Studies On The Effect Of Naoshuning On Nerve Regeneration In Ischemic Stroke And Its Mechanism

Objective:To investigate the effects of Naoshuning,a traditional Chinese medicine,on nerve regeneration in acute ischemic stroke and the possible molecular mechanism of its effect on nerve regeneration.In order to enrich the basic theory of Chinese medicine treatment knowledge,and provide a new treatment for ischemic stroke.Methods:Sorting out the research progress of plant ecological characteristics and pharmacology of"Jun" medicine Leonurus artemisia in Naoshuning by literature research;As a research object,rat embryonic neural stem cells(NSCs)were isolated,cultured and identified in vitro,the effects of Nao-shuning(21.4 mg/mL?2.14 mg/mL?214 ?g/mL?21.4 ?g/mL?2.14 ?g/mL?0)on NSCs were measured by WST-1 method after 24 h and 48 h intervention with gradient concentration of Naoshuning respectively;NSCs were treated with 50 ?M inhibitor U0126 and LY294002 for 6 h,and then Naoshuning(21.4?g/mL)was added into the medium for 24 h,the effects of Naoshuning on the injury model of NSCS were examined by WST-1 and Image J nerve ball counting methods;U0126 and LY294002 were used to prepare the models of MAPK/ERK and PI3K/Akt signal pathway blocked in NSCs,and then the models were treated with Naoshuning(21.4 ?g/mL),the expression of p-ERK1/2,p-Raf-1 Ser259 protein in MAPK/ERK signal pathway and the expression of p-Akt protein in Pi3k/Akt signal pathway were detected by western blot.Results:Sort out a total of 157 compounds including alkaloids,diterpenes,flavonoids,etc.in Leonurus artemisia;Compared with the control group,Naoshuning could promote the proliferation of NSCs in a certain concentration range,while the high concentration of Naoshuning had cytotoxicity and inhibited the proliferation of NSCS,the Naoshuning treatment of NSCs at 21.4 ?g/mL for 24 h could significantly promote the proliferation of NSCs(p<0.05);The concentration of 21.4?g/mL Naoshuning significantly improved the injury of NSCs induced by U0126 and LY294002(p<0.05);Western Blot showed,compared with the control group,the expression of p-ERK1/2 protein was significantly increased in the Naoshuning-treated group(p<0.05),but p-Akt protein was not changed,the Naoshuning promoted the expression of p-Akt protein and p-Raf-1 Ser259 protein in total protein Raf-1 in the MAPK/ERK signal pathway blocking model(p<0.05),the Naoshuning promoted the expression of p-ERK1/2 protein(p<0.05),but the expression level of p-Raf-1 Ser259 protein in total protein Raf-1 decreased in the PI3K/Akt signal pathway blocking model(p<0.05).Conclusion:The proliferation of NSCs could be promoted by appropriate concentration and time treatment of Naoshuning,which may be realized by activating ERK1/2 protein in MAPK/ERK signal pathway;The possible mechanism of the effect of Naoshuning on nerve regeneration in ischemic stroke is:during ischemic,MAPK/ERK signal pathway was activated by inhibiting the phosphorylation of Ser259 site of Raf-1 and stimulating the phosphorylation of ERK1/2,during reperfusion,the inhibition of Akt in PI3K/Akt signaling pathway was alleviated by activating Akt and inhibiting phosphorylation of Raf-1.