Article

Synthetic surfaces for human embryonic stem cell culture

Biomolecular Science and Engineering, University of California, Santa Barbara, CA 93106, USA.
Journal of Biotechnology (Impact Factor: 2.87). 04/2010; 146(3):143-6. DOI: 10.1016/j.jbiotec.2010.01.016
Source: PubMed

ABSTRACT

Human embryonic stem cells (hESCs) have numerous potential biomedical applications owing to their unique abilities for self-renewal and pluripotency. Successful clinical application of hESCs and derivatives necessitates the culture of these cells in a fully defined environment. We have developed a novel peptide-based surface that uses a high-affinity cyclic RGD peptide for culture of hESCs under chemically defined conditions.

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Available from: Sherry Hikita, Aug 20, 2014
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    • "There was a 3-fold increase in hPSC numbers compared with feeder-based culture (Saha et al., 2011). Several groups have also designed new surface substrates based on a high-affinity cyclic arginine-glycine-aspartate (RGD) peptide that contains the RGD integrin recognition sequence (Kolhar et al., 2010), synthetic peptide-acrylate surfaces (Melkoumian et al., 2010), and synthetic polymer coating with poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl)ammonium hydroxide] (PMEDSAH) (Villa-Diaz et al., 2010) for long-term maintenance of hPSCs. Thus, advanced material technology could offer fully synthetic environmental cues that sustain long-term culture of clinicalgrade hPSCs. "
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    ABSTRACT: Human pluripotent stem cells (hPSCs) provide powerful resources for application in regenerative medicine and pharmaceutical development. In the past decade, various methods have been developed for large-scale hPSC culture that rely on combined use of multiple growth components, including media containing various growth factors, extracellular matrices, 3D environmental cues, and modes of multicellular association. In this Protocol Review, we dissect these growth components by comparing cell culture methods and identifying the benefits and pitfalls associated with each one. We further provide criteria, considerations, and suggestions to achieve optimal cell growth for hPSC expansion, differentiation, and use in future therapeutic applications.
    Full-text · Article · Jan 2014 · Cell stem cell
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    • "In feederrindependent ESC cultures, the substrate is composed of natural or recombinant ECM proteins, as well as artificial peptides with ECM domains (Klim et al., 2010; Kolhar et al., 2010; Melkoumian et al., 2010; Meng et al., 2010b). Recently, a new approach was proposed. "
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    ABSTRACT: We developed a feeder-free system for human embryonic stem cells (ESCs) based on extracellular matrix protein (ECM) as the substrate. ECM was synthesized by mesenchymal stem cells (SC5-MSC) derived from an original ESC line, SC5. The ECM proteins fibronectin and laminin facilitate ESC growth in the feeder-free system. An important component of this system is a conditioned medium from SC5-MSC cells. Two ESC sublines were obtained: SC5-FF cells were cultured in an autogenic, and SC7-FF in an allogenic, feeder-free system. SC5-FF and SC7-FF underwent more than 300 and 115 population doublings, respectively, and retain a normal diploid karyotype. Histochemical and immunofluorescence assays showed that both sublines express undifferentiated ESC markers—alkaline phosphatase, Oct-4, SSEA-4, and TRA-1-81—as well as multidrug resistance transporter ABCG2. PCR assay revealed that undifferentiated SC5-FF cells, like the original SC5 line, maintained on feeder cells express OCT4 and NANOG genes common for somatic cells and DPPA3/STELLA and DAZL genes common for germ line cells. Expression of these genes was gradually diminished during differentiation of embryoid bodies, whereas expression of genes specific for early differentiated cells increased: GATA4, AFP (extraembryonic and embryonic endoderm), PAX6 (neuroectoderm), and BRY (mesoderm). ESC properties (karyotype structure, average time of population doubling, undifferentiated cell number in population) of the SC5 and SC7 and SC5-FF and SC7-FF sublines derived from original ESCs were not altered. It shows that the feeder-free systems, which are more stable than any feeder systems, maintain key ESC properties and may be recommended for fundamental, biomedical, and pharmacological studies performed with human ESCs.
    Full-text · Article · Jan 2013 · Cell and Tissue Biology
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    • "Ïîýòîìó ïîäáîð îïòèìàëüíûõ ñóáñòðàòîâ äëÿ áåñôèäåðíûõ ñèñòåì êóëüòèâèðîâàíèÿ îñíîâàí íà âçàèìîäåéñòâèè îïðåäåëåííûõ äîìåíîâ ñ ïîâåðõíîñòíûìè ðåöåïòîðàìè — èíòåãðèíàìè, ýêñïðåññèðóåìûìè ñàìèìè ÝÑÊ ïðè êóëüòèâèðîâàíèè èõ â ñðåäàõ ñ äîáàâêàìè ðîñòîâûõ ôàêòîðîâ èëè â õèìè÷åñêè îïðåäåëåííûõ ñðåäàõ (Miyazaki et al., 2008; Evseenko et al., 2009; Vuoristo et al., 2009; Hongisto et al., 2012). Ïîìèìî èñïîëüçîâàíèÿ â êà÷åñòâå áåñôèäåðíûõ ñóáñòðàòîâ ñìåñè èëè îòäåëüíûõ ïðèðîäíûõ èëè ðåêîìáèíàíòíûõ áåëêîâ ÂÊÌ â ðÿäå ðàáîò ñèíòåçèðîâàëè èñêóññòâåííûå ïåïòèäû, ñîäåðaeàùèå äîìåíû ÂÊÌ, íà êîòîðûõ óñïåøíî êóëüòèâèðîâàëè ÝÑÊ (Klim et al., 2010; Kolhar et al., 2010; Melkoumian et al., 2010; Meng et al., 2010b).  ïîñëåäíåå âðåìÿ ïîÿâèëñÿ íîâûé ïîäõîä ê ïîëó÷åíèþ áåñôèäåðíûõ ñóáñòðàòîâ èç ñèíòåòè÷åñêèõ ïîëèìåðîâ . "
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    ABSTRACT: A new feeder-free culture system for human embryonic stem cells (hESC) was developed. It consist of extracellular matrix proteins synthesized by feeder cells--mesenchymal stem cell line SC5-MSC, which was derived from initial hESC line SC5. The major ECM proteins--fibronectin and laminin--that maintain hESC growth in feeder-free system were identified. An essential component of this system is a SC5-MSC-conditioned medium. Two hESC sublines were derived. The subline SC5-FF was cultured in autogenic and subline SC7-FF in allogenic system. Sublines SC5-FF and SC7-FF passed through more than 300 and 115 cell population doublings, retained normal diploid karyotype and an ability of in vitro differentiation into derivates of three germ layers. These sublines express markers of undifferentiated hESC: alkaline phosphatase, Oct-4, SSEA-4, TRA-1-81 and multidrug resistance transporter--ABCG2. The RT-PCR analysis revealed that undifferentiated cells SC5-FF subline, like cells of initial feeder-maintained hESC line SC5, expressed genes OCT4 and NANOG, and germ line specific genes such as DPPA3/STELLA and DAZL. An expression of OCT4, NANOG, DPPA3/STELLA ans DAZL was down-regulated during embryonic bodies differentiation, whereas expression of somatic lineages specific genes like GATA4 and AFP (extra embryonic and embryonic endoderm), PAX6 (neuroectoderm) and BRY (mesoderm) was up-regulated. The comparative analysis of some typical features (karyotype structure, the average population doubling time and the number of undifferentiated cells in populations) did not reveal essential differences between initial SC5 and SC7 lines and their sublines SC5-FF and SC7-FF. This shows that feeder-free culture systems, which are much more stable than any feeder systems, do not break main hESC features during long cultivation and can be recommended for fundamental, biomedicine and pharmacological investigations, using hESCs.
    Full-text · Article · Oct 2012 · Tsitologiia
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