Inhibition of myosin light chain kinase reduces NADPH oxidase-mediated oxidative injury in rat brain following cerebral ischemia/reperfusion.
Previous studies have demonstrated that nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX)-mediated oxidative stress plays a key role in brain injury following cerebral ischemia/reperfusion (I/R) and myosin regulatory light chain kinase (MLCK) has been reported to be involved in NOX activation in lung endothelium. This study was performed to explore the correlation between MLCK and NOX following cerebral I/R and the underlying mechanisms. Sprague-Dawley (SD) rats were subjected to 2 h middle cerebral artery occlusion and 24 h reperfusion to establish a model of focal cerebral I/R injury. At the end of experiments, neurological function, infarct volume, cellular apoptosis, activities of MLCK and NOX, messenger RNA (mRNA) and protein expression of NOX (NOX1-NOX4), phosphorylation level of myosin regulatory light chain (MLC20) and hydrogen peroxide (H2O2) level were determined. The results showed that I/R treatment led to increase in neurological deficit score, infarct volume and cellular apoptosis, accompanied by the elevated activities of MLCK and NOX, expressions of NOX2 and NOX4, levels of phosphorylation MLC20 and H2O2, these effects were attenuated by MLCK specific inhibitor (ML-7). NOX inhibitors (diphenylene iodonium (DPI) or apocynin) were able to achieve similar results to that of ML-7 except no effect on MLCK activity and MLC20 phosphorylation. These results suggest that activation of MLCK contributes to cerebral I/R oxidative injury through upregulation of NOX2 and NOX4 expression, which is involved in phosphorylation of MLC20.
Department of Pharmacology, School of Pharmaceutical Sciences, Central South University, Changsha, 410078, China.
Hong-Feng
HF
Zhang
Ting-Bo
TB
Li
Bin
B
Liu
Zheng
Z
Lou
Jie-Jie
JJ
Zhang
Jing-Jie
JJ
Peng
Xiao-Jie
XJ
Zhang
Qi-Lin
QL
Ma
Jun
J
Peng
Xiu-Ju
XJ
Luo
D000818:Animals; D001381:Azepines / Q000494:pharmacology; D001921:Brain / Q000503:physiopathology; D002545:Brain Ischemia / Q000503:physiopathology* / Q000517:prevention & control; D004195:Disease Models, Animal; D006861:Hydrogen Peroxide / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics; D018994:Myosin Light Chains / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D009281:Naphthalenes / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D010766:Phosphorylation / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000503:physiopathology* / Q000517:prevention & control; D015854:Up-Regulation
D001381:Azepines; D008562:Membrane Glycoproteins; D018994:Myosin Light Chains; D009281:Naphthalenes; C070571:ML 7; D006861:Hydrogen Peroxide; C500593:Cybb protein, rat; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498911:Nox4 protein, rat; D009219:Myosin-Light-Chain Kinase