M. Ohji et al., BASEMENT-MEMBRANE SYNTHESIS BY HUMAN CORNEAL EPITHELIAL-CELLS IN-VITRO, Investigative ophthalmology & visual science, 35(2), 1994, pp. 479-485
Purpose. Collagen gels may prove to be potential carriers for transpla
ntation of cultured corneal epithelial cells. The purpose of this stud
y was to evaluate the suitability of collagen gels in comparison with
corneal stromal blocks as the substrate to support the growth of human
corneal epithelial cells in culture and the synthesis and deposition
of the basement membrane components by these cells. Methods. Corneal e
pithelial sheets, freed from the culture dishes using Dispase II (Boeh
ringer Mannheim, Indianapolis, IN), were cultured on corneal stromal b
locks. Deposition of laminin, type IV collagen, type VII collagen, and
perlecan (heparan sulfate proteoglycan) were evaluated immunohistoche
mically after 4 days, 7 days, 2 weeks, and 3 weeks. Human limbal expla
nt cultures were established on collagen gels prepared from bovine typ
e I collagen with or without addition of cultured human corneal fibrob
lasts. After 1, 2, 3, and 4 weeks, the deposition of the basement memb
rane components was evaluated immunohistochemically. Results. Corneal
epithelial cells, cultured on corneal stromal blocks as well as on col
lagen gels with or without fibroblasts, deposited laminin, type IV col
lagen, perlecan, and type VII collagen at the interface of the cells a
nd the substrates. However, different substrates differentially influe
nced the temporal pattern of the deposition of various basement membra
ne components. On the stromal blocks, deposition of laminin, type IV c
ollagen, and perlecan by the epithelial cells was evident at 1 week. T
ype VII collagen was detected at 2 weeks. On the collagen gels with fi
broblasts, deposition of laminin, type IV collagen and perlecan was de
tectable at 1 week. In the epithelial cultures on the collagen gels wi
thout fibroblasts, only perlecan was detectable at 1 week. At 2 weeks,
all of the basement membrane components, including type VII collagen
were detectable on the collagen gels, either with or without fibroblas
ts. Conclusion. Human corneal epithelium cultured on collagen gels or
on corneal stromal blocks can synthesize and deposit basement membrane
components, including laminin, type IV collagen, type VII collagen, a
nd perlecan within 2 weeks in culture. Therefore, collagen gels may se
rve as potential carriers for human corneal epithelial transplantation
.