All issues > Volume 43(12); 2000

Original Article

J Korean Pediatr Soc. 2000;43(12):1576-1582. Published online December 15, 2000.

The Changes of Apoptosis and Viability in Stem Cells after Thawing with Lapse of the Time

Seun Joo SJ Park¹, Sang Young SY Bae¹, Kyung Ha KH Ryu¹, Il Tae IT Whang¹, Kyung Hyo KH Kim¹, Young Mi YM Hong¹, Kyung Hee KH Kim¹, Keun K Lee¹, Joo Young JY Seoh²

¹Department of Pediatrics, College of Medicine, Ewha Womans University, Seoul, Korea
²Department of Microbiology, College of Medicine, Ewha Womans University, Seoul, Korea

Abstract

Purpose
: Thawed stem cells should be infused as early as possible because delay of infusion leads to decrease of cell viability and formation of DNA clumping. The procedure of 10% Dimethyl sulfoxide(DMSO) removal and a long distance from the thawing location to the patient are the main causes of delay of infusion that results in the loss of cell viability and apoptosis. Therefore, we investigated the changes of cell viability and apoptosis after thawing with lapse of time.
Methods
: Five samples of mobilized peripheral blood were evaluated. We measured cell viability, colony forming unit(CFU) and apoptosis at 0, 1, 2, 4, and 24 hours after thawing. The state of stem cells were divided into live, apoptotic and dead with double staining using annexin V and 7-aminoactinomycin D(7-AAD) in flow cytometry.
Results
: Viability of the total cells after thawing was 77.3(53.3-97.7)%. The percentage which recovered to initial CFU at 1, 2, 4 and 24 hours after thawing decreased to 63.9%, 50.2%, 45.8% and 11.6%, respectively. The proportion of apoptotic cells among CD34＋ cells after thawing were increased from 0.2% at 0 hour to 16.5% at 1 hour, 21.9% at 2 hours, and then decreased to 15.0% at 4 hours, 2.7% at 24 hours because they were replaced by dead cells.
Conclusion
: Thawed cells changed to apoptotic and had less colony forming capacity from 1 hour after thawing, and were then replaced by dead cells from 4 hours after thawing.

Keywords :Thawing, Cell viability, Apoptosis, Time, CFU