The impact of human placental mesenchymal stem cells on the ability to maintain and amplify the limbal stem cells

Abstract

Objective To look for maintaining limbal stem cells (LSCs) undifferentiated growth of human-derived cells as the feeder layer and investigating placenta derived mesenchymal stem cells (HPMSCs) can be used in vitro amplification LSCs and maintain their stem cell properties. Methods In vitro isolation and culture HPMSCs, induce adipogenic, osteogenic differentiation, flow cytometry cell surface markers. Rabbits’ LSCs respectively co-cultured with HPMSCs, NIH-3T3 to calculate and compare the group cloning efficiency (CFE). Immunofluorescence examination can detect that if IPO13, CK3/12 existed. Results In vitro culture HPMSCs had the potency to differentiate into adipocytes and osteoblasts. They had strong proliferation ability and showed positive expression of CD29, CD44 and negative of CD34, CD45, HLA-DR. After a total of 8 days of co-culturing rabbits’ LSCs, the two feeder layers can form a larger LSCs clone group. The LSCs CFE of HPMSCs, NIH-3T3 feeder and feeder-free group was (3.02±0.73)%, (4.11±0.69)%, (0.24±0.08)%, respectively. The overall difference among the three groups was statistically significant (F=34.87, P 0.05); Among HPMSCs, NIH-3T3 feeder group and cell-free group were different (t=5.82, 8.10, P <0.01). Immunofluorescence detection of two feeder layers LSCs clone group markers IPO13 showed high expression, CK3/12 showed low expression, each group had no significant differences in the expression of stem cell markers. Conclusions HPMSCs can amplify LSCs and maintain their stem characteristics, which can be used as an ideal substitute of 3T3 feeder layer. Key words: Placental mesenchymal stem cells; Feeder layer; Co-culture