Abstract
Purpose :
Although many integrins have been previously identified in native corneal endothelium, integrin expression between native and cultured cells remains incomplete. The goal of this study is to compare the integrin expression profile of native corneal endothelium and of cultured corneal endothelial cells (CECs), using both primary cultures and an immortalized cell line.
Methods :
Human CECs were isolated from healthy eye bank corneas (n=1) and cultured on plastic until first passage. Immortalized CECs (HCEnC21T; P26) and primary cultures of CECs were seeded on glass coverslips at an initial seeding density of 10,000 cells per cm2 and cultured for 7 days before fixation (90% acetone). Fixed cells, and fixed native cornea (n=1), were immunostained against integrins subunits (α1, α2, α4, α5, α6, α10, α11, αv, β1, β4 and β5), and photographed (Zeiss LSM-800 confocal microscope).
Results :
Collagen-binding integrin subunit α1 was expressed in CEnC21T cells, whereas α1 and α2 were present in primary cultures of CECs and native corneal endothelium. Subunits α10 and α11 were absent from CEnC21T, native CECs and native endothelium. Laminin-binding integrin subunits α6 and β4 were absent from cultured CECs, in both the cell line and the primary cultures, but were expressed in native endothelium. For the RGD-binding integrin subunits, α4, α5, αv and β5 were absent in cultured CECs (in both the cell line and the primary cultures), whereas native corneal endothelium expressed the four of them. Integrin subunit β1, which can pair with a variety of different α subchains to form 12 different known integrins, was expressed in cultured CECs as well as in native corneal endothelium.
Conclusions :
The integrin expression profile of cultured CECs was different from the profile of native corneal endothelium, namely the loss of α4, α5, α6, αv, β4 and β5 with cultured CECs, demonstrating that cell culture conditions affect integrin expression. While much has been learned about integrins, additional knowledge on the roles of integrins in the adhesion of CECs is needed, as loss of adhesion may be implicated in corneal diseases.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.