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Protoporphyrin biosynthesis in Melanoma B16 cells stimulated by 5‐aminolevulinic acid and chemical inducers: Characterization of photodynamic inactivation
Author(s) -
Schoenfeld N.,
Mamet R.,
Nordenberg Y.,
Shafran M.,
Babushkin T.,
Malik Z.
Publication year - 1994
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910560119
Subject(s) - protoporphyrin ix , protoporphyrin , inducer , biosynthesis , melanoma , biochemistry , chemistry , photodynamic therapy , cancer research , biology , porphyrin , enzyme , gene , organic chemistry
Abstract The stimulation of protoporphyrin (PP) biosynthesis in BI6 melanoma cells in order to facilitate photodynamic cell killing was studied. Biosynthesis and accumulation of PP in the melanoma cells was increased from 8 to 15 pmol/mg protein by the use of dimethyl‐sulfoxide (DMSO), a differentiation‐inducer. Treatment of the cells with the porphyrogenic agent alh/l‐isopropyl‐acetamide (AIA) stimulated an additional PP increase. The most remarkable enhancement of intracellular PP was achieved by the supplementation of 5‐aminolevulinic acid (5‐ALA) to the growth medium following the addition of DMSO and AIA during the induction phase. The intracellular concentration of PP exceeded 21 950 pmol/mg protein following combined stimulation by DMSO/AIA and 5‐ALA. The porphyrins produced in the incubated cells, in serum‐depleted medium, consisted of 95% PP; 88% of it was recovered from the cells and only 7% was excreted into the medium. Photosensitization of the B16 melanoma cells containing high PP concentrations was effective even at low light doses. Potassium (K) efflux was the first measurable sign of cell damage determined by X‐ray microanalysis (XRMA) following fast liquid‐nitrogen fixation. During a I min interval, 70% of cellular K was lost. After 5 min illumination, complete cell destruction was detected by scanning electron microscopy (SEM) and XRMA. The photodam‐aged cells showed influx of Na, CI and Ca ions accompanying the immediate K losses. Ultrastructural cell damage was manifested by disintegration of the outer membrane. Total cell death of B16 melanoma cells was achieved by chemical induction of endogenous PP and photosensitization.