Generation of human embryonic stem cell line expressing green fluorescent protein

Shetty, Ronak; Shetty, Deeti K.; Inamdar, Maneesha S.

doi:10.1016/j.scr.2015.12.022

Cited by 1 publication

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“…Further, they are also valuable tools to control stem cell fate and study cell behaviour during differentiation. Hence, various transgenic hESC lines with constitutive or inducible tissue, cell-specific, or gene-specific fluorescent reporters have been generated by various groups [12–16]. Recently, lentiviral-based transduction and targeted gene knock-in methods, like zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regulatory interspaced short palindromic repeats system (CRISPR/CAS9), are increasingly used as effective tools for gene delivery to hESCs.…”

Section: Introductionmentioning

confidence: 99%

In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell‐Derived Osteoprogenitors

Islam

Sriram

et al. 2016

Stem Cells International

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Cellular therapy using stem cells in bone regeneration has gained increasing interest. Various studies suggest the clinical utility of osteoprogenitors-like mesenchymal stem cells in bone regeneration. However, limited availability of mesenchymal stem cells and conflicting evidence on their therapeutic efficacy limit their clinical application. Human embryonic stem cells (hESCs) are potentially an unlimited source of healthy and functional osteoprogenitors (OPs) that could be utilized for bone regenerative applications. However, limited ability to track hESC-derived progenies in vivo greatly hinders translational studies. Hence, in this study, we aimed to establish hESC-derived OPs (hESC-OPs) expressing green fluorescent protein (GFP) and to investigate their osteogenic differentiation potential in vitro. We fluorescently labelled H9-hESCs using a plasmid vector encoding GFP. The GFP-expressing hESCs were differentiated into hESC-OPs. The hESC-OPsGFP+ stably expressed high levels of GFP, CD73, CD90, and CD105. They possessed osteogenic differentiation potential in vitro as demonstrated by increased expression of COL1A1, RUNX2, OSTERIX, and OPG transcripts and mineralized nodules positive for Alizarin Red and immunocytochemical expression of osteocalcin, alkaline phosphatase, and collagen-I. In conclusion, we have demonstrated that fluorescently labelled hESC-OPs can maintain their GFP expression for the long term and their potential for osteogenic differentiation in vitro. In future, these fluorescently labelled hESC-OPs could be used for noninvasive assessment of bone regeneration, safety, and therapeutic efficacy.

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Section: Introductionmentioning

confidence: 99%

In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell‐Derived Osteoprogenitors

Islam

Sriram

et al. 2016

Stem Cells International

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Generation of human embryonic stem cell line expressing green fluorescent protein

Abstract: Human embryonic stem cell line BJNhem20 pCAG-EGFP was generated using non-viral method (Fig. 1). The construct pCAG-EGFP was transfected using microporation procedure.

Cited by 1 publication

References 3 publications

In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell‐Derived Osteoprogenitors

In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell‐Derived Osteoprogenitors

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