PKA activity exacerbates hypoxia-induced ROS formation and hypoxic injury in PC-12 cells

Abstract

Hypoxia is a primary factor in many pathological conditions. Hypoxic cell death is commonly attributed to metabolic failure and oxidative injury. cAMP-dependent protein kinase A (PKA) is activated in hypoxia and regulates multiple enzymes of the mitochondrial electron transport chain, thus may be implicated in cellular energy depletion and hypoxia-induced cell death. Wild type (WT) PC-12 cells and PKA activity-deficient 123.7 PC-12 cells were exposed to 3, 6, 12 and 24h hypoxia (0.1% or 5% O2). Hypoxia, at 24 h 0.1% O2, induced cell death and increased reactive oxygen species (ROS) in WT PC-12 cells. Despite lower ATP levels in normoxic 123.7 cells than in WT cells, hypoxia only decreased ATP levels in WT cells. However, menadione-induced oxidative stress similarly affected both cell types. While mitochondrial COX IV expression remained consistently higher in 123.7 cells, hypoxia decreased COX IV expression in both cell types. N-acetyl cysteine antioxidant treatment blocked hypoxia-induced WT cell death without preventing ATP depletion. Transient PKA catα expression in 123.7 cells partially restored hypoxia-induced ROS but did not alter ATP levels or COX IV expression. We conclude that PKA signaling contributes to hypoxic injury, by regulating oxidative stress rather than by depleting ATP levels. Therapeutic strategies targeting PKA signaling may improve cellular adaptation and recovery in hypoxic pathologies.

Hypoxia with inflammation and reperfusion alters membrane resistance by dynamically regulating voltage-gated potassium channels in hippocampal CA1 neurons

“…Therefore, in the present study, we focused on the role of K V channels in determining the input resistance during Hypo-Inf and reperfusion. K V channels are a prime candidate because the current density and gating kinetics are dynamically regulated by phosphorylation states [51][52][53][54], which are altered in response to ischemia [55][56][57].…”

The Role of Reversible Protein Phosphorylation in Regulation of the Mitochondrial Electron Transport System During Hypoxia and Reoxygenation Stress in Marine Bivalves

“…Earlier studies in humans and model organisms demonstrated an important role of PKA and PKC in responses to H/R stress such as occurs during ischemia-reperfusion (Armstrong, 2004;Sanada et al, 2004;Hüttemann et al, 2012). In hypoxiasensitive terrestrial mammals, PKA and PKC are activated by hypoxia (Srinivasan et al, 2013;Gozal et al, 2017;Lucia et al, 2020), whereas in a nematode Caenorhabditis elegans, hypoxia suppresses PKA activity (Han et al, 2019). Our present study showed a strong species-specific modulation of the protein kinase activities by H/R stress in marine bivalves ranging from no change (except for a transient decrease in PKC during shortterm hypoxia) in M. edulis, an increase during hypoxia and 1 26 1 3 5 8 2 3 2 31 50 2 54 188 ND1 5 1 7 10 12 35 ND2 1 1 1 5 10 8 26 ND3 2 1 3 3 9 ND4 6 1 2 2 1 1 7 10 5 35 ND4L 1 1 1 3 2 1 9 ND5 8 2 3 1 2 2 6 12 1 19 56 ND6 3 1 1 3 3 1 6 18 Names of the kinases are given in the head row of the table.…”

“…reoxygenation in A. islandica, and a strong suppression after the long-term hypoxia in C. gigas. In mammals, activation of PKA and PKC may have protective or damaging effects during H/R stress depending on the timing of activation (i.e., during preconditioning vs. hypoxic exposure) and the isoform of the protein kinase that becomes activated (Armstrong, 2004;Sanada et al, 2004;Miura et al, 2010;Duquesnes et al, 2011;Yang et al, 2013;Gozal et al, 2017). Physiological implications of the H/Rinduced modulation of the PKA and PKC activity are not known in marine bivalves, and further studies are needed to determine the consequences of the modulation of these enzymes for cellular signaling during hypoxia.…”

Proapoptotic effects of 2,5‑hexanedione on pheochromocytoma cells via oxidative injury

“…The clonal line PC12, originally derived from a solid rat adrenal medulla tumor, has been widely used as a dopaminergic neuronal model for in vitro studies, and also serves as a principal cell model for investigations in molecular neuroscience (12,13). In addition, PC12 cells present as an excellent in vitro model to investigate certain aspects of various neurological disorders, including glutamate excitotoxicity (14), Parkinson's disease (15), Alzheimer's disease (16), and epilepsy (17); these cells have been reported to be useful in analyzing oxidative-stress-associated effects on neuronal cell survival (18–20). The production of reactive oxygen species (ROS) induced by environmental toxicants may elicit oxidative stress, which subsequently induces a state of imbalance between the antioxidant defense system and the generation of free radicals (21).…”