A comparison of three methods of decellularization of pig corneas to reduce immunogenicity.

Lee, Whayoung and Miyagawa, Yuko and Long, Cassandra and Cooper, David KC and Hara, Hidetaka (2014) A comparison of three methods of decellularization of pig corneas to reduce immunogenicity. Int J Ophthalmol, 7 (4). 587 - 593. ISSN 2222-3959

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Abstract

AIM: To investigate whether decellularization using different techniques can reduce immunogenicity of the cornea, and to explore the decellularized cornea as a scaffold for cultured corneal endothelial cells (CECs). Transplantation of decellularized porcine corneas increases graft transparency and survival for longer periods compared with fresh grafts. METHODS: Six-month-old wild-type pig corneas were cut into 100-200 µm thickness, and then decellularized by three different methods: 1) 0.1% sodium dodecyl sulfate (SDS); 2) hypoxic nitrogen (N2); and 3) hypertonic NaCl. Thickness and transparency were assessed visually. Fresh and decellularized corneas were stained with hematoxylin/eosin (H&E), and for the presence of galactose-α1,3-galactose (Gal) and N-glycolylneuraminic acid (NeuGc, a nonGal antigen). Also, a human IgM/IgG binding assay was performed. Cultured porcine CECs were seeded on the surface of the decellularized cornea and examined after H&E staining. RESULTS: All three methods of decellularization reduced the number of keratocytes in the stromal tissue by >80% while the collagen structure remained preserved. No remaining nuclei stained positive for Gal or NeuGc, and expression of these oligosaccharides on collagen was also greatly decreased compared to expression on fresh corneas. Human IgM/IgG binding to decellularized corneal tissue was considerably reduced compared to fresh corneal tissue. The cultured CECs formed a confluent monolayer on the surface of decellularized tissue. CONCLUSION: Though incomplete, the significant reduction in the cellular component of the decellularized cornea should be associated with a significantly reduced in vivo immune response compared to fresh corneas.

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Details

Item Type:	Article
Status:	Published
Creators/Authors:	Creators Email Pitt Username ORCID Lee, Whayoung Miyagawa, Yuko Long, Cassandra cal46@pitt.edu CAL46 Cooper, David KC cooperdk@pitt.edu COOPERDK Hara, Hidetaka hih3@pitt.edu HIH3
Centers:	Other Centers, Institutes, Offices, or Units > Thomas E. Starzl Transplantation Institute
Date:	30 May 2014
Date Type:	Acceptance
Access Restriction:	No restriction; Release the ETD for access worldwide immediately.
Journal or Publication Title:	Int J Ophthalmol
Volume:	7
Number:	4
Page Range:	587 - 593
DOI or Unique Handle:	10.3980/j.issn.2222-3959.2014.04.01
Institution:	University of Pittsburgh
Schools and Programs:	School of Medicine > Surgery
Refereed:	Yes
Uncontrolled Keywords:	cornea, decellularization, immune response, pig, xenotransplantation
ISSN:	2222-3959
Funders:	NHLBI NIH HHS (P01 HL107152)
Date Deposited:	01 Jun 2015 21:11
Last Modified:	13 Oct 2017 18:55
URI:	http://d-scholarship.pitt.edu/id/eprint/24787

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