Regulation of 5-aminolevulinic acid-mediated protoporphyrin IX accumulation in human urothelial carcinomas.
OBJECTIVE
The purpose of this study was to clarify the regulatory mechanism of protoporphyrin IX (PpIX) synthesis mediated by 5-aminolevulinic acid (ALA) in human urothelial carcinoma (UC), leading to improved accuracy in photodynamic diagnosis and therapy using ALA.


METHODS
PpIX accumulation in cultured UC cells after incubation for 1-5 h with 0.5-5 mM ALA was analyzed by fluorescence analysis using fluorescence microscopy and flow cytometry technique.


RESULTS
PpIX fluorescence mediated by ALA was increased, and the intensity of PpIX fluorescence was time-dependently increased in UC cells compared to noncancerous cells. The distribution of endogenous PpIX fluorescence primarily coincided with mitochondria, and then increased at a specific perinuclear region in the cells during the time of incubation. The ALA-mediated PpIX synthesis in UC cells was suppressed by beta-alanine, an inhibitor of beta-transporters of cell membrane, and carbonylcyanide p-trifluoromethoxyphenyl hydrazone, an uncoupler of mitochondrial oxidative phosphorylation. In contrast, the ALA-mediated PpIX accumulation was increased by deferoxamine, an iron chelator, manganese and nitric oxide, which is contributed to PpIX metabolism by inhibiting ferrochelatase activity, generated by a nitric oxide-generating reagent NOC-18. As observed above, ALA-mediated PpIX synthesis in human UC cells was regulated by the process of ALA uptake, ALA conversion to PpIX and metabolism of accumulated PpIX to heme.


CONCLUSIONS
This shows that the suppression of ferrochelatase increased PpIX accumulation in UC cells using small amount of ALA, thus leading to an improved clinical practicability of photodynamic diagnosis and therapy.
Inoue
Keiji
K
Department of Urology, Kochi Medical School, Nankoku, Kochi 783-8505, Japan. keiji@kochi-u.ac.jp
Karashima
Takashi
T
Kamada
Masayuki
M
Shuin
Taro
T
Kurabayashi
Atsushi
A
Furihata
Mutsuo
M
Fujita
Hirofumi
H
Utsumi
Kozo
K
Sasaki
Junzo
J
D000622:Aminolevulinic Acid / Q000494:pharmacology*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002295:Carcinoma, Transitional Cell / Q000188:drug therapy / Q000378:metabolism*; D045744:Cell Line, Tumor; D003676:Deferoxamine / Q000494:pharmacology; D005294:Ferrochelatase / Q000037:antagonists & inhibitors / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009603:Nitroso Compounds / Q000494:pharmacology; D010085:Oxidative Phosphorylation / Q000187:drug effects; D017319:Photosensitizing Agents / Q000494:pharmacology*; D011524:Protoporphyrins / Q000096:biosynthesis*; D013997:Time Factors; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy / Q000378:metabolism*; D019459:Urothelium / Q000187:drug effects / Q000378:metabolism; D015091:beta-Alanine / Q000494:pharmacology
C093285:NOC 18; D009603:Nitroso Compounds; D017319:Photosensitizing Agents; D011524:Protoporphyrins; D015091:beta-Alanine; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D000622:Aminolevulinic Acid; C028025:protoporphyrin IX; D005294:Ferrochelatase; D003676:Deferoxamine