Abstract
Purpose:
To test whether human bone marrow-derived mesenchymal stem cells (BM-MSCs) can serve as feeder cells to expand human limbal epithelial progenitor/stem cells (LSCs).
Methods:
Primary human LSCs were isolated and seeded as single cells and cell clusters in the presence of BM-MSCs. LSCs were cultured directly on a BM-MSC monolayer (standard method), in a 3-dimensional (3-D) method, and 3-D fibrin method. Single LSCs cultured on a 3T3-J2 monolayer were used as a control. Cultured LSCs were analyzed for cell morphology, cell growth rate, and expression level of putative stem cell markers.
Results:
LSCs did not grow efficiently using the direct or standard method. LSCs in the cell cluster form showed a 2.5-fold higher cell growth rate using the 3-D method than in the control. LSCs cultured by 3-D and 3-D fibrin methods had a similar K14 mRNA expression level and a significantly lower K12 mRNA expression compared to the control (decreased by 34% and 95%, respectively; p<0.05). There was no difference in the percentage of K14+ and K12+ cells among all culture methods. The percentage of p63α bright cells in the 3-D (13.6%) and 3-D fibrin method (11%) was comparable to that in the control (12.8%, p>0.05).
Conclusions:
BM-MSCs could support the growth and maintenance of LSCs. The expansion rate was the highest in the novel 3-D culture method.
Keywords: 482 cornea: epithelium •
721 stem cells