Purpose: We analyse effectiveness of corneal limbal cells' culture prepared from heart-beating organ donors, that include living donors and from cadaver donors buttons following 3 days storage in 4 degrees C in Likorol.
Material and methods: For experiment 12 adults (living and heart-beating organ donors) aged 28-63 (mean 46.3) and 12 corneal buttons of cadaver (aged 18-51, mean 34.1) were qualified. Tissue samples (1 mm2) were taken from superior corneal limbus. Sample from living donor obtained during routine operation was sent immediately to laboratory, as well as from heart-beating organ donor. The limbal biopsy of preserved cornea was taken after 3 days storage in 4 degrees C (Likorol). Samples were treated with trypsin/EDTA solution before culture. Collected cells in similar density in 1 ml of medium laid on dishes inserts, covered with fibrin (Tissucol) and cultured in presence of feeder layer of fibroblasts (L929 line). Epithelial cells were cultured for 14 days at 37 degrees C in humidified 5% CO2 atmosphere in supplemented 2:1 mixture of DMEM and Ham's F12. On the 14th day cells were collected. Number of cells per 1 ml of medium was counted by cytometer. Immunostaining for epithelium type (Keratin 3) was performed.
Results: The number of cells obtained from cadaver donors reached 184.2+/-14.9% whereas from living donors revealed 1013.1+/-104.2%, increase in relation to number of delivered cells. We observed only 0.83+/-0.3 colonies per microscopic area in cultures from preserved tissue versus 6.67+/-0.6 colonies in cultures from living donor.
Conclusions: The preservation in 4 degrees C in Likorol significantly decreases proliferative potential of the corneal limbus.