| Cerebral ischemic stroke is the pathological change in braintissue caused by focal cerebral blood flow broken off and hypoxia.This disease has high incidence rate, mortality, deformity rate,recrudesce rate. It is the main reason for death and deformity ofhuman being. This disease make heavy burden for our society.Nowadays people pay more and more attention to the prevention andcure of cerebral ischemic stroke. Focus of cerebral ischemic strokeincluding two parts, infarction core and ischemia penumbra.Angiogenesis occurs after cerebral ischemia, and the extent ofangiogenesis has been correlated with survival in stroke patients. It isthe common standpoint that retrieval of the neuron in ischemiapenumbra through angiogenesis is the key to cure cerebral ischemicstroke. But now it seems that there is no effective medicine ortherapeutics for this disease. Therapeutic angiogenesis, reported recentyears, is a kind of the therapeutics to make vessels develope intosmaller capillary vessels, promoting the process of revascularizationvia sprouting and intussusception to improve the blood flow by usingsome pharmacological or genic embedding therapy. Maybe this kindof therapeutics will become a new effective way to cure cerebralischemic stroke. This study was undertaken to investigate whether the expressionof angiopoietin-1, -2mRNA changed or not in the ischemic brain andtried to find the regulation. We discuss the mechanism ofangiopoietin-1 and angiopoietin-2 acting on vessels, speculating theirfunction after cerebral ischemic stroke. Our research will provide newscientific theory for this kind of therapeutics-therapeutic angiogenesis.Angiopoietin belong to a novel specific vessel growth factoracting on vascular endothelial cells that as ligand for theendothelial-specific receptor tyrosine kinase, Tie. Ang-1 and Ang-2play important roles in process of angiogenesis. They aredistinguished by theirspecificity for endothelial cells with theircommon receptor Tie-2 expressing primarily on endothelial cells.Ang-1 and Ang-2 have similar constructure, 60% is the same. LikeAng-1, Ang-2 has an NH2-terminal coiled-coil domain and aCOOH-terminal fibrinogen-like domain. Eigth of the nine cysteines inmature Ang-1 are conversed in mature Ang-2;one cysteine betweenthe coiled-coil and fibrinogen-like domains in Ang-1 is absent fromAng-2. In adult Ang-1 is widely expressed on smooth muscle cells,pericytes and tumor cells. Ang-2 is expressed primarily on endothelialcells. Alought Ang-1 and Ang-2 has similar constructure, but Ang-2 isnaturally occurring endogenous antagonist of the action of Ang-1 atTie-2. The main reason for this maybe their fibrinogen-like domainsconnecting with Tie-2 have different function. By combinating withTie-2, Ang-1 make Tie-2 phosphorylate, and show some complicatedphysiological function by series of signal transduction. Angiopoietin-1appears to be required for communication of endothelial cells with thesurrounding mesenchyme in order to establish stable celluar andbiochemical interactions. Ang-1 reduces endothelial permeability ofvessels and has a major role in vascular stabilization and maturation,on the other hand, there is experiment proved Ang-1 can restrainneuron apoptosis, mostly neuron apoptosis was detected in ischemicpenumbra, whereas Ang-2 is thought to be an endogenous antagonistof the action of Ang-1 at Tie-2. Ang-2 bind but does not activate Tie-2,Ang-2 might block Ang-1 activation of Tie-2,leading vessels tounstabilization. Ang-1, Ang-2 and other specific vessel growth factors,like VEGF, regulate the process of vessels formation in different phase.Positive or negative regulation of Ang-1 and Ang-2 is likely to resultin different outcomes depenging on the combination of simultaneouslyacting angiogenic signals. For example, although Ang-1 may providea maturation or stabilization signal through Tie-2 that can be blockedby Ang-2, such inhibition may result in continued remodeling or theinitiation of vascular sprouting in the context of simultaneous VEGFexposure but may result in frank vessel regression in the absence ofVEGF. Foreign research speculated that In ischemic and hypoxiacondition, angiopoietin act on vascular endothelial cells, smoothmuscle cells, pericytes regulating their interaction, promoting vesselsto sprout and remodel, enhanceing the cerebral blood flow in ischemicfocus.In our experiment we detected the altered expression ofangiopoietin-1, angiopoietin-2 at the mRNA level in MCAO model,and in rats by RT-PCR analysis respectively, during a time course of1hours to 3 days after onset of cerebral ischemia. all experiments wereperformed in male Wistar rats, 3-4 months old, weighing 250-280g.Animals were housed under standard conditions with free access torats to chow and tap water before and after surgery. All the rats weredivded into 3 groups: control group (group C), sham-operated group(group S), and ischemia group (group I). Irreversible occlusion of theright middle cerebral artery was performed by ameliorated methods,which based on the methods of Zealonga's and Nagasawa's reports.After middle cerebral artery occlusion (MCAO), the animals survivedfor 1 hours, 3 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours(n=7 per time). Thereafter the animals were decapitated under deepanesthesia, brains were removed within 5 minutes after decapitated,frozen in -80 ℃ . We chosed the infarction core, ischemic penumbraarea in surgery animals and corresponding area in control animals andsham-operated animals to detect angiopoietin-1 mRNA andangiopoietin-2 mRNA in brain. The mRNA levels of Ang-1 and Ang-2were measured by reverse transcription polymerase chain reaction(RT-PCR).Data from our experiment was compute by statistical softwareSPSS in computer, with its result written in X ±s. Means of eachgroup were compared by ANVOA program. Means of sham-operatedgroup and ischemia group compared with control group by Dunett tmethod.Result: normal mRNA levels of Ang-1 were constitutivelyexpressed in the brain of control animals and sham-operated animals(group N 0.88±0.03;group S 0.87±0.05. group S compared with groupN, p>0.05, revealed no significant change). During 3~6 hours afteronset of ischemia, mRNA levels of Ang-1 went down to lower level inthe infarction core and ischemic penumbra (I3h 0.62±0.08;I6h0.60±0.03, compared with group N, I3h and I6h, P<0.01, revealed asignificant decrease). 48hours to 3 days after MCAO, high mRNAlevels of Ang-1 were detected in the infarction core and ischemicpenumbra (I48h 1.23±0.16;I72h 1.29±0.10, compared with group C, I48hand I72h, P<0.01, revealed a significant increase). Extreme low mRNAlevels of Ang-2 expressed in the brain of control animals andsham-operated animals (group C 0.26±0.07;group S 0.25±0.05. groupS compared with group N, p>0.05, revealed no significant change).After MCAO, the expressions of Ang-2 were dramatically increased ininfarction core and ischemic penumbra. The increase started at 3 hoursafter onset of ischemia, peaking at 12 hours, and remain elevated forup to 72 hours after ischemia (I3h 0.60±0.06;I12h 1.31±0.05, I72h1.18±0.05 compared with group C, I3h, I12h and I72h, P<0.01, revealed asignificant increase). Uniting current theory and our experiment result,we speculate that early time postischemia, expression of Ang-1 wentdown, whereas expression of Ang-2 elevated, maybe these coursewould lead to the vessels' breakage in the infarction core, and makethe ischemic lesion more serious. But the coming increased expressionof Ang-1, Ang-2 and VEGF, interacting with each other, regulated newvessels' formation in temporal and dimensional, enhancing blood flowin ischemic penumbra, promoting nerve function to recover.From our study, we can make conclusions: 1. we have made theintraluminal thread occlusion model in the middle cerebral artery ofrats. 2. focal cerebral ischemia lead to altered expression ofAng-1mRNA in onfarction core and ischemia penumbra. During 1~72hours, expression of Ang-1mRNA fluctuated, declined in early time,elevated to higher level than normal later. 3. focal cerebral ischemialead to ascending expression of Ang-2mRNA. 4. Ang-1, Ang-2 andother specific vessel growth factors may regulate the process ofvessels formation together after focal cerebral ischemia, enhancingblood flow restraining neuron apoptosis in ischemic penumbra,promoting nerve function to recover.Coming up with the progress of our science and update oftechnology, therapeutic angiogenesis may become a new rational,effective way to cure cerebral ischemic stroke in the future. Our studyprovided some positive evidence for it. |
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