Cryopreservation of Porcine Fetal Ventral Mesencephalic Tissue for Intrastriatal Transplantation in Parkinson's Disease

Abstract

In this study we examined the efficacy of cryopreserving porcine fetal mesencephalic tissue. After micro-scopical dissection of the ventral mesencephalon (VM) from E28 pig fetuses, the collection of explants was randomly divided into two equal parts. One part was directly prepared as cell suspension. The other part was stored in hibernation medium for less than 2 days and then cryopreserved as tissue fragments and stored in liquid nitrogen. After 2 weeks up to 1 year, these tissue fragments were thawed and processed as cell suspensions. After cell counting and assessment of viability, these cell suspensions were used to examine survival, morphology, and neurite formation of the dopaminergic neurons in cell culture as well as after intrastriatal implantation in 6-OHDA-lesioned rats. Comparison of cryopreserved with fresh VM cell suspensions showed no significant difference with respect to cell viability and the average number of living cells per VM explant. The morphology of cultured dopaminergic neurons after cryopreservation was identical to that of fresh cells. After intrastriatal implantation, survival and outgrowth of cryopreserved dopaminergic neurons as well as functional effects did not differ from those of fresh cells. In conclusion, the cryopreservation technique we used proves to be a reliably effective method for storing porcine fetal VM tissue.