1 6235 132 THE M(6)A DEMETHYLASE FTO PROMOTES RENAL EPITHELIAL-MESENCHYMAL TRANSITION BY REDUCING THE M(6)A MODIFICATION OF LNCRNA GAS5. BACKGROUND: RENAL INTERSTITIAL FIBROSIS (RIF) IS THE MAIN PATHOLOGICAL CHANGE OF A VARIETY OF CHRONIC KIDNEY DISEASES (CKD). EPIGENETIC MODIFICATIONS OF FIBROSIS-PRONE GENES REGULATE RIF PROGRESSION. THIS STUDY AIMED TO INVESTIGATE LONG NON-CODING RNA (LNCRNA) N6-METHYLADENOSINE (M(6)A) MODIFICATION AND ITS ROLE IN REGULATING RIF PROGRESSION. METHODS: UNILATERAL URETERAL OCCLUSION (UUO) WAS EMPLOYED TO CONSTRUCT THE RIF IN VIVO MODEL; AND TGF-BETA1-TREATED HK-2 AND HKC-8 CELLS WERE USED FOR IN VITRO EXPERIMENTS. THE MRNA AND PROTEIN EXPRESSIONS WERE ASSESSED USING QRT-PCR AND WESTERN BLOT. THE PROLIFERATION AND MIGRATION WERE EVALUATED BY EDU ASSAY AND TRANSWELL ASSAY, RESPECTIVELY. IN ADDITION, LEVELS OF INFLAMMATORY CYTOKINES WERE DETERMINED BY ELISA ASSAY AND QRT-PCR. MOREOVER, LNCRNA GAS5 M(6)A LEVEL WAS DETECTED USING ME-RIP ASSAY. HE AND MASSON STAINING WERE EMPLOYED TO EVALUATE FIBROTIC LESIONS OF THE KIDNEY. RESULTS: FTO EXPRESSION WAS ELEVATED IN HK-2 AND HKC-8 CELLS AFTER TGF-BETA1 TREATMENT AND MOUSE KIDNEY TISSUE FOLLOWING UUO, AND LNCRNA GAS5 WAS DOWNREGULATED. LNCRNA GAS5 OVEREXPRESSION OR FTO SILENCING SUPPRESSED TGF-BETA1-INDUCED THE INCREASE OF EMT-RELATED PROTEINS (VIMENTIN, SNAIL AND N-CADHERIN) AND INFLAMMATORY CYTOKINES (IL-6, IL-1BETA AND TNF-ALPHA) LEVELS IN HK-2 CELLS. FTO SUPPRESSED LNCRNA GAS5 EXPRESSION BY REDUCING THE M6A MODIFICATION OF LNCRNA GAS5. ADDITIONALLY, FTO KNOCKDOWN COULD SUPPRESS EMT PROCESS AND INFLAMMATION RESPONSE INDUCED BY TGF-BETA1 AND UUO IN VITRO AND IN VIVO. AS EXPECTED, FTO KNOCKDOWN ABROGATED THE PROMOTION EFFECTS OF LNCRNA GAS5 SILENCING ON TGF-BETA1-INDUCED EMT PROCESS AND INFLAMMATION RESPONSE IN HK-2 AND HKC-8 CELLS. CONCLUSION: FTO PROMOTED EMT PROCESS AND INFLAMMATION RESPONSE THROUGH REDUCING THE M(6)A MODIFICATION OF LNCRNA GAS5. 2022 2 5850 45 SUBEROYLANILIDE HYDROXAMIC ACID (SAHA) REDUCES FIBROSIS MARKERS AND DEACTIVATES HUMAN STELLATE CELLS VIA THE EPITHELIAL-MESENCHYMAL TRANSITION (EMT). HEPATIC FIBROSIS IS KNOWN AS THE ACCUMULATION OF CONNECTIVE TISSUE SECONDARY TO CHRONIC DAMAGE TO THE LIVER. EPITHELIAL-MESENCHYMAL TRANSITION (EMT) CORRESPONDING INCREASE IN LIVER FIBROGENESIS WAS SHOWN WITH IMMUNOHISTOCHEMISTRY AND PCR-BASED STUDIES. SUBEROYLANILIDE HYDROXAMIC ACID (SAHA), A SYNTHETIC COMPOUND APPROVED AS A HISTONE DEACETYLASE INHIBITOR (HDAC) BY THE FDA TO TREAT CUTANEOUS T-CELL LYMPHOMA IS UNDER INVESTIGATION FOR THE TREATMENT OF LUNG AND RENAL FIBROSIS. EXPERIMENTAL MODELING FOR HEPATIC FIBROSIS CAN BE CONSTRUCTED WITH AN LX2 CELL LINE ISOLATED FROM HUMAN HEPATIC STELLATE CELLS (HSCS). IN THIS STUDY, WE AIMED TO INVESTIGATE THE MODULATION OF SAHA IN THE PATHOGENESIS OF LIVER FIBROSIS BY DETECTING THE LEVELS OF PROTEINS; (E-CADHERIN (E-CAD), N-CADHERIN (N-CAD), VIMENTIN (VIM), AND GENES; E-CAD, N-CAD, VIM, TRANSFORMING GROWTH FACTOR-BETA (TGF-BETA), ALPHA-SMOOTH MUSCLE ACTIN (ALPHA-SMA), TYPE 1 COLLAGEN (COL1A1), TYPE 3 COLLAGEN (COL3A1)) THAT PLAY A SIGNIFICANT ROLE IN EMT WITH THE LX2 CELL LINE. WE ALSO EVALUATED THE ACTION OF SAHA WITH CELL PROLIFERATION, CLONOGENIC, AND MIGRATION ASSAY. CELL PROLIFERATION WAS PERFORMED BY FLOW CYTOMETRY. ALL THE PROTEIN LEVELS WERE DETERMINED BY WESTERN BLOT ANALYSIS, AND GENE EXPRESSION LEVELS WERE MEASURED BY REAL-TIME PCR. OUR STUDY OBSERVED THAT SAHA TREATMENT DECREASED CELL VIABILITY, COLONY FORMATION AND MIGRATION IN LX2 CELLS. WE FOUND THAT SAHA INCREASED E-CAD EXPRESSION LEVEL, WHILE IT DECREASED N-CAD, VIM, COL1A1, COL3A1, ALPHA-SMA TGF-BETA GENES EXPRESSION LEVELS. SAHA DECREASED THE LEVEL OF E-CAD, N-CAD, AND VIM PROTEIN LEVELS. WE THOUGHT THAT SAHA POSSESSES POTENT ANTIFIBROTIC AND ANTI-EMT PROPERTIES IN LX2. 2021 3 1632 36 DNMTS ARE INVOLVED IN TGF-BETA1-INDUCED EPITHELIAL-MESENCHYMAL TRANSITIONS IN AIRWAY EPITHELIAL CELLS. CHRONIC RHINOSINUSITIS (CRS) PATHOGENESIS IS CLOSELY RELATED TO TISSUE REMODELING, INCLUDING EPITHELIAL-MESENCHYMAL TRANSITION (EMT). EPIGENETIC MECHANISMS PLAY KEY ROLES IN EMT. DNA METHYLATION, MEDIATED BY DNA METHYLTRANSFERASES (DNMTS), IS AN EPIGENETIC MARKER THAT IS CRITICAL TO EMT. THE GOAL OF THIS STUDY WAS TO DETERMINE WHETHER DNMTS WERE INVOLVED IN TGF-BETA1-INDUCED EMT AND ELUCIDATE THE UNDERLYING MECHANISMS IN NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. GLOBAL DNA METHYLATION AND DNMT ACTIVITY WERE QUANTIFIED. DNMT EXPRESSION WAS MEASURED USING REAL-TIME PCR (QRT-PCR) IN HUMAN CRS TISSUES. MRNA AND PROTEIN LEVELS OF DNMTS, E-CADHERIN, VIMENTIN, ALPHA-SMA, AND FIBRONECTIN WERE DETERMINED USING RT-PCR AND WESTERN BLOTTING, RESPECTIVELY. DNMT1, DNMT3A, AND DNMT3B GENE EXPRESSION WERE KNOCKED DOWN USING SIRNA TRANSFECTION. MAPK PHOSPHORYLATION AND EMT-RELATED TRANSCRIPTION FACTOR LEVELS WERE DETERMINED USING WESTERN BLOTTING. SIGNALING PATHWAYS WERE ANALYZED USING SPECIFIC INHIBITORS OF MAPK. WE DEMONSTRATED THESE DATA IN PRIMARY NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. GLOBAL DNA METHYLATION, DNMT ACTIVITY, AND DNMT EXPRESSION INCREASED IN CRS TISSUES. DNMT EXPRESSION WAS POSITIVELY CORRELATED WITH LUND-MCKAY CT SCORES. TGF-BETA1 DOSE-DEPENDENTLY INDUCED DNMT EXPRESSION. FURTHER, 5-AZA INHIBITED TGF-BETA1-INDUCED DNMT, SNAIL, AND SLUG EXPRESSION RELATED TO EMT, AS WELL AS P38 AND JNK PHOSPHORYLATION IN A549 CELLS AND TGF-BETA1-INDUCED DNMT EXPRESSION AND EMT IN PRIMARY NASAL EPITHELIAL CELLS AND AIR-LIQUID INTERFACE CULTURES. TGF-BETA1-INDUCED DNMT EXPRESSION LEADS TO DNA METHYLATION AND EMT VIA P38, JNK, SNAIL, AND SLUG SIGNALING PATHWAYS. INHIBITION OF DNMT SUPPRESSED THE EMT PROCESS AND THEREFORE IS POTENTIALLY A CRS THERAPEUTIC STRATEGY. 2022 4 6758 24 WNT SIGNALING IN STEM CELL BIOLOGY AND REGENERATIVE MEDICINE. WNT FAMILY MEMBERS ARE SECRETED-TYPE GLYCOPROTEINS TO ORCHESTRATE EMBRYOGENESIS, TO MAINTAIN HOMEOSTASIS, AND TO INDUCE PATHOLOGICAL CONDITIONS. FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, AND ROR2 ARE TRANSMEMBRANE RECEPTORS TRANSDUCING WNT SIGNALS BASED ON LIGAND-DEPENDENT PREFERENTIALITY FOR CAVEOLIN- OR CLATHRIN-MEDIATED ENDOCYTOSIS. WNT SIGNALS ARE TRANSDUCED TO CANONICAL PATHWAY FOR CELL FATE DETERMINATION, AND TO NON-CANONICAL PATHWAYS FOR REGULATION OF PLANAR CELL POLARITY, CELL ADHESION, AND MOTILITY. MYC, CCND1, AXIN2, FGF20, WISP1, JAG1, DKK1 AND GLUCAGON ARE TARGET GENES OF CANONICAL WNT SIGNALING CASCADE, WHILE CD44, VIMENTIN AND STX5 ARE TARGET GENES OF NON-CANONICAL WNT SIGNALING CASCADES. HOWEVER, TARGET GENES OF WNT SIGNALING CASCADES ARE DETERMINED IN A CONTEXT-DEPENDENT MANNER DUE TO EXPRESSION PROFILE OF TRANSCRIPTION FACTORS AND EPIGENETIC STATUS. WNT SIGNALING CASCADES NETWORK WITH NOTCH, FGF, BMP AND HEDGEHOG SIGNALING CASCADES TO REGULATE THE BALANCE OF STEM CELLS AND PROGENITOR CELLS. HERE WNT SIGNALING IN EMBRYONIC STEM CELLS, NEURAL STEM CELLS, MESENCHYMAL STEM CELLS, HEMATOPOIETIC STEM CELLS, AND INTESTINAL STEM CELLS WILL BE REVIEWED. WNT3, WNT5A AND WNT10B ARE EXPRESSED IN UNDIFFERENTIATED HUMAN EMBRYONIC STEM CELLS, WHILE WNT6, WNT8B AND WNT10B IN ENDODERM PRECURSOR CELLS. WNT6 IS EXPRESSED IN INTESTINAL CRYPT REGION FOR STEM OR PROGENITOR CELLS. TNF/ALPHA-WNT10B SIGNALING IS A NEGATIVE FEEDBACK LOOP TO MAINTAIN HOMEOSTASIS OF ADIPOSE TISSUE AND GASTROINTESTINAL MUCOSA WITH CHRONIC INFLAMMATION. RECOMBINANT WNT PROTEIN OR WNT MIMETIC (CIRCULAR PEPTIDE, SMALL MOLECULE COMPOUND, OR RNA APTAMER) IN COMBINATION WITH NOTCH MIMETIC, FGF PROTEIN, AND BMP PROTEIN OPENS A NEW WINDOW TO TISSUE ENGINEERING FOR REGENERATIVE MEDICINE. 2008 5 1826 39 EFFECTS OF HISTONE DEACETYLASE INHIBITOR ON EXTRACELLULAR MATRIX PRODUCTION IN HUMAN NASAL POLYP ORGAN CULTURES. BACKGROUND: NASAL POLYPOSIS IS ASSOCIATED WITH A CHRONIC INFLAMMATORY CONDITION OF THE SINONASAL MUCOSA AND INVOLVES MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX (ECM) ACCUMULATION. EPIGENETIC MODULATION BY HISTONE DEACETYLASE (HDAC) INHIBITORS INCLUDING TRICHOSTATIN A (TSA) HAS BEEN REPORTED TO HAVE INHIBITORY EFFECTS ON MYOFIBROBLAST DIFFERENTIATION IN LUNG AND RENAL FIBROBLASTS. THE PURPOSE OF THIS STUDY WAS TO INVESTIGATE THE INHIBITORY EFFECT OF TSA ON MYOFIBROBLAST DIFFERENTIATION AND ECM PRODUCTION IN NASAL POLYP ORGAN CULTURES. METHODS: NASAL POLYP TISSUES FROM 18 PATIENTS WERE ACQUIRED DURING ENDOSCOPIC SINUS SURGERY. AFTER ORGAN CULTURE, NASAL POLYPS WERE STIMULATED WITH TGF-BETA1 AND THEN TREATED WITH TSA. ALPHA-SMOOTH MUSCLE ACTIN (ALPHA-SMA), FIBRONECTIN, AND COLLAGEN TYPE I EXPRESSION LEVELS WERE EXAMINED BY REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION (PCR), REAL-TIME PCR, WESTERN BLOT, AND IMMUNOFLUORESCENT STAINING. HDAC2, HDAC4, AND ACETYLATED H4 EXPRESSION LEVELS WERE ASSAYED BY WESTERN BLOT. CYTOTOXICITY WAS ANALYZED BY THE TERMINAL DEOXYNUCLEOTIDYL TRANSFERASE BIOTIN-DUTP NICK END LABELING ASSAY. RESULTS: THE EXPRESSION LEVELS OF ALPHA-SMA, FIBRONECTIN, AND COLLAGEN TYPE 1 WERE INCREASED IN NASAL POLYP AFTER TRANSFORMING GROWTH FACTOR (TGF) BETA1 TREATMENT. TSA-INHIBITED TGF-BETA1 INDUCED THESE GENE AND PROTEIN EXPRESSION LEVELS. FURTHERMORE, TSA SUPPRESSED PROTEIN EXPRESSION LEVELS OF HDAC2 AND HDAC4. HOWEVER, TSA INDUCED HYPERACETYLATION OF HISTONES H4. TREATMENT WITH TGF-BETA1 WITH OR WITHOUT TSA DID NOT HAVE CYTOTOXIC EFFECT. CONCLUSION: THESE FINDINGS PROVIDE NOVEL INSIGHTS INTO THE EPIGENETIC REGULATION IN MYOFIBROBLAST DIFFERENTIATION AND ECM PRODUCTION OF NASAL POLYP. TSA COULD BE A CANDIDATE OF A THERAPEUTIC AGENT FOR REVERSING THE TGF-BETA1-INDUCED ECM SYNTHESIS THAT LEADS TO NASAL POLYP DEVELOPMENT. 2013 6 1393 33 DIAGNOSTIC VALUE OF THE HYPOMETHYLATION OF THE WISP1 PROMOTER IN PATIENTS WITH HEPATOCELLULAR CARCINOMA ASSOCIATED WITH HEPATITIS B VIRUS. WNT1-INDUCIBLE SIGNALING PATHWAY PROTEIN 1 (WISP1) REGULATES CELL PROLIFERATION, DIFFERENTIATION, ADHESION, MIGRATION AND SURVIVAL. ABNORMAL WISP1 EXPRESSION IS ASSOCIATED WITH THE CARCINOGENESIS OF HEPATOCELLULAR CARCINOMA (HCC). ABERRANT DNA METHYLATION IS ONE OF THE MAJOR EPIGENETIC ALTERATIONS IN HCC. HOWEVER, THE METHYLATION STATUS OF THE WISP1 PROMOTER IS STILL UNCLEAR. WE THEREFORE AIMED TO DETERMINE THE METHYLATION STATUS OF THE WISP1 PROMOTER AND EVALUATE ITS CLINICAL VALUE IN HCC. THE STUDY ENROLLED 251 PARTICIPANTS, INCLUDING 123 PARTICIPANTS WITH HCC, 90 PARTICIPANTS WITH CHRONIC HEPATITIS B (CHB) AND 38 HEALTHY CONTROLS (HCS). WISP1 METHYLATION STATUS, MRNA LEVELS AND PLASMA SOLUBLE WISP1 WERE DETECTED BY METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MSP), QUANTITATIVE REAL-TIME PCR (RT-QPCR) AND ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA), RESPECTIVELY. WE FOUND THAT THE METHYLATION FREQUENCY OF WISP1 IN PATIENTS WITH HCC WAS SIGNIFICANTLY LOWER THAN THAT IN PATIENTS WITH CHB AND HCS, WHILE THE RELATIVE EXPRESSION LEVELS OF WISP1 MRNA WERE MARKEDLY HIGHER IN PATIENTS WITH HCC THAN IN PATIENTS WITH CHB AND HCS. FURTHERMORE, THE PLASMA SOLUBLE WISP1 IN PATIENTS WITH HCC WAS OBVIOUSLY LOWER THAN IN THAT IN PATIENTS WITH CHB AND HCS. ALPHA-FETOPROTEIN (AFP) IS A WIDELY RECOGNIZED BIOMARKER TO DIAGNOSE HCC WHICH LACKS ENOUGH SENSITIVITY AND SPECIFICITY. WISP1 PROMOTER METHYLATION STATUS COMBINED WITH AFP SIGNIFICANTLY IMPROVED THE DIAGNOSTIC ABILITY IN DISCRIMINATING HCC FROM CHB COMPARED WITH AFP OR WISP1 METHYLATION STATUS ALONE. IN CONCLUSION, HYPOMETHYLATION OF THE WISP1 GENE PROMOTER MAY SERVE AS A NONINVASIVE BIOMARKER FOR DETECTING HBV-ASSOCIATED HCC. 2020 7 3445 35 HYPERMETHYLATION OF ITGA4, TFPI2 AND VIMENTIN PROMOTERS IS INCREASED IN INFLAMED COLON TISSUE: PUTATIVE RISK MARKERS FOR COLITIS-ASSOCIATED CANCER. PURPOSE: EPIGENETIC SILENCING OF TUMOR SUPPRESSOR GENES IS INVOLVED IN EARLY TRANSFORMING EVENTS AND HAS A HIGH IMPACT ON COLORECTAL CARCINOGENESIS. LIKEWISE, COLON CANCERS THAT DERIVE FROM CHRONICALLY INFLAMED BOWEL DISEASES FREQUENTLY EXHIBIT EPIGENETIC CHANGES. BUT THERE IS LITTLE DATA ABOUT EPIGENETIC ABERRATIONS CAUSING COLORECTAL CANCER IN CHRONICALLY INFLAMED TISSUE. THE AIM OF THE PRESENT STUDY WAS TO EVALUATE THE ABERRANT GAIN OF METHYLATION IN THE GENE PROMOTERS OF VIM, TFPI2 AND ITGA4 AS PUTATIVE EARLY MARKERS IN THE DEVELOPMENT FROM INFLAMED TISSUE VIA PRECANCEROUS LESIONS TOWARD COLORECTAL CANCER. METHODS: INITIAL SCREENING OF DIFFERENT CANCER CELL LINES BY USING METHYLATION-SPECIFIC PCR REVEALED A PUTATIVE COLON CANCER-SPECIFIC METHYLATION PATTERN. ADDITIONALLY, A DEMETHYLATION ASSAY WAS PERFORMED TO INVESTIGATE THE METHYLATION-DEPENDENT GENE SILENCING OF ITGA4. THE CANDIDATE MARKERS WERE ANALYZED IN COLONIC TISSUE SPECIMENS FROM PATIENTS WITH COLORECTAL CANCER (N = 15), ADENOMAS (N = 76), SERRATED LESIONS (N = 13), CHRONIC INFLAMMATION (N = 10) AND NORMAL MUCOSAL SAMPLES (N = 9). RESULTS: A HIGH METHYLATION FREQUENCY OF VIM (55.6 %) WAS OBSERVED IN NORMAL COLON TISSUE, WHEREAS ITGA4 AND TFPI2 WERE COMPLETELY UNMETHYLATED IN CONTROLS. A SIGNIFICANT GAIN OF METHYLATION FREQUENCY WITH PROGRESSION OF DISEASE AS WELL AS AN AGE-DEPENDENT EFFECT WAS DETECTABLE FOR TFPI2. ITGA4 METHYLATION FREQUENCY WAS HIGH IN PRECANCEROUS AND CANCEROUS TISSUES AS WELL AS IN INFLAMMATORY BOWEL DISEASES (IBD). CONCLUSION: THE ALREADY ESTABLISHED METHYLATION MARKER VIM DOES NOT PERMIT A SPECIFIC AND SENSITIVE DISCRIMINATION OF HEALTHY AND NEOPLASTIC TISSUE. THE METHYLATION MARKERS ITGA4 AND TFPI2 SEEM TO BE SUITABLE RISK MARKERS FOR INFLAMMATION-ASSOCIATED COLON CANCER. 2015 8 766 54 CCL5 SUPPRESSES KLOTHO EXPRESSION VIA P-STAT3/DNA METHYLTRANSFERASE1-MEDIATED PROMOTER HYPERMETHYLATION. BACKGROUND: ENHANCED INFLAMMATION AND REDUCED KLOTHO ARE COMMON FEATURES IN CHRONIC KIDNEY DISEASE (CKD). INFLAMMATION INDUCES DNA HYPERMETHYLATION. THIS STUDY ASSESSED THE PERFORMANCE OF INFLAMMATORY MARKER C-C MOTIF CHEMOKINE 5 (CCL5) IN EPIGENETIC REGULATION OF KLOTHO EXPRESSION. METHODS: FIFTY CKD PATIENTS AND 25 MATCHED CONTROLS WERE ENROLLED, AND SERUM CCL5 LEVEL, SKLOTHO LEVEL, AND DNA METHYLATION WERE EVALUATED IN THESE SUBJECTS. A RENAL INTERSTITIAL FIBROSIS (RIF) MODEL WITH CKD WAS INDUCED IN MICE VIA UNILATERAL URETERAL OBSTRUCTION (UUO) IN VIVO AND HUMAN PROXIMAL TUBULAR EPITHELIAL (HK-2) CELLS TREATED WITH CCL5 IN VITRO. 5-AZA-2'-DEOXYCYTIDINE (5-AZA), A DNA METHYLTRANSFERASE INHIBITOR WAS GIVEN TO UUO MICE. HEMATOXYLIN AND EOSIN (HE) AND MASSON TRICHROME STAINING WERE ADOPTED TO EVALUATE RENAL PATHOLOGICAL CHANGES. METHYLATION-SPECIFIC PCR WAS PERFORMED TO ASSESS DNA METHYLATION OF KLOTHO PROMOTER IN THE PERIPHERAL BLOOD LEUCOCYTES (PBLS) FROM CKD PATIENTS AND OBSTRUCTIVE KIDNEY FROM UUO MICE. CCL5, KLOTHO, AND DNA METHYLTRANSFERASES (DNMTS) WERE DETERMINED BY ELISAS, IMMUNOFLUORESCENCE, OR WESTERN BLOTTING. HK-2 CELLS WERE EXPOSED TO CCL5 WITH OR WITHOUT 5-AZA AND STATTIC, A P-SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION 3 (STAT3) INHIBITOR, AND EXPRESSIONS OF P-STAT3, DNMT1, AND KLOTHO WERE DETERMINED BY WESTERN BLOTTING. RESULTS: CCL5 UPREGULATION CONCOMITANT WITH KLOTHO DOWNREGULATION IN SERUM AND GLOBAL DNA METHYLATION IN PBLS WERE OBSERVED IN CKD SAMPLES. UUO CONTRIBUTED TO SEVERE RENAL INTERSTITIAL FIBROSIS AND ENHANCED EXPRESSIONS OF FIBROTIC MARKERS. MOREOVER, UUO INCREASED THE CCL5 LEVEL, INDUCED KLOTHO PROMOTER METHYLATION, SUPPRESSED KLOTHO LEVEL, ACTIVATED P-STAT3 SIGNALING, AND UPREGULATED DNMT1 LEVEL. A SIMILAR OBSERVATION WAS MADE IN HK-2 CELLS TREATED WITH CCL5. MORE IMPORTANTLY, 5-AZA INHIBITED UUO-INDUCED KLOTHO HYPERMETHYLATION, REVERSED KLOTHO, DOWNREGULATED P-STAT3 EXPRESSIONS, AND AMELIORATED RIF IN VIVO. THE CONSISTENT FINDINGS IN VITRO WERE ALSO OBTAINED IN HK-2 CELLS EXPOSED TO 5-AZA AND STATTIC. CONCLUSION: THE CCL5/P-STAT3/DNMT1 AXIS IS IMPLICATED IN EPIGENETIC REGULATION OF KLOTHO EXPRESSION IN CKD. THIS STUDY PROVIDES NOVEL THERAPEUTIC POSSIBILITIES FOR REVERSAL OF KLOTHO SUPPRESSION BY CKD. 2022 9 878 49 CHRONIC CADMIUM EXPOSURE AGGRAVATES MALIGNANT PHENOTYPES OF NASOPHARYNGEAL CARCINOMA BY ACTIVATING THE WNT/BETA-CATENIN SIGNALING PATHWAY VIA HYPERMETHYLATION OF THE CASEIN KINASE 1ALPHA PROMOTER. BACKGROUND: OUR PREVIOUS STUDY HAS SHOWN THAT CADMIUM (CD) EXPOSURE IS NOT ONLY A RISK FACTOR FOR NASOPHARYNGEAL CARCINOMA (NPC), BUT ALSO CORRELATED WITH THE CLINICAL STAGE AND LYMPH NODE METASTASIS. HOWEVER, THE UNDERLYING MOLECULAR EVENTS OF CD INVOLVED IN NPC PROGRESSION REMAIN TO BE ELUCIDATED. PURPOSE: THE OBJECTIVE OF THIS STUDY WAS TO DECIPHER HOW CD IMPACTS THE MALIGNANT PHENOTYPES OF NPC CELLS. METHODS: NPC CELL LINES CNE-1 AND CNE-2 WERE CONTINUOUSLY EXPOSED WITH 1 MUM CD CHLORIDE FOR 10 WEEKS, DESIGNATING AS CHRONIC CD TREATED NPC CELLS (CCT-NPC). MTT ASSAY, COLONY FORMATION ASSAY AND XENOGRAFT TUMOR GROWTH WERE USED TO ASSESS CELL VIABILITY IN VITRO AND IN VIVO. TRANSWELL ASSAYS WERE PERFORMED TO DETECT CELL INVASION AND MIGRATION. THE PROTEIN LEVELS OF E-CADHERIN, N-CADHERIN, VIMENTIN AS WELL AS BETA-CATENIN AND CASEIN KINASE 1ALPHA(CK1ALPHA) WERE MEASURED BY WESTERN BLOT. IMMUNOFLUORESCENCE STAINING WAS USED TO OBSERVE THE DISTRIBUTION OF FILAMENT ACTIN (F-ACTIN), BETA-CATENIN AND CK1ALPHA. THE MRNA LEVELS OF DOWNSTREAM TARGET GENES OF BETA-CATENIN WERE DETECTED BY RT-PCR. WNT/BETA-CATENIN SIGNALING ACTIVITY WAS ASSESSED BY TOPFLASH/FOPFLASH DUAL LUCIFERASE REPORT SYSTEM. MS-PCR WAS USED TO DETECT THE METHYLATION STATUS OF CK1ALPHA. FINALLY, THE ACTIVATION OF WNT/BETA-CATENIN PATHWAY AND CELL BIOLOGICAL PROPERTIES WERE EXAMINED FOLLOWING TREATMENT OF CCT-NPC CELLS WITH 5-AZA-2-DEOXY-CYTIDINE(5-AZA-CDR). RESULTS: CCT-NPC CELLS SHOWED AN INCREASE IN CELL PROLIFERATION, COLONY FORMATION, INVASION AND MIGRATION COMPARED TO THE PARENTAL CELLS. CD ALSO INDUCED CYTOSKELETON REORGANIZATION AND EPITHELIAL-TO-MESENCHYMAL TRANSITION. UPREGULATION AND NUCLEAR TRANSLOCATION OF BETA-CATENIN AND INCREASED LUCIFERASE ACTIVITY ACCOMPANIED WITH TRANSCRIPTION OF DOWNSTREAM TARGET GENES WERE FOUND IN CCT-NPC CELLS. TREATMENT OF CCT-CNE1 CELLS WITH 5-AZA-CDR COULD REVERSE THE HYPERMETHYLATION OF CK1ALPHA AND ATTENUATE THE CELL MALIGNANCY. CONCLUSION: THESE RESULTS SUPPORT A ROLE FOR CHRONIC CD EXPOSURE AS A DRIVING FORCE FOR THE MALIGNANT PROGRESSION OF NPC VIA EPIGENETIC ACTIVATION OF THE WNT/BETA-CATENIN PATHWAY. 2019 10 2326 37 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 11 6662 37 UPREGULATION OF FZD5 IN EOSINOPHILIC CHRONIC RHINOSINUSITIS WITH NASAL POLYPS BY EPIGENETIC MODIFICATION. EOSINOPHILIC CHRONIC RHINOSINUSITIS WITH NASAL POLYPS (CRSWNP) IS ONE OF THE MOST CHALLENGING PROBLEMS IN CLINICAL RHINOLOGY. FZD5 IS A RECEPTOR FOR WNT5A, AND ITS COMPLEX WITH WNT5A CONTRIBUTES TO ACTIVATING INFLAMMATION AND TISSUE MODIFICATION. NASAL POLYPS AND EOSINOPHIL/NON-EOSINOPHIL COUNTS ARE REPORTED TO BE DIRECTLY CORRELATED. THIS STUDY INVESTIGATED THE EXPRESSION AND DISTRIBUTION OF FZD5, AND THE ROLE OF EOSINOPHIL INFILTRATION AND FZD5 IN EOSINOPHILIC CRSWNP PATHOGENESIS. THE PROGNOSTIC ROLE OF EOSINOPHIL LEVELS WAS EVALUATED IN SEVEN PATIENTS WITH CRSWNP. FIFTEEN PATIENTS WITH CRS WERE CLASSIFIED BASED ON THE PERCENTAGE OF EOSINOPHILS IN NASAL POLYP TISSUE. METHYLATED GENES WERE DETECTED USING METHYL-CPG-BINDING DOMAIN SEQUENCING, AND QRT-PCR AND IMMUNOHISTOCHEMISTRY WERE USED TO DETECT FZD5 EXPRESSION IN NASAL POLYP TISSUE SAMPLES. THE RESULTS SHOWED THAT MRNA EXPRESSION OF FZD5 WAS UPREGULATED IN NASAL POLYPS. FZD5 EXPRESSION WAS SIGNIFICANTLY HIGHER IN NASAL POLYP SAMPLES FROM PATIENTS WITH EOSINOPHILIC CRSWNP THAN IN THOSE FROM PATIENTS WITH NON-EOSINOPHILIC CRSWNP, AS INDICATED BY IMMUNOHISTOCHEMISTRY. FURTHERMORE, INFLAMMATORY CYTOKINE LEVELS WERE HIGHER IN EOSINOPHILIC CRSWNP-DERIVED EPITHELIAL CELLS THAN IN NORMAL TISSUES. IN CONCLUSION, FZD5 EXPRESSION IN NASAL MUCOSAL EPITHELIAL CELLS IS CORRELATED WITH INFLAMMATORY CELLS AND MIGHT PLAY A ROLE IN THE PATHOGENESIS OF EOSINOPHILIC CRSWNP. 2019 12 2349 40 EPIGENETIC REGULATION OF MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX PRODUCTION IN NASAL POLYP-DERIVED FIBROBLASTS. BACKGROUND: NASAL POLYPOSIS IS A MULTI-FACTORIAL DISEASE ASSOCIATED WITH CHRONIC INFLAMMATORY CONDITION OF THE PARANASAL SINUSES. MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLULAR MATRIX (ECM) ACCUMULATION ARE INVOLVED IN THE PATHOGENESIS OF NASAL POLYPOSIS. OBJECTIVE: THE AIM OF THIS STUDY WAS TO STUDY THE EFFECT OF TRICHOSTATIN A (TSA), A HISTONE DEACETYLASE (HDAC) INHIBITOR, ON TRANSFORMING GROWTH FACTOR (TGF)-BETA1-INDUCED MYOFIBROBLAST DIFFERENTIATION AND ECM ACCUMULATION IN NASAL POLYP-DERIVED FIBROBLASTS (NPDFS). METHODS: NASAL POLYP-DERIVED FIBROBLASTS WERE ISOLATED FROM NASAL POLYPS OF PATIENTS WHO HAVE CHRONIC RHINOSINUSITIS WITH NASAL POLYP. TSA WAS TREATED IN TGF-BETA1-INDUCED NPDFS. EXPRESSION LEVELS OF HDAC2, ALPHA-SMOOTH MUSCLE ACTIN (SMA), TGF-BETA1, COLLAGEN TYPE I, ACETYLATED HISTONE H3, ACETYLATED HISTONE H4, PHOSPHORYLATED SMAD2/3 AND SMAD7 WERE DETERMINED BY RT-PCR, WESTERN BLOT AND/OR IMMUNOFLUORESCENT STAINING. THE TOTAL COLLAGEN AMOUNT PRODUCTION WAS ANALYSED BY SIRCOL SOLUBLE COLLAGEN ASSAY AND CONTRACTILE ACTIVITY WAS MEASURED BY COLLAGEN GEL CONTRACTION ASSAY. HDAC2 INHIBITION BY TSA OR HDAC2 SILENCING WAS ESTABLISHED BY RT-PCR AND WESTERN BLOT. THE EPIGENETIC EFFECT ON ALPHA-SMA GENE INACTIVATION WAS EXAMINED BY CHROMATIN IMMUNOPRECIPITATION ASSAY. PROLIFERATION WAS DETERMINED BY KI67-POSITIVE CELL STAINING AND CYTOTOXICITY WAS ASSESSED BY 3-(4,5- DIMETHYLTHIAZOL-2YL)-2,5-DIPHENYL-2H-TETRAZOLIUM BROMIDE (MTT) ASSAY. RESULTS: THE EXPRESSION LEVELS OF HDAC2, ALPHA-SMA AND TGF-BETA1 WERE INCREASED IN NASAL POLYP TISSUES COMPARED TO NORMAL INFERIOR TURBINATE TISSUES. TSA AND HDAC2 SILENCING INHIBITED EXPRESSION LEVELS ALPHA-SMA, COLLAGEN AND HDAC2. TSA INDUCED HYPERACETYLATION OF HISTONE AND SUPPRESSED OPENING OF ALPHA-SMA GENE PROMOTER IN TGF-BETA1-INDUCED NPDFS. TSA INHIBITED TGF-BETA1-INDUCED SMAD 2/3 AND RESCUED TGF-BETA1-SUPPRESSED SMAD7 SIGNALLING PATHWAY. FINALLY, TSA BLOCKED PROLIFERATION IN TGF-BETA1-INDUCED NPDFS AND HAS NO CYTOTOXIC EFFECT IN NPDFS. CONCLUSIONS AND CLINICAL RELEVANCE: THESE RESULTS SUGGEST THAT HDAC INHIBITION IS ASSOCIATED WITH MYOFIBROBLAST DIFFERENTIATION AND EXTRACELLUAR MATRIX ACCUMULATION IN NASAL POLYPOSIS. TSA MAY BE USEFUL AS AN INHIBITOR OF NASAL POLYP GROWTH, AND THUS HAS POTENTIAL TO BE USED AS A NOVEL TREATMENT OPTION FOR NASAL POLYPOSIS. 2012 13 1730 26 DYSREGULATION OF STEM CELL SIGNALING NETWORK DUE TO GERMLINE MUTATION, SNP, HELICOBACTER PYLORI INFECTION, EPIGENETIC CHANGE AND GENETIC ALTERATION IN GASTRIC CANCER. GENETIC FACTORS, HELICOBACTER PYLORI INFECTION, SALT OVER-UPTAKE, DECREASED VEGETABLE/FRUIT CONSUMPTION, SMOKING, AND METABOLIC SYNDROME ARE RISK FACTORS OF HUMAN GASTRIC CANCER. GERMLINE MUTATIONS OF CDH1 GENE, AND SNPS OF PTPN11 (SHP2), TLR4, IL1B, TNFA, BMP6, GDF15 AND RUNX3 GENES ARE ASSOCIATED WITH GASTRIC CANCER. HELICOBACTER PYLORI ACTIVATES CAGA-SHP2-ERK AND PEPTIDOGLYCAN-NOD1-NFKAPPAB SIGNALING CASCADES IN GASTRIC EPITHELIAL CELLS USING TYPE IV SECRETION SYSTEM, AND ALSO TRAF6-MAP3K7-NFKAPPAB AND TRAF6-MAP3K7-AP-1 SIGNALING CASCADES IN EPITHELIAL AND IMMUNE CELLS THROUGH LIPOPOLYSACCHARIDE RECOGNITION BY TLR2 OR TLR4. IL-1BETA, IL-6, IL-8, TNFALPHA AND IFNGAMMA ARE ELEVATED IN GASTRIC MUCOSA WITH HELICOBACTER PYLORI INFECTION. IL-6 AND TNFALPHA INDUCE UPREGULATION OF WNT5A AND WNT10B, RESPECTIVELY. WNT SIGNALS ARE TRANSDUCED TO BETA-CATENIN-TCF/LEF, RHOA, JNK, PKC, NFAT, AND NLK SIGNALING CASCADES. WNT-BETA-CATENIN-TCF/LEF SIGNALING INDUCES UPREGULATION OF MYC, CCND1, WISP1, FGF20, JAG1 AND DKK1 GENES. NOTCH SIGNALS ARE TRANSDUCED TO CSL-NICD-MAML AND NFKAPPAB SIGNALING CASCADES. FGF SIGNALS ARE TRANSDUCED TO ERK, PI3K-AKT, PKC, AND NFAT SIGNALING CASCADES. HELICOBACTER PYLORI INFECTION INDUCES SHH UPREGULATION IN PARIETAL CELL LINEAGE, WHILE BMP SIGNALS INDUCE IHH UPREGULATION IN PIT CELL LINEAGE. HEDGEHOG SIGNALS INDUCE UPREGULATION OF GLI1, PTCH1, CCND2, FOXL1, JAG2 AND SFRP1 GENES. JAG1 AND JAG2 ACTIVATE NOTCH SIGNALING, WHILE DKK1 AND SFRP1 INHIBIT WNT SIGNALING. STEM CELL SIGNALING NETWORK, CONSISTING OF WNT, NOTCH, FGF, HEDGEHOG AND BMP SIGNALING PATHWAYS, IS ACTIVATED DURING CHRONIC HELICOBACTER PYLORI INFECTION. EPIGENETIC SILENCING OF SFRP1 GENE OCCURS IN THE EARLIER STAGE OF CARCINOGENESIS IN THE STOMACH, WHILE AMPLIFICATION AND OVEREXPRESSION OF FGFR2 GENE IN THE LATER STAGE. DYSREGULATION OF THE STEM CELL SIGNALING NETWORK DUE TO THE ACCUMULATION OF GERMLINE MUTATION, SNP, HELICOBACTER PYLORI INFECTION, EPIGENETIC CHANGE AND GENETIC ALTERATION GIVES RISE TO GASTRIC CANCER. SNP TYPING AND CUSTOM-MADE MICROARRAY ANALYSES ON GENES ENCODING STEM CELL SIGNALING MOLECULES COULD BE UTILIZED FOR THE PERSONALIZED MEDICINE. 2007 14 3036 55 GENISTEIN AMELIORATES RENAL FIBROSIS THROUGH REGULATION SNAIL VIA M6A RNA DEMETHYLASE ALKBH5. RENAL TUBULE-INTERSTITIAL FIBROSIS IS RELATED TO CHRONIC KIDNEY DISEASE PROGRESSION AND A TYPICAL FEATURE OF THE AGING KIDNEY. EPIGENETIC MODIFICATIONS OF FIBROSIS-PRONE GENES REGULATE THE DEVELOPMENT OF RENAL FIBROSIS. AS A KIND OF "EPIGENETIC DIET", SOY ISOFLAVONE GENISTEIN WAS REPORTED TO HAVE RENAL PROTECTIVE ACTION AND EPIGENETIC-MODULATING EFFECTS. HOWEVER, ITS RENAL PROTECTION ROLE AND UNDERLYING MECHANISMS ARE YET TO BE FULLY CLARIFIED. HEREIN, WE SHOWED THAT GENISTEIN EXHIBITS A DEMONSTRABLE ANTI-FIBROTIC EFFECT ON KIDNEY IN VIVO UUO (UNILATERAL URETERAL OCCLUSION) MODEL AND RENAL EPITHELIAL CELLS IN VITRO MODEL. THE MECHANISM IS STRONGLY ASSOCIATED WITH EPITHELIAL-TO-MESENCHYMAL TRANSITION AND M6A RNA DEMETHYLASE ALKBH5. MOUSE FIBROTIC KIDNEYS INDUCED BY UUO EXHIBITED ADVERSE EXPRESSION OF RENAL FIBROSIS-RELATED PROTEINS AND SIGNIFICANT INCREASES IN THE TOTAL M6A LEVEL. AS AN ERASER, ALKBH5 SHOWED SEVERER SUPPRESSION IN THE RENAL FIBROSIS PROCESS. HOWEVER, GENISTEIN PRETREATMENT RESTORED ALKBH5 LOSS REMARKABLY AND REDUCED RENAL FIBROSIS, ABNORMAL PROTEIN, AND INFLAMMATORY MARKERS. THE EXAMINATION OF POSSIBLE MECHANISMS REVEALED THAT GENISTEIN PROMOTED ALKBH5 AND MAYBE INDUCED THE LEVEL OF MRNA M6A METHYLATION IN SOME EPITHELIAL-TO-MESENCHYMAL TRANSITION-RELATED TRANSCRIPTION FACTORS. WE FOUND SNAIL WAS THE CRITICAL REGULATOR AND CRITICAL FOR THE PROTECTIVE ROLE OF GENISTEIN. TO VERIFY THE RELATIONSHIP BETWEEN ALKBH5 AND SNAIL, WE GENERATED KNOCKDOWN AND OVEREXPRESSION OF ALKBH5 CELLS IN VITRO. ALKBH5 KNOCKDOWN ENHANCED THE MESENCHYMAL PHENOTYPE MARKER ALPHA-SMOOTH MUSCLE ACTIN AND SNAIL EXPRESSION. IN AGREEMENT, OVEREXPRESSION ALKBH5 INCREASED EPITHELIAL ADHESION MOLECULE E-CADHERIN AND REDUCED SNAIL EXPRESSION. IN CONCLUSION, GENISTEIN INCREASED RENAL ALKBH5 EXPRESSION IN UUO-INDUCED RENAL FIBROSIS AND REDUCED RNA M6A LEVELS AND AMELIORATES RENAL DAMAGES. 2020 15 922 29 CHRONIC IL-1BETA-INDUCED INFLAMMATION REGULATES EPITHELIAL-TO-MESENCHYMAL TRANSITION MEMORY PHENOTYPES VIA EPIGENETIC MODIFICATIONS IN NON-SMALL CELL LUNG CANCER. CHRONIC INFLAMMATION FACILITATES TUMOR PROGRESSION. WE DISCOVERED THAT A SUBSET OF NON-SMALL CELL LUNG CANCER CELLS UNDERWENT A GRADUALLY PROGRESSING EPITHELIAL-TO-MESENCHYMAL (EMT) PHENOTYPE FOLLOWING A 21-DAY EXPOSURE TO IL-1BETA, AN ABUNDANT PROINFLAMMATORY CYTOKINE IN THE AT-RISK FOR LUNG CANCER PULMONARY AND THE LUNG TUMOR MICROENVIRONMENTS. PATHWAY ANALYSIS OF THE GENE EXPRESSION PROFILE AND IN VITRO FUNCTIONAL STUDIES REVEALED THAT THE EMT AND EMT-ASSOCIATED PHENOTYPES, INCLUDING ENHANCED CELL INVASION, PD-L1 UPREGULATION, AND CHEMORESISTANCE, WERE SUSTAINED IN THE ABSENCE OF CONTINUOUS IL-1BETA EXPOSURE. WE REFERRED TO THIS PHENOMENON AS EMT MEMORY. UTILIZING A DOXYCYCLINE-CONTROLLED SLUG EXPRESSION SYSTEM, WE FOUND THAT HIGH EXPRESSION OF THE TRANSCRIPTION FACTOR SLUG WAS INDISPENSABLE FOR THE ESTABLISHMENT OF EMT MEMORY. HIGH SLUG EXPRESSION IN TUMORS OF LUNG CANCER PATIENTS WAS ASSOCIATED WITH POOR SURVIVAL. CHEMICAL OR GENETIC INHIBITION OF SLUG UPREGULATION PREVENTED EMT FOLLOWING THE ACUTE IL-1BETA EXPOSURE BUT DID NOT REVERSE EMT MEMORY. CHROMATIN IMMUNOPRECIPITATION AND METHYLATION-SPECIFIC PCR FURTHER REVEALED A SLUG-MEDIATED TEMPORAL REGULATION OF EPIGENETIC MODIFICATIONS, INCLUDING ACCUMULATION OF H3K27, H3K9, AND DNA METHYLATION, IN THE CDH1 (E-CADHERIN) PROMOTER FOLLOWING THE CHRONIC IL-1BETA EXPOSURE. CHEMICAL INHIBITION OF DNA METHYLATION NOT ONLY RESTORED E-CADHERIN EXPRESSION IN EMT MEMORY, BUT ALSO PRIMED CELLS FOR CHEMOTHERAPY-INDUCED APOPTOSIS. 2020 16 6756 24 WNT ANTAGONIST, SFRP1, IS HEDGEHOG SIGNALING TARGET. HEDGEHOG AND WNT SIGNALING PATHWAYS NETWORK TOGETHER DURING EMBRYOGENESIS AND CARCINOGENESIS. HEDGEHOG SIGNALING IN INTESTINAL EPITHELIUM REPRESSES CANONICAL WNT SIGNALING TO RESTRICT EXPRESSION OF WNT TARGET GENES TO STEM OR PROGENITOR CELLS; HOWEVER, THE MECHANISM REMAINS UNCLEAR. THE HEDGEHOG SIGNAL IS TRANSDUCED TO GLI FAMILY TRANSCRIPTION FACTORS THOUGH PATCHED RECEPTOR, SMOOTHENED SIGNAL TRANSDUCER, AND OTHER SIGNALING COMPONENTS, SUCH AS KIF27, KIF7, STK36, SUFU, AND DZIP1. HERE, WE SEARCHED FOR THE GLI-BINDING SITE WITHIN THE PROMOTER REGION OF GENES ENCODING SECRETED-TYPE WNT SIGNAL INHIBITORS, INCLUDING SFRP1, SFRP2, SFRP3, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, AND WIF1. THE GLI-BINDING SITE WAS IDENTIFIED WITHIN THE HUMAN SFRP1 PROMOTER BASED ON BIOINFORMATICS AND HUMAN INTELLIGENCE. THE CHIMPANZEE SFRP1 GENE WAS IDENTIFIED WITHIN THE NW_110515.1 GENOME SEQUENCE. THE GLI-BINDING SITE OF THE HUMAN SFRP1 PROMOTER WAS CONSERVED IN CHIMPANZEE SFRP1, MOUSE SFRP1, AND RAT SFRP1 PROMOTERS. SFRP1 IS THE EVOLUTIONARILY CONSERVED TARGET OF THE HEDGEHOG-GLI SIGNALING PATHWAY. EXPRESSION DOMAIN ANALYSES BASED ON TEXT MINING REVEALED THAT INDIAN HEDGEHOG (IHH), SFRP1, AND WNT6 ARE EXPRESSED IN DIFFERENTIATED INTESTINAL EPITHELIAL CELLS, MESENCHYMAL CELLS, AND STEM/PROGENITOR CELLS, RESPECTIVELY. HEDGEHOG IS SECRETED FROM DIFFERENTIATED EPITHELIAL CELLS TO INDUCE SFRP1 EXPRESSION IN MESENCHYMAL CELLS, WHICH KEEPS DIFFERENTIATED EPITHELIAL CELLS AWAY FROM THE EFFECTS OF CANONICAL WNT SIGNALING. THESE FACTS INDICATE THAT SFRP1 IS THE HEDGEHOG TARGET TO CONFINE CANONICAL WNT SIGNALING WITHIN STEM OR PROGENITOR CELLS. THEREFORE, EPIGENETIC CPG HYPERMETHYLATION OF THE SFRP1 PROMOTER DURING CHRONIC PERSISTENT INFLAMMATION AND AGING LEADS TO THE OCCURRENCE OF GASTROINTESTINAL CANCERS, SUCH AS COLORECTAL CANCER AND GASTRIC CANCER, THROUGH THE BREAKDOWN OF HEDGEHOG-DEPENDENT WNT SIGNAL INHIBITION. 2006 17 1483 29 DKK1 IS EPIGENETICALLY DOWNREGULATED BY PROMOTER METHYLATION AND INHIBITS BILE ACID-INDUCED GASTRIC INTESTINAL METAPLASIA. DICKKOPF-RELATED PROTEIN 1 (DKK1) IS ESSENTIAL TO GASTRIC CANCER AS AN INHIBITOR OF WNT SIGNALING. GASTRIC INTESTINAL METAPLASIA (GIM) IS AN IMPORTANT PRECANCEROUS LESION OF GASTRIC CANCER THAT CAN BE ACTIVATED BY BILE ACID REFLUX AND CHRONIC INFLAMMATION. HOWEVER, THE EXACT MECHANISM OF DKK1 IN BILE ACID-INDUCED GIM HAS NOT BEEN COMPLETELY ELUCIDATED. WE AIMED TO EXPLORE THE EPIGENETIC ALTERATIONS AND BIOLOGICAL FUNCTIONS OF DKK1 IN THE DEVELOPMENT OF GIM. IN THE PRESENT STUDY, BILE ACID WAS FOUND TO INDUCE THE EXPRESSION OF INTESTINAL MARKERS IN GASTRIC EPITHELIAL CELLS, WHEREAS DKK1 WAS DOWNREGULATED IN RESPONSE TO BILE ACID STIMULATION. THE MRNA AND PROTEIN EXPRESSION LEVELS OF DKK1 WERE DECREASED IN GIM TISSUES AS EVIDENCED BY QRT-PCR AND IMMUNOHISTOCHEMICAL STAINING. SURPRISINGLY, THE METHYLATION OF THE DKK1 PROMOTER INCREASED IN GIM TISSUES, AND WE DISCOVERED 28 DIFFERENTIAL METHYLATION SITES OF THE DKK1 PROMOTER IN GIM TISSUES. BILE ACID WAS ABLE TO INDUCE THE PARTIAL METHYLATION OF THE DKK1 PROMOTER, WHILE 5-AZA COULD INCREASE DKK1 EXPRESSION AS WELL AS DECREASE INTESTINAL MARKERS EXPRESSION IN GASTRIC EPITHELIAL CELLS. IN CONCLUSION, THE PROMOTER METHYLATION AND DOWNREGULATION OF DKK1 MIGHT PLAY IMPORTANT ROLES IN THE DEVELOPMENT OF GIM, ESPECIALLY BILE ACID-INDUCED GIM. 2020 18 5479 39 RESVERATROL ATTENUATES CIGARETTE SMOKE EXTRACT INDUCED CELLULAR SENESCENCE IN HUMAN AIRWAY EPITHELIAL CELLS BY REGULATING THE MIR-34A/SIRT1/NF-KAPPAB PATHWAY. CHRONIC OBSTRUCTIVE PULMONARY DISEASE (COPD) IS CHARACTERIZED BY ACCELERATED LUNG AGING. SMOKING IS THE CRITICAL RISK FACTOR FOR COPD. CELLULAR SENESCENCE OF AIRWAY EPITHELIAL CELLS IS THE CYTOLOGICAL BASIS OF ACCELERATED LUNG AGING IN COPD, AND THE REGULATION OF MICRORNAS (MIRNAS) IS THE CENTRAL EPIGENETIC MECHANISM OF CELLULAR SENESCENCE. RESVERATROL (RES) IS A POLYPHENOL WITH ANTI-AGING PROPERTIES. THIS STUDY INVESTIGATED WHETHER RES ATTENUATES CIGARETTE SMOKE EXTRACT (CSE)-INDUCED CELLULAR SENESCENCE IN HUMAN AIRWAY EPITHELIAL CELLS (BEAS-2B) THROUGH THE MIR-34A/SIRT1/NUCLEAR FACTOR-KAPPAB (NF-KAPPAB) PATHWAY. BEAS-2B CELLS WERE TREATED WITH RES, CSE AND TRANSFECTED WITH MIR-34A-5P MIMICS. CELLULAR SENESCENCE WAS EVALUATED BY SENESCENCE -RELATED BETA-GALACTOSIDASE (SA-BETA-GAL) STAINING AND EXPRESSION OF SENESCENCE-RELATED GENES (P16, P21, AND P53). THE EXPRESSIONS OF MIR-34A-5P, SIRT1, AND NF-KAPPAB P65 WERE EXAMINED USING QUANTITATIVE REAL TIME POLYMERASE CHAIN REACTION AND WESTERN BLOTTING. THE SENESCENCE-ASSOCIATED SECRETORY PHENOTYPE (SASP) CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) WERE ASSESSED BY ENZYME-LINKED IMMUNOSORBENT ASSAY. THE BINDING BETWEEN MIR-34A-5P AND SIRT1 WAS CONFIRMED BY DUAL-LUCIFERASE REPORTER ASSAY. THE RESULTS SHOWED THAT CSE DOSE-DEPENDENTLY DECREASED CELL VIABILITY AND ELEVATED CELLULAR SENESCENCE, CHARACTERIZED BY INCREASED SA-BETA-GAL STAINING AND SENESCENCE-RELATED GENE EXPRESSIONS (P16, P21, AND P53). FURTHER, CSE DOSE-DEPENDENTLY INCREASED THE EXPRESSION OF MIR-34A-5P AND SASP CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) IN BEAS-2B CELLS. PRETREATMENT WITH RES INHIBITED CSE-INDUCED CELLULAR SENESCENCE AND SECRETION OF SASP CYTOKINES (IL-1BETA, IL-6, IL-8, TNF-ALPHA) IN A DOSE-DEPENDENT MANNER. MOREOVER, RES REVERSED THE CSE-INDUCED DOWN-REGULATION OF SIRT1 AND UP-REGULATION OF MIR-34A-5P AND NF-KAPPAB P65. SIRT1 IS A TARGET OF MIR-34A-5P. OVEREXPRESSION OF MIR-34A-5P VIA TRANSFECTION WITH MIR-34A-5P MIMIC IN BEAS-2B CELLS ATTENUATED THE INHIBITORY EFFECT OF RES ON CELLULAR SENESCENCE, ACCOMPANIED BY REVERSING THE EXPRESSION OF SIRT1 AND NF-KAPPAB P65. IN CONCLUSION, RES ATTENUATED CSE-INDUCED CELLULAR SENESCENCE IN BEAS-2B CELLS BY REGULATING THE MIR-34A/SIRT1/NF-KAPPAB PATHWAY, WHICH MAY PROVIDE A NEW APPROACH FOR COPD TREATMENT. 2022 19 2842 30 FREQUENT CONCOMITANT EPIGENETIC SILENCING OF SOX1 AND SECRETED FRIZZLED-RELATED PROTEINS (SFRPS) IN HUMAN HEPATOCELLULAR CARCINOMA. BACKGROUND AND AIM: EXCEPT FOR GENETIC MUTATIONS, EPIGENETIC CHANGES ARE ALSO INVOLVED IN THE DEVELOPMENT OF HUMAN CANCERS. RECENTLY, WE HAVE IDENTIFIED SOX1, SRY (SEX DETERMINING REGION Y)-BOX 1, IS HYPERMETHYLATED IN CERVICAL CANCER AND OVARIAN CANCER. THEREFORE, WE INVESTIGATED WHETHER PROMOTER HYPERMETHYLATION OF SOX1 IS COMMON IN HEPATOCELLULAR CARCINOMA (HCC). METHODS: WE USED METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MS-PCR) AND BISULFITE SEQUENCING TO ANALYZE THE METHYALTION LEVEL OF THE SOX1 PROMOTER IN SEVEN HCC CELL LINES, 54 CLINICAL HCCS, 42 CIRRHOTIC LIVERS, 21 LIVERS WITH CHRONIC HEPATITIS, AND 15 CONTROL LIVERS. THEN, WE EMPLOYED QUANTITATIVE MS-PCR (QMSP) TO VALIDATE IN AN INDEPENDENT SET OF SAMPLES (60 PAIRED HCCS AND 30 CONTROL LIVERS). FINALLY, WE USED LUCIFERASE REPORTER AND COLONY FORMATION ASSAY TO CHECK THE EFFECT OF SOX1 IN HCC. RESULTS: PROMOTER METHYLATION OF SOX1 WAS SIGNIFICANTLY FREQUENT IN HCC CELL LINES AND CLINICAL HCCS, CIRRHOTIC LIVERS, BUT NOT IN CONTROL LIVERS (P < 0.0001). THERE IS A SIGNIFICANT CORRELATION BETWEEN DOWNREGULATION OF SOX1 EXPRESSION AND PROMOTER METHYLATION. QMSP RESULTS CONFIRMED THAT PROMOTER HYPERMETHYLATION OF SOX1 IS SIGNIFICANTLY MORE FREQUENT IN HCCS THAN CONTROL LIVERS (P < 0.0001). THE FREQUENCY OF SOX1 METHYLATION IN PATIENTS WITH SECRETED FRIZZLED-RELATED PROTEINS (SFRPS) METHYLATION IS SIGNIFICANTLY HIGHER THAN IN PATIENTS WITHOUT SFRPS METHYLATION (P < 0.0001). FURTHERMORE, ECTOPIC EXPRESSION OF SOX1 COULD SUPPRESS T-CELL FACTOR-DEPENDENT TRANSCRIPTIONAL ACTIVITY AND COLONY FORMATION NUMBER IN HCCS. CONCLUSIONS: CONCOMITANT EPIGENETIC SILENCING OF SOX1 AND SFRPS THROUGH PROMOTER HYPERMETHYLATION IS FREQUENT IN HCCS, AND THIS MIGHT CONTRIBUTE TO ABNORMAL ACTIVATION OF CANONICAL WNT SIGNAL PATHWAY. 2013 20 4180 34 MERCURY EXPOSURE INDUCES CYTOSKELETON DISRUPTION AND LOSS OF RENAL FUNCTION THROUGH EPIGENETIC MODULATION OF MMP9 EXPRESSION. MERCURY IS ONE OF THE MAJOR HEAVY METAL POLLUTANTS OCCURRING IN ELEMENTAL, INORGANIC AND ORGANIC FORMS. DUE TO BAN ON MOST INORGANIC MERCURY CONTAINING PRODUCTS, HUMAN EXPOSURE TO MERCURY GENERALLY OCCURS AS METHYLMERCURY (MEHG) BY CONSUMPTION OF CONTAMINATED FISH AND OTHER SEA FOOD. ANIMAL AND EPIDEMIOLOGICAL STUDIES INDICATE THAT MEHG AFFECTS NEURAL AND RENAL FUNCTION. OUR STUDY IS FOCUSED ON NEPHROTOXIC POTENTIAL OF MEHG. IN THIS STUDY, WE HAVE SHOWN FOR THE FIRST TIME HOW MEHG COULD EPIGENETICALLY MODULATE MATRIX METALLOPROTEINASE 9(MMP9) TO PROMOTE NEPHROTOXICITY USING AN ANIMAL MODEL OF SUB CHRONIC MEHG EXPOSURE. MEHG CAUSED RENAL TOXICITY AS WAS SEEN BY INCREASED LEVELS OF SERUM CREATININE AND EXPRESSION OF EARLY NEPHROTOXICITY MARKERS (KIM-1, CLUSTERIN, IP-10, AND TIMP). MEHG EXPOSURE ALSO CORRELATED STRONGLY WITH INDUCTION OF MMP9 MRNA AND PROTEIN IN A DOSE DEPENDENT MANNER. FURTHER, WHILE INDUCTION OF MMP9 PROMOTED CYTOSKELETON DISRUPTION AND LOSS OF CELL-CELL ADHESION (LOSS OF F-ACTIN, VIMENTIN AND FIBRONECTIN), INHIBITION OF MMP9 WAS FOUND TO REDUCE THESE DISRUPTIONS. MECHANISTIC STUDIES BY CHIP ANALYSIS SHOWED THAT MEHG MODULATED MMP9 BY PROMOTING DEMETHYLATION OF ITS REGULATORY REGION TO INCREASE ITS EXPRESSION. BISULFITE SEQUENCING IDENTIFIED CRITICAL CPGS IN THE FIRST EXON OF MMP9 WHICH WERE DEMETHYLATED FOLLOWING MEHG EXPOSURE. CHIP STUDIES ALSO SHOWED LOSS OF METHYL BINDING PROTEIN, MECP2 AND TRANSCRIPTION FACTOR PEA3 AT THE DEMETHYLATED SITE CONFIRMING DECREASED CPG METHYLATION. OUR STUDIES THUS SHOW HOW MEHG COULD EPIGENETICALLY MODULATE MMP9 TO PROMOTE CYTOSKELETON DISRUPTION LEADING TO LOSS OF RENAL FUNCTION. 2017