Endothelial cell preservation at hypothermic to normothermic conditions using clinical and experimental organ preservation solutions

Abstract

Introduction: Endothelial barrier function is pivotal for the outcome of organ transplantation. Since hypothermic preservation (gold standard) is associated with cold-induced endothelial damage, endothelial barrier function may benefit from organ preservation at warmer temperatures. We therefore assessed endothelial barrier integrity and viability as function of preservation temperature and perfusion solution, and hypothesized that endothelial cell preservation at subnormothermic conditions using metabolism-supporting solutions constitute optimal preservation conditions.

Methods: Human umbilical vein endothelial cells (HUVEC) were preserved at 4-37°C for up to 20 h using Ringer's lactate, histidine-tryptophan-ketoglutarate solution, University of Wisconsin (UW) solution, Polysol, or endothelial cell growth medium (ECGM). Following preservation, the monolayer integrity, metabolic capacity, and ATP content were determined as positive parameters of endothelial cell viability. As negative parameters, apoptosis, necrosis, and cell activation were assayed. A viability index was devised on the basis of these parameters.

Results: HUVEC viability and barrier integrity was compromised at 4°C regardless of the preservation solution. At temperatures above 20°C, the cells' metabolic demands outweighed the preservation solutions' supporting capacity. Only UW maintained HUVEC viability up to 20°C. Despite high intracellular ATP content, none of the solutions were capable of sufficiently preserving HUVEC above 20°C except for ECGM.

Conclusion: Optimal HUVEC preservation is achieved with UW up to 20°C. Only ECGM maintains HUVEC viability at temperatures above 20°C.

Keywords: (sub)normothermic (20–37°C) preservation; 1% FCS-enriched PBS; ATP; DW; ECGM; ECIS; FACS; FCS; FLICA; HTK; HUVEC; Histidine–tryptophan–ketoglutarate solution; Human umbilical vein endothelial cells; ICAM-1; PBS; PBS+; PS; Polysol; RL; RT; Ringer's lactate; SNP; TRIS; Transplantation; UW; University of Wisconsin solution; WE; WST-1; Williams medium E; adenosine triphosphate; demineralized water; electric cell–substrate impedance sensing; endothelial cell growth medium; fetal calf serum; fluorescence-activated cell sorting; fluorescently labeled inhibitor of caspases; histidine–tryptophan–ketoglutarate solution; human umbilical vein endothelial cells; intercellular adhesion molecule-1; phosphate buffered saline; room temperature; tris(hydroxymethyl)aminomethane buffer; water-soluble tetrazolium-1.