Mitochondrial Alterations in Fanconi Anemia Fibroblasts Following Ultraviolet A or Psoralen Photoactivation(PARA)
Received 1 August 2001; accepted 14 November 2001 ABSTRACT
The genetic disease Fanconi anemia (FA), generally considered to be a DNA repair defect, has also been related to a deficiency in cellular defense against reactive oxygen species (ROS). Results show that mitochondrial matrix densification occurs rapidly and transiently in FA fibroblasts following 8-methoxypsoralen (8-MOP) photoreaction or ultraviolet A (320 to 380 nm) (UVA) irradiation. This effect is oxygen dependent because it is more important under 20 than under 5 % oxygen tension. In contrast, in normal fibroblasts very little, if any, densification of mitochondrial matrix is induced by treatments even at the highest oxygen tension. The changes in matrix density in FA cells are accompanied by some modifications in transmembrane potential, linked to a Fenton-like reaction, and in mitochondrial cardiolipin content, differing from the responses of normal cells. These data are indicative of some sort of membrane damage induced by 8-MOP photoreaction and UVA irradiation, to which FA cells appear to be particularly sensitive.
Abbreviations: DESF, iron chelator deferoxamine; DiOC^sub 6^(3), 3,3'dihexyloxacarbocyanine iodide; FA, Fanconi anemia; mCICCP, carbamoyl cyanide m-chlorophenylhydrazone; 8-MOP, 8-methoxypsoralen; NAO, 10-N-nonyl-3,6-bis(dimethylamino)acridine (10-N-nonyl acridine orange); ^sup 1^O^sub 2^ singlet molecular oxygen; OD, optical density; OH, hydroxyl radical; PBS, phosphate-buffered saline; ROS, reactive oxygen species; TNF(alpha), tumor necrosis factor(alpha); UVA, ultraviolet radiation in the A region (320 to 380 nm); Delta Psi ^sub m^ mitochondrial transmembrane potential.
INTRODUCTION Fanconi anemia (FA) is an autosomal recessive disorder characterized by progressive pancytopenia associated with multiple developmental abnormalities, and cancer proneness (1). Eight different complementation groups, FA-A to FA-G, are actually distinguished (2,3), and six genes are cloned: FANCA, FANCC, FANCD2, FANCE, FANCG and FANCF. However, the precise functions) of the multiprotein FA complex are still unknown (4). FA cells demonstrate an increased sensitivity to DNA cross-linking agents such as psoralen plus ultraviolet A (320 to 380 nm) (UVA), chromosomal instability, a G2 cell cycle delay, modifications of some cytokine expression and of the apoptotic response (for review, see D'Andrea and Grompe [51).
Although FA is generally considered to be a DNA repair defect (for reviews, see Clarke et al. [61 and Buchwald and Moustacchi [71), some of the metabolic disturbances reported also suggest anomalies in oxygen metabolism (8,9). For instance, FA cells demonstrate an increased activity of the antioxidant phospholipid-hydroperoxide-gluthatione-peroxidase and a constitutive induction of the factor nuclear factor-K B, despite a reduced oxygen consumption (10), indicating an elevated cellular level of reactive oxygen species (ROS), either by overproduction or by diminished detoxification. Moreover, an oxidative stress to FA cells results in an increased production of ROS and accumulation of 8-hydroxy-2'-deoxyguanosine in the DNA (11).
Acknowledgements-We thank Dr. Patrice Xavier Petit for helpful
advice about specific mitochondrion fluorochromes. We are very grateful to Dr. Pierre Rustin for critically reading the manuscript and to Yuval Cohen for English language advice. We are very grateful to Prof. M. Grompe and Dr. B. Cox for determination of the complementation group of FA 150 fibroblasts. The technical contributions of Elly Efthymiou and Michele Guggiari are gratefully acknowledged. S.R. thanks particularly Drs. Giuseppe Baldacci and Evelyne Sage for their welcome in UMR 2027 CNRS/IC. This work was supported by grants from CNRS, Institut Curie, ACC SV n(deg) 8 (Ministere de la Recherche, France).
(PARA)Posted on the web site on November 28, 2001.
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Solange Rousset*1, Silvano Nocentini1, Danielle Rouillard2, Christiane Baroche1 and Ethel Moustacchi1
1UMR 218 CNRS, Institut Curie-Recherche, Paris, France and 2Laboratoire de Cytometrie, Institut Curie-Recherche, Paris, France
*To whom correspondence should be addressed at: UMR 2027 CNRS, Institut Curie-Recherche, Batiment 110, Centre Universitaire, 91405 Orsay Cedex, France. Fax: 33-1-6986-9429; e-mail: solange.rousset@ curie.u-psud.fr
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