Doxorubicin-induced MAPK activation in hepatocyte cultures is independent of oxidant damage.
Doxorubicin (DOX) is a potent anticancer drug, whose clinical use is limited on account of its toxicity. DOX cytotoxic effects have been associated with reactive oxygen species (ROS) generated during drug metabolism. ROS induce signaling cascades leading to changes in the phosphorylation status of target proteins, which are keys for cell survival or apoptosis. The mitogen-activated protein kinase (MAPK) cascades are routes activated in response to oxidative stress. In this work, the effects of DOX on cytotoxicity, indicators of oxidative stress (malondialdehyde -MDA- and GSH), and the phosphorylation status of extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs), and p38 kinases were analyzed in primary cultures of rat hepatocytes. DOX (1-50 microM) did not modify lactate dehydrogenase (LDH) release into the medium, the levels of MDA (determined by high-performance liquid chromatography [HPLC]) or the intracellular GSH during the incubation time up to 6 h. GSH levels from mitochondria extracted by Percoll gradient from cultured hepatocytes were not modified by DOX, thus excluding its depletion or any impaired mitochondrial uptake. Characterization of proteins by Western blot analysis revealed that DOX increased phosphorylation of p38 kinases and JNK1 and JNK2 in a dose- and time-dependent manner. DOX also increased ERK2 phosphorylation at latter time points. In conclusion, DOX triggers activation of ERK, JNK, and p38 kinases in primary cultures of rat hepatocytes independently of oxidant damage.
Navarro
Rosaura
R
Department of Physiology, Faculty of Medicine and Dentistry, University of the Basque Country, 48080-Bilbao, Spain.
Martínez
Rosa
R
Busnadiego
Idoia
I
Ruiz-Larrea
M Begoña
MB
Ruiz-Sanz
José Ignacio
JI
D000818:Animals; D004317:Doxorubicin / Q000494:pharmacology*; D004789:Enzyme Activation; D005978:Glutathione / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology; D015227:Lipid Peroxidation; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology*; D018384:Oxidative Stress; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley
D016877:Oxidants; D004317:Doxorubicin; D020928:Mitogen-Activated Protein Kinases; D005978:Glutathione