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Three-dimensional characterization of collagen remodeling in cell-seeded collagen scaffolds via polarization second harmonic generation

Authors
  • Xydias, Dionysios
  • Ziakas, Georgios
  • Psilodimitrakopoulos, Sotiris
  • Lemonis, Andreas
  • Bagli, Eleni
  • Fotsis, Theodore
  • Gravanis, Achille
  • Tzeranis, Dimitrios S.
  • Stratakis, Emmanuel
Type
Published Article
Journal
Biomedical Optics Express
Publisher
The Optical Society
Publication Date
Jan 28, 2021
Volume
12
Issue
2
Pages
1136–1153
Identifiers
DOI: 10.1364/BOE.411501
PMID: 33680563
PMCID: PMC7901316
Source
PubMed Central
License
Unknown

Abstract

In this study, we use non-linear imaging microscopy to characterize the structural properties of porous collagen-GAG scaffolds (CGS) seeded with human umbilical vein endothelial cells (HUVECs), as well as human mesenchymal stem cells (hMSCs), a co-culture previously reported to form vessel-like structures inside CGS. The evolution of the resulting tissue construct was monitored over 10 days via simultaneous two- and three-photon excited fluorescence microscopy. Time-lapsed 2- and 3-photon excited fluorescence imaging was utilized to monitor the temporal evolution of the vascular-like structures up to 100 µm inside the scaffold up to 10 days post-seeding. 3D polarization-dependent second harmonic generation (PSHG) was utilized to monitor collagen-based scaffold remodeling and determine collagen fibril orientation up to 200 µm inside the scaffold. We demonstrate that polarization-dependent second harmonic generation can provide a novel way to quantify the reorganization of the collagen architecture in CGS simultaneously with key biomechanical interactions between seeded cells and CGS that regulate the formation of vessel-like structures inside 3D tissue constructs. A comparison between samples at different days in vitro revealed that gradually, the scaffolds developed an orthogonal net-like architecture, previously found in real skin.

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