{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T02:48:33Z","timestamp":1788922113188,"version":"build-2803163510"},"reference-count":81,"publisher":"MDPI AG","issue":"15","license":[{"start":{"date-parts":[[2025,7,22]],"date-time":"2025-07-22T00:00:00Z","timestamp":1753142400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Spanish Government","award":["TED2021-132681B-I00"],"award-info":[{"award-number":["TED2021-132681B-I00"]}]},{"name":"Spanish Government","award":["CPP2021-008458"],"award-info":[{"award-number":["CPP2021-008458"]}]},{"name":"Spanish Government","award":["PI20\/01131"],"award-info":[{"award-number":["PI20\/01131"]}]},{"name":"Spanish Government","award":["PI24\/01600"],"award-info":[{"award-number":["PI24\/01600"]}]},{"name":"Spanish Government","award":["IT1547-22"],"award-info":[{"award-number":["IT1547-22"]}]},{"name":"European Union","award":["TED2021-132681B-I00"],"award-info":[{"award-number":["TED2021-132681B-I00"]}]},{"name":"European Union","award":["CPP2021-008458"],"award-info":[{"award-number":["CPP2021-008458"]}]},{"name":"European Union","award":["PI20\/01131"],"award-info":[{"award-number":["PI20\/01131"]}]},{"name":"European Union","award":["PI24\/01600"],"award-info":[{"award-number":["PI24\/01600"]}]},{"name":"European Union","award":["IT1547-22"],"award-info":[{"award-number":["IT1547-22"]}]},{"name":"Basque Government Research Groups","award":["TED2021-132681B-I00"],"award-info":[{"award-number":["TED2021-132681B-I00"]}]},{"name":"Basque Government Research Groups","award":["CPP2021-008458"],"award-info":[{"award-number":["CPP2021-008458"]}]},{"name":"Basque Government Research Groups","award":["PI20\/01131"],"award-info":[{"award-number":["PI20\/01131"]}]},{"name":"Basque Government Research Groups","award":["PI24\/01600"],"award-info":[{"award-number":["PI24\/01600"]}]},{"name":"Basque Government Research Groups","award":["IT1547-22"],"award-info":[{"award-number":["IT1547-22"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Epigenetic regulation, particularly DNA methylation, plays a crucial role in embryonic development by controlling gene expression patterns. The disruption of this regulation by environmental factors can have long-lasting consequences. Opioid drugs, such as morphine, are known to cross the placental barrier and affect the developing central nervous system, yet their precise epigenetic effects during early development remain unclear. This study aimed to elucidate the impact of chronic morphine exposure on the DNA methylation landscape and gene expression in mouse embryonic stem cells (mESCs). mESCs were chronically exposed to morphine (10 \u03bcM for 24 h). Genome-wide bisulfite sequencing was performed to identify DNA methylation changes, while RNA sequencing (RNA-Seq) assessed corresponding gene expression alterations. Global levels of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) were quantified using mass spectrometry. Morphine exposure induced global DNA hypomethylation and identified 16,808 differentially methylated genes (DMGs) related to development, cell signalling, metabolism, and transcriptional regulation. Integrative transcriptomic analysis with RNA-Seq data revealed 651 overlapping genes, including alterations in key epigenetic regulators involved on DNA methylation machinery. Specifically, Tet1 was upregulated with promoter hypomethylation, while Dnmt1 was downregulated, without changes in promoter methylation after morphine exposiure. Mass spectrometry results confirmed a global decrease in 5mC levels alongside increased 5hmC, indicating the involvement of both passive and active demethylation pathways. These findings demonstrate for the first time that morphine disrupts the epigenetic homeostasis of mESCs by promoting global and gene-specific DNA demethylation, which might be key to the phenotypic changes that occur in adulthood. This work provides novel mechanistic insights into how opioid exposure during early development may lead to persistent epigenetic alterations, with potential long-term implications for neurodevelopment and disease susceptibility.<\/jats:p>","DOI":"10.3390\/ijms26157056","type":"journal-article","created":{"date-parts":[[2025,7,23]],"date-time":"2025-07-23T08:02:06Z","timestamp":1753257726000},"page":"7056","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Chronic Morphine Treatment Leads to a Global DNA Hypomethylation via Active and Passive Demethylation Mechanisms in mESCs"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1838-4594","authenticated-orcid":false,"given":"Manu","family":"Araolaza","sequence":"first","affiliation":[{"name":"Department of Physiology, Faculty of Medicine and Nursery, University of the Basque Country, 48940 Leioa, Spain"},{"name":"Bizkaia Health Research Institute, 48903 Barakaldo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5107-6991","authenticated-orcid":false,"given":"Iraia","family":"Mu\u00f1oa-Hoyos","sequence":"additional","affiliation":[{"name":"Department of Physiology, Faculty of Medicine and Nursery, University of the Basque Country, 48940 Leioa, Spain"},{"name":"Bizkaia Health Research Institute, 48903 Barakaldo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6390-7887","authenticated-orcid":false,"given":"Itziar","family":"Urizar-Arenaza","sequence":"additional","affiliation":[{"name":"Department of Physiology, Faculty of Medicine and Nursery, University of the Basque Country, 48940 Leioa, Spain"},{"name":"Bizkaia Health Research Institute, 48903 Barakaldo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1787-6792","authenticated-orcid":false,"given":"Irune","family":"Calzado","sequence":"additional","affiliation":[{"name":"Department of Physiology, Faculty of Medicine and Nursery, University of the Basque Country, 48940 Leioa, Spain"},{"name":"Bizkaia Health Research Institute, 48903 Barakaldo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9202-1287","authenticated-orcid":false,"given":"Nerea","family":"Subir\u00e1n","sequence":"additional","affiliation":[{"name":"Department of Physiology, Faculty of Medicine and Nursery, University of the Basque Country, 48940 Leioa, Spain"},{"name":"Bizkaia Health Research Institute, 48903 Barakaldo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,7,22]]},"reference":[{"key":"ref_1","first-page":"763","article-title":"Epigenetic Biomarkers and Cardiovascular Disease: Circulating MicroRNAs","volume":"70","year":"2017","journal-title":"Rev. Espa\u00f1ola Cardiol. (Engl. Ed.)"},{"key":"ref_2","first-page":"246","article-title":"Role of Epigenetics in Biology and Human Diseases","volume":"20","author":"Moosavi","year":"2016","journal-title":"Iran. Biomed. J."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.reprotox.2019.08.006","article-title":"Prenatal Toxicity of the Environmental Pollutants on Neuronal and Cardiac Development Derived from Embryonic Stem Cells","volume":"90","author":"Ko","year":"2019","journal-title":"Reprod. Toxicol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"819","DOI":"10.1038\/383819a0","article-title":"Loss of Morphine-Induced Analgesia, Reward Effect and Withdrawal Symptoms in Mice Lacking the Mu-Opioid-Receptor Gene","volume":"383","author":"Matthes","year":"1996","journal-title":"Nature"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"321","DOI":"10.1038\/nature10954","article-title":"Crystal Structure of the \u03bc-Opioid Receptor Bound to a Morphinan Antagonist","volume":"485","author":"Manglik","year":"2012","journal-title":"Nature"},{"key":"ref_6","first-page":"294","article-title":"Opioid Therapy and Management of Side Effects Associated with Opioids","volume":"44","author":"Nakatani","year":"2017","journal-title":"Gan Kagaku Ryoho"},{"key":"ref_7","first-page":"71","article-title":"The Effect of Morphine Consumption on Plasma Corticosteron Concentration and Placenta Development in Pregnant Rats","volume":"9","author":"Kazemi","year":"2011","journal-title":"Iran. J. Reprod. Med."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1016\/S0376-8716(98)00070-2","article-title":"Prenatal Effects of Drugs of Abuse on Brain Development","volume":"51","author":"Levitt","year":"1998","journal-title":"Drug Alcohol. Depend."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"187","DOI":"10.1016\/0376-8716(89)90055-0","article-title":"Effects of Prenatal Morphine Treatment of Rats on Mortality, Bodyweight and Analgesic Response in the Offspring","volume":"24","author":"Eriksson","year":"1989","journal-title":"Drug Alcohol. Depend."},{"key":"ref_10","first-page":"130","article-title":"Evaluating the Effects of Oral Morphine on Embryonic Development of Cerebellum in Wistar Rats","volume":"4","author":"Niknam","year":"2013","journal-title":"Basic Clin. Neurosci."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"200","DOI":"10.1016\/j.bbr.2010.11.059","article-title":"Adolescent Opioid Exposure in Female Rats: Transgenerational Effects on Morphine Analgesia and Anxiety-like Behavior in Adult Offspring","volume":"218","author":"Byrnes","year":"2011","journal-title":"Behav. Brain Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"667","DOI":"10.1038\/nn.3695","article-title":"Implication of Sperm RNAs in Transgenerational Inheritance of the Effects of Early Trauma in Mice","volume":"17","author":"Gapp","year":"2014","journal-title":"Nat. Neurosci."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"846","DOI":"10.2119\/molmed.2010.00268","article-title":"Regulation of Male Fertility by the Opioid System","volume":"17","author":"Casis","year":"2011","journal-title":"Mol. Med."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1615\/CritRevImmunol.2020033728","article-title":"DNA Methylation in T-Cell Development and Differentiation","volume":"40","author":"Correa","year":"2020","journal-title":"Crit. Rev. Immunol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"204","DOI":"10.1038\/nrg3354","article-title":"DNA Methylation: Roles in Mammalian Development","volume":"14","author":"Smith","year":"2013","journal-title":"Nat. Rev. Genet."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1111\/dgd.12716","article-title":"Pluripotent Stem Cells in the Research for Extraembryonic Cell Differentiation","volume":"63","author":"Toyooka","year":"2021","journal-title":"Dev. Growth Differ."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"4149","DOI":"10.1128\/MCB.18.7.4149","article-title":"Cis-Acting Signal for Inheritance of Imprinted DNA Methylation Patterns in the Preimplantation Mouse Embryo","volume":"18","author":"Howell","year":"1998","journal-title":"Mol. Cell. Biol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"829","DOI":"10.1016\/S0092-8674(01)00280-X","article-title":"Genomic Imprinting Disrupted by a Maternal Effect Mutation in the Dnmt1 Gene","volume":"104","author":"Howell","year":"2001","journal-title":"Cell"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"McGraw, S., Oakes, C.C., Martel, J., Cirio, M.C., de Zeeuw, P., Mak, W., Plass, C., Bartolomei, M.S., Chaillet, J.R., and Trasler, J.M. (2013). Loss of DNMT1o Disrupts Imprinted X Chromosome Inactivation and Accentuates Placental Defects in Females. PLoS Genet., 9.","DOI":"10.1371\/journal.pgen.1003873"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"915","DOI":"10.1016\/0092-8674(92)90611-F","article-title":"Targeted Mutation of the DNA Methyltransferase Gene Results in Embryonic Lethality","volume":"69","author":"Li","year":"1992","journal-title":"Cell"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1670","DOI":"10.1038\/cr.2011.189","article-title":"Generation and Replication-Dependent Dilution of 5fC and 5caC during Mouse Preimplantation Development","volume":"21","author":"Inoue","year":"2011","journal-title":"Cell Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1038\/jhg.2013.63","article-title":"Tet Family of 5-Methylcytosine Dioxygenases in Mammalian Development","volume":"58","author":"Zhao","year":"2013","journal-title":"J. Hum. Genet."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1016\/j.cell.2013.12.019","article-title":"Reversing DNA Methylation: Mechanisms, Genomics, and Biological Functions","volume":"156","author":"Wu","year":"2014","journal-title":"Cell"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1016\/j.stem.2013.08.005","article-title":"Dna Demethylation in Pluripotency and Reprogramming: The Role of Tet Proteins and Cell Division","volume":"13","author":"Bagci","year":"2013","journal-title":"Cell Stem Cell"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1016\/j.neulet.2018.12.046","article-title":"Mesenchymal Stem Cells from Bone Marrow Attenuated the Chronic Morphine-Induced CAMP Accumulation In Vitro","volume":"698","author":"Yang","year":"2019","journal-title":"Neurosci. Lett."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Cain, J.A., Montibus, B., and Oakey, R.J. (2022). Intragenic CpG Islands and Their Impact on Gene Regulation. Front. Cell Dev. Biol., 10.","DOI":"10.3389\/fcell.2022.832348"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1010","DOI":"10.1101\/gad.2037511","article-title":"CpG Islands and the Regulation of Transcription","volume":"25","author":"Deaton","year":"2011","journal-title":"Genes Dev."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1016\/j.febslet.2009.04.012","article-title":"CpG Islands\u2014\u201cA Rough Guide\u201d","volume":"583","author":"Illingworth","year":"2009","journal-title":"FEBS Lett."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"457","DOI":"10.1038\/ng1990","article-title":"Distribution, Silencing Potential and Evolutionary Impact of Promoter DNA Methylation in the Human Genome","volume":"39","author":"Weber","year":"2007","journal-title":"Nat. Genet."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"170","DOI":"10.1186\/s13148-020-00955-w","article-title":"Morphine Leads to Global Genome Changes in H3K27me3 Levels via a Polycomb Repressive Complex 2 (PRC2) Self-Regulatory Mechanism in MESCs","volume":"12","author":"Halsall","year":"2020","journal-title":"Clin. Epigenetics"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Zeng, Y., and Chen, T. (2019). DNA Methylation Reprogramming during Mammalian Development. Genes, 10.","DOI":"10.3390\/genes10040257"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"25893","DOI":"10.1074\/jbc.M603140200","article-title":"Reconstitution and Mechanism of the Stimulation of de Novo Methylation by Human DNMT3L","volume":"281","author":"Kareta","year":"2006","journal-title":"J. Biol. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"562","DOI":"10.1242\/dev.089268","article-title":"Plasticity in Dnmt3L-Dependent and -Independent Modes of de Novo Methylation in the Developing Mouse Embryo","volume":"140","author":"Guenatri","year":"2013","journal-title":"Development"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"866","DOI":"10.1161\/01.RES.0000041435.95082.84","article-title":"Derivation and Potential Applications of Human Embryonic Stem Cells","volume":"91","author":"Gepstein","year":"2002","journal-title":"Circ. Res."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"929","DOI":"10.1016\/j.bpobgyn.2004.06.005","article-title":"Human Embryonic Stem Cells as a Model for Early Human Development","volume":"18","author":"Dvash","year":"2004","journal-title":"Best Pract. Res. Clin. Obstet. Gynaecol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"111","DOI":"10.1203\/01.pdr.0000228349.24676.17","article-title":"Human Embryonic Stem Cells as a Powerful Tool for Studying Human Embryogenesis","volume":"60","author":"Dvash","year":"2006","journal-title":"Pediatr. Res."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Tian, W., Zhao, M., Li, M., Song, T., Zhang, M., Quan, L., Li, S., and Sun, Z.S. (2012). Reversal of Cocaine-Conditioned Place Preference through Methyl Supplementation in Mice: Altering Global DNA Methylation in the Prefrontal Cortex. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0033435"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Novikova, S.I., He, F., Bai, J., Cutrufello, N.J., Lidow, M.S., and Undieh, A.S. (2008). Maternal Cocaine Administration in Mice Alters DNA Methylation and Gene Expression in Hippocampal Neurons of Neonatal and Prepubertal Offspring. PLoS ONE, 3.","DOI":"10.1371\/journal.pone.0001919"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2993","DOI":"10.1038\/npp.2015.155","article-title":"Genome-Wide DNA Methylation Profiling Reveals Epigenetic Changes in the Rat Nucleus Accumbens Associated With Cross-Generational Effects of Adolescent THC Exposure","volume":"40","author":"Watson","year":"2015","journal-title":"Neuropsychopharmacology"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1016\/j.toxlet.2016.01.010","article-title":"Morphine Sulfate Concomitantly Decreases Neuronal Differentiation and Opioid Receptor Expression in Mouse Embryonic Stem Cells","volume":"247","author":"Dholakiya","year":"2016","journal-title":"Toxicol. Lett."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1431","DOI":"10.1111\/j.1471-4159.2009.06479.x","article-title":"Mu and Kappa Opioids Modulate Mouse Embryonic Stem Cell-Derived Neural Progenitor Differentiation via MAP Kinases","volume":"112","author":"Hahn","year":"2010","journal-title":"J. Neurochem."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"109215","DOI":"10.1016\/j.celrep.2021.109215","article-title":"Contiguous Erosion of the Inactive X in Human Pluripotency Concludes with Global DNA Hypomethylation","volume":"35","author":"Bansal","year":"2021","journal-title":"Cell Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"848","DOI":"10.1016\/j.molcel.2016.04.025","article-title":"Impairment of DNA Methylation Maintenance Is the Main Cause of Global Demethylation in Naive Embryonic Stem Cells","volume":"62","author":"Iurlaro","year":"2016","journal-title":"Mol. Cell"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"311","DOI":"10.1038\/nsmb.2510","article-title":"Naive pluripotency is associated with global DNA hypomethylation","volume":"20","author":"Leitch","year":"2013","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Montgomery, T., Uh, K., and Lee, K. (2024). TET Enzyme Driven Epigenetic Reprogramming in Early Embryos and Its Implication on Long-Term Health. Front. Cell Dev. Biol., 12.","DOI":"10.3389\/fcell.2024.1358649"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"2150","DOI":"10.1016\/j.celrep.2020.01.065","article-title":"Regulatory Dynamics of Tet1 and Oct4 Resolve Stages of Global DNA Demethylation and Transcriptomic Changes in Reprogramming","volume":"30","author":"Bartoccetti","year":"2020","journal-title":"Cell Rep."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1186\/s13072-019-0278-5","article-title":"MYC deregulates TET1 and TET2 expression to control global DNA (hydroxy)methylation and gene expression to maintain a neoplastic phenotype in T-ALL","volume":"12","author":"Poole","year":"2019","journal-title":"Epigenetics Chromatin"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"474","DOI":"10.1016\/j.bbagen.2017.10.016","article-title":"Morphine delays neural stem cells differentiation by facilitating Nestin overexpression","volume":"1862","author":"Rodriguez","year":"2018","journal-title":"Biochim. Biophys. Acta (BBA)-Gen. Subj."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"18347","DOI":"10.1074\/jbc.R110.205286","article-title":"DNA Methylation and Demethylation in Mammals","volume":"286","author":"Chen","year":"2011","journal-title":"J. Biol. Chem."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"18542","DOI":"10.1074\/jbc.M117.810457","article-title":"Passive DNA Demethylation Preferentially Up-Regulates Pluripotency-Related Genes and Facilitates the Generation of Induced Pluripotent Stem Cells","volume":"292","author":"He","year":"2017","journal-title":"J. Biol. Chem."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1016\/j.biocel.2008.09.016","article-title":"Histone H2A Ubiquitination in Transcriptional Regulation and DNA Damage Repair","volume":"41","author":"Zhou","year":"2009","journal-title":"Int. J. Biochem. Cell Biol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"107778","DOI":"10.1016\/j.neuropharm.2019.107778","article-title":"Reversal of Oxycodone Conditioned Place Preference by Oxytocin: Promoting Global DNA Methylation in the Hippocampus","volume":"160","author":"Fan","year":"2019","journal-title":"Neuropharmacology"},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Fan, X.Y., Shi, G., He, X.J., Li, X.Y., Wan, Y.X., and Jian, L.Y. (2021). Oxytocin Prevents Cue-Induced Reinstatement of Oxycodone Seeking: Involvement of DNA Methylation in the Hippocampus. Addict. Biol., 26.","DOI":"10.1111\/adb.13025"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"6595","DOI":"10.1523\/JNEUROSCI.0695-19.2019","article-title":"Contribution of DNMT1 to Neuropathic Pain Genesis Partially through Epigenetically Repressing Kcna2 in Primary Afferent Neurons","volume":"39","author":"Sun","year":"2019","journal-title":"J. Neurosci."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"11435","DOI":"10.1073\/pnas.172091899","article-title":"CREB activity in the nucleus accumbens shell controls gating of behavioral responses to emotional stimuli","volume":"99","author":"Barrot","year":"2002","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"1300","DOI":"10.1126\/science.1210597","article-title":"Tet Proteins Can Convert 5-Methylcytosine to 5-Formylcytosine and 5-Carboxylcytosine","volume":"333","author":"Ito","year":"2011","journal-title":"Science"},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"1698","DOI":"10.4161\/cc.8.11.8580","article-title":"Prediction of Novel Families of Enzymes Involved in Oxidative and Other Complex Modifications of Bases in Nucleic Acids","volume":"8","author":"Iyer","year":"2009","journal-title":"Cell Cycle"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1038\/nrm3589","article-title":"TETonic Shift: Biological Roles of TET Proteins in DNA Demethylation and Transcription","volume":"14","author":"Pastor","year":"2013","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"930","DOI":"10.1126\/science.1170116","article-title":"Conversion of 5-Methylcytosine to 5-Hydroxymethylcytosine in Mammalian DNA by MLL Partner TET1","volume":"324","author":"Tahiliani","year":"2009","journal-title":"Science"},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"624","DOI":"10.1038\/nature01075","article-title":"The Polycomb Group Protein EZH2 Is Involved in Progression of Prostate Cancer","volume":"419","author":"Varambally","year":"2002","journal-title":"Nature"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"1456","DOI":"10.1158\/1541-7786.MCR-09-0121","article-title":"Polycomb Group Protein Enhancer of Zeste 2 Is an Oncogene That Promotes the Neoplastic Transformation of a Benign Prostatic Epithelial Cell Line","volume":"7","author":"Karanikolas","year":"2009","journal-title":"Mol. Cancer Res."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"1246","DOI":"10.2353\/ajpath.2009.090042","article-title":"Targeted Overexpression of EZH2 in the Mammary Gland Disrupts Ductal Morphogenesis and Causes Epithelial Hyperplasia","volume":"175","author":"Li","year":"2009","journal-title":"Am. J. Pathol."},{"key":"ref_63","unstructured":"Andrews, S. (2020, January 26). FastQC: A Quality Control Tool for High Throughput Sequence Data. Available online: https:\/\/www.bioinformatics.babraham.ac.uk\/projects\/fastqc\/."},{"key":"ref_64","unstructured":"Krueger, F., James, F., Ewels, P., Afyounian, E., and Schuster-Boeckler, B. (2020, January 26). FelixKrueger\/: v0.6.2. Available online: https:\/\/www.bioinformatics.babraham.ac.uk\/projects\/trim_galore\/."},{"key":"ref_65","unstructured":"Granlund, T., and Stallman, R.M. (2020, January 26). Cat. Available online: https:\/\/github.com\/Batch-Man\/cat."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"357","DOI":"10.1038\/nmeth.1923","article-title":"Fast gapped-read alignment with Bowtie 2","volume":"9","author":"Langmead","year":"2012","journal-title":"Nat. Methods"},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1093\/bioinformatics\/bty648","article-title":"Scaling read aligners to hundreds of threads on general-purpose processors","volume":"35","author":"Langmead","year":"2019","journal-title":"Bioinformatics"},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1038\/nmeth.1828","article-title":"DNA methylome analysis using short bisulfite sequencing data","volume":"9","author":"Krueger","year":"2012","journal-title":"Nat. Methods."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","article-title":"The Sequence Alignment\/Map format and SAMtools","volume":"25","author":"Li","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"2055","DOI":"10.12688\/f1000research.13196.1","article-title":"Differential methylation analysis of reduced representation bisulfite sequencing experiments using edgeR","volume":"6","author":"Chen","year":"2017","journal-title":"F1000Research"},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"139","DOI":"10.1093\/bioinformatics\/btp616","article-title":"edgeR: A Bioconductor package for differential expression analysis of digital gene expression data","volume":"26","author":"Robinson","year":"2010","journal-title":"Bioinformatics"},{"key":"ref_72","doi-asserted-by":"crossref","unstructured":"Akalin, A., Kormaksson, M., Li, S., Garrett-Bakelman, F.E., Figueroa, M.E., Melnick, A., and Mason, C.E. (2012). methylKit: A comprehensive R package for the analysis of genome-wide DNA methylation profiles. Genome Biol., 13.","DOI":"10.1186\/gb-2012-13-10-r87"},{"key":"ref_73","unstructured":"Oliveros, J.C. (2025, May 25). Venny. An Interactive Tool for Comparing Lists with Venn\u2019s Diagrams. 2007\u20132015. Available online: https:\/\/bioinfogp.cnb.csic.es\/tools\/venny\/index.html."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1038\/75556","article-title":"Gene ontology: Tool for the unification of biology. The Gene Ontology Consortium","volume":"25","author":"Ashburner","year":"2000","journal-title":"Nat. Genet."},{"key":"ref_75","doi-asserted-by":"crossref","unstructured":"The Gene Ontology Consortium (2019). The Gene Ontology Resource: 20 years and still GOing strong. Nucleic Acids Res., 47, D330\u2013D338.","DOI":"10.1093\/nar\/gky1055"},{"key":"ref_76","doi-asserted-by":"crossref","first-page":"996","DOI":"10.1101\/gr.229102","article-title":"The human genome browser at UCSC","volume":"12","author":"Kent","year":"2002","journal-title":"Genome Res."},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1016\/j.pbiomolbio.2015.03.002","article-title":"Multigenerational and Transgenerational Inheritance of Drug Exposure: The Effects of Alcohol, Opiates, Cocaine, Marijuana, and Nicotine","volume":"118","author":"Yohn","year":"2015","journal-title":"Prog. Biophys. Mol. Biol."},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"72","DOI":"10.1016\/j.bbr.2007.07.041","article-title":"Effect of Parental Morphine Addiction on Hippocampal Long-Term Potentiation in Rats Offspring","volume":"186","author":"Sarkaki","year":"2008","journal-title":"Behav. Brain Res."},{"key":"ref_79","doi-asserted-by":"crossref","first-page":"258","DOI":"10.5055\/jom.2011.0067","article-title":"Elevated Levels of DNA Methylation at the OPRM1 Promoter in Blood and Sperm from Male Opioid Addicts","volume":"7","author":"Chorbov","year":"2011","journal-title":"J. Opioid Manag."},{"key":"ref_80","doi-asserted-by":"crossref","unstructured":"Saeidinezhad, M., Razban, V., Safizadeh, H., and Ezzatabadipour, M. (2021). Effects of Maternal Consumption of Morphine on Rat Skeletal System Development. BMC Musculoskelet. Disord., 22.","DOI":"10.1186\/s12891-021-04321-6"},{"key":"ref_81","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.brainres.2008.09.052","article-title":"Effects of Maternal Oral Administration of Morphine Sulfate on Developing Rat Fetal Cerebrum: A Morphometrical Evaluation","volume":"1245","author":"Sadraie","year":"2008","journal-title":"Brain Res."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/15\/7056\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T18:14:12Z","timestamp":1760033652000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/15\/7056"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,22]]},"references-count":81,"journal-issue":{"issue":"15","published-online":{"date-parts":[[2025,8]]}},"alternative-id":["ijms26157056"],"URL":"https:\/\/doi.org\/10.3390\/ijms26157056","relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/2025.03.16.643512","asserted-by":"object"}]},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,7,22]]}}}