In our experiment we digested and incubated the LNCs and LSCs in 10% FBS-DMEM medium, which is not suitable for corneal epithelial cell growth but benefits mesenchymal cell growth. Isolated cells showed fibroblast cell morphology, as well as negative expression of K3 and K4, to make sure that LNCs and LSCs were not contaminated by corneal epithelial cells or conjunctival cells. In additional, reverse transcriptase-PCR analysis showed both LNC and LSC expression of various growth factors (EGF, FGF2, EPR, HGF, KGF, NGF, GDNF, and BDNF) and the potential corneal epithelial stem/progenitor cell marker N-cadherin
11 and importin 13.
30 However, LNCs expressed a higher level of E-cadherin and a lower level of NT3 compared with LSCs. Growth factors play an important role in the maintenance and normal wound healing of the corneal epithelium
31 ; among these, most are predominantly expressed by fibroblasts and affect the proliferation and differentiation of the surrounding epithelial cells in a paracrine manner through their cognate receptors.
23–26 Our results confirmed that both LNCs and LSCs secrete various growth factors. Although LSCs expressed higher levels of NT3 than did LNCs, NT3 is considered to have a weak effect on limbal stem/progenitor cells because its receptor TrkC is weakly expressed by limbal epithelial stem/progenitor cells.
32 Our findings also verified this. Cadherin mediates cell–cell adhesion and plays a major role in embryonic development, tissue formation, and cellular growth and differentiation.
33 Previous studies had found that N-cadherin (neuronal cadherin) plays an important role in the interactions between hematopoietic/limbal stem/progenitor cells with their niche cells.
11,34 N-cadherin in limbal epithelial stem/progenitor cells likely acts as an anchor molecule for their attachment to N-cadherin+ niche cells.
11 Limbal niche cells located in the basal layer of the limbal epithelium express N-cadherin, suggesting that they might have direct contact with epithelial stem/progenitor cells in the limbus via N-cadherin. E-cadherin (epithelial cadherin), which is located on the lateral surfaces of most epithelial cells, plays an important role in transformation of mesenchyme to epithelium in embryos
35 and plays an essential role in the collective directional migration of large epithelial sheets as mediate wound epithelium healing.
36 It is noteworthy that LNCs express higher levels of E-cadherin than LSCs, which might be why LNCs had a greater capacity to support the limbal epithelial stem/progenitor cells than the LSCs. It should be mentioned that the LNCs have the potential to different when cultured in serum-containing medium for a long time. According to a previous report, a modified embryonic stem cell medium can maintain the undifferentiated state and the expression of embryonic stem cell markers of LNCs,
17 so further study should be performed to compare LNCs and LSCs under the embryonic stem cell medium condition.