Human corneal epithelial cell-transformed (HCE-T) cell line is used as a widely accepted
barrier model for pharmacological investigations in the context of eye application.
The differentiation of (limbal) corneal epithelial into mature corneal epithelium
coincides with the expression of established differentiation markers. If these differentiation
mechanisms are disturbed, it will lead to ocular surface disease. In this study, we
want to compare the expression of differentiation markers in the HCE-T cell line to
differentiated primary epithelial cells (pCECs) and primary limbal epithelial cell
(LEC) culture. This is necessary in order to decide whether HCE-T cells could be a
tool to study the differentiation process and its regulatory networks in corneal epithelium.Primary
limbal epithelial cells (LECs) for cell culture and primary corneal epithelial cells
(pCECs) as differentiated tissue samples were obtained from the limbus or central
cornea region of corneal donors. HCE-T cell line was purchased from RIKEN Institute
RCB-2280.Expression levels of conjunctival- and corneal-specific keratin and adhesion
markers (KRT3, KRT12, KRT13, KRT19, DSG1), stem cell and differentiation markers (PAX6,
ABCG2, ADH7, TP63, ALDH1A1), and additional (unvalidated) putative differentiation
and stem cell markers (CTSV, SPINK7, DKK1) were analyzed with qPCR. Additionally,
KRT3, KRT12, DSG1, and PAX6 protein levels were analyzed with Western blot.KRT3, KRT12,
DSG1, PAX6, ADH7, and ALDH1A1 mRNA expressions were higher in LECs and magnitudes
higher in pCECs compared to HCE-T cells. KRT3, KRT12, PAX6, ALDH1A1, ADH7, TP63, and
CTSV mRNAs have shown increasing mRNA expression from HCE-T < HCE-T cultured in keratinocyte
serum-free medium (KSFM) < LEC < to pCEC.KRT3 and KRT12 protein expressions were only
slightly increased in LEC compared to HCE-T samples, and the strongest signals were
seen in pCEC samples. DSG1 protein expression was only detected in pCECs. PAX6 protein
expression was hardly detected in HCE-T cells, and no difference could be seen between
LECs and pCECs.The HCE-T cell line is even less differentiated than LECs regarding
the investigated markers and therefore might also lack the ability to express differentiation
markers at protein level. Hence, this cell line is not suitable to study corneal differentiation
processes. Primary LECs in the way cultured here are not an ideal system compared
to differentiated epithelium in organ culture but should be preferred to HCE-T cells
if corneal differentiation markers are investigated. Other cell models or differentiation
protocols should be developed in the future to gain new tools for research on ocular
surface diseases.
