Fluctuations in Corneal Endothelial LAP2 Expression Levels Correlate with Passage Dependent Declines in Their Cell Proliferative Activity

Abstract

The corneal endothelium is the inner corneal mono-layered epithelium, fundamental for preserving corneal hydration and transparency. However, molecular mechanisms that regulate corneal endothelial cells (CEnCs), in particular regarding their proliferative capacity, have been only partially elucidated. CEnCs are quiescent in vivo and they easily undergo endothelial to mesenchymal transition (EnMT) in vitro. This study aims to analyze CEnCs behavior and expression in vitro, either in sub-confluent growing (S) or confluent (C) CEnCs cultures. Primary rabbit and human CEnCs were cultured and used for RT-PCR, immunofluorescence or western blot analysis. These methods allowed identifying a novel molecular marker, LAP2, that is upregulated in S while downregulated in C human or rabbit CEnCs. Those results were observed for several subsequent passages in culture and this, together with the correlation between ki67 and LAP2 expression, suggested LAP2 as a novel possible indicator for culture ageing. Finally, treatment with FGF and TGFβ in rCEnCs highlighted how LAP2 can vary as the cells regulate their proliferative state. In conclusion, we have identified a novel marker for CEnCs, LAP2, that regulates its expression depending on the cells sub/confluent state and that correlates with CEnCs proliferation.