Mass Spectrometry Reveals α-2-HS-Glycoprotein as a Key Early Extracellular Matrix Protein for Conjunctival Cells



Makuloluwa, Aruni K, Stewart, Rosalind MK, Kaye, Stephen B ORCID: 0000-0003-0390-0592, Williams, Rachel L ORCID: 0000-0002-1954-0256 and Hamill, Kevin J ORCID: 0000-0002-7852-1944
(2020) Mass Spectrometry Reveals α-2-HS-Glycoprotein as a Key Early Extracellular Matrix Protein for Conjunctival Cells. INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE, 61 (3). 44-.

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Abstract

<h4>Purpose</h4>To determine the composition of extracellular matrix (ECM) proteins secreted by a conjunctival epithelial cell line and to identify components that aid conjunctival epithelial cell culture.<h4>Methods</h4>Human conjunctival epithelial cell line (HCjE-Gi) cells were cultured in serum-free media and their ECM isolated using ammonium hydroxide. Growth characteristics were evaluated for fresh HCjE-Gi cells plated onto ECMs obtained from 3- to 28-day cell cultures. Mass spectrometry was used to characterize the ECM composition over 42 culture days. Cell adhesion and growth on pre-adsorbed fibronectin and α-2-HS-glycoprotein (α-2-HS-GP) were investigated.<h4>Results</h4>Day 3 ECM provided the best substrate for cell growth compared to ECM obtained from 5- to 28-day cell cultures. Mass spectrometry identified a predominantly laminin 332 matrix throughout the time course, with progressive changes to matrix composition over time: proportional decreases in matrix-bound growth factors and increases in proteases. Fibronectin and α-2-HS-GP were 5- and 200-fold enriched as a proportion of the early ECM relative to the late ECM, respectively. Experiments on these proteins in isolation demonstrated that fibronectin supported rapid cell adhesion, whereas fibronectin and α-2-HS-GP both supported enhanced cell growth compared to tissue culture polystyrene.<h4>Conclusions</h4>These data reveal α-2-HS-GP as a candidate protein to enhance the growth of conjunctival epithelial cells and raise the possibility of exploiting these findings for targeted improvement to synthetic tissue engineered conjunctival substrates.

Item Type: Article
Uncontrolled Keywords: ocular surface, conjunctiva, extracellular matrix, tissue engineering, mass spectrometry
Depositing User: Symplectic Admin
Date Deposited: 17 Feb 2020 10:23
Last Modified: 15 Mar 2024 04:23
DOI: 10.1167/iovs.61.3.44
Related URLs:
URI: https://livrepository.liverpool.ac.uk/id/eprint/3075032