{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,1]],"date-time":"2025-11-01T09:32:16Z","timestamp":1761989536777,"version":"3.40.5"},"reference-count":54,"publisher":"Public Library of Science (PLoS)","issue":"11","license":[{"start":{"date-parts":[[2020,11,12]],"date-time":"2020-11-12T00:00:00Z","timestamp":1605139200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>The efficiency of cell reprogramming in two-dimensional (2D) cultures is limited. Given that cellular stemness is intimately related to microenvironmental changes, 3D cell cultures have the potential of overcoming this limited capacity by allowing cells to self-organize by aggregation. In 3D space, cells interact more efficiently, modify their cellular topology, gene expression, signaling, and metabolism. It is yet not clear as how 3D culture environments modify the reprogramming potential of fibroblasts. We demonstrate that 3D spheroids from dermal fibroblasts formed under ultra-low attachment conditions showed increased lactate production. This is a requisite for cell reprogramming, increase their expression of pluripotency genes, such as <jats:italic>OCT4<\/jats:italic>, <jats:italic>NANOG<\/jats:italic> and <jats:italic>SOX2<\/jats:italic>, and display upregulated cystathionine-\u03b2-synthase (<jats:italic>CBS<\/jats:italic>) and hydrogen sulfide (H<jats:sub>2<\/jats:sub>S) production. Knockdown of <jats:italic>CBS<\/jats:italic> by RNAi suppresses lactic acid and H<jats:sub>2<\/jats:sub>S production and concomitantly decreases the expression of <jats:italic>OCT4<\/jats:italic> and <jats:italic>NANOG<\/jats:italic>. On the contrary, H<jats:sub>2<\/jats:sub>S donors, NaHS and garlic-derived diallyl trisulfide (DATS), promote the expression of <jats:italic>OCT4<\/jats:italic>, and support osteogenic trans-differentiation of fibroblasts. These results demonstrate that <jats:italic>CBS<\/jats:italic> mediated release of H<jats:sub>2<\/jats:sub>S regulates the reprogramming of dermal fibroblasts grown in 3D cultures and supports their trans-differentiation.<\/jats:p>","DOI":"10.1371\/journal.pone.0241685","type":"journal-article","created":{"date-parts":[[2020,11,12]],"date-time":"2020-11-12T18:41:04Z","timestamp":1605206464000},"page":"e0241685","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":3,"title":["Hydrogen sulfide facilitates reprogramming and trans-differentiation in 3D dermal fibroblast"],"prefix":"10.1371","volume":"15","author":[{"given":"Elena A.","family":"Ostrakhovitch","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shin","family":"Akakura","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8419-7223","authenticated-orcid":true,"given":"Siamak","family":"Tabibzadeh","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"340","published-online":{"date-parts":[[2020,11,12]]},"reference":[{"issue":"5","key":"pone.0241685.ref001","doi-asserted-by":"crossref","first-page":"861","DOI":"10.1016\/j.cell.2007.11.019","article-title":"Induction of pluripotent stem cells from adult human fibroblasts by defined factors","volume":"131","author":"K Takahashi","year":"2007","journal-title":"Cell"},{"issue":"4","key":"pone.0241685.ref002","doi-asserted-by":"crossref","first-page":"663","DOI":"10.1016\/j.cell.2006.07.024","article-title":"Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors","volume":"126","author":"K Takahashi","year":"2006","journal-title":"Cell"},{"issue":"5858","key":"pone.0241685.ref003","doi-asserted-by":"crossref","first-page":"1917","DOI":"10.1126\/science.1151526","article-title":"Induced pluripotent stem cell lines derived from human somatic cells","volume":"318","author":"J Yu","year":"2007","journal-title":"Science"},{"issue":"7","key":"pone.0241685.ref004","doi-asserted-by":"crossref","first-page":"795","DOI":"10.1038\/nbt1418","article-title":"Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds","volume":"26","author":"D Huangfu","year":"2008","journal-title":"Nat Biotechnol"},{"issue":"7","key":"pone.0241685.ref005","doi-asserted-by":"crossref","first-page":"1617","DOI":"10.1016\/j.cell.2012.11.039","article-title":"A molecular roadmap of reprogramming somatic cells into iPS cells","volume":"151","author":"JM Polo","year":"2012","journal-title":"Cell"},{"issue":"12","key":"pone.0241685.ref006","doi-asserted-by":"crossref","first-page":"1154","DOI":"10.1038\/nmat3777","article-title":"Biophysical regulation of epigenetic state and cell reprogramming","volume":"12","author":"TL Downing","year":"2013","journal-title":"Nat Mater"},{"issue":"7","key":"pone.0241685.ref007","doi-asserted-by":"crossref","first-page":"769","DOI":"10.1038\/nbt.3270","article-title":"Transient acquisition of pluripotency during somatic cell transdifferentiation with iPSC reprogramming factors","volume":"33","author":"I Maza","year":"2015","journal-title":"Nat Biotechnol"},{"key":"pone.0241685.ref008","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1016\/j.ceb.2014.06.013","article-title":"Modeling human development in 3D culture","volume":"31","author":"M Ader","year":"2014","journal-title":"Curr Opin Cell Biol"},{"issue":"1","key":"pone.0241685.ref009","doi-asserted-by":"crossref","first-page":"9086","DOI":"10.1038\/s41598-019-45352-3","article-title":"Soft culture substrates favor stem-like cellular phenotype and facilitate reprogramming of human mesenchymal stem\/stromal cells (hMSCs) through mechanotransduction","volume":"9","author":"H Gerardo","year":"2019","journal-title":"Sci Rep"},{"key":"pone.0241685.ref010","doi-asserted-by":"crossref","first-page":"13113","DOI":"10.1038\/srep13113","article-title":"Human cell dedifferentiation in mesenchymal condensates through controlled autophagy","volume":"5","author":"R Pennock","year":"2015","journal-title":"Sci Rep"},{"key":"pone.0241685.ref011","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1186\/s12861-015-0065-x","article-title":"Human dental pulp stem cells (hDPSCs): isolation, enrichment and comparative differentiation of two sub-populations","volume":"15","author":"A Pisciotta","year":"2015","journal-title":"BMC Dev Biol"},{"key":"pone.0241685.ref012","doi-asserted-by":"crossref","first-page":"547","DOI":"10.3389\/fphys.2018.00547","article-title":"Use of a 3D Floating Sphere Culture System to Maintain the Neural Crest-Related Properties of Human Dental Pulp Stem Cells","volume":"9","author":"A Pisciotta","year":"2018","journal-title":"Front Physiol"},{"issue":"2","key":"pone.0241685.ref013","doi-asserted-by":"crossref","first-page":"264","DOI":"10.1016\/j.cmet.2011.06.011","article-title":"Somatic oxidative bioenergetics transitions into pluripotency-dependent glycolysis to facilitate nuclear reprogramming","volume":"14","author":"CD Folmes","year":"2011","journal-title":"Cell Metab"},{"issue":"2","key":"pone.0241685.ref014","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1159\/000445096","article-title":"Fibroblasts and Mesenchymal Stromal\/Stem Cells Are Phenotypically Indistinguishable","volume":"136","author":"RA Denu","year":"2016","journal-title":"Acta Haematol"},{"issue":"12","key":"pone.0241685.ref015","doi-asserted-by":"crossref","first-page":"9099","DOI":"10.1002\/jcp.26860","article-title":"Fibroblasts and mesenchymal stem cells: Two sides of the same coin?","volume":"233","author":"M Soundararajan","year":"2018","journal-title":"J Cell Physiol"},{"issue":"10","key":"pone.0241685.ref016","first-page":"11","article-title":"Growing Human Dermal Fibroblasts as Spheroids Renders Them Susceptible for Early Expression of Pluripotency Genes","volume":"3","author":"M Lo LMR","year":"2019","journal-title":"Advanced Biosystems"},{"issue":"4","key":"pone.0241685.ref017","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1006\/meth.2001.1262","article-title":"Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method","volume":"25","author":"KJ Livak","year":"2001","journal-title":"Methods"},{"issue":"1","key":"pone.0241685.ref018","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1002\/ar.21487","article-title":"Real-time reverse transcription polymerase chain reaction to measure mRNA: use, limitations, and presentation of results","volume":"295","author":"EC Jensen","year":"2012","journal-title":"Anat Rec (Hoboken)"},{"issue":"1","key":"pone.0241685.ref019","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1016\/j.yexcr.2014.09.027","article-title":"Dedifferentiation of cancer cells following recovery from a potentially lethal damage is mediated by H2S-Nampt","volume":"330","author":"EA Ostrakhovitch","year":"2015","journal-title":"Exp Cell Res"},{"issue":"1","key":"pone.0241685.ref020","first-page":"10","article-title":"A simple colorimetric assay for muramic acid and lactic acid(Applied Biochemistry and Biotechnology\u2014Part A Enzyme Engineering and Biotechnology","volume":"56","author":"KACC Taylor","year":"1996"},{"issue":"39","key":"pone.0241685.ref021","doi-asserted-by":"crossref","first-page":"4767","DOI":"10.1039\/c2cc30730h","article-title":"Selective turn-on fluorescent probes for imaging hydrogen sulfide in living cells","volume":"48","author":"LA Montoya","year":"2012","journal-title":"Chem Commun (Camb)"},{"key":"pone.0241685.ref022","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1007\/978-1-4939-3584-0_8","article-title":"Protocols for in vitro Differentiation of Human Mesenchymal Stem Cells into Osteogenic, Chondrogenic and Adipogenic Lineages","volume":"1416","author":"MC Ciuffreda","year":"2016","journal-title":"Methods Mol Biol"},{"key":"pone.0241685.ref023","doi-asserted-by":"crossref","first-page":"25438","DOI":"10.1038\/srep25438","article-title":"Nanog, Oct4 and Tet1 interplay in establishing pluripotency","volume":"6","author":"V Olariu","year":"2016","journal-title":"Sci Rep"},{"issue":"6047","key":"pone.0241685.ref024","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":"S Ito","year":"2011","journal-title":"Science"},{"issue":"19","key":"pone.0241685.ref025","doi-asserted-by":"crossref","DOI":"10.1242\/dev.174722","article-title":"Loss of DNA methyltransferase activity in primed human ES cells triggers increased cell-cell variability and transcriptional repression","volume":"146","author":"AM Tsankov","year":"2019","journal-title":"Development"},{"issue":"7","key":"pone.0241685.ref026","doi-asserted-by":"crossref","first-page":"630","DOI":"10.1089\/ars.2014.6186","article-title":"Hydrogen Sulfide Maintains Mitochondrial DNA Replication via Demethylation of TFAM","volume":"23","author":"S Li","year":"2015","journal-title":"Antioxid Redox Signal"},{"issue":"30","key":"pone.0241685.ref027","doi-asserted-by":"crossref","first-page":"10573","DOI":"10.1021\/ja505371z","article-title":"Understanding hydrogen sulfide storage: probing conditions for sulfide release from hydrodisulfides","volume":"136","author":"TS Bailey","year":"2014","journal-title":"J Am Chem Soc"},{"issue":"46","key":"pone.0241685.ref028","doi-asserted-by":"crossref","first-page":"17977","DOI":"10.1073\/pnas.0705710104","article-title":"Hydrogen sulfide mediates the vasoactivity of garlic","volume":"104","author":"GA Benavides","year":"2007","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"5309","key":"pone.0241685.ref029","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1126\/science.276.5309.71","article-title":"Marrow stromal cells as stem cells for nonhematopoietic tissues","volume":"276","author":"DJ Prockop","year":"1997","journal-title":"Science"},{"issue":"5411","key":"pone.0241685.ref030","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1126\/science.284.5411.143","article-title":"Multilineage potential of adult human mesenchymal stem cells","volume":"284","author":"MF Pittenger","year":"1999","journal-title":"Science"},{"issue":"1","key":"pone.0241685.ref031","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1006\/dbio.1994.1022","article-title":"Induction of rapid osteoblast differentiation in rat bone marrow stromal cell cultures by dexamethasone and BMP-2","volume":"161","author":"DJ Rickard","year":"1994","journal-title":"Dev Biol"},{"issue":"8","key":"pone.0241685.ref032","first-page":"797","article-title":"Cell-binding peptides conjugated to poly(ethylene glycol) promote neural cell aggregation","volume":"12","author":"W Dai","year":"1994","journal-title":"Biotechnology (N Y)"},{"key":"pone.0241685.ref033","doi-asserted-by":"crossref","first-page":"141","DOI":"10.1007\/978-1-59745-443-8_8","article-title":"Generation of multicellular tumor spheroids by the hanging-drop method","volume":"140","author":"NE Timmins","year":"2007","journal-title":"Methods Mol Med"},{"issue":"12","key":"pone.0241685.ref034","doi-asserted-by":"crossref","first-page":"2164","DOI":"10.1016\/j.biomaterials.2008.12.084","article-title":"Organ printing: tissue spheroids as building blocks","volume":"30","author":"V Mironov","year":"2009","journal-title":"Biomaterials"},{"issue":"5","key":"pone.0241685.ref035","doi-asserted-by":"crossref","first-page":"1038","DOI":"10.1002\/bit.25156","article-title":"Scaffold-free formation of a millimeter-scale multicellular spheroid with an internal cavity from magnetically levitated 3T3 cells that ingested iron oxide-containing microspheres","volume":"111","author":"JH Lee","year":"2014","journal-title":"Biotechnol Bioeng"},{"issue":"5","key":"pone.0241685.ref036","doi-asserted-by":"crossref","first-page":"1601","DOI":"10.1182\/blood-2005-03-0987","article-title":"Forced aggregation of defined numbers of human embryonic stem cells into embryoid bodies fosters robust, reproducible hematopoietic differentiation","volume":"106","author":"ES Ng","year":"2005","journal-title":"Blood"},{"issue":"2","key":"pone.0241685.ref037","doi-asserted-by":"crossref","first-page":"430","DOI":"10.1006\/excr.1993.1265","article-title":"Characterization of morphology and cellular metabolism during the spheroid formation by fibroblasts","volume":"208","author":"T Takezawa","year":"1993","journal-title":"Exp Cell Res"},{"issue":"6","key":"pone.0241685.ref038","doi-asserted-by":"crossref","first-page":"769","DOI":"10.1016\/j.stem.2012.11.008","article-title":"A late transition in somatic cell reprogramming requires regulators distinct from the pluripotency network","volume":"11","author":"A Golipour","year":"2012","journal-title":"Cell Stem Cell"},{"issue":"22","key":"pone.0241685.ref039","doi-asserted-by":"crossref","first-page":"5514","DOI":"10.1016\/j.biomaterials.2012.04.001","article-title":"Promotion of the induction of cell pluripotency through metabolic remodeling by thyroid hormone triiodothyronine-activated PI3K\/AKT signal pathway","volume":"33","author":"M Chen","year":"2012","journal-title":"Biomaterials"},{"issue":"6","key":"pone.0241685.ref040","doi-asserted-by":"crossref","first-page":"e99665","DOI":"10.1371\/journal.pone.0099665","article-title":"mTORC2 phosphorylation of Akt1: a possible mechanism for hydrogen sulfide-induced cardioprotection","volume":"9","author":"Y Zhou","year":"2014","journal-title":"PLoS One"},{"issue":"4","key":"pone.0241685.ref041","doi-asserted-by":"crossref","first-page":"627","DOI":"10.1016\/j.molcel.2012.08.030","article-title":"Reciprocal regulation of Akt and Oct4 promotes the self-renewal and survival of embryonal carcinoma cells","volume":"48","author":"Y Lin","year":"2012","journal-title":"Mol Cell"},{"issue":"51","key":"pone.0241685.ref042","doi-asserted-by":"crossref","first-page":"20853","DOI":"10.1073\/pnas.1219181110","article-title":"Identification of Oct4-activating compounds that enhance reprogramming efficiency","volume":"109","author":"W Li","year":"2012","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"5","key":"pone.0241685.ref043","doi-asserted-by":"crossref","first-page":"1543","DOI":"10.1091\/mbc.e07-01-0029","article-title":"Epigenetic reprogramming of OCT4 and NANOG regulatory regions by embryonal carcinoma cell extract","volume":"18","author":"CT Freberg","year":"2007","journal-title":"Mol Biol Cell"},{"issue":"9","key":"pone.0241685.ref044","doi-asserted-by":"crossref","first-page":"e106973","DOI":"10.1371\/journal.pone.0106973","article-title":"The cultural divide: exponential growth in classical 2D and metabolic equilibrium in 3D environments","volume":"9","author":"K Wrzesinski","year":"2014","journal-title":"PLoS One"},{"issue":"21","key":"pone.0241685.ref045","doi-asserted-by":"crossref","first-page":"5741","DOI":"10.1158\/0008-5472.CAN-16-3480","article-title":"Upregulation of Cystathionine-beta-Synthase in Colonic Epithelia Reprograms Metabolism and Promotes Carcinogenesis","volume":"77","author":"CM Phillips","year":"2017","journal-title":"Cancer Res"},{"key":"pone.0241685.ref046","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1016\/j.niox.2014.12.007","article-title":"Hydrogen sulfide improves glucose metabolism and prevents hypertrophy in cardiomyocytes","volume":"46","author":"M Liang","year":"2015","journal-title":"Nitric Oxide"},{"issue":"16","key":"pone.0241685.ref047","doi-asserted-by":"crossref","first-page":"1463","DOI":"10.1089\/ars.2017.7046","article-title":"Hydrogen Sulfide and Glucose Homeostasis: A Tale of Sweet and the Stink","volume":"28","author":"A Untereiner","year":"2018","journal-title":"Antioxid Redox Signal"},{"issue":"12","key":"pone.0241685.ref048","volume":"8","author":"NE Ryu","year":"2019","journal-title":"Cells"},{"issue":"6","key":"pone.0241685.ref049","doi-asserted-by":"crossref","first-page":"737","DOI":"10.1097\/SHK.0000000000001420","article-title":"Stem Cell Therapy and Hydrogen Sulfide: Conventional or Nonconventional Mechanisms of Action?","volume":"53","author":"AR Jensen","year":"2020","journal-title":"Shock"},{"issue":"1","key":"pone.0241685.ref050","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1016\/j.stem.2014.03.005","article-title":"Hydrogen sulfide maintains mesenchymal stem cell function and bone homeostasis via regulation of Ca(2+) channel sulfhydration","volume":"15","author":"Y Liu","year":"2014","journal-title":"Cell Stem Cell"},{"issue":"13","key":"pone.0241685.ref051","doi-asserted-by":"crossref","first-page":"1445","DOI":"10.1177\/0022034516659041","article-title":"Hydrogen Sulfide Regulates Homeostasis of Mesenchymal Stem Cells and Regulatory T Cells","volume":"95","author":"R Yang","year":"2016","journal-title":"J Dent Res"},{"issue":"2","key":"pone.0241685.ref052","doi-asserted-by":"crossref","first-page":"176","DOI":"10.1080\/1061186X.2019.1624969","article-title":"Hydrogen sulphide promotes osteoclastogenesis by inhibiting autophagy through the PI3K\/AKT\/mTOR pathway","volume":"28","author":"J Ma","journal-title":"J Drug Target. 2020"},{"issue":"11","key":"pone.0241685.ref053","doi-asserted-by":"crossref","first-page":"742","DOI":"10.1089\/ars.2016.6826","article-title":"Cystathionine gamma-Lyase-Hydrogen Sulfide Induces Runt-Related Transcription Factor 2 Sulfhydration, Thereby Increasing Osteoblast Activity to Promote Bone Fracture Healing","volume":"27","author":"Y Zheng","year":"2017","journal-title":"Antioxid Redox Signal"},{"issue":"5","key":"pone.0241685.ref054","doi-asserted-by":"crossref","first-page":"755","DOI":"10.1016\/S0092-8674(00)80258-5","article-title":"Targeted disruption of Cbfa1 results in a complete lack of bone formation owing to maturational arrest of osteoblasts","volume":"89","author":"T Komori","year":"1997","journal-title":"Cell"}],"container-title":["PLOS ONE"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0241685","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2020,11,12]],"date-time":"2020-11-12T18:41:36Z","timestamp":1605206496000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0241685"}},"subtitle":[],"editor":[{"given":"Gianpaolo","family":"Papaccio","sequence":"first","affiliation":[],"role":[{"role":"editor","vocabulary":"crossref"}]}],"short-title":[],"issued":{"date-parts":[[2020,11,12]]},"references-count":54,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2020,11,12]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0241685","relation":{},"ISSN":["1932-6203"],"issn-type":[{"type":"electronic","value":"1932-6203"}],"subject":[],"published":{"date-parts":[[2020,11,12]]}}}