{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,30]],"date-time":"2026-04-30T05:58:51Z","timestamp":1777528731069,"version":"3.51.4"},"reference-count":55,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2015,10,18]],"date-time":"2015-10-18T00:00:00Z","timestamp":1445126400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"name":"NSF-CBET CAREER","award":["NSF-CBET-1150854"],"award-info":[{"award-number":["NSF-CBET-1150854"]}]},{"name":"AU-CMB\/NSF EPSCoR","award":["NSF-EPS-1158862"],"award-info":[{"award-number":["NSF-EPS-1158862"]}]},{"name":"U.S. Department of Education GAANN Fellowship","award":["P00A120228"],"award-info":[{"award-number":["P00A120228"]}]},{"name":"Alabama EPSCoR Graduate Research Scholarship Program"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2016,4]]},"abstract":"<jats:title>ABSTRACT<\/jats:title><jats:sec><jats:label\/><jats:p>The development of cell\u2010based treatments for heart disease relies on the creation of functionally mature stem cell\u2010derived cardiomyocytes employing in vitro culture suspension systems, a process which remains a formidable and expensive endeavor. The use of nitric oxide as a signaling molecule during differentiation has demonstrated the potential for creating increased numbers of spontaneously contracting embryoid bodies in culture; however, the effects of nitric oxide signaling on the function and maturation of stem cell\u2010derived cardiomyocytes is not well understood. In this study, the effects of nitric oxide on mouse embryonic stem cell\u2010derived cardiomyocyte contractile activity, protein, and gene expression, and calcium handling were quantified. Embryoid bodies (EBs) formed using the hanging drop method, were treated with the soluble nitric oxide donor <jats:italic>S<\/jats:italic>\u2010nitrosocysteine (CysNO) over a period of 18 days in suspension culture and spontaneous contractile activity was assessed. On day 8, selected EBs were dissociated to form monolayers for electrophysiological characterization using calcium transient mapping. Nitric oxide treatment led to increased numbers of stem cell\u2010derived cardiomyocytes (SC\u2010CMs) relative to non\u2010treated EBs after 8 days in suspension culture. Increased incidence of spontaneous contraction and frequency of contraction were observed from days 8\u201314 in EBs receiving nitric oxide treatment in comparison to control. Expression of cardiac markers and functional proteins was visualized using immunocytochemistry and gene expression was assessed using qPCR. Cardiac\u2010specific proteins were present in both CysNO\u2010treated and control SC\u2010CMs; however, CysNO treatment during differentiation significantly increased \u03b2MHC gene expression in SC\u2010CMs relative to control SC\u2010CMs. Furthermore, increased calcium transient velocity and decreased calcium transient duration was observed for CysNO\u2010treated SC\u2010CMs in comparison to control SC\u2010CMs. Soluble nitric oxide donors, including <jats:italic>S<\/jats:italic>\u2010nitrosocysteine, have advantages over other bioactive molecules for use in scalable culture systems in driving cardiac differentiation, since they are inexpensive and the diffusivity of nitric oxide is relatively high. By enabling maintenance of spontaneous contraction in suspension culture and progressing electrophysiological function of resulting SC\u2010CMs toward a more mature phenotype, long\u2010term application of <jats:italic>S<\/jats:italic>\u2010nitrosocysteine was shown to be beneficial during cardiac differentiation. Taken together, these results demonstrate the efficiency of nitric oxide as a signaling compound, with implications in the improvement of pluripotent stem cell\u2010derived cardiomyocyte maturation in large\u2010scale culture systems. Biotechnol. Bioeng. 2016;113: 882\u2013894. \u00a9 2015 Wiley Periodicals, Inc.<\/jats:p><\/jats:sec>","DOI":"10.1002\/bit.25849","type":"journal-article","created":{"date-parts":[[2015,10,7]],"date-time":"2015-10-07T06:29:58Z","timestamp":1444199398000},"page":"882-894","source":"Crossref","is-referenced-by-count":14,"title":["Enhanced stem cell\u2010derived cardiomyocyte differentiation in suspension culture by delivery of nitric oxide using <i>S<\/i>\u2010nitrosocysteine"],"prefix":"10.1002","volume":"113","author":[{"given":"Alexander J.","family":"Hodge","sequence":"first","affiliation":[{"name":"Department of Chemical Engineering Auburn University 212 Ross Hall Auburn 36849 Alabama"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Juming","family":"Zhong","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine Auburn University Auburn Alabama"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elizabeth 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