Submitted on September 12, 2008
Revised on January 20, 2009
Accepted on January 28, 2009
HYPOXIA STIMULATES VIA SEPARATE PATHWAYS ERK PHOSPHORYLATION AND NF-
B ACTIVATION IN SKELETAL MUSCLE CELLS IN PRIMARY CULTURE
Cesar Osorio-Fuentealba1, Juan Antonio Valdes2, Denise Riquelme1, Jorge Hidalgo1, Cecilia Hidalgo1, and Maria Angelica Carrasco1*
1 University of Chile, Faculty of Medicine
2 Universidad Andres Bello, Faculty of Ecology and Natural Resources and University of Chile, Faculty of Medicine
* To whom correspondence should be addressed. E-mail: maccarras{at}gmail.com.
Mammalian cells sense oxygen levels and respond to hypoxic conditions through the regulation of multiple signaling pathways and transcription factors. Here, we investigated the effects of hypoxia on the activity of two transcriptional regulators, ERK1/2 and NF-
B, in skeletal muscle cells in primary culture. We found that hypoxia significantly enhanced ERK1/2 phosphorylation, and stimulated NF-B-dependent gene transcription as well as nuclear translocation of a GFP-labeled p65 NF-B isoform. Phosphorylation of ERK1/2 and NF-B dependent transcription by hypoxia required calcium entry through L-type calcium channels. Calcium release from ryanodine sensitive stores was also necessary for ERK1/2 activation but not for NF-B-dependent transcription. N-acetylcysteine, a general scavenger of reactive oxygen species (ROS), blocked hypoxia-induced ROS generation but did not affect the stimulation of ERK1/2 phosphorylation induced by hypoxia. In contrast, NF-B activation was significantly inhibited by N-acetylcysteine and did not depend on ERK1/2 stimulation, as shown by the lack of effect of the upstream ERK inhibitor U0126. These separate pathways of activation of ERK1/2 and NF-B by hypoxia may contribute to muscle adaptation in response to hypoxic conditions.
