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Originally published In Press as doi:10.1074/jbc.M107678200 on September 28, 2001

J. Biol. Chem., Vol. 276, Issue 50, 47379-47386, December 14, 2001
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Photodynamic Therapy-induced Apoptosis in Epidermoid Carcinoma Cells
REACTIVE OXYGEN SPECIES AND MITOCHONDRIAL INNER MEMBRANE PERMEABILIZATION*

Minh LamDagger §, Nancy L. OleinickDagger , and Anna-Liisa Nieminen||

From the Departments of  Anatomy and Dagger  Radiation Oncology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106

Photodynamic therapy (PDT), a novel and promising cancer treatment that employs a combination of a photosensitizing chemical and visible light, induces apoptosis in human epidermoid carcinoma A431 cells. However, the precise mechanism of PDT-induced apoptosis is not well characterized. To dissect the pathways of PDT-induced apoptosis, we investigated the involvement of mitochondrial damage by examining a second generation photosensitizer, the silicon phthalocyanine 4 (Pc 4). By using laser-scanning confocal microscopy, we found that Pc 4 localized to cytosolic membranes primarily, but not exclusively, in mitochondria. Formation of mitochondrial reactive oxygen species (ROS) was detected within minutes when cells were exposed to Pc 4 and 670-675 nm light. This was followed by mitochondrial inner membrane permeabilization, depolarization and swelling, cytochrome c release, and apoptotic death. Desferrioxamine prevented mitochondrial ROS production and the events thereafter. Cyclosporin A plus trifluoperazine, blockers of the mitochondrial permeability transition, inhibited mitochondrial inner membrane permeabilization and depolarization without affecting mitochondrial ROS generation. These data indicate that the mitochondrial ROS are critical in initiating mitochondrial inner membrane permeabilization, which leads to mitochondrial swelling, cytochrome c release to the cytosol, and apoptotic death during PDT with Pc 4.

* This work was supported in part by National Institutes of Health Grants RO1 NS39469 (to A.-L. N.), PO1 CA48735 (to N. L. O.), and P30 CA43703.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ Supported by National Institutes of Health Research Oncology Training Grant T32 CA59366.

|| To whom correspondence should be addressed: Case Western Reserve University, Dept. of Anatomy, School of Medicine W520, 10900 Euclid Ave., Cleveland, OH 44106. Tel.: 216-844-8204; Fax: 216-844-8209; E-mail: axn25@po.cwru.edu.

Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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