Mr Hortman Stem Cell Laboratory, Dubai, United Arab Emirates
Abstract: The isolation and cryopreservation of Wharton's Jelly mesenchymal stem cells (WJ-MSC) under Good Manufacturing Practice (GMP) conditions is an important prerequisite for the future clinical implementation of WJ-MSC. Although the isolation of WJ-MSC from fresh human umbilical cord (UC) has become a standard procedure in most laboratories, the quality of WJ-MSC after cryopreservation of UC tissue remains a topic of debate. Human UCs were transported from the hospitals to the class 5 cleanroom facility, samples were cleaned of residual blood, vessels were removed, and UCs were minced into small pieces 1-2 mm in size. Finally, the minced UCs were cryopreserved in 1.5 cryo-vials, which were stored in -80°C for 24 hours. The cryo-vials were then transferred to liquid nitrogen. After three-months of cryopreservation, WJ-MSCs were isolated from UC tissue of different donors. At the three-month mark, the cord from one donor was used for isolation, while at the six and nine-month marks, the cord from two donors were used, including the cord from the three-month isolation and one additional cord. The enzymatic isolation of WJ-MSCs was performed according to current GMP guidelines. After the isolation process, the sterility test was negative. Bright field images showed the typical spindle shape of adherent cells. Flow cytometry revealed expression of MSC-specific markers CD73, CD 90 and CD105 at both passage 0 and 1 in 99% of the cells. Less than 1% of cells expressed negative MSC markers (CD34:CD45). In conclusion, cryopreservation of minced human UC tissue fragments is a feasible and safe method for preserving UC tissue for the post-stored isolation of WJ-MSCs.
