Intracellular glutathione levels determine cell sensitivity to apoptosis induced by the antineoplasic agent N-(4-hydroxyphenyl) retinamide.
BACKGROUND
We have previously demonstrated that the synthetic retinoid N-(4-hydroxyphenyl) retinamide (4-HPR) induces the overproduction of reactive oxygen species (ROS) in human leukemia cells, which in turn triggers the intrinsic (mitochondrial) apoptotic pathway. In order to study the role of glutathione in 4-HPR-induced apoptosis, the levels of this antioxidant were analyzed in cell lines which are sensitive and resistant to the effects of 4-HPR, and the effect of the modulation of glutathione levels on 4-HPR cytotoxicity was characterized.


METHODS
Mitochondrial membrane potential (deltaPsim) and the levels of glutathione were measured by flow cytometry. A fluorometric assay was used to measure intracellular ROS generation and Western blot was employed to analyze tissue transglutaminase expression.


RESULTS
4-HPR generated large quantities of ROS in cell lines which expressed low glutathione levels, these cells being the most sensitive to the retinoid. The sensitivity of leukemia cells to 4-HPR could be modulated, either by increasing intracellular glutathione contents using all-trans retinoic acid (ATRA), or by decreasing it using DL-buthionine-S,R-sulfoximine (BSO). ATRA increased the level of expression of tissue transglutaminase, whereas inhibition of this enzyme led to enhanced apoptosis.


CONCLUSIONS
Our findings indicate that the glutathione content contributes to determining the sensitivity of cells to 4-HPR and points to the potential application of glutathione-inhibiting agents as enhancers in 4-HPR-based therapies.
Morales
María-Celia
MC
Department of Cell Biology and Histology, Faculty of Medicine and Dentistry, University of the Basque Country, Leioa- 48940, Bizkaia, Spain.
Pérez-Yarza
Gorka
G
Nieto-Rementeria
Naiara
N
Boyano
María-Dolores
MD
Jangi
Muhialdin
M
Atencia
Rafael
R
Asumendi
Aintzane
A
D000970:Antineoplastic Agents / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D019328:Buthionine Sulfoximine / Q000008:administration & dosage / Q000494:pharmacology; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017313:Fenretinide / Q000008:administration & dosage / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D015458:Leukemia, T-Cell / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D018384:Oxidative Stress / Q000187:drug effects; D011503:Transglutaminases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D014212:Tretinoin / Q000008:administration & dosage / Q000494:pharmacology
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D017313:Fenretinide; D019328:Buthionine Sulfoximine; D014212:Tretinoin; D011503:Transglutaminases; D005978:Glutathione; C008542:monodansylcadaverine; D002103:Cadaverine