Fluorescent imaging of endothelial cells in bioengineered blood vessels: the impact of crosslinking of the scaffold

Authors

  • Guoguang Niu,

    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
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  • Etai Sapoznik,

    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
    2. Virginia Tech-Wake Forest University School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA, USA
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  • Peng Lu,

    1. Bradley Department of Electrical and Computer Engineering, Wake Forest University School of Biomedical Engineering and Sciences, Blacksburg, VA, USA
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  • Tracy Criswell,

    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
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  • Aaron M. Mohs,

    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
    2. Virginia Tech-Wake Forest University School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA, USA
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  • Ge Wang,

    1. Virginia Tech-Wake Forest University School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA, USA
    Current affiliation:
    1. Department of Biomedical Engineering, Room 3209, CBIS/BME, Rensselaer Polytechnic Institute, Troy, NY, USA
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  • Sang-Jin Lee,

    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
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  • Yong Xu,

    1. Bradley Department of Electrical and Computer Engineering, Wake Forest University School of Biomedical Engineering and Sciences, Blacksburg, VA, USA
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  • Shay Soker

    Corresponding author
    1. Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Medical Center, Medical Center Boulevard, Winston-Salem, NC
    2. Virginia Tech-Wake Forest University School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA, USA
    • Correspondence to: Shay Soker, Wake Forest Institute for Regenerative Medicine, Wake Forest Baptist Health Medical Center Boulevard, Winston-Salem, NC 27157, USA. E-mail: ssoker@wfubmc.edu

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Abstract

Fluorescent imaging is a useful tool to monitor and evaluate bioengineered tissues and organs. However, autofluorescence emitted from the scaffold can be comparable or even overwhelm signals generated by fluorescently labelled cells and biomarkers. Using standard fluorescent microscopy techniques, a simple and easy-to-measure signal to noise ratio metric was developed, which can facilitate the selection of fluorescent biomarkers and the respective biomaterials for tissue engineering studies. Endothelial cells (MS1) expressing green-fluorescent protein and red fluorescent protein (mKate) were seeded on poly(epsilon-caprolactone)–collagen hybrid scaffolds that were prepared by crosslinking with glutaraldehyde, genipin and ethyl(dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide. All scaffolds had comparable mechanical properties, which could meet the requirements for vascular graft applications. ethyl(dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide crosslinked scaffolds had a high signal to noise ratio value because of its low autofluorescence in green and red channels. Genipin crosslinked scaffolds had a high signal to noise ratio only in the green channel, while glutaraldehyde crosslinked scaffolds had a low signal to noise ratio in both green and red channels. The signal to noise ratio was independent of the exposure time. The data show that although similar in their mechanical properties and ability to support cell growth, scaffolds crosslinked with different agents have significant differences in causing autofluorescence of the scaffolds. This result indicates that scaffold's preparation method may have a significant impact on direct imaging of fluorescently labelled cells on scaffolds used for tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

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