Date on Master's Thesis/Doctoral Dissertation
Physiology and Biophysics
Protein kinase A; Hypoxia; Metabolism; Energy; Cell death; Reactive oxygen species; PC-12
Protein kinases; Anoxemia
Hypoxia is characterized by an inadequate oxygen supply to the tissues in proportion to their metabolic needs, and is a primary factor in traumatic CNS injury, strokes, cardiopulmonary diseases, and obstructive sleep apnea. The cAMP-dependent protein kinase (PKA) has been attributed a role as an antiapoptotic kinase as well as a pro-apoptotic signal. Thus the role of PKA in hypoxia-induced cell survival is currently unclear. We show here that hypoxia induces cell death in WT PC-12 cells, pheochromocytoma cells used as a model of the oxygen-sensitive carotid body glomus cells. The onset of cell death following 24 hours of severe, sustained hypoxia also correlated with an increase in oxidative stress and a drop in ATP levels. Inactivation of PKA in the 123.7 PC-12 cell line prevented the hypoxiainduced ROS surge, energy depletion, and cell death. PKA activity thus contributes to hypoxia~induced cell death in WT cells via enhanced ROS production and energy depletion, but was not shown to playa role in the regulation of cellular antioxidant mechanisms. 123.7 cells had lower levels of hypoxia-induced glucose utilization and lactate release than WT cells, thus PKA appears to stimulate increases in glycolytic flux during hypoxia. Mitochondrial potentials were higher in 123.7 cells than in WT cells, indicating that PKA inhibits mitochondrial metabolism. Furthermore, the expression of COX IV was decreased by hypoxic exposure in both cell types, but was significantly greater in 123.7 cells. The greater expression of COX IV may allow 123.7 cells to maintain mitochondrial electron flow and energy production under drastically reduced oxygen conditions. Antioxidant treatment blocked hypoxia-induced cell death, but was not sufficient to prevent depletion of ATP or to modulate the metabolic pathways. Thus, this finding indicates that hypoxia-induced metabolic regulation and energy depletion occur independently of oxidative stress. Consequently, the PKA signaling pathway appears to contribute to hypoxia-induced cell death through its regulatory effects on oxidative stress and metabolic pathways. Modulation of the PKA signaling pathway could provide novel therapeutic strategies to improve cellular adaptation and recovery in the many pathologies characterized by periods of hypoxia.
Miller, Cynthia Jayne, "The role of protein kinase A in hypoxia-induced PC-12 cell death." (2008). Electronic Theses and Dissertations. Paper 976.