1 3147 143 GLP OVEREXPRESSION IS ASSOCIATED WITH POOR PROGNOSIS IN CHRONIC LYMPHOCYTIC LEUKEMIA AND ITS INHIBITION INDUCES LEUKEMIC CELL DEATH. BACKGROUND HETERODIMERIC METHYLTRANSFERASES GLP (EHMT1/KMT1D) AND G9A (EHMT2/KMT1C) ARE TWO CLOSELY RELATED ENZYMES THAT PROMOTE THE MONOMETHYLATION AND DIMETHYLATION OF HISTONE H3 LYSINE 9. DYSREGULATION OF THEIR ACTIVITY HAS BEEN IMPLICATED IN SEVERAL TYPES OF HUMAN CANCER. PATIENTS AND METHODS HERE, IN ORDER TO INVESTIGATE WHETHER GLP/G9A EXERTS ANY IMPACT ON CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), GLP/G9A EXPRESSION LEVELS WERE ASSESSED IN A COHORT OF 50 PATIENTS AND THE EFFECTS OF THEIR INHIBITION WERE VERIFIED FOR THE VIABILITY OF CLL CELLS. ALSO, QRT-PCR WAS USED TO INVESTIGATE THE TRANSCRIPTIONAL LEVELS OF GLP/G9A IN CLL PATIENTS. IN ADDITION, PATIENT SAMPLES WERE CLASSIFIED ACCORDING TO ZAP-70 PROTEIN EXPRESSION BY FLOW CYTOMETRY AND ACCORDING TO KARYOTYPE INTEGRITY BY CYTOGENETICS ANALYSIS. FINALLY, A SELECTIVE SMALL MOLECULE INHIBITOR FOR GLP/G9A WAS USED TO ASCERTAIN WHETHER THESE METHYLTRANSFERASES INFLUENCED THE VIABILITY OF MEC-1 CLL CELL LINEAGE. RESULTS MRNA ANALYSIS REVEALED THAT CLL SAMPLES HAD HIGHER LEVELS OF GLP, BUT NOT G9A, WHEN COMPARED TO NON-LEUKEMIC CONTROLS. INTERESTINGLY, PATIENTS WITH UNFAVORABLE CYTOGENETICS SHOWED HIGHER EXPRESSION LEVELS OF GLP COMPARED TO PATIENTS WITH FAVORABLE KARYOTYPES. MORE IMPORTANTLY, GLP/G9A INHIBITION MARKEDLY INDUCED CELL DEATH IN CLL CELLS. CONCLUSION TAKEN TOGETHER, THESE RESULTS INDICATE THAT GLP IS ASSOCIATED WITH A WORSE PROGNOSIS IN CLL, AND THAT THE INHIBITION OF GLP/G9A INFLUENCES CLL CELL VIABILITY. ALTOGETHER, THE PRESENT DATA DEMONSTRATE THAT THESE METHYLTRANSFERASES CAN BE POTENTIAL MARKERS FOR DISEASE PROGRESSION, AS WELL AS A PROMISING EPIGENETIC TARGET FOR CLL TREATMENT AND THE PREVENTION OF DISEASE EVOLUTION. 2018 2 2063 32 EPIGENETIC CONTROL OF IL-23 EXPRESSION IN KERATINOCYTES IS IMPORTANT FOR CHRONIC SKIN INFLAMMATION. THE CHRONIC SKIN INFLAMMATION PSORIASIS IS CRUCIALLY DEPENDENT ON THE IL-23/IL-17 CYTOKINE AXIS. ALTHOUGH IL-23 IS EXPRESSED BY PSORIATIC KERATINOCYTES AND IMMUNE CELLS, ONLY THE IMMUNE CELL-DERIVED IL-23 IS BELIEVED TO BE DISEASE RELEVANT. HERE WE USE A GENETIC MOUSE MODEL TO SHOW THAT KERATINOCYTE-PRODUCED IL-23 IS SUFFICIENT TO CAUSE A CHRONIC SKIN INFLAMMATION WITH AN IL-17 PROFILE. FURTHERMORE, WE REVEAL A CELL-AUTONOMOUS NUCLEAR FUNCTION FOR THE ACTIN POLYMERIZING MOLECULE N-WASP, WHICH CONTROLS IL-23 EXPRESSION IN KERATINOCYTES BY REGULATING THE DEGRADATION OF THE HISTONE METHYLTRANSFERASES G9A AND GLP, AND H3K9 DIMETHYLATION OF THE IL-23 PROMOTER. THIS MECHANISM MEDIATES THE INDUCTION OF IL-23 BY TNF, A KNOWN INDUCER OF IL-23 IN PSORIASIS. FINALLY, IN KERATINOCYTES OF PSORIATIC LESIONS A DECREASE IN H3K9 DIMETHYLATION CORRELATES WITH INCREASED IL-23 EXPRESSION, SUGGESTING RELEVANCE FOR DISEASE. TAKEN TOGETHER, OUR DATA DESCRIBE A MOLECULAR PATHWAY WHERE EPIGENETIC REGULATION OF KERATINOCYTES CAN CONTRIBUTE TO CHRONIC SKIN INFLAMMATION. 2018 3 1384 26 DIABETES AND KIDNEY DISEASE: EMPHASIS ON TREATMENT WITH SGLT-2 INHIBITORS AND GLP-1 RECEPTOR AGONISTS. KIDNEY DISEASE IS A FREQUENT MICROVASCULAR COMPLICATION OF BOTH TYPE 1 AND TYPE 2 DIABETES. HISTORIC TRIALS HAVE DEMONSTRATED THAT A TIGHT GLYCAEMIC CONTROL IS THE MOST POWERFUL APPROACH TO DECREASE THE CHANCES OF DEVELOPING DIABETIC NEPHROPATHY. HOWEVER, HAVING AN HBA1C < 7% DOES NOT COMPLETELY SUPPRESS THE RISK OF KIDNEY DISEASE. THE OBSERVED RESIDUAL RISK IS LIKELY ASCRIBABLE TO TWO PHENOMENA: 1- THE PRESENCE OF RISK FACTORS AND ALTERATIONS ADDITIVE TO AND INDEPENDENT OF GLYCAEMIA, AND 2- THE ACTIVATION OF LONG-LASTING IMBALANCES BY PERIODS OF EXPOSURE TO UNCONTROLLED GLYCEMIA, A PHENOMENON REFERRED TO AS METABOLIC MEMORY OR LEGACY EFFECT. LONG-LASTING OXIDATIVE STRESS, EPIGENETIC ALTERATIONS, CELLULAR SENESCENCE, AND THE RESULTING CHRONIC LOW-GRADE INFLAMMATION ARE ALL CANDIDATE MECHANISMS EXPLAINING THE DEVELOPMENT OF NEPHROPATHY DESPITE PROPER CONTROL OF RISK FACTORS. RECENTLY, TWO CLASSES OF DRUGS, I.E. GLUCAGON-LIKE PEPTIDE (GLP) 1 RECEPTOR AGONISTS (RA) AND SODIUM-GLUCOSE TRANSPORTER 2 INHIBITORS (SGLT-I) HAVE CHANGED THIS SCENARIO. INDEED, CARDIOVASCULAR OUTCOME AND OTHER TRIALS HAVE CLEARLY SHOWN A RENOPROTECTIVE EFFECT FOR THESE DRUGS, WELL-BEYOND THEIR GLUCOSE-LOWERING PROPERTIES. IN THIS REVIEW, WE SUMMARIZE: 1- SELECTED KEY TRIALS AND MECHANISMS UNDERLYING THE DEVELOPMENT OF DIABETIC KIDNEY DISEASE AND 2- THE RESULTS RELATIVE TO RENAL ENDPOINTS IN CLINICAL TRIALS OF GLP-1 RA AND SGLT-2I. THEN, WE BRIEFLY DISCUSS SOME OF THE HYPOTHESES POSITED TO EXPLAIN THE MARKED RENOPROTECTIVE PROPERTIES OF THESE TWO CLASSES, EVIDENCING THE STILL EXISTING GAPS IN KNOWLEDGE AND PROPOSING FUTURE DIRECTIONS TO FURTHER IMPLEMENT THE USE OF THESE POWERFUL, DISEASE-MODIFYING DRUGS. 2021 4 2126 44 EPIGENETIC INACTIVATION OF MIR-34B/C IN ADDITION TO MIR-34A AND DAPK1 IN CHRONIC LYMPHOCYTIC LEUKEMIA. BACKGROUND: TP53 MUTATION/DELETION IS UNCOMMON IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). WE POSTULATED THAT COMPONENTS OF TP53-CENTERED TUMOR SUPPRESSOR NETWORK, MIR-34B/C, IN ADDITION TO DAPK1 AND MIR-34A MIGHT BE INACTIVATED BY DNA HYPERMETHYLATION. MOREOVER, WE TESTED IF MIR-34B/C METHYLATION MIGHT CORRELATE WITH MIR-203 OR MIR-124-1 METHYLATION IN CLL. METHODS: MIR-34B/C, MIR-34A AND DAPK1 METHYLATION WAS STUDIED IN 11 NORMAL CONTROLS, 7 CLL CELL LINES, AND 78 DIAGNOSTIC CLL SAMPLES BY METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION. MEC-1 CELLS WERE TREATED WITH 5-AZA-2'-DEOXYCYTIDINE FOR REVERSAL OF METHYLATION-ASSOCIATED MIRNA SILENCING. TUMOR SUPPRESSOR PROPERTIES OF MIR-34B WERE DEMONSTRATED BY OVER-EXPRESSION OF PRECURSOR MIR-34B IN MEC-1 CELLS. RESULTS: MIR-34B/C PROMOTER WAS UNMETHYLATED IN NORMAL CONTROLS, BUT COMPLETELY METHYLATED IN 4 CLL CELL LINES. MIR-34B/C EXPRESSION WAS INVERSELY CORRELATED WITH MIR-34B/C METHYLATION. DIFFERENT MSP STATUSES OF MIR-34B/C, INCLUDING COMPLETE METHYLATION AND COMPLETE UNMETHYLATION, WERE VERIFIED BY QUANTITATIVE BISULFITE PYROSEQUENCING. 5-AZA-2'-DEOXYCYTIDINE TREATMENT RESULTED IN PROMOTER DEMETHYLATION AND MIR-34B RE-EXPRESSION IN MEC1 CELLS. MOREOVER, OVER-EXPRESSION OF MIR-34B RESULTED IN INHIBITION OF CELLULAR PROLIFERATION AND INCREASED CELL DEATH. IN PRIMARY CLL SAMPLES, MIR-34A, MIR-34B/C AND DAPK1 METHYLATION WAS DETECTED IN 2.6%, 17.9% AND 34.6% OF PATIENTS AT DIAGNOSIS RESPECTIVELY. FURTHERMORE, 39.7%, 3.8% AND 2.6% PATIENTS HAD METHYLATION OF ONE, TWO OR ALL THREE GENES RESPECTIVELY. OVERALL, 46.2% PATIENTS HAD METHYLATION OF AT LEAST ONE OF THESE THREE GENES. BESIDES, MIR-34B/C METHYLATION WAS ASSOCIATED WITH METHYLATION OF MIR-34A (P = 0.03) AND MIR-203 (P = 0.012) IN CLL. CONCLUSIONS: TAKEN TOGETHER, MIR-34B/C IS A TUMOR SUPPRESSOR MIRNA FREQUENTLY METHYLATED, AND HENCE SILENCED IN CLL. TOGETHER WITH DAPK1 METHYLATION, MIR-34B/C METHYLATION IS IMPLICATED IN THE DISRUPTION OF THE TP53-CENTERED TUMOR SUPPRESSOR NETWORK. MOREOVER, THE ASSOCIATION OF MIRNA METHYLATION WARRANTS FURTHER STUDY. 2014 5 273 40 AGE-INDUCED SUPPRESSION OF EZH2 MEDIATES INJURY OF PODOCYTES BY REDUCING H3K27ME3. BACKGROUND: CHRONIC HYPERGLYCEMIA, A PIVOTAL FEATURE OF DIABETES MELLITUS (DM), INITIATES THE FORMATION OF ADVANCED GLYCATION END PRODUCTS (AGES) AND THE DYSREGULATION OF EPIGENETIC MECHANISMS, WHICH MAY CAUSE INJURY TO RENAL PODOCYTES, A CENTRAL FEATURE OF DIABETIC KIDNEY DISEASE (DKD). PREVIOUS DATA OF OUR GROUP SHOWED THAT AGES SIGNIFICANTLY REDUCE THE EXPRESSION OF NIPP1 (NUCLEAR INHIBITOR OF PROTEIN PHOSPHATASE 1) IN PODOCYTES IN VITRO AS WELL AS IN HUMAN AND MURINE DKD. NIPP1 WAS SHOWN BY OTHERS TO INTERACT WITH ENHANCER OF ZESTE HOMOLOG 2 (EZH2), WHICH CATALYZES THE REPRESSIVE METHYLATION OF H3K27ME3 ON HISTONE 3. THEREFORE, WE HYPOTHESIZED THAT AGES CAN DIRECTLY INDUCE EPIGENETIC CHANGES IN PODOCYTES. METHODS: WE ANALYZED THE RELEVANCE OF AGES ON EZH2 EXPRESSION AND ACTIVITY IN A MURINE PODOCYTE CELL LINE. CELLS WERE TREATED WITH 5 MG/ML GLYCATED BSA FOR 24 H. TO DETERMINE THE MEANING OF EZH2 SUPPRESSION, EZH2 ACTIVITY WAS INHIBITED BY INCUBATING THE CELLS WITH THE PHARMACOLOGICAL METHYLTRANSFERASE INHIBITOR 3-DEAZANEPLANOCIN A; EZH2 EXPRESSION WAS REPRESSED WITH SIRNA. MRNA EXPRESSION WAS ANALYZED WITH REAL-TIME PCR, AND PROTEIN EXPRESSION WITH WESTERN BLOT. EZH2 EXPRESSION AND LEVEL OF H3K27 TRIMETHYLATION IN PODOCYTES OF DIABETIC DB/DB MICE, A MOUSE MODEL FOR TYPE 2 DM, WERE ANALYZED USING IMMUNOFLUORESCENCE. RESULTS: OUR DATA DEMONSTRATED THAT AGES DECREASE EZH2 EXPRESSION IN PODOCYTES AND CONSEQUENTLY REDUCE H3K27ME3. THIS SUPPRESSION OF EZH2 MIMICKED THE AGE EFFECTS AND CAUSED AN UPREGULATED EXPRESSION OF PATHOLOGICAL FACTORS THAT CONTRIBUTE TO PODOCYTE INJURY IN DKD. IN ADDITION, ANALYSES OF DB/DB MICE SHOWED SIGNIFICANTLY REDUCED H3K27ME3 AND EZH2 EXPRESSION IN PODOCYTES. MOREOVER, THE SUPPRESSION OF NIPP1 AND EZH2 SHOWED SIMILAR EFFECTS REGARDING PODOCYTE INJURY. CONCLUSIONS: OUR STUDIES PROVIDE A NOVEL PATHWAY HOW AGES CONTRIBUTE TO PODOCYTE INJURY AND THE FORMATION OF THE SO-CALLED METABOLIC MEMORY IN DKD. 2020 6 6171 41 THE HDAC INHIBITOR VALPROATE INDUCES A BIVALENT STATUS OF THE CD20 PROMOTER IN CLL PATIENTS SUGGESTING DISTINCT EPIGENETIC REGULATION OF CD20 EXPRESSION IN CLL IN VIVO. TREATMENT WITH ANTI-CD20 ANTIBODIES IS ONLY MODERATELY EFFICIENT IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), A FEATURE WHICH HAS BEEN EXPLAINED BY THE INHERENTLY LOW CD20 EXPRESSION IN CLL. IT HAS BEEN SHOWN THAT CD20 IS EPIGENETICALLY REGULATED AND THAT HISTONE DEACETYLASE INHIBITORS (HDACIS) CAN INCREASE CD20 EXPRESSION IN VITRO IN CLL. TO ASSESS WHETHER HDACIS CAN UPREGULATE CD20 ALSO IN VIVO IN CLL, THE HDACI VALPROATE WAS GIVEN TO THREE DEL13Q/NOTCH1WT CLL PATIENTS AND CD20 LEVELS WERE ANALYSED (THE PREVAIL STUDY). VALPROATE TREATMENT RESULTED IN EXPECTED GLOBAL ACTIVATING HISTONE MODIFICATIONS SUGGESTING HDAC INHIBITORY EFFECTS. HOWEVER, ALTHOUGH VALPROATE INDUCED EXPRESSION OF CD20 MRNA AND PROTEIN IN THE DEL13Q/NOTCH1WT I83-E95 CLL CELL LINE, NO SUCH EFFECTS WERE OBSERVED IN THE PATIENTS STUDIED. IN CONTRAST TO THE CELL LINE, IN PATIENTS VALPROATE TREATMENT RESULTED IN TRANSIENT RECRUITMENT OF THE TRANSCRIPTIONAL REPRESSOR EZH2 TO THE CD20 PROMOTER, CORRELATING TO AN INCREASE OF THE REPRESSIVE HISTONE MARK H3K27ME3. THIS SUGGESTS THAT VALPROATE-MEDIATED INDUCTION OF CD20 MAY BE HAMPERED BY EZH2 MEDIATED H3K27ME3 IN VIVO IN CLL. MOREOVER, VALPROATE TREATMENT RESULTED IN INDUCTION OF EZH2 AND GLOBAL H3K27ME3 IN PATIENT CELLS, SUGGESTING TRANSCRIPTIONALLY REPRESSIVE EFFECTS OF VALPROATE IN CLL. OUR RESULTS SUGGEST NEW IN VIVO MECHANISMS OF HDACIS WHICH MAY HAVE IMPLICATIONS ON THE DESIGN OF FUTURE CLINICAL TRIALS IN B-CELL MALIGNANCIES. 2017 7 66 41 A KEY ROLE FOR EZH2 IN EPIGENETIC SILENCING OF HOX GENES IN MANTLE CELL LYMPHOMA. THE CHROMATIN MODIFIER EZH2 IS OVEREXPRESSED AND ASSOCIATED WITH INFERIOR OUTCOME IN MANTLE CELL LYMPHOMA (MCL). RECENTLY, WE DEMONSTRATED PREFERENTIAL DNA METHYLATION OF HOX GENES IN MCL COMPARED WITH CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), DESPITE THESE GENES NOT BEING EXPRESSED IN EITHER ENTITY. SINCE EZH2 HAS BEEN SHOWN TO REGULATE HOX GENE EXPRESSION, TO GAIN FURTHER INSIGHT INTO ITS POSSIBLE ROLE IN DIFFERENTIAL SILENCING OF HOX GENES IN MCL VS. CLL, WE PERFORMED DETAILED EPIGENETIC CHARACTERIZATION USING REPRESENTATIVE CELL LINES AND PRIMARY SAMPLES. WE OBSERVED SIGNIFICANT OVEREXPRESSION OF EZH2 IN MCL VS. CLL. CHROMATIN IMMUNE PRECIPITATION (CHIP) ASSAYS REVEALED THAT EZH2 CATALYZED REPRESSIVE H3 LYSINE 27 TRIMETHYLATION (H3K27ME3), WHICH WAS SUFFICIENT TO SILENCE HOX GENES IN CLL, WHEREAS IN MCL H3K27ME3 IS ACCOMPANIED BY DNA METHYLATION FOR A MORE STABLE REPRESSION. MORE IMPORTANTLY, HYPERMETHYLATION OF THE HOX GENES IN MCL RESULTED FROM EZH2 OVEREXPRESSION AND SUBSEQUENT RECRUITMENT OF THE DNA METHYLATION MACHINERY ONTO HOX GENE PROMOTERS. THE IMPORTANCE OF EZH2 UPREGULATION IN THIS PROCESS WAS FURTHER UNDERSCORED BY SIRNA TRANSFECTION AND EZH2 INHIBITOR EXPERIMENTS. ALTOGETHER, THESE OBSERVATIONS IMPLICATE EZH2 IN THE LONG-TERM SILENCING OF HOX GENES IN MCL, AND ALLUDE TO ITS POTENTIAL AS A THERAPEUTIC TARGET WITH CLINICAL IMPACT. 2013 8 826 37 CHARACTERIZATION OF K562 CELLS: UNCOVERING NOVEL CHROMOSOMES, ASSESSING TRANSFERRIN RECEPTOR EXPRESSION, AND PROBING PHARMACOLOGICAL THERAPIES. HUMAN ERYTHROLEUKEMIC K562 CELLS REPRESENT THE PROTOTYPICAL CELL CULTURE MODEL OF CHRONIC MYELOID LEUKEMIA (CML). THE CELLS ARE PSEUDO-TRIPLOID AND POSITIVE FOR THE PHILADELPHIA CHROMOSOME. THEREFORE, K562 CELLS HAVE BEEN WIDELY USED FOR INVESTIGATING THE BCR/ABL1 ONCOGENE AND THE TYROSINE KINASE INHIBITOR, IMATINIB-MESYLATE. FURTHER, K562 CELLS OVEREXPRESS TRANSFERRIN RECEPTORS (TFR) AND HAVE BEEN USED AS A MODEL FOR TARGETING CYTOTOXIC THERAPIES, VIA RECEPTOR-MEDIATED ENDOCYTOSIS. HERE, WE HAVE CHARACTERIZED K562 CELLS FOCUSING ON THE KARYOTYPE OF CELLS IN PROLONGED CULTURE, REGULATION OF EXPRESSION OF TFR IN WILDTYPE (WT) AND DOXORUBICIN-RESISTANT CELLS, AND RESPONSES TO HISTONE DEACETYLASE INHIBITION (HDACI). KARYOTYPE ANALYSIS INDICATES NOVEL CHROMOSOMES AND GENE EXPRESSION ANALYSIS SUGGESTS A SHIFT OF CULTURED K562 CELLS AWAY FROM PATIENT-DERIVED LEUKEMIC CELLS. WE CONFIRM THE HIGH EXPRESSION OF TFR ON K562 CELLS USING IMMUNOFLUORESCENCE AND CELL-SURFACE RECEPTOR BINDING RADIOASSAYS. IMPORTANTLY, HIGH TFR EXPRESSION IS OBSERVED IN PATIENT-DERIVED CELLS, AND WE HIGHLIGHT THE PERSISTENT EXPRESSION OF TFR FOLLOWING DOXORUBICIN ACQUIRED RESISTANCE. EPIGENETIC ANALYSIS INDICATES THAT PERMISSIVE HISTONE ACETYLATION AND METHYLATION AT THE PROMOTER REGION REGULATES THE TRANSCRIPTION OF TFR IN K562 CELLS. FINALLY, WE SHOW RELATIVELY HIGH EXPRESSION OF HDAC ENZYMES IN K562 CELLS AND DEMONSTRATE THE CHEMOTOXIC EFFECTS OF HDACI, USING THE FDA-APPROVED HYDROXAMIC ACID, VORINOSTAT. TOGETHER WITH A DESCRIPTION OF MORPHOLOGY, INFRARED SPECTRAL ANALYSIS, AND EXAMINATION OF METABOLIC PROPERTIES, WE PROVIDE A COMPREHENSIVE CHARACTERIZATION OF K562 CELLS. OVERALL, K562 CELL CULTURE SYSTEMS REMAIN WIDELY USED FOR THE INVESTIGATION OF NOVEL THERAPEUTICS FOR CML, WHICH IS PARTICULARLY IMPORTANT IN CASES OF IMATINIB-MESYLATE RESISTANCE. 2023 9 6177 34 THE HISTONE METHYLTRANSFERASE G9A MEDIATES STRESS-REGULATED ALCOHOL DRINKING. THE EPIGENETIC ENZYME G9A IS A HISTONE METHYLTRANSFERASE THAT DIMETHYLATES LYSINE 9 ON HISTONE H3 (H3K9ME2), AND IN THE ADULT NUCLEUS ACCUMBENS (NAC), G9A REGULATES MULTIPLE BEHAVIORS ASSOCIATED WITH SUBSTANCE USE DISORDER. WE SHOW HERE THAT CHRONIC INTERMITTENT ETHANOL (CIE) EXPOSURE IN MALE MICE REDUCED BOTH G9A AND H3K9ME2 LEVELS IN THE ADULT NAC, BUT NOT DORSAL STRIATUM. VIRAL-MEDIATED REDUCTION OF G9A IN THE NAC HAD NO EFFECTS ON BASELINE VOLITIONAL ETHANOL DRINKING OR ESCALATED ALCOHOL DRINKING PRODUCED BY CIE EXPOSURE; HOWEVER, NAC G9A WAS REQUIRED FOR STRESS-REGULATED CHANGES IN ETHANOL DRINKING, INCLUDING POTENTIATED ALCOHOL DRINKING PRODUCED BY ACTIVATION OF THE KAPPA-OPIOID RECEPTOR. IN ADDITION, WE OBSERVED THAT CHRONIC SYSTEMIC ADMINISTRATION OF A G9A INHIBITOR, UNC0642, ALSO BLOCKED STRESS-POTENTIATED ALCOHOL DRINKING. TOGETHER, OUR FINDINGS SUGGEST THAT CHRONIC ALCOHOL USE, SIMILAR TO OTHER ABUSED SUBSTANCES, PRODUCES A NAC-SELECTIVE REDUCTION IN G9A LEVELS THAT SERVES TO LIMIT STRESS-REGULATED ALCOHOL DRINKING. MOREOVER, OUR FINDINGS SUGGEST THAT PHARMACOLOGICAL INHIBITION OF G9A MIGHT PROVIDE A NOVEL THERAPEUTIC APPROACH TO TREAT STRESS-INDUCED ALCOHOL DRINKING, WHICH IS A MAJOR TRIGGER OF RELAPSE IN INDIVIDUALS SUFFERING FROM AUD. 2022 10 5716 34 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 11 3531 31 IMATINIB CAUSES EPIGENETIC ALTERATIONS OF PTEN GENE VIA UPREGULATION OF DNA METHYLTRANSFERASES AND POLYCOMB GROUP PROTEINS. WE HAVE RECENTLY REPORTED THE POSSIBLE IMATINIB-RESISTANT MECHANISM; LONG-TERM EXPOSURE OF LEUKEMIA CELLS TO IMATINIB DOWNREGULATED LEVELS OF PHOSPHATASE AND TENSIN HOMOLOG DELETED ON CHROMOSOME 10 (PTEN) VIA HYPERMETHYLATION OF ITS PROMOTER REGION (LEUKEMIA 2010; 24: 1631). THE PRESENT STUDY EXPLORED THE MOLECULAR MECHANISMS BY WHICH IMATINIB CAUSED METHYLATION ON THE PROMOTER REGION OF THIS TUMOR SUPPRESSOR GENE IN LEUKEMIA CELLS. REAL-TIME REVERSE TRANSCRIPTION PCR FOUND THAT LONG-TERM EXPOSURE OF CHRONIC EOSINOPHILIC LEUKEMIA EOL-1 CELLS EXPRESSING FIP1L1/PLATELET-DERIVED GROWTH FACTOR RECEPTOR-ALPHA TO IMATINIB INDUCED EXPRESSION OF DNA METHYLTRANSFERASE 3A (DNMT3A) AND HISTONE-METHYLTRANSFERASE ENHANCER OF ZESTE HOMOLOG 2 (EZH2), A FAMILY OF POLYCOMB GROUP, THEREBY INCREASING METHYLATION OF THE GENE. IMMUNOPRECIPITATION ASSAY FOUND THE INCREASED COMPLEX FORMATION OF DNMT3A AND EZH2 PROTEINS IN THESE CELLS. MOREOVER, CHROMATIN IMMUNOPRECIPITATION ASSAY SHOWED THAT AMOUNTS OF BOTH DNMT3A AND EZH2 PROTEINS BOUND AROUND THE PROMOTER REGION OF PTEN GENE WERE INCREASED IN EOL-1 CELLS AFTER EXPOSURE TO IMATINIB. FURTHERMORE, WE FOUND THAT LEVELS OF DNMT3A AND EZH2 WERE STRIKINGLY INCREASED IN LEUKEMIA CELLS ISOLATED FROM INDIVIDUALS WITH CHRONIC MYELOGENOUS LEUKEMIA (N=1) AND PHILADELPHIA CHROMOSOME-POSITIVE ACUTE LYMPHOBLASTIC LEUKEMIA (N=2), WHO RELAPSED AFTER TREATMENT WITH IMATINIB COMPARED WITH THOSE ISOLATED AT THEIR INITIAL PRESENTATION. TAKEN TOGETHER, IMATINIB COULD CAUSE DRUG-RESISTANCE VIA RECRUITMENT OF POLYCOMB GENE COMPLEX TO THE PROMOTER REGION OF THE PTEN AND DOWNREGULATION OF THIS GENE'S TRANSCRIPTS IN LEUKEMIA PATIENTS. 2011 12 1669 45 DOWNREGULATION OF THE HISTONE METHYLTRANSFERASE SETD2 PROMOTES IMATINIB RESISTANCE IN CHRONIC MYELOID LEUKAEMIA CELLS. OBJECTIVES: EPIGENETIC MODIFIERS WERE IMPORTANT PLAYERS IN THE DEVELOPMENT OF HAEMATOLOGICAL MALIGNANCIES AND SENSITIVITY TO THERAPY. MUTATIONS OF SET DOMAIN-CONTAINING 2 (SETD2), A METHYLTRANSFERASE THAT CATALYSES THE TRIMETHYLATION OF HISTONE 3 ON LYSINE 36 (H3K36ME3), WERE FOUND IN VARIOUS MYELOID MALIGNANCIES. HOWEVER, THE DETAILED MECHANISMS THROUGH WHICH SETD2 CONFERS CHRONIC MYELOID LEUKAEMIA PROGRESSION AND RESISTANCE TO THERAPY TARGETING ON BCR-ABL REMAIN UNCLEAR. MATERIALS AND METHODS: THE LEVEL OF SETD2 IN IMATINIB-SENSITIVE AND IMATINIB-RESISTANT CHRONIC MYELOID LEUKAEMIA (CML) CELLS WAS EXAMINED BY IMMUNOBLOTTING AND QUANTITATIVE REAL-TIME PCR. WE ANALYSED CD34(+) CD38(-) LEUKAEMIC STEM CELLS BY FLOW CYTOMETRY AND COLONY FORMATION ASSAYS UPON SETD2 KNOCKDOWN OR OVEREXPRESSION. THE IMPACT OF SETD2 EXPRESSION ALTERATIONS OR SMALL-MOLECULE INHIBITOR JIB-04 TARGETING H3K36ME3 LOSS ON IMATINIB SENSITIVITY WAS ASSESSED BY IC50, CELL APOPTOSIS AND PROLIFERATION ASSAYS. FINALLY, RNA SEQUENCING AND CHIP-QUANTITATIVE PCR WERE PERFORMED TO VERIFY PUTATIVE DOWNSTREAM TARGETS. RESULTS: SETD2 WAS FOUND TO ACT AS A TUMOUR SUPPRESSOR IN CML. THE NOVEL ONCOGENIC TARGETS MYCN AND ERG WERE SHOWN TO BE THE DIRECT DOWNSTREAM TARGETS OF SETD2, WHERE THEIR OVEREXPRESSION INDUCED BY SETD2 KNOCKDOWN CAUSED IMATINIB INSENSITIVITY AND LEUKAEMIC STEM CELL ENRICHMENT IN CML CELL LINES. TREATMENT WITH JIB-04, AN INHIBITOR THAT RESTORES H3K36ME3 LEVELS THROUGH BLOCKADE OF ITS DEMETHYLATION, SUCCESSFULLY IMPROVED THE CELL IMATINIB SENSITIVITY AND ENHANCED THE CHEMOTHERAPEUTIC EFFECT. CONCLUSIONS: OUR STUDY NOT ONLY EMPHASIZES THE REGULATORY MECHANISM OF SETD2 IN CML, BUT ALSO PROVIDES PROMISING THERAPEUTIC STRATEGIES FOR OVERCOMING THE IMATINIB RESISTANCE IN PATIENTS WITH CML. 2019 13 6704 29 VHL GENE METHYLATION CONTRIBUTES TO EXCESSIVE ERYTHROCYTOSIS IN CHRONIC MOUNTAIN SICKNESS RAT MODEL BY UPREGULATING THE HIF-2ALPHA/EPO PATHWAY. AIMS: HYPOXIA-INDUCIBLE FACTORS (HIFS) PLAY IMPORTANT ROLES IN THE PATHOGENESIS OF ERYTHROCYTOSIS IN CHRONIC MOUNTAIN SICKNESS (CMS). VON HIPPEL-LINDAU (VHL) IS A KEY REGULATOR OF HYPOXIA THAT CAN DIRECT THE POLY-UBIQUITYLATION AND DEGRADATION OF HIFS. EPIGENETIC MECHANISMS ARE BELIEVED TO CONTRIBUTE TOWARD ADAPTION TO CHRONIC HYPOXIA. HERE, WE INVESTIGATED THE CONTRIBUTION AND MECHANISM OF VHL METHYLATION IN RATS WITH ERYTHROCYTOSIS IN CMS. MAIN METHODS: THE METHYLATION STATUS OF VHL WAS MEASURED VIA BISULFITE SEQUENCING PCR, WHILE VHL, DNMT1, DNMT3ALPHA, AND DNMT3BETA EXPRESSION WERE ASSESSED USING REAL-TIME REVERSE TRANSCRIPTION PCR AND WESTERN BLOTTING. HIF-2ALPHA AND EPO EXPRESSION LEVELS IN BONE MARROW WERE DETERMINED VIA IMMUNOHISTOCHEMICAL STAINING, AND ERYTHROID HYPERPLASIA IN BONE MARROW SECTIONS WERE OBSERVED WITH HEMATOXYLIN AND EOSIN STAINING. KEY FINDINGS: WE FOUND THAT CHRONIC HYPOXIA TRIGGERED ERYTHROID HYPERPLASIA IN THE BONE MARROW AND INCREASED THE QUANTITY OF PERIPHERAL RED BLOOD CELLS IN CMS RATS. CHRONIC HYPOXIA SIGNIFICANTLY INDUCED METHYLATION AT THE CPG SITE IN THE VHL PROMOTER, DECREASED VHL EXPRESSION, AND INCREASED HIF-2ALPHA AND EPO EXPRESSION. CHRONIC HYPOXIA INCREASED DNMT3ALPHA AND DNMT3BETA EXPRESSION, CONSISTENT WITH THE DECREASE IN VHL EXPRESSION. THE DNA METHYLTRANSFERASE INHIBITOR 5-AZACYTIDINE REDUCED CHRONIC HYPOXIA-INDUCED ERYTHROID PROLIFERATION IN THE BONE MARROW OF RATS WITH CMS BY SUPPRESSING VHL METHYLATION AND DNMTS EXPRESSION. SIGNIFICANCE: OUR STUDY SUGGESTS THAT VHL METHYLATION CONTRIBUTES TOWARD EXCESSIVE ERYTHROCYTOSIS IN CMS BY UPREGULATING THE HIF-2ALPHA/EPO PATHWAY IN THE BONE MARROW OF RATS. WE DEMONSTRATED THAT THE DNMT INHIBITOR 5-AZACYTIDINE CAN ATTENUATE ERYTHROID HYPERPLASIA IN THE BONE MARROW BY DEMETHYLATING THE VHL PROMOTER. 2021 14 3323 36 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 15 2326 43 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 16 2787 41 EZH2-MEDIATED EPIGENETIC MODIFICATION IS REQUIRED FOR ALLOGENEIC T CELL-INDUCED LUPUS DISEASE. BACKGROUND: THE MECHANISMS INVOLVED IN THE PATHOGENESIS OF AUTOIMMUNE DISORDERS, INCLUDING SYSTEMIC LUPUS ERYTHEMATOSUS (SLE), HAVE NOT BEEN FULLY ELUCIDATED. SOME OF THESE MECHANISMS INVOLVE EPIGENETIC REGULATION OF GENE EXPRESSION. THE HISTONE METHYLTRANSFERASE EZH2 CONTRIBUTES TO EPIGENETIC REGULATION OF GENE EXPRESSION, IS HIGHLY EXPRESSED IN GERMINAL CENTER (GC) B CELLS AND FOLLICULAR T HELPER (T(FH)) CELLS, AND MAY BE INVOLVED IN LUPUS PATHOGENESIS. METHODS: THE MURINE BM12 MODEL OF LUPUS-LIKE CHRONIC GRAFT VERSUS HOST DISEASE (CGVHD) WAS INDUCED BY INTRA-PERITONEAL INJECTION OF NEGATIVELY ISOLATED ALLOGENEIC CD4(+) T CELLS. LUPUS-LIKE DISEASE DEVELOPMENT WAS MONITORED BY ELISA DETERMINATION OF SERUM ANTI-DSDNA AND ANTI-CHROMATIN ANTIBODY TITERS. IMMUNE CELL ACTIVATION AND EZH2 EXPRESSION WERE EVALUATED BY FLOW CYTOMETRY AND WESTERN BLOTTING. RESULTS: DECREASED AUTOANTIBODY PRODUCTION AND GC FORMATION ARE OBSERVED WHEN EZH2-DEFICIENT CD4(+) T CELLS ARE USED INSTEAD OF WILD-TYPE (WT) TO INDUCE CGVHD AND WHEN MICE THAT RECEIVE ALLOGENEIC WT DONOR T CELLS TO INDUCE CGVHD ARE TREATED WITH GSK503, AN EZH2-SPECIFIC INHIBITOR. IN THE BM12 CGVHD MODEL, WT DONOR T CELLS ARE NORMALLY FULLY ACTIVATED 1 WEEK AFTER INFUSION INTO AN ALLOGENEIC HOST, EXHIBIT A T(FH) CELL (PD-1(HI)/CXCR5(HI)) PHENOTYPE WITH UPREGULATED EZH2, AND ACTIVATE B CELLS TO FORM GERMINAL CENTERS (GCS). IN CONTRAST, EZH2-DEFICIENT DONOR T CELLS GENERATE FEWER T(FH) CELLS THAT FAIL TO ACTIVATE B CELLS OR PROMOTE GC FORMATION. DESPITE SIMILAR T-INDEPENDENT, LPS-INDUCED B CELL RESPONSES, OVA-IMMUNIZED CD4.EZH2-KO MICE HAD A SKEWED LOW-AFFINITY IGM PHENOTYPE IN COMPARISON TO SIMILARLY TREATED WT MICE. IN ADDITION, EARLY AFTER OVA IMMUNIZATION, MORE CD4(+) T CELLS FROM B6.CD4.EZH2-KO MICE HAD A CD44(LO)/CD62L(LO) PHENOTYPE, WHICH SUGGESTS ARRESTED OR DELAYED ACTIVATION, THAN CD4(+) T CELLS FROM OVALBUMIN-IMMUNIZED B6.WT MICE. CONCLUSION: EZH2 GENE DELETION OR PHARMACOLOGICAL EZH2 INHIBITION SUPPRESSES AUTOANTIBODY PRODUCTION AND GC FORMATION IN BM12 LUPUS-LIKE CGVHD AND DECREASES AFFINITY MATURATION AND ISOTYPE SWITCHING IN RESPONSE TO IMMUNIZATION WITH A T CELL-DEPENDENT ANTIGEN. EZH2 INHIBITION MAY BE USEFUL FOR THE TREATMENT OF LUPUS AND OTHER AUTOIMMUNE DISORDERS. 2020 17 2825 44 FLOW-DEPENDENT EPIGENETIC REGULATION OF IGFBP5 EXPRESSION BY H3K27ME3 CONTRIBUTES TO ENDOTHELIAL ANTI-INFLAMMATORY EFFECTS. RATIONALE: ATHEROSCLEROSIS IS A CHRONIC INFLAMMATORY AND EPIGENETIC DISEASE THAT IS INFLUENCED BY DIFFERENT PATTERNS OF BLOOD FLOW. HOWEVER, THE EPIGENETIC MECHANISM WHEREBY ATHEROPROTECTIVE FLOW CONTROLS ENDOTHELIAL GENE PROGRAMMING REMAINS ELUSIVE. HERE, WE INVESTIGATED THE POSSIBILITY THAT FLOW ALTERS ENDOTHELIAL GENE EXPRESSION THROUGH EPIGENETIC MECHANISMS. METHODS: EN FACE STAINING AND WESTERN BLOT WERE USED TO DETECT PROTEIN EXPRESSION. REAL-TIME PCR WAS USED TO DETERMINE RELATIVE GENE EXPRESSION. RNA-SEQUENCING OF HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS TREATED WITH SIRNA OF ENHANCER OF ZESTE HOMOLOG 2 (EZH2) OR LAMINAR FLOW WAS USED FOR TRANSCRIPTIONAL PROFILING. RESULTS: WE FOUND THAT TRIMETHYLATION OF HISTONE 3 LYSINE 27 (H3K27ME3), A REPRESSIVE EPIGENETIC MARK THAT ORCHESTRATES GENE REPRESSION, WAS REDUCED IN LAMINAR FLOW AREAS OF MOUSE AORTA AND FLOW-TREATED HUMAN ENDOTHELIAL CELLS. THE DECREASE OF H3K27ME3 PARALLELED A REDUCTION IN THE EPIGENETIC "WRITER"-EZH2, THE CATALYTIC SUBUNIT OF THE POLYCOMB REPRESSIVE COMPLEX 2 (PRC2). MOREOVER, LAMINAR FLOW DECREASED EXPRESSION OF EZH2 VIA MECHANOSENSITIVE MIR101. GENOME-WIDE TRANSCRIPTOME PROFILING STUDIES IN ENDOTHELIAL CELLS TREATED WITH EZH2 SIRNA AND FLOW REVEALED THE UPREGULATION OF NOVEL MECHANOSENSITIVE GENE IGFBP5 (INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN 5), WHICH IS EPIGENETICALLY SILENCED BY H3K27ME3. FUNCTIONALLY, INHIBITION OF H3K27ME3 BY EZH2 SIRNA OR GSK126 (A SPECIFIC EZH2 INHIBITOR) REDUCED H3K27ME3 LEVELS AND MONOCYTE ADHESION TO ENDOTHELIAL CELLS. ADENOVIRAL OVEREXPRESSION OF IGFBP5 ALSO RECAPITULATED THE ANTI-INFLAMMATORY EFFECTS OF H3K27ME3 INHIBITION. MORE IMPORTANTLY, WE OBSERVED EZH2 UPREGULATION, AND IGFBP5 DOWNREGULATION, IN ADVANCED ATHEROSCLEROTIC PLAQUES FROM HUMAN PATIENTS. CONCLUSION: TAKEN TOGETHER, OUR FINDINGS REVEAL THAT ATHEROPROTECTIVE FLOW REDUCES H3K27ME3 AS A CHROMATIN-BASED MECHANISM TO AUGMENT THE EXPRESSION OF GENES THAT CONFER AN ANTI-INFLAMMATORY RESPONSE IN THE ENDOTHELIUM. OUR STUDY EXEMPLIFIES FLOW-DEPENDENT EPIGENETIC REGULATION OF ENDOTHELIAL GENE EXPRESSION, AND ALSO SUGGESTS THAT TARGETING THE EZH2/H3K27ME3/IGFBP5 PATHWAY MAY OFFER NOVEL THERAPEUTICS FOR INFLAMMATORY DISORDERS SUCH AS ATHEROSCLEROSIS. 2018 18 5675 26 SHIFTS IN PODOCYTE HISTONE H3K27ME3 REGULATE MOUSE AND HUMAN GLOMERULAR DISEASE. HISTONE PROTEIN MODIFICATIONS CONTROL FATE DETERMINATION DURING NORMAL DEVELOPMENT AND DEDIFFERENTIATION DURING DISEASE. HERE, WE SET OUT TO DETERMINE THE EXTENT TO WHICH DYNAMIC CHANGES TO HISTONES AFFECT THE DIFFERENTIATED PHENOTYPE OF ORDINARILY QUIESCENT ADULT GLOMERULAR PODOCYTES. TO DO THIS, WE EXAMINED THE CONSEQUENCES OF SHIFTING THE BALANCE OF THE REPRESSIVE HISTONE H3 LYSINE 27 TRIMETHYLATION (H3K27ME3) MARK IN PODOCYTES. ADRIAMYCIN NEPHROTOXICITY AND SUBTOTAL NEPHRECTOMY (SNX) STUDIES INDICATED THAT DELETION OF THE HISTONE METHYLATING ENZYME EZH2 FROM PODOCYTES DECREASED H3K27ME3 LEVELS AND SENSITIZED MICE TO GLOMERULAR DISEASE. H3K27ME3 WAS ENRICHED AT THE PROMOTER REGION OF THE NOTCH LIGAND JAG1 IN PODOCYTES, AND DEREPRESSION OF JAG1 BY EZH2 INHIBITION OR KNOCKDOWN FACILITATED PODOCYTE DEDIFFERENTIATION. CONVERSELY, INHIBITION OF THE JUMONJI C DOMAIN-CONTAINING DEMETHYLASES JMJD3 AND UTX INCREASED THE H3K27ME3 CONTENT OF PODOCYTES AND ATTENUATED GLOMERULAR DISEASE IN ADRIAMYCIN NEPHROTOXICITY, SNX, AND DIABETES. PODOCYTES IN GLOMERULI FROM HUMANS WITH FOCAL SEGMENTAL GLOMERULOSCLEROSIS OR DIABETIC NEPHROPATHY EXHIBITED DIMINISHED H3K27ME3 AND HEIGHTENED UTX CONTENT. ANALOGOUS TO HUMAN DISEASE, INHIBITION OF JMJD3 AND UTX ABATED NEPHROPATHY PROGRESSION IN MICE WITH ESTABLISHED GLOMERULAR INJURY AND REDUCED H3K27ME3 LEVELS. TOGETHER, THESE FINDINGS INDICATE THAT OSTENSIBLY STABLE CHROMATIN MODIFICATIONS CAN BE DYNAMICALLY REGULATED IN QUIESCENT CELLS AND THAT EPIGENETIC REPROGRAMMING CAN IMPROVE OUTCOMES IN GLOMERULAR DISEASE BY REPRESSING THE REACTIVATION OF DEVELOPMENTAL PATHWAYS. 2018 19 5940 32 TARGETING METHYLTRANSFERASE PRMT5 ELIMINATES LEUKEMIA STEM CELLS IN CHRONIC MYELOGENOUS LEUKEMIA. IMATINIB-INSENSITIVE LEUKEMIA STEM CELLS (LSCS) ARE BELIEVED TO BE RESPONSIBLE FOR RESISTANCE TO BCR-ABL TYROSINE KINASE INHIBITORS AND RELAPSE OF CHRONIC MYELOGENOUS LEUKEMIA (CML). IDENTIFYING THERAPEUTIC TARGETS TO ERADICATE CML LSCS MAY BE A STRATEGY TO CURE CML. IN THE PRESENT STUDY, WE DISCOVERED A POSITIVE FEEDBACK LOOP BETWEEN BCR-ABL AND PROTEIN ARGININE METHYLTRANSFERASE 5 (PRMT5) IN CML CELLS. OVEREXPRESSION OF PRMT5 WAS OBSERVED IN HUMAN CML LSCS. SILENCING PRMT5 WITH SHRNA OR BLOCKING PRMT5 METHYLTRANSFERASE ACTIVITY WITH THE SMALL-MOLECULE INHIBITOR PJ-68 REDUCED SURVIVAL, SERIAL REPLATING CAPACITY, AND LONG-TERM CULTURE-INITIATING CELLS (LTC-ICS) IN LSCS FROM CML PATIENTS. FURTHER, PRMT5 KNOCKDOWN OR PJ-68 TREATMENT DRAMATICALLY PROLONGED SURVIVAL IN A MURINE MODEL OF RETROVIRAL BCR-ABL-DRIVEN CML AND IMPAIRED THE IN VIVO SELF-RENEWAL CAPACITY OF TRANSPLANTED CML LSCS. PJ-68 ALSO INHIBITED LONG-TERM ENGRAFTMENT OF HUMAN CML CD34+ CELLS IN IMMUNODEFICIENT MICE. MOREOVER, INHIBITION OF PRMT5 ABROGATED THE WNT/BETA-CATENIN PATHWAY IN CML CD34+ CELLS BY DEPLETING DISHEVELLED HOMOLOG 3 (DVL3). THIS STUDY SUGGESTS THAT EPIGENETIC METHYLATION MODIFICATION ON HISTONE PROTEIN ARGININE RESIDUES IS A REGULATORY MECHANISM TO CONTROL SELF-RENEWAL OF LSCS AND INDICATES THAT PRMT5 MAY REPRESENT A POTENTIAL THERAPEUTIC TARGET AGAINST LSCS. 2016 20 3347 42 HISTONE DEACETYLASES MEDIATE THE SILENCING OF MIR-15A, MIR-16, AND MIR-29B IN CHRONIC LYMPHOCYTIC LEUKEMIA. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) DEMONSTRATES A GLOBAL DOWN-REGULATION OF MIR-15A AND MIR-16 AND A SELECTIVE SILENCING OF THE RELATED MIR-29B IN AGGRESSIVE DISEASE. DELETIONS IN CHROMOSOME 13 [DEL(13Q14)] PARTIALLY ACCOUNT FOR THE LOSS OF EXPRESSION OF MIR-15A AND MIR-16, BUT THE MECHANISMS BY WHICH MIR-29B BECOMES SILENCED IS UNKNOWN. IN THE PRESENT STUDY, WE SHOW THAT THE HISTONE DEACETYLASES (HDACS) ARE OVEREXPRESSED IN CLL AND MEDIATE THE EPIGENETIC SILENCING OF MIR-15A, MIR-16, AND MIR-29B. HDAC INHIBITION TRIGGERED THE ACCUMULATION OF THE TRANSCRIPTIONALLY ACTIVATING CHROMATIN MODIFICATION H3K4ME2 AND RESTORED THE EXPRESSION OF MIR-15A, MIR-16, AND MIR-29B IN APPROXIMATELY 35% OF SAMPLES. ECTOPIC EXPRESSION OF MIR-15A AND MIR-16 AND HDAC INHIBITION-INDUCED EXPRESSION OF MIR-15A, MIR-16, OR MIR-29B IN PRIMARY CLL CELLS WAS ASSOCIATED WITH DECLINES IN THE LEVELS OF MCL-1, BUT NOT BCL-2, MITOCHONDRIAL DYSFUNCTION, AND INDUCTION OF CELL DEATH. THEREFORE, OUR RESULTS SHOW THAT HDACS ABERRANTLY SILENCE THE EXPRESSION OF THE CRITICAL TUMOR SUPPRESSORS MIR-15A, MIR-16, AND MIR-29B IN CLL. DEACETYLASE INHIBITION MAY BE A THERAPEUTIC STRATEGY THAT RESTORES THE EXPRESSION OF THESE MIRS TO ANTAGONIZE MCL-1, AN IMPORTANT SURVIVAL PROTEIN IN THESE CELLS. CONSEQUENTLY, CLL PATIENTS WHO EXHIBIT SUCH EPIGENETIC SILENCING MAY BENEFIT FROM HDAC INHIBITOR-BASED THERAPY. 2012