title: Limb Remote Ischemic Postconditioning Reduces Ischemia-Reperfusion Injury by Inhibiting NADPH Oxidase Activation and MyD88-TRAF6-P38MAP-Kinase Pathway of Neutrophils.
abstract: Limb remote ischemic postconditioning (LRIP) has been confirmed to reduce the ischemia-reperfusion injury but its mechanisms are still not clear. This study clarified the mechanism of LRIP based on the nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase and Myeloid differentiation factor 88 (MyD88)-Tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6)-P38 pathway of neutrophils. Rat middle cerebral artery occlusion (MCAO) model was used in this study. Ischemia-reperfusion injury was carried out by MCAO 1.5 h followed by 24 h reperfusion. LRIP operation was performed to the left femoral artery at 0, 1 or 3 h after reperfusion. Behavioral testing, including postural reflex test, vibrissae-elicited forelimb placing test and tail hang test, showed that LRIP operated at 0 h of reperfusion could significantly ameliorate these behavioral scores. Pathological examinations, infarct size, Myeloperoxidase (MPO) activity showed that LRIP operated at 0 h of reperfusion could significantly ameliorate the pathological scores, reduce the infarct size and MPO activity in the brain and increase the MPO activity in the left leg. By using Neutrophil counting, immunofluorescence and real-time PCR techniques, we found that LRIP operated at 0 h of reperfusion could reduce neutrophil counts in the peripheral blood and downregulate the activation of neutrophil in the peripheral blood and rat brain. Western blots revealed that MyD88, TRAF6, p38 mitogen-activated protein kinase (p38-MAPK) in neutrophils and the phosphorylation of p47phox (Ser 304 and Ser 345) in neutrophil could be downregulated by LRIP. Our study suggests that LRIP inhibits the number and activation of neutrophils in the rat brain and peripheral blood linked to down-regulating the activation of NADPH oxidase in neutrophils by MyD88/TRAF6/p38-MAPK pathway.
authors.affiliation: Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. chengangling@cpu.edu.cn.|Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. chengangling@cpu.edu.cn.
authors.forename: Gangling
authors.initials: G
authors.lastname: Chen
authors.affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. yexinyi1991@126.com.
authors.forename: Xinyi
authors.initials: X
authors.lastname: Ye
authors.affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. 15651671978@163.com.
authors.forename: Jiangwei
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. 15651648635@163.com.
authors.forename: Tingli
authors.initials: T
authors.lastname: Tang
authors.affiliation: Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. 15850670086@163.com.
authors.forename: Lin
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Pharmacology of Chinese Materia Medica, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. lupeirong321@163.com.
authors.forename: Peirong
authors.initials: P
authors.lastname: Lu
authors.affiliation: State Key Laboratory of Natural Medicines, Research Department of Pharmacognosy, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. christina_576636@126.com.
authors.forename: Qi
authors.initials: Q
authors.lastname: Wu
authors.affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. boyangyu59@163.com.
authors.forename: Boyang
authors.initials: B
authors.lastname: Yu
authors.affiliation: Jiangsu Key Laboratory of TCM Evaluation and Translational Research, Department of Complex Prescription of TCM, China Pharmaceutical University, 639 Longmian Road, Nanjing 211198, China. junpingkou@cpu.edu.cn.
authors.forename: Junping
authors.initials: J
authors.lastname: Kou
mesh_terms: D000818:Animals; D002545:Brain Ischemia / Q000201:enzymology / Q000378:metabolism; D005121:Extremities / Q000098:blood supply*; D020244:Infarction, Middle Cerebral Artery / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D057775:Ischemic Postconditioning; D008297:Male; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism*; D019255:NADPH Oxidases / Q000378:metabolism*; D009504:Neutrophils / Q000201:enzymology / Q000378:metabolism*; D009195:Peroxidase / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction; D048029:TNF Receptor-Associated Factor 6 / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
chemical_list: D053594:Myeloid Differentiation Factor 88; D048029:TNF Receptor-Associated Factor 6; D009195:Peroxidase; D019255:NADPH Oxidases; D048051:p38 Mitogen-Activated Protein Kinases
keywords: MyD88/TRAF6/p38-MAPK pathway; NADPH oxidase; ischemic stroke; limb; neutrophil; reperfusion injury