Photofrin and light induces microtubule depolymerization in cultured human endothelial cells.

Endothelial cells were cultured from human umbilical veins and incubated with Photofrin (1 microgram/ml). Cells were then exposed to light, and cytoplasmic microtubule (MT) status was monitored by immunofluorescence microscopy using alpha-tubulin antibody. As early as 15 min following irradiation, a light dose-dependent depolymerization of MT was observed. At sublethal light doses, this effect was transient, with MT repolymerizing within 2-3 h. Cellular ATP levels were monitored to determine whether diminished ATP levels were correlated with MT depolymerization. No correlation was found, since ATP levels remained at a constant value near 50% of unirradiated controls during a time interval in which transient MT depolymerization was observed. Cell viability was monitored by trypan blue exclusion. Transient MT depolymerization occurred at photodynamic doses that produced essentially no decrease in cell viability, while at higher doses, irreversible MT depolymerization was observed prior to loss of viability. Since MT are unstable at intracellular calcium levels greater than 1 microM, we postulate that MT depolymerization results from increases in intracellular calcium caused by photodynamic insult. MT are important in maintaining cell shape. Disruption of MT in endothelial cells due to photodynamic therapy could result in or contribute to exposure of the thrombogenic subendothelium or could alter vascular permeability in the treatment area.