF-KAPPAB) AND P38 SIGNALING PATHWAYS IN HDPCS, THE INHIBITION OF WHICH REVERSED THE INDUCTION OF MMPS AND THE SUPPRESSION OF TYPE I COLLAGEN. EZH2 CAN DOWNREGULATE THE TYPE I COLLAGEN LEVELS IN AN EXPERIMENTAL MODEL OF DENTAL PULPITIS IN RATS. EZH2 PROMOTES EXTRACELLULAR MATRIX DEGRADATION VIA NUCLEAR FACTOR-KAPPAB (NF-KAPPAB) AND P38 SIGNALING PATHWAYS IN PULPITIS. EZH2 CAN DECREASE THE TYPE I COLLAGEN LEVELS IN VIVO AND IN VITRO. 2021 12 3048 26 GENOME-WIDE ANALYSIS REVEALED THAT DZNEP REDUCES TUBULOINTERSTITIAL FIBROSIS VIA DOWN-REGULATION OF PRO-FIBROTIC GENES. TUBULOINTERSTITIAL FIBROSIS HAS BEEN RECENTLY REPORTED TO BE CAUSED BY THE COLLAPSE OF THE EPIGENETIC REGULATION OF KIDNEY DISEASES. WE EXAMINED WHETHER PHARMACOLOGICAL INHIBITION OF HISTONE MODIFICATION IS EFFECTIVE AGAINST RENAL FIBROSIS. DZNEP (3-DEAZANEPLANOCIN A) WAS ORIGINALLY DEVELOPED AS AN ANTI-CANCER DRUG TO INHIBIT THE REPRESSIVE HISTONE MARK, H3K27ME3. WE USED A MODEL OF CHRONIC TUBULOINTERSTITIAL FIBROSIS INDUCED BY UNILATERAL ISCHAEMIA/REPERFUSION AND ADMINISTERED DZNEP INTRAVENOUSLY TO THE MICE FOR 8 WEEKS. WE FOUND DZNEP CONTRIBUTES TO THE REDUCTION OF TUBULOINTERSTITIAL FIBROSIS. WE SELECTED ONLY TUBULAR CELLS FROM IN VIVO SAMPLES USING LASER-CAPTURE MICRODISSECTION BECAUSE EPIGENETIC REGULATION IS SPECIFIC TO THE CELL TYPES, AND WE FOCUSED ON THE CHANGES IN THE TUBULAR CELLS. WE PERFORMED A GENOME-WIDE ANALYSIS OF TUBULAR CELLS USING HIGH-THROUGHPUT SEQUENCING (RNA-SEQ) TO IDENTIFY NOVEL EPIGENETIC FACTORS ASSOCIATED WITH RENAL FIBROSIS. WE FOUND THAT PRO-FIBROTIC GENES SUCH AS COL3A1 (COLLAGEN TYPE 3A1) AND TIMP2 (TISSUE INHIBITOR OF METALLOPROTEINASE 2) WERE SUPPRESSED BY DZNEP IN VIVO. IN ADDITION, PRO-FIBROTIC GENES SUCH AS COL4A1 (COLLAGEN TYPE 4A1), TIMP2 AND MMP14 WERE DOWN-REGULATED BY DZNEP IN VITRO. IN CONCLUSION, WE FOUND THAT PHARMACOLOGICAL EPIGENETIC MODIFICATION BY DZNEP DECREASED THE EXPRESSION LEVELS OF FIBROGENIC GENES IN TUBULAR CELLS AND INHIBITED TUBULOINTERSTITIAL FIBROSIS. 2018 13 3323 26 HISTONE DEACETYLASE 1 (HDAC1): A KEY PLAYER OF T CELL-MEDIATED ARTHRITIS. RHEUMATOID ARTHRITIS (RA) REPRESENTS A CHRONIC T CELL-MEDIATED INFLAMMATORY AUTOIMMUNE DISEASE. STUDIES HAVE SHOWN THAT EPIGENETIC MECHANISMS CONTRIBUTE TO THE PATHOGENESIS OF RA. HISTONE DEACETYLASES (HDACS) REPRESENT ONE IMPORTANT GROUP OF EPIGENETIC REGULATORS. HOWEVER, THE ROLE OF INDIVIDUAL HDAC MEMBERS FOR THE PATHOGENESIS OF ARTHRITIS IS STILL UNKNOWN. IN THIS STUDY WE DEMONSTRATE THAT MICE WITH A T CELL-SPECIFIC DELETION OF HDAC1 (HDAC1-CKO) ARE RESISTANT TO THE DEVELOPMENT OF COLLAGEN-INDUCED ARTHRITIS (CIA), WHEREAS THE ANTIBODY RESPONSE TO COLLAGEN TYPE II WAS UNDISTURBED, INDICATING AN UNALTERED T CELL-MEDIATED B CELL ACTIVATION. THE INFLAMMATORY CYTOKINES IL-17 AND IL-6 WERE SIGNIFICANTLY DECREASED IN SERA OF HDAC1-CKO MICE. IL-6 TREATED HDAC1-DEFICIENT CD4(+) T CELLS SHOWED AN IMPAIRED UPREGULATION OF CCR6. SELECTIVE INHIBITION OF CLASS I HDACS WITH THE HDAC INHIBITOR MS-275 UNDER TH17-SKEWING CONDITIONS INHIBITED THE UPREGULATION OF CHEMOKINE RECEPTOR 6 (CCR6) IN MOUSE AND HUMAN CD4(+) T CELLS. ACCORDINGLY, ANALYSIS OF HUMAN RNA-SEQUENCING (RNA-SEQ) DATA AND HISTOLOGICAL ANALYSIS OF SYNOVIAL TISSUE SAMPLES FROM HUMAN RA PATIENTS REVEALED THE EXISTENCE OF CD4(+)CCR6(+) CELLS WITH ENHANCED HDAC1 EXPRESSION. OUR DATA INDICATE A KEY ROLE FOR HDAC1 FOR THE PATHOGENESIS OF CIA AND SUGGEST THAT HDAC1 AND OTHER CLASS I HDACS MIGHT BE PROMISING TARGETS OF SELECTIVE HDAC INHIBITORS (HDACI) FOR THE TREATMENT OF RA. 2020 14 4582 23 N-TERMINAL BET BROMODOMAIN INHIBITORS DISRUPT A BRD4-P65 INTERACTION AND REDUCE INDUCIBLE NITRIC OXIDE SYNTHASE TRANSCRIPTION IN PANCREATIC BETA-CELLS. CHRONIC INFLAMMATION OF PANCREATIC ISLETS IS A KEY DRIVER OF BETA-CELL DAMAGE THAT CAN LEAD TO AUTOREACTIVITY AND THE EVENTUAL ONSET OF AUTOIMMUNE DIABETES (T1D). IN THE ISLET, ELEVATED LEVELS OF PROINFLAMMATORY CYTOKINES INDUCE THE TRANSCRIPTION OF THE INDUCIBLE NITRIC OXIDE SYNTHASE (INOS) GENE, NOS2, ULTIMATELY RESULTING IN INCREASED NITRIC OXIDE (NO). EXCESSIVE OR PROLONGED EXPOSURE TO NO CAUSES BETA-CELL DYSFUNCTION AND FAILURE ASSOCIATED WITH DEFECTS IN MITOCHONDRIAL RESPIRATION. RECENT STUDIES SHOWED THAT INHIBITION OF THE BROMODOMAIN AND EXTRATERMINAL DOMAIN (BET) FAMILY OF PROTEINS, A DRUGGABLE CLASS OF EPIGENETIC READER PROTEINS, PREVENTS THE ONSET AND PROGRESSION OF T1D IN THE NON-OBESE DIABETIC MOUSE MODEL. WE HYPOTHESIZED THAT BET PROTEINS CO-ACTIVATE TRANSCRIPTION OF CYTOKINE-INDUCED INFLAMMATORY GENE TARGETS IN BETA-CELLS AND THAT SELECTIVE, CHEMOTHERAPEUTIC INHIBITION OF BET BROMODOMAINS COULD REDUCE SUCH TRANSCRIPTION. HERE, WE INVESTIGATED THE ABILITY OF BET BROMODOMAIN SMALL MOLECULE INHIBITORS TO REDUCE THE BETA-CELL RESPONSE TO THE PROINFLAMMATORY CYTOKINE INTERLEUKIN 1 BETA (IL-1BETA). BET BROMODOMAIN INHIBITION ATTENUATED IL-1BETA-INDUCED TRANSCRIPTION OF THE INFLAMMATORY MEDIATOR NOS2 AND CONSEQUENT INOS PROTEIN AND NO PRODUCTION. REDUCED NOS2 TRANSCRIPTION IS CONSISTENT WITH INHIBITION OF NF-KAPPAB FACILITATED BY DISRUPTING THE INTERACTION OF A SINGLE BET FAMILY MEMBER, BRD4, WITH THE NF-KAPPAB SUBUNIT, P65. USING RECENTLY REPORTED SELECTIVE INHIBITORS OF THE FIRST AND SECOND BET BROMODOMAINS, INHIBITION OF ONLY THE FIRST BROMODOMAIN WAS NECESSARY TO REDUCE THE INTERACTION OF BRD4 WITH P65 IN BETA-CELLS. MOREOVER, INHIBITION OF THE FIRST BROMODOMAIN WAS SUFFICIENT TO MITIGATE IL-1BETA-DRIVEN DECREASES IN MITOCHONDRIAL OXYGEN CONSUMPTION RATES AND BETA-CELL VIABILITY. BY IDENTIFYING A ROLE FOR THE INTERACTION BETWEEN BRD4 AND P65 IN CONTROLLING THE RESPONSE OF BETA-CELLS TO PROINFLAMMATORY CYTOKINES, WE PROVIDE MECHANISTIC INFORMATION ON HOW BET BROMODOMAIN INHIBITION CAN DECREASE INFLAMMATION. THESE STUDIES ALSO SUPPORT THE POTENTIAL THERAPEUTIC APPLICATION OF MORE SELECTIVE BET BROMODOMAIN INHIBITORS IN ATTENUATING BETA-CELL INFLAMMATION. 2022 15 6658 33 UPREGULATED LNCRNA H19 SPONGES MIR-106A-5P AND CONTRIBUTES TO ALDOSTERONE-INDUCED VASCULAR CALCIFICATION VIA ACTIVATING THE RUNX2-DEPENDENT PATHWAY. BACKGROUND: EXCESS ALDOSTERONE IS IMPLICATED IN VASCULAR CALCIFICATION (VC), BUT THE MECHANISM BY WHICH ALDOSTERONE-MR (MINERALOCORTICOID RECEPTOR) COMPLEX PROMOTES VC IS UNCLEAR. EMERGING EVIDENCE INDICATES THAT LONG-NONCODING RNA H19 (H19) PLAYS A CRITICAL ROLE IN VC. WE EXAMINED WHETHER ALDOSTERONE-INDUCED OSTEOGENIC DIFFERENTIATION OF VASCULAR SMOOTH MUSCLE CELLS (VSMCS) THROUGH H19 EPIGENETIC MODIFICATION OF RUNX2 (RUNT-RELATED TRANSCRIPTION FACTOR-2) IN A MR-DEPENDENT MANNER. METHODS: WE INDUCED IN VIVO RAT MODEL OF CHRONIC KIDNEY DISEASE USING A HIGH ADENINE AND PHOSPHATE DIET TO EXPLORE THE RELATIONSHIP AMONG ALDOSTERONE, MR, H19, AND VC. WE ALSO CULTURED HUMAN AORTIC VSMCS TO EXPLORE THE ROLES OF H19 IN ALDOSTERONE-MR COMPLEX-INDUCED OSTEOGENIC DIFFERENTIATION AND CALCIFICATION OF VSMCS. RESULTS: H19 AND RUNX2 WERE SIGNIFICANTLY INCREASED IN ALDOSTERONE-INDUCED VSMC OSTEOGENIC DIFFERENTIATION AND VC, BOTH IN VITRO AND IN VIVO, WHICH WERE SIGNIFICANTLY BLOCKED BY THE MR ANTAGONIST SPIRONOLACTONE. MECHANISTICALLY, OUR FINDINGS REVEAL THAT THE ALDOSTERONE-ACTIVATED MR BOUND TO H19 PROMOTER AND INCREASED ITS TRANSCRIPTIONAL ACTIVITY, AS DETERMINED BY CHROMATIN IMMUNOPRECIPITATION, ELECTROPHORETIC MOBILITY SHIFT ASSAY, AND LUCIFERASE REPORTER ASSAY. SILENCING H19 INCREASED MICRORNA-106A-5P (MIR-106A-5P) EXPRESSION, WHICH SUBSEQUENTLY INHIBITED ALDOSTERONE-INDUCED RUNX2 EXPRESSION AT THE POSTTRANSCRIPTIONAL LEVEL. IMPORTANTLY, WE OBSERVED A DIRECT INTERACTION BETWEEN H19 AND MIR-106A-5P, AND DOWNREGULATION OF MIR-106A-5P EFFICIENTLY REVERSED THE SUPPRESSION OF RUNX2 INDUCED BY H19 SILENCING. CONCLUSIONS: OUR STUDY CLARIFIES A NOVEL MECHANISM BY WHICH UPREGULATION OF H19 CONTRIBUTES TO ALDOSTERONE-MR COMPLEX-PROMOTED RUNX2-DEPENDENT VSMC OSTEOGENIC DIFFERENTIATION AND VC THROUGH SPONGING MIR-106A-5P. THESE FINDINGS HIGHLIGHT A POTENTIAL THERAPEUTIC TARGET FOR ALDOSTERONE-INDUCED VC. 2023 16 5868 32 SUPPRESSIVE EFFECTS OF METFORMIN ON T-HELPER 1-RELATED CHEMOKINES EXPRESSION IN THE HUMAN MONOCYTIC LEUKEMIA CELL LINE THP-1. PURPOSE OF THE STUDY: TYPE 1 AND TYPE 2 DIABETES MELLITUS (DM) ARE CHRONIC T-CELL-MEDIATED INFLAMMATORY DISEASES. METFORMIN IS A WIDELY USED DRUG FOR TYPE 2 DM THAT REDUCES THE NEED FOR INSULIN IN TYPE 1 DM. HOWEVER, WHETHER METFORMIN HAS AN ANTI-INFLAMMATORY EFFECT FOR TREATING DM IS UNKNOWN. WE INVESTIGATED THE ANTI-INFLAMMATORY MECHANISM OF METFORMIN IN THE HUMAN MONOCYTIC LEUKEMIA CELL LINE THP-1. MATERIALS AND METHODS: THE HUMAN MONOCYTIC LEUKEMIA CELL LINE THP-1 WAS PRETREATED WITH METFORMIN AND STIMULATED WITH LIPOPOLYSACCHARIDE (LPS). THE PRODUCTION OF T-HELPER (TH)-1-RELATED CHEMOKINES INCLUDING INTERFERON-GAMMA-INDUCED PROTEIN-10 (IP-10) AND MONOCYTE CHEMOATTRACTANT PROTEIN-1 (MCP-1), TH2-RELATED CHEMOKINE MACROPHAGE-DERIVED CHEMOKINE, AND THE PROINFLAMMATORY CHEMOKINE TUMOR NECROSIS FACTOR-ALPHA WAS MEASURED USING ENZYME-LINKED IMMUNOSORBENT ASSAY. INTRACELLULAR SIGNALING PATHWAYS WERE INVESTIGATED USING WESTERN BLOT ANALYSIS AND CHROMATIN IMMUNOPRECIPITATION ASSAY. RESULTS: METFORMIN SUPPRESSED LPS-INDUCED IP-10 AND MCP-1 PRODUCTION AS WELL AS LPS-INDUCED PHOSPHORYLATION OF C-JUN N-TERMINAL KINASE (JNK), P38, EXTRACELLULAR SIGNAL-REGULATED KINASE (ERK), AND NUCLEAR FACTOR-KAPPA B (NF-KAPPAB). MOREOVER, METFORMIN SUPPRESSED LPS-INDUCED ACETYLATION OF HISTONES H3 AND H4 AT THE IP-10 PROMOTER. CONCLUSIONS: METFORMIN SUPPRESSED THE PRODUCTION OF TH1-RELATED CHEMOKINES IP-10 AND MCP-1 IN THP-1 CELLS. SUPPRESSIVE EFFECTS OF METFORMIN ON IP-10 PRODUCTION MIGHT BE ATTRIBUTED AT LEAST PARTIALLY TO THE JNK, P38, ERK, AND NF-KAPPAB PATHWAYS AS WELL AS TO EPIGENETIC REGULATION THROUGH THE ACETYLATION OF HISTONES H3 AND H4. THESE RESULTS INDICATED THE THERAPEUTIC ANTI-INFLAMMATORY POTENTIAL OF METFORMIN. 2018 17 2748 31 EXPRESSION AND FUNCTION OF EZH2 IN SYNOVIAL FIBROBLASTS: EPIGENETIC REPRESSION OF THE WNT INHIBITOR SFRP1 IN RHEUMATOID ARTHRITIS. OBJECTIVES: TO STUDY THE EXPRESSION, REGULATION AND FUNCTION OF THE HISTONE METHYLTRANSFERASE ENHANCER OF ZESTE HOMOLOGUE 2 (EZH2) IN SYNOVIAL FIBROBLASTS (SF) FROM PATIENTS WITH RHEUMATOID ARTHRITIS (RA) AND OSTEOARTHRITIS (OA). METHODS: SF WERE OBTAINED FROM RA AND OA PATIENTS UNDERGOING JOINT SURGERY. EXPRESSION LEVELS WERE ASSESSED BY QUANTITATIVE REAL-TIME PCR AND WESTERN BLOT. KINASE INHIBITORS AND REPORTER GENE ASSAYS WERE EMPLOYED TO STUDY SIGNALLING PATHWAYS. FUNCTIONAL ANALYSES INCLUDED EZH2 OVEREXPRESSION BY PLASMID TRANSFECTION AND GENE SILENCING BY SMALL INTERFERING RNA. CHROMATIN IMMUNOPRECIPITATION ASSAY WAS USED TO ANALYSE HISTONE METHYLATION WITHIN DISTINCT PROMOTER REGIONS. RESULTS: BY STUDYING THE EXPRESSION AND FUNCTION OF EZH2 IN SF THE AUTHORS FOUND THAT EZH2 IS OVEREXPRESSED IN RHEUMATOID ARTHRITIS SYNOVIAL FIBROBLASTS (RASF) AND FURTHER INDUCED BY TUMOUR NECROSIS FACTOR ALPHA THROUGH THE NUCLEAR FACTOR KAPPA B AND JUN KINASE PATHWAYS. AS A TARGET GENE OF EZH2 THE AUTHORS IDENTIFIED SECRETED FRIZZLED-RELATED PROTEIN 1 (SFRP1), AN INHIBITOR OF WNT SIGNALLING, WHICH IS ASSOCIATED WITH THE ACTIVATION OF RASF, AND SHOW THAT SFRP1 EXPRESSION CORRELATES WITH THE OCCUPATION OF ITS PROMOTER WITH ACTIVATING AND SILENCING HISTONE MARKS. CONCLUSIONS: THESE DATA STRONGLY SUGGEST THAT THE CHRONIC INFLAMMATORY ENVIRONMENT OF THE RA JOINT INDUCES EZH2 AND THUS MIGHT CAUSE CHANGES IN THE EPIGENETIC PROGRAMMES OF SF. 2011 18 2782 21 EZH2 INHIBITION CONFERS PIK3CA-DRIVEN LUNG TUMORS ENHANCED SENSITIVITY TO PI3K INHIBITION. MEMBERS OF THE PI3K SIGNALING PATHWAY, ESPECIALLY PIK3CA, THE GENE ENCODING THE CATALYTIC SUBUNIT OF THE PI3K COMPLEX, ARE HIGHLY MUTATED AND AMPLIFIED IN VARIOUS CANCER TYPES, INCLUDING NON-SMALL CELL LUNG CANCER. ALTHOUGH PI3K INHIBITORS HAVE BEEN USED IN CLINICS FOR FOLLICULAR LYMPHOMA AND CHRONIC LYMPHOCYTIC LEUKEMIA, NO AGENTS TARGETING PI3K ABERRATIONS IN LUNG CANCER HAVE BEEN APPROVED BY THE FDA SO FAR. IN THIS STUDY, WE OBSERVED THAT PIK3CA-E545K, THE MOST COMMON MUTATION IN LUNG CANCER, HARBORED A MODEST INDUCTION OF STEM-LIKE PROPERTIES IN LUNG EPITHELIAL CELLS, AND DROVE DEVELOPMENT OF ADENOCARCINOMA AUTOCHTHONOUSLY WHEN PAIRED WITH P53 LOSS IN A MURINE MOUSE MODEL. WE ALSO FOUND THAT PIK3CA-MUTANT OF AMPLIFIED LUNG CANCER CELLS WERE SENSITIVE TO EZH2 INHIBITION. EZH2 INHIBITION SYNERGIZED WITH PI3K INHIBITION IN HUMAN CANCER CELLS IN VITRO AND WORKED TOGETHER EFFICIENTLY IN VIVO. MECHANISTICALLY, EZH2 INHIBITION COOPERATED WITH PI3K INHIBITION TO PRODUCE A MORE POTENT SUPPRESSION OF PHOSPHO-AKT DOWNSTREAM OF PI3K. THIS STUDY SUGGESTS A PROMISING COMBINATION THERAPY TO COMBAT LUNG CANCERS WITH PIK3CA MUTATION OR AMPLIFICATION. BOTH COPANLISIB, THE PI3K INHIBITOR, AND TAZEMETOSTAT, THE EZH2 INHIBITOR, ARE FDA-APPROVED, WHICH SHOULD ENHANCE THE CLINICAL TRANSLATION OF THIS WORK. 2022 19 6231 30 THE LONG NONCODING RNA MEG3 AND ITS TARGET MIR-147 REGULATE JAK/STAT PATHWAY IN ADVANCED CHRONIC MYELOID LEUKEMIA. BACKGROUND: LONG NON-CODING (LNC) RNAS PLAYS AN IMPORTANT ROLE IN CHRONIC MYELOID LEUKEMIA (CML). IN THIS STUDY, WE AIMED TO UNCOVER THE MECHANISM OF THE LNCRNA MATERNALLY EXPRESSED 3 (MEG3) AND ITS TARGET MICRORNA-147 (MIR-147) IN CML. METHODS: SIXTY CML PATIENTS AND 10 HEALTHY DONORS WERE INCLUDED IN THE STUDY. THE METHYLATION OF MEG3 AND MIR-147 PROMOTER WAS DETERMINED BY METHYLATION-SPECIFIC PCR. THE RELATIONSHIP OF MEG3 AND MIR-147 WAS EXPLORED BY LUCIFERASE ASSAY. THE INTERACTIONS OF PROTEINS WERE STUDIED BY RNA PULL-DOWN ASSAY, RNA IMMUNOPRECIPITATION AND CO-IMMUNOPRECIPITATION. FINDINGS: PATIENTS IN ACCELERATED PHASE CML (CML-AP) AND BLAST PHASE CML (CML-BP) SHOWED LOWER EXPRESSIONS OF MEG3 AND MIR-147 AND HIGHER EXPRESSIONS OF DNMT1, DNMT3B, MBD2, MECP2 AND HDAC1 COMPARED TO THE CONTROLS. THESE PATIENTS ALSO SHOWED A HIGHER DEGREE OF METHYLATION OF MEG3 AND MIR-147 WHILE THERE WAS A REDUCTION AFTER CHIDAMIDE TREATMENT. FURTHERMORE, THE OVEREXPRESSION OF MEG3 AND MIR-147 INHIBITED CELL PROLIFERATION BOTH IN VIVO AND IN VITRO, PROMOTED APOPTOSIS AND DECREASED THE EXPRESSIONS OF DNMT1, DNMT3A, DNMT3B, MBD2, HDAC1 AND MECP2. WE ALSO FOUND MEG3 INTERACTED WITH DNMT1, JAK2, STAT3, HDAC1, AND TYK2, AND JAK2 WAS BOUND TO STAT3, STAT5 AND MYC. MORE INTERESTINGLY, JAK2 WAS BOUND TO TYK2 BY THE BRIDGE OF MEG3. INTERPRETATION: LNCRNA MEG3 AND ITS TARGET MIR-147 MAY SERVE AS A NOVEL THERAPEUTIC TARGET FOR CML BLAST CRISIS, AND CHIDAMIDE MIGHT HAVE A POTENTIAL CLINICAL APPLICATION IN TREATING CML BLAST CRISIS. 2018 20 2326 31 EPIGENETIC REGULATION OF HOTAIR IN ADVANCED CHRONIC MYELOID LEUKEMIA. PURPOSE: CHRONIC MYELOID LEUKEMIA (CML) ACCOUNTS FOR ~10% OF LEUKEMIA CASES, AND ITS PROGRESSION INVOLVES EPIGENETIC GENE REGULATION. THIS STUDY INVESTIGATED EPIGENETIC REGULATION OF HOTAIR AND ITS TARGET MICRORNA, MIR-143, IN ADVANCED CML. PATIENTS AND METHODS: WE FIRST ISOLATED BONE MARROW MONONUCLEAR CELLS FROM 70 PATIENTS WITH DIFFERENT PHASES OF CML AND FROM HEALTHY DONORS AS NORMAL CONTROL; WE ALSO CULTURED K562 AND KCL22 CELLS, TREATED WITH DEMETHYLATION DRUG; MTT ASSAY, FLOW CYTOMETRY, QUANTITATIVE REAL-TIME POLYMERASE CHAIN REACTION (QPCR), METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MSP), WESTERN BLOT, LUCIFERASE ASSAY, RNA PULL-DOWN ASSAY AND RNA-BINDING PROTEIN IMMUNOPRECIPITATION (RIP) ASSAY WERE PERFORMED. RESULT: AS MEASURED BY QPCR, HOTAIR EXPRESSION IN K562 CELLS, KCL22 CELLS, AND SAMPLES FROM CASES OF ADVANCED-STAGE CML INCREASED WITH LEVELS OF SEVERAL DNA METHYLTRANSFERASES AND HISTONE DEACETYLATES, INCLUDING DNMT1, DNMT3A, HDAC1, EZH2, AND LSD1, AND MIR-143 LEVELS WERE DECREASED AND HOTAIR LEVELS WERE INCREASED. TREATMENT WITH 5-AZACYTIDINE, A DNA METHYLATION INHIBITOR, DECREASED DNMT1, DNMT3A, HDAC1, EZH2, LSD1 MRNA, PROTEIN LEVELS, AND HOTAIR MRNA LEVELS BUT INCREASED MIR-143 LEVELS. HOTAIR KNOCKDOWN AND MIR-143 OVEREXPRESSION BOTH INHIBITED PROLIFERATION AND PROMOTED APOPTOSIS IN KCL22 AND K562 CELLS THROUGH THE PI3K/AKT PATHWAY. RNA PULL-DOWN, MASS SPECTROMETRY, AND RIP ASSAYS SHOWED THAT HOTAIR INTERACTED WITH EZH2 AND LSD1. A DUAL-LUCIFERASE ASSAY DEMONSTRATED THAT HOTAIR INTERACTED WITH MIR-143. CONCLUSION: OUR FINDINGS DEMONSTRATE THE KEY EPIGENETIC INTERACTIONS OF HOTAIR RELATED TO CML PROGRESSION AND SUGGEST HOTAIR AS A POTENTIAL THERAPEUTIC TARGET FOR ADVANCED CML. FURTHERMORE, OUR RESULTS SUPPORT THE USE OF DEMETHYLATION DRUGS AS A CML TREATMENT STRATEGY. 2018