Study On The Intervention Effect On Animal Models Of Cerebral Ischemia Tolerance Of Total Flavonoids From Rabdosia Rubescens

Objective: By establishing rat and mouse brain ischemic tolerance model to study theprotective effect of total flavonoids from Rabdosia rubescens on rat and mouse cerebralischemia tolerance model, and by determining contents of neuron specific enolase (NSE) inserum, inflammatory cytokine-nitric oxide (NO), nitric oxide synthase (NOS), tumornecrosis factor TNF-α, IL-1β，IL-8，apoptosis related protein B cell lymphoma factor2(Bcl-2) and its associated X protein (Bax), cysteine proteinase3(Caspase-3); Observed theexpression of nuclear transcription factor NF-κBp65and neurotrophic factors GDNF、BDNF;calculated cerebral infarction area of model;observed the change of brain tissuemorphology,ect,so as to investigate the protective effect of total flavonoids from Rabdosiarubescenson brain ischemic tolerance model.Methods: By Blocking bilateral common carotid artery blood flow for10min and restoringperfusion for120h later, blocking bilateral common carotid artery for30min again andreperfusing for22h, established cerebral ischemic tolerance model of mouse. After BITmodel group mice preconditioned with transient ischemic for10min,Nimodipine group,Naoluotong capsule group, rubescens total flavonoids in each dose (300mg/kg,150mg/kg,75mg/kg) groups were given corresponding drugs by gavage;Sham operation group,ischemia reperfusion group, BIT model group were fed with the same volume of0.5%sodium carboxymethyl cellulose (CMC),1time a day, continuous administration for5days;after administrated1h in the fifth day(120h)from modeling, except the sham operationgroup, the rest of the groups of mice respectively blocked blood flow for30min, thenreperfused for22h, detected serum NSE levels in serum and changes of NO content andNOS activity in brain organization, observed the pathological morphological changes inbrain tissue.By using the method of shortly blocking common carotid artery for10min aspretreatment time and restoring perfusion for72h later, used suture method to block the leftside of the middle cerebral artery for2h, reperfused for22h, which is named2VO+MCAO,to established ischemia tolerance model of rat. After BIT model group rats preconditioned with transient ischemic for10min,Nimodipine group, Naoluotong capsule group, rubescenstotal flavonoids in each dose (200mg/kg and100mg/kg,50mg/kg) groups were givencorresponding drugs by gavage;Sham operation group, ischemia reperfusion group, BITmodel group were fed with the same volume of0.5%sodium carboxymethyl cellulose(CMC),1time a day, continuous administration for3days;after administrated1hlater,except the sham operation group,the rest of the groups all were performed MCAOsurgery, after22h reperfusion, assessed rats’ nerve disfunction,measured the NSE level inserum, calculated infarction area by brain tissue TTC staining, determinated levels of TNF-α,Bcl-2, Bax, Casp-3in brain homogenate,determinated the contents of TNF-α、IL-1β、IL-8、Bcl-2、Bax、Casp-3in brain homogenate, Conventional HE staining to observe the changeof brain tissue pathologic morphology and immunohistochemical method to observe theGDNF, BDNF and NF-κBp65proteins’ expressions.Results:Ischemia tolerance model of mice and rats were both copied successfully.Comparedwith sham operation group and ischemia-reperfusion group, each indicator of models ofmice and rats had differences.Total flavonoids from Rabdosia rubescens can reduce themortality in both mice and rats;significantly reduced rat neurological deficit score, reducedthe cerebral infarction area of rats;can significantly reduce the serum NSE levels of mice andrats;can improve NO content and NOS activity in brain tissue in mice; can increase contentsof TNF-α、IL-1βand Bcl-2in rat brain tissue and decrease levels of IL-8、Bax and Casp-3of rats, can increase expressions of neurotrophic factors GDNF, BDNF protein, reduce theexpression of NF-κBp65protein; Improve the cortex and hippocampus pathological damagein mice and rats brain tissues.Conclusion: Total flavonoids from Rabdosia rubescens can induce low concentration of NO,NOS, TNF-α, IL-1βto stimulate the body to produce endogenous protective mechanism,improve neurons’ ability to resist damage; increased the Bcl-2protein content in braintissue;reduced the Bcl-2related proteins Bax levels; reduced the NF-κBp65proteinexpression,inhibited cysteine proteinase3(Casp-3) activity and neurons apoptosis; increaseexpressions of neurotrophic factors BDNF, GDNF proteins in brain tissue,reduced serumNSE release to protect neurons; improved neurological function score, reduced theinfarction area. All that stated dong ling grass flavonoids had obvious protective effect onneurons;further improved the protective effect of ischemic preconditioning on brain injury.