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Susumu Hara, Ryuhei Hayashi, Tomofumi Kageyama, Motokazu Tsujikawa, Kohji Nishida; Isolation and characterization of p75NTR positive and high-proliferative corneal endothelial cells from the human corneal endothelium. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1018. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The corneal endothelium is believed to be developmentally originated from periocular mesenchyme via neural crest. The human corneal endothelial progenitor cells (HCEPs) have been investigated because of their potential availability for the tissue regenerative medicine. However, the existence and the properties of HCEPs have not been elucidated yet. We attempted to isolate the HCEPs from the human corneal endothelium by using the specified culture system and p75 neurotrophin receptor (p75NTR).
The Descemet's membranes were stripped from the human cornea, then, treated with a cell dissociated reagent. To isolate the HCEPs, the endothelial cells were seeded on the dish coated with laminin and cultured in the serum-free media containing basic fibroblast growth factor. Expression of neural crest markers in the isolated HCEPs was examined by real-time PCR and immunostaining.
The proliferating cells were appeared at around 14 days after the seeding, exhibited a bipolar, spindle-shaped morphology, similar to neural crest cells. Interestingly, the proliferating cells expressed neural crest markers, p75NTR and Sox9. The colony forming efficiency was approximately 0.31±0.04%, showed no significant relation to donor ages. The proliferating cells were able to undergo passage several times in younger donors below 60 years old, and the proliferative capability was higher than that of human corneal endothelial cells cultivated by the conventional method with fetal bovine serum containing media.
We succeeded in the isolation of HCEPs which had high p75NTR expression and proliferative capability.
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