casein-kinase-ii and Nerve-Degeneration

NADPH oxidase is a major complex that produces reactive oxygen species (ROSs) during the ischemic period and aggravates brain damage and cell death after ischemic injury. Although many approaches have been tested for preventing production of ROSs by NADPH oxidase in ischemic brain injury, the regulatory mechanisms of NADPH oxidase activity after cerebral ischemia are still unclear. In this study, we identified casein kinase 2 (CK2) as a critical modulator of NADPH oxidase and elucidated the role of CK2 as a neuroprotectant after oxidative insults to the brain. We found that the protein levels of the catalytic subunits CK2alpha and CK2alpha', as well as the total activity of CK2, are significantly reduced after transient focal cerebral ischemia (tFCI). We also found this deactivation of CK2 caused by ischemia/reperfusion increases expression of Nox2 and translocation of p67(phox) and Rac1 to the membrane after tFCI. Interestingly, we found that the inactive status of Rac1 was captured by the catalytic subunit CK2alpha under normal conditions. However, binding between CK2alpha and Rac1 was immediately diminished after tFCI, and Rac1 activity was markedly increased after CK2 inhibition. Moreover, we found that deactivation of CK2 in the mouse brain enhances production of ROSs and neuronal cell death via increased NADPH oxidase activity. The increased brain infarct volume caused by CK2 inhibition was restored by apocynin, a NADPH oxidase inhibitor. This study suggests that CK2 can be a direct molecular target for modulation of NADPH oxidase activity after ischemic brain injury.

Topics: Acetophenones; Animals; Brain; Brain Ischemia; Casein Kinase II; Cytoprotection; Disease Models, Animal; Down-Regulation; Enzyme Activation; Enzyme Inhibitors; Male; Membrane Glycoproteins; Mice; NADPH Oxidase 2; NADPH Oxidases; Nerve Degeneration; Neuroprotective Agents; Phosphoproteins; Protein Binding; rac1 GTP-Binding Protein; Reactive Oxygen Species; Reperfusion Injury

The heme oxygenase (HO) enzymes catalyze the rate-limiting step of heme breakdown, and may accelerate oxidative injury to neurons exposed to heme or hemoglobin. HO-1 and HO-2 are activated in vitro by the phos-phatidylinositol 3-kinase (PI3K)/Akt and protein kinase C (PKC)/CK2 pathways, respectively. The present study tested the hypotheses that CK2, PKC, and PI3K inhibitors would reduce both HO activity and neuronal vulnerability to hemoglobin in murine cortical cultures. Oxidative cell injury was quantified by LDH release and malondialdehyde assays. HO activity was assessed by carbon monoxide assay. Consistent with prior observations, treating primary cortical cultures with hemoglobin for 16h resulted in release of approximately half of neuronal LDH and a seven-fold increase in malondialdehyde. Both endpoints were significantly reduced by the CK2 inhibitors 4,5,6,7-tetrabromobenzotriazole (TBB) and 2-dimethyl-amino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT), and by the PKC inhibitor GF109203X; the PI3K inhibitors LY294002 and wortmannin had no effect. None of these inhibitors altered basal HO activity. The 1.9-fold activity increase observed after hemoglobin treatment was largely prevented by LY294002 and LY303511, a structural analog of LY294002 that does not inhibit PI3K activity. It was not reduced by wortmannin, TBB or GF109203X. These results suggest that the protective effect of CK2 and PKC inhibitors in this model is not dependent on reduction in HO activity. In this culture system that expresses both HO-1 and HO-2, HO activity does not appear to be primarily regulated by the PKC/CK2 or PI3K pathways.

Topics: Animals; Casein Kinase II; Cells, Cultured; Cerebral Hemorrhage; Coculture Techniques; Cytoprotection; Heme Oxygenase (Decyclizing); Hemoglobins; Isoenzymes; L-Lactate Dehydrogenase; Malondialdehyde; Mice; Nerve Degeneration; Neurons; Oxidative Stress; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Protein Kinase C; Protein Kinase Inhibitors; Signal Transduction

Neurofibrillary tangles (NFT) are pathological cytoskeletal structures composed of paired helical filaments (PHF), and are found in neurons of patients afflicted with many neurodegenerative disorders, including Alzheimer's disease (AD). We previously found that an antiserum against casein kinase II (CK-II) stained NFT intensely in the brain tissue of AD patients. In the current study, we found that the anti-CK-II antiserum stains NFT and neuronal inclusions in many other neurodegenerative diseases as well, including Guam-Parkinson dementia complex, chromosome 18 deletion syndrome, progressive supranuclear palsy, Kufs' disease, and Pick's disease. This antiserum reacted, in crude brain homogenates, with both a doublet of Mr 43,000 and a Mr 27,000 Da protein which could correspond to the alpha, alpha', and beta chains of CK-II. The staining of these bands was adsorbed by preincubating anti-CK-II antiserum with purified CK-II. Preincubation of brain sections with purified CK-II strongly intensified the immunostaining of NFT with anti-CK-II, suggesting that NFT may bind CK-II. In the AD brain homogenates, the particulate CK-II levels are increased whereas the cytosolic levels are decreased without a change in total CK-II levels, consistent with the idea that CK-II binds to the particulate PHF, a major constituent of NFT. In accord with these findings, purified PHF bound CK-II, but purified PHF did not contain CK-II as its component. These results suggest that CK-II might be an extraneously deposited component of NFT. Thus, the altered CK-II compartmentalization might have significant consequences in the pathogenesis of AD.

Topics: Brain; Casein Kinase II; Cytoplasm; Humans; Immunoblotting; Immunohistochemistry; Intermediate Filaments; Nerve Degeneration; Nervous System Diseases; Neurofibrillary Tangles; Protein Serine-Threonine Kinases