June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Establishment of a bi-layer tissue engineered conjunctiva construct using a three-dimensional (3D) printed scaffold
Author Affiliations & Notes
  • Jiajun Xie
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
    Eye Center, Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China
  • QI GAO
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
    Eye Center, Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China
  • Alfonso L. Sabater
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
    Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, Florida, United States
  • Enrique Salero
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
    Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, Florida, United States
  • Gabriel Gaidosh
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
  • Elena De Juan-Pardo
    Institute for Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland, Australia
  • Dietmar W Hutmacher
    Institute for Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland, Australia
  • Juan Ye
    Eye Center, Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China
  • Victor L Perez
    Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, Florida, United States
    Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, Florida, United States
  • Footnotes
    Commercial Relationships   Jiajun Xie, None; QI GAO, None; Alfonso Sabater, None; Enrique Salero, None; Gabriel Gaidosh, None; Elena De Juan-Pardo, None; Dietmar Hutmacher, None; Juan Ye, None; Victor Perez, Allergan (C), Baush & Lomb (C), Eleven Biotherapeutics (C), EyeGate Pharma (C), Genentech (C), Parion Sciences (C), Rigel (C)
  • Footnotes
    Support  NIH/NEI NIH:  R01EY024484; Center Core Grant P30EY01480; RPB Unrestricted Award and Career Development Awards; Department of Defense Grant# W81XWH-13-1-0048; Walter G. Ross Chair in Ophthalmic Research
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 4356. doi:
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      Jiajun Xie, QI GAO, Alfonso L. Sabater, Enrique Salero, Gabriel Gaidosh, Elena De Juan-Pardo, Dietmar W Hutmacher, Juan Ye, Victor L Perez; Establishment of a bi-layer tissue engineered conjunctiva construct using a three-dimensional (3D) printed scaffold. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4356.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Conjunctiva is an essential component of the ocular surface. To better address the conjunctiva diseases and other ocular surface diseases which may result in local lesions of conjunctiva, it is ideal to create a 3D model instead of conventional 2D model for an in vitro study. In this study, we established a bi-layer tissue engineered conjunctiva construct using a 3D printed poly(ε-caprolactone) (PCL) scaffold to mimic the biological structure and microenvironment of conjunctiva tissue for in vitro study.

Methods : Human conjunctival tissue was digested with dispase to isolate the conjunctival epithelial cells (CjECs), followed by collagenase A digestion to obtain conjunctival stromal cells (CjSCs). A 2-step co-culture method was employed to fabricate a bi-layer tissue engineered conjunctiva construct. The CjSCs and CjECs were characterized by qPCR and immunofluorescence staining. The construct was assessed under a confocal microscope and a scanning electronic microscope (SEM).

Results : A bi-layer tissue engineered conjunctiva construct was successfully established using a 3D printed scaffold. As revealed in Figure A, proliferating CjSCs and the extracellular matrix they produced gradually filled the 3D printed scaffold to form a stromal layer during step 1, while CjECs formed a confluent epithelial layer on a suitable surface provided by the pre-seeded scaffold during the second co-culture step. The intact epithelial layer was also illustrated by CK13 expression and cell junctions indicated in Figure B.

Conclusions : Compared with other conjunctiva substitute, this 3D printed scaffold presented as a platform which allowed cells to behave following their intrinsic biological properties and better mimicked the local microenvironment and cell interaction in vivo. The tissue engineered conjunctiva construct with a stromal layer and an intact epithelial layer was well established, and could be an option for conjunctival reconstruction as well as an in vitro model for the study of conjunctival diseases.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

 

(A) Confocal microscopic images of bi-layer conjunctiva construct. Red: CjSCs stained with Vimentin. Green: CjECs stained with CK13. Blue: nuclei stained with DAPI. (B) SEM images of bi-layer conjunctiva construct. Top: epithelial layer. Bottom: stromal layer. Black arrow: scaffold fibers. White arrow: cell junction.

(A) Confocal microscopic images of bi-layer conjunctiva construct. Red: CjSCs stained with Vimentin. Green: CjECs stained with CK13. Blue: nuclei stained with DAPI. (B) SEM images of bi-layer conjunctiva construct. Top: epithelial layer. Bottom: stromal layer. Black arrow: scaffold fibers. White arrow: cell junction.

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