Rock Inhibitors In Limbal Stem Cells In Vitro Amplification And Save The Role Of Research

ObjectiveThrough adding ROCK inhibitor Y-27632in corneal preservation solution to observe it’s influence to the organizational structure of the cornea of the saved, the cell colony forming capacity, the number of endothelial cells and morphological changes.MethodsPut fifteen fresh rabbit corneal ring and the central corneal into cornea medium-term preservation solution supplemented with10μM of Y-27632for1d,4d,7d and14d, the normal corneal preservation solution as a control group. Though detecting mean cell volume calculated, cell colony formation ability to analysis the effects of ROCK inhibitor Y-27632on the preservation of the cornea material limbal stem cells. Using the Bolster Blue-Alizarin red staining to observed the morphology of corneal endothelial cell, and count the number of endothelial cells to assessment the influence of Y-27632on the corneal endothelium cells.ResultsWith the limbal stem cells stored in the medium-term preservation solution from7to14days, the epithelial cells compared with the rules; Limbal epithelial cells was still capable of the formation of colony forming and stratified ability. The corneal endothelial cells can maintain its normal form, no significant changes in activity when the cornea limbal stored in the medium-term preservation solution for14days.ConclusionsThe results of the preliminary test showed that Y-27632was suitable to be added into the medium-term preservation solution to effectively extend the retention time of the corneal epithelium and corneal endothelial cells in vitro. ObjectiveTo observe the influence of the activity and function of the limbal stem cells through adding the ROCK inhibitor Y-27632to the cryopreservation and amplification of the limbal stem cells.MethodsPut cornea limbal stem cells or after amplification cryopreserved, make different groups according add Y-27632or not when the cells were cryopreserved and cultured, and its’concentration was10uM. Through detecting the mean cell volume and the cell colony formation ability to evaluate the function of the Y-27632on the cryopreservation and the culture of the limbal stem cells.ResultsY-27632can significantly increase the cell proliferation and stem cell colony formation ability when it was added to the culture of limbal epithelial cells. Only Y-27632was added in the cryopreservation of the limbal stem cells, there was no significant increase on the mean cell volume and the cell colony formation ability when the limbal stem cells were cryopreservation for a short time. But there was a significantly increase when they were cryopreservation for a long time (>14days). Added Y-27632both in the cryopreservation and in the culturvation of the limbal stem cells could significantly increase the mean cell volume and the cell colony formation ability as the same as before cryopreserved.ConclusionsY-27632has a significant effect during the cryopreservation and the culture of the limbal stem cells.It can maintain the activity of limbal stem cells and function. It also can improve the Cell proliferation and colony formation ability.