Photodynamically induced effects in colon carcinoma cells (WiDr) by endogenous photosensitizers generated by incubation with 5-aminolaevulinic acid.
Human adenocarcinoma cells of the line WiDr have been treated with 2 mM 5-aminolaevulinic acid (5-ALA) in the presence of 10% foetal calf serum. The treatment induces a linear accumulation of protoporphyrin IX (PpIX) for at least 7.5 h. After 7.5 h of incubation about 45% of the PpIX accumulated is cell-bound, while the rest is found in the medium (25%) or lost from the cells during washing with phosphate-buffered saline (30%). Exposure to white light at an intensity of 30 W/m2 for 18 min results in 95% reduction of clonogenicity in cells treated with 2 mM 5-ALA for 3.5 h. The enzymatic activities of enzymes located in cytosol (glyceraldehyde 3-phosphate dehydrogenase and lactate dehydrogenase) and lysosomes (acid phosphatase and beta-glucuronidase) are not influenced by a 5-ALA and light treatment inactivating about 35% of the cells. The MTT assay, which reflects mitochondrial dehydrogenase activity, but not succinate dehydrogenase, is partly inhibited by the same treatment. Treatment with 5-ALA in the absence of light increases O2 consumption by a factor of two, while the O2 consumption is inhibited when 5-ALA treatment is combined with exposure to light. In addition, 5-ALA and light exposure enhance accumulation of rhodamine 123 by 40% and reduce the intracellular ATP level by 25%. Confocal laser scanning microscopical analysis indicates granular perinuclear localization of the PpIX formed by 5-ALA treatment. In conclusion, photodynamic treatment using 5-ALA as a prodrug induces damage to mitochondrial function without inhibiting lysosomal and cytosolic marker enzymes.
Department of Clinical Chemistry, University Hospital, Trondheim, Norway.
O A
OA
Gederaas
M H
MH
Rasch
K
K
Berg
J W
JW
Lagerberg
T M
TM
Dubbelman
D000255:Adenosine Triphosphate / Q000378:metabolism; D000622:Aminolevulinic Acid / Q000627:therapeutic use*; D014408:Biomarkers, Tumor; D003110:Colonic Neoplasms / Q000188:drug therapy*; D005456:Fluorescent Dyes / Q000493:pharmacokinetics; D006537:Heptanoates / Q000494:pharmacology; D006801:Humans; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010101:Oxygen Consumption / Q000187:drug effects; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000627:therapeutic use*; D011524:Protoporphyrins / Q000378:metabolism; D020112:Rhodamine 123 / Q000493:pharmacokinetics; D013997:Time Factors; D014407:Tumor Cells, Cultured
D014408:Biomarkers, Tumor; D005456:Fluorescent Dyes; D006537:Heptanoates; D017319:Photosensitizing Agents; D011524:Protoporphyrins; D020112:Rhodamine 123; C020804:succinylacetone; D000622:Aminolevulinic Acid; D000255:Adenosine Triphosphate; C028025:protoporphyrin IX