Effects of Perfluorocarbon Emulsions on Cultured Human Endothelial Cells

Abstract

Perfluorocarbons (PFCs) and their emulsions (PFCEs) were used in organ preservation before transplantation, but not in organ perfusion. Our purpose was to achieve organ perfusion with a PFCE at room temperature or at 37°C, i. e. with oxygenation, to prevent damages related to reoxygenation after hypoxia. Therefore, we first investigated the effect of such emulsions on endothelial cells, the first cells to be in contact with the emulsion. A stem emulsion was prepared from perfluorooctyl bromide (90%w/v), emulsified with egg yolk phospholipids (2%w/v) and stabilized with a mixed fluorocarbon-hydrocarbon “molecular dowel” (1,4%w/v) (droplets of ca 0.2 μm in diameter). This emulsion was found to be stable when diluted with cell culture media or organ preservation fluids. Endothelial cells from human umbilical vein (HUVECs) were cultured in multiwell plates in M199 medium (with growth factors, 10% foetal calf serum and 5% human serum). Confluent cells were incubated overnight with 51Cr, washed and overlayed with M199 (control) or the above PFCE diluted 2 × or 4 × with M199 (test). After incubation, the cytotoxicity of the PFCEs was estimated by measuring 51Cr release and observing cell morphology by electron and light microscopy.The percentages of released 51Cr were identical to those of the control cells for the 2 × 3 × or 4 × diluted PFCEs at 4, 25 or 37°C. After return to the M199 medium, the cells grew and multiplied normally. We conclude that the diluted PFCEs were devoid of cytotoxicity. The 2 × diluted PFCE was however partially taken up by the cells: by microscopy, we observed intracellular PFC droplets and by density gradient analysis we found a slight increase in cellular density. The diluted PFCEs were compared to classical organ preservation solutions: HUVECs were incubated with UW (University of Wisconsin) or EC (EuroCollins) solutions at +4 and 37°C (3, 17 or 24 h of incubation). The solutions were observed to be toxic to the cells under these conditions, with cell mortality after return to the M199 medium. This cytotoxicity may be attributed to the high K+ concentration of UW and EC, since similar assays performed on HUVECs with Hank's solution adjusted to 100 mM K⁺ showed a similar % of ⁵¹Cr release. UW and EC are therefore not acceptable as dilution media for PFCEs.

Additional information

Notes on contributors

M. Mathy-Hartert

Joyce Laing works in the Department of Child and Family Psychiatry, Playfield House, Cupar, Fife, and is a Consultant Art Therapist to Psychiatric Hospitals and Prisons and Chairwoman of the Scottish Society of Art and Psychology.