title: Dynamic Assessment of the Endothelialization of Tissue-Engineered Blood Vessels Using an Optical Coherence Tomography Catheter-Based Fluorescence Imaging System.
abstract: BACKGROUND
  Lumen endothelialization of bioengineered vascular scaffolds is essential to maintain small-diameter graft patency and prevent thrombosis postimplantation. Unfortunately, nondestructive imaging methods to visualize this dynamic process are lacking, thus slowing development and clinical translation of these potential tissue-engineering approaches. To meet this need, a fluorescence imaging system utilizing a commercial optical coherence tomography (OCT) catheter was designed to visualize graft endothelialization.
  
  
  METHODS
  C7 DragonFly™ intravascular OCT catheter was used as a channel for delivery and collection of excitation and emission spectra. Poly-dl-lactide (PDLLA) electrospun scaffolds were seeded with endothelial cells (ECs). Seeded cells were exposed to Calcein AM before imaging, causing the living cells to emit green fluorescence in response to blue laser. By positioning the catheter tip precisely over a specimen using high-fidelity electromechanical components, small regions of the specimen were excited selectively. The resulting fluorescence intensities were mapped on a two-dimensional digital grid to generate spatial distribution of fluorophores at single-cell-level resolution. Fluorescence imaging of endothelialization on glass and PDLLA scaffolds was performed using the OCT catheter-based imaging system as well as with a commercial fluorescence microscope. Cell coverage area was calculated for both image sets for quantitative comparison of imaging techniques. Tubular PDLLA scaffolds were maintained in a bioreactor on seeding with ECs, and endothelialization was monitored over 5 days using the OCT catheter-based imaging system.
  
  
  RESULTS
  No significant difference was observed in images obtained using our imaging system to those acquired with the fluorescence microscope. Cell area coverage calculated using the images yielded similar values. Nondestructive imaging of endothelialization on tubular scaffolds showed cell proliferation with cell coverage area increasing from 15 ± 4% to 89 ± 6% over 5 days.
  
  
  CONCLUSIONS
  In this study, we showed the capability of an OCT catheter-based imaging system to obtain single-cell resolution and to quantify endothelialization in tubular electrospun scaffolds. We also compared the resulting images with traditional microscopy, showing high fidelity in image capability. This imaging system, used in conjunction with OCT, could potentially be a powerful tool for in vitro optimization of scaffold cellularization, ensuring long-term graft patency postimplantation.
authors.affiliation: 1 School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University , Blacksburg, Virginia.
authors.forename: Abhijit Achyut
authors.initials: AA
authors.lastname: Gurjarpadhye
authors.affiliation: 1 School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University , Blacksburg, Virginia.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: DeWitt
authors.affiliation: 2 Department of Electrical and Computer Engineering, Virginia Polytechnic Institute and State University , Blacksburg, Virginia.
authors.forename: Yong
authors.initials: Y
authors.lastname: Xu
authors.affiliation: 3 Biomedical Imaging Cluster, Rensselaer Polytechnic Institute , Troy, New York.
authors.forename: Ge
authors.initials: G
authors.lastname: Wang
authors.affiliation: 4 Department of Mechanical Engineering, The University of Texas at Austin , Austin, Texas.
authors.forename: Marissa Nichole
authors.initials: MN
authors.lastname: Rylander
authors.affiliation: 4 Department of Mechanical Engineering, The University of Texas at Austin , Austin, Texas.
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Rylander
mesh_terms: D001808:Blood Vessels; D057785:Catheters; D002461:Cell Line, Transformed; D005453:Fluorescence; D006801:Humans; D023822:Tissue Engineering; D054457:Tissue Scaffolds; D041623:Tomography, Optical Coherence / Q000295:instrumentation*