Experimental study on the cryopreservation of LLC-PK1 epithelial cells with hypoxic UW solution

Abstract

The effects of oxygen partial pressure on cryopreservation of the cells with organ preservation solution were explored. Hypoxic UW solution was made by purging the UW solution with argon. The pig proximal tubule epithelial cells (LLC-PK1 cells) were cryopreserved in hypoxic UW solution (Ar-UW group) or standard UW solution (UW group) at 4°C for 48 h. Trypan blue staining and LDH detection were performed to evaluate the injury of the cells. The results showed that the oxygen partial pressure in Ar-UW group was significantly declined from 242±6 mmHg to 83±10 mmHg. After cryopreservation at 4°C for 48 h, LDH leakage rate and Trypan blue-stained rate in Ar-UW group were (11.3±3.4)% and (10.5±4.7)%, respectively, which were significantly lower than in UW group [(49.5±6.9)% and (47.6±9.3)% respectively, both P<0.01]. It was concluded that lower oxygen partial pressure of UW solution was more beneficial to the cryopreservation of LLC.

Keywords

organ preservation solution / oxygen partial pressure / cryopreservation

References

[1] LeeP. J., ChoiA. M.. Pathways of cell signaling in hyperoxia. Free Radic Biol Med, 2003, 35: 341-350

[2] SouthardJ. H.. Viability assays in organ preservation. Cryobiology, 1989, 26: 232-238

[3] PoovalaV. S., HuangH., SalahudeenA. K.. Role of reactive oxygen metabolites in organophosphate-bidrin-induced renal tubular cytotoxicity. J Am Soc Nephrol, 1999, 10: 1746-1752

[4] BelzerF. O., SouthardJ. H.. Principles of solid organ preservation by cold storage. Transplantation, 1988, 45: 673-676

[5] HorowitzS.. Pathways to cell death in hyperoxia. Chest, 1999, 116: 64-67

[6] DreherD., RochatT.. Hyperoxia induces alkalinzation and dome formation in MDCK epithelial cells. Am J Physiol, 1992, 262(2Pt1): C358-364

[7] BaligaR., UedaN., WalkerP. D., et al.. Oxidant mechanisms in toxic acute renal failure. Am J Kidney Dis, 1997, 29: 465-477