Previously, we have shown that cultured liver endothelial cells are affected by an energy-dependent injury when incubated in cold University of Wisconsin (UW) or histidine-tryptophan-ketoglutarate solution. Here, we studied the susceptibility of other endothelial cells to this type of injury. Aortic endothelial cells in early-confluent, i.e., still proliferating, monolayer cultures were damaged more quickly during cold incubation in UW solution than during cold incubation in Krebs-Henseleit buffer. At this stage the addition of KCN did not alter the loss of viability in UW solution, but when the culture period was prolonged, cells were protected by the addition of cyanide. A paradoxical, protective effect of KCN could also be observed in late-confluent, i.e., nonproliferating, cultures of coronary endothelial cells incubated in UW solution. Similarly, liver endothelial cells in subconfluent, growing cultures were damaged by the addition of cyanide (loss of viability after 48 h, 3 +/- 1% in UW, 65 +/- 19% in UW + KCN), whereas in late-confluent cultures the addition of cyanide to UW solution was protective (loss of viability after 48 h, 100 +/- 0% in UW, 31 +/- 15% in UW + KCN). Variations of culture period and seeding density and the use of inhibitors of cell proliferation demonstrated that liver endothelial cells acquire their susceptibility to energy-dependent injury along with confluence. Subcultured cells retained this susceptibility for some hours. These results suggest that the energy-dependent injury described previously is not confined to liver endothelial cells and that the occurrence of energy-dependent injury requires a capacity of the cells that develops only after cultures have grown to confluence.