Human mesenchymal stem cells differentiate into keratocyte-like cells in keratocyte-conditioned medium

Culturing corneal keratocytes is difficult because keratocytes growing in a monolayer rapidly lose their stellate morphology and cease to express keratocyte markers such as keratocan, lumican and aldehyde dehydrogenase 1 family, member A1 (ALDH1A1). Conversely, mesenchymal stem cells (MSCs) can be easily expanded in cell culture, and they have a variety of differentiation pathways. We studied the feasibility of using MSCs as a source for corneal tissue engineering. Based on the observation that keratocytes have MSC-like properties, similar to bone marrow-derived MSCs (BM-MSCs), we hypothesized that MSCs would differentiate into corneal keratocyte-like cells in keratocyte-conditioned medium (KCM). We measured changes in the expression of keratocyte markers through quantitative real-time polymerase chain reaction (qRT-PCR) and found that human MSC's cultured in KCM expressed both keratocan and ALDH1A1. Western blot analysis demonstrated that human MSCs cultured in KCM steadily increased their expression of lumican and ALDH1A1, while they lost expression of α-smooth muscle actin (α-SMA). Immunocytochemistry indicated that human MSCs grown in KCM acquired characteristics similar to those of keratocytes. These results suggest that KCM can direct human MSCs to differentiate into keratocyte-like cells.