THE EFFECT OF ALA AND RADIATION ON PORPHYRIN/HEME BIOSYNTHESIS IN ENDOTHELIAL CELLS

Abstract

To study porphyrin biosynthesis in human microvascular endothelial cells, HMEC-1 cells, a transformed human microvascular endothelial cell line, were incubated with 5-aminolevulinic acid (ALA), the precursor of endogenous porphyrins, and porphyrin accumulation was measured spectro-fluorometrically. The HMEC-1 cells accumulated porphyrin in a concentration-related and a time-dependent fashion. Protoporphyrin was the predominant porphyrin accumulated in the cells. The effect of light on protoporphyrin accumulation was evaluated by exposing the ALA-loaded HMEC-1 cells to ultraviolet-A (UVA) and blue light, followed by another incubation with ALA for 2-24 h. Enhancement of protoporphyrin accumulation in irradiated HMEC-1 cells was observed 2-24 h after irradiation, which was associated with a decrease in ferrochelatase protein and activity. Porphyrin accumulation from ALA after irradiation was significantly decreased when catalase (750-3000 U/mL, 29.3-44.3% suppression) or superoxide dismutase (270 U/mL, 36.4% suppression) was present during irradiation. These data demonstrate that HMEC-1 cells were capable of porphyrin biosynthesis, and that exposure of protoporphyrin-containing HMEC-1 cells to UVA and blue light, which includes the Soret band spectrum, decreased the ferrochelatase activity and its protein. These changes were mediated, at least in part, by reactive oxygen species.