The iron regulatory protein can determine the effectiveness of 5-aminolevulinic acid in inducing protoporphyrin IX in human primary skin fibroblasts

The level of endogenous photosensitiser, protoporphyrin IX (PPIX), can be enhanced in the cells by 5-aminolevulinic acid (ALA). We investigated the effect of critical parameters such as growth state of the cells and availability of intracellular iron in modulating the level of PPIX, in human primary cultured skin fibroblasts (FEK4) maintained either in exponentially growing or growth-arrested phase, following treatment with ALA. The addition of ALA to exponentially growing cells increased the level of PPIX 6-fold relative to control cells; however, in growth-arrested cells the same treatment increased the level of PPIX up to 34-fold. The simultaneous addition of the hydrophilic iron-chelator Desferal with ALA, boosted the level of PPIX up to 47-fold in growing cells and up to 42-fold in growth-arrested cells, suggesting that iron is limiting under the latter conditions. The strict dependence of PPIX enhancement on free available iron levels was examined by the level of activation of iron regulatory protein in band shift assays. This analysis revealed that the basal level of iron regulatory protein in growth-arrested cells was 6-fold higher than in growing cells, reflecting the influence of the free available iron pool in exponentially growing cells. Interestingly, the same ratio was found between the basal level concentration of PPIX in growing and growth-arrested cells. We propose that iron regulatory protein activation could serve as a marker for developing photodynamic therapy protocols because it identifies cells and tissues with a propensity to accumulate PPIX and it is therefore likely to predict the effectiveness of such therapies.