| BackgroundIschemic cerebrovascular disease as a clinical common disease, its highmorbidity and high mortality is a serious threat to human health.Aftercerebral ischemia with or against cerebral ischemia reperfusion injury,cerebrovascular generating mechanism was started, but the body of theresponse ability is very weak, it is not enough to improve vascularremodeling after cerebral ischemia and tissue perfusion status. Therefore,selecting effective drugs or other interventions to promote after ischemiacerebrovascular generated is particularly important. In recent years, throughthe in-depth study of ICVD pathophysiological mechanism, found in younganimals as the research object of conclusions far from the clinical actualICVD happens in the elderly population. Mechanism and treatment of ICVDtreatment of traditional Chinese medicine research has made some progress,but the angiogenesis after cerebral ischemia/reperfusion injury effects ofresearch are reported. Therefore, this topic on the basis of previous research,from p-Akt1and F Ⅷ R:Ag expression changes to explore aging inischemia/reperfusion rats brain microvascular generation mechanism, and asa breakthrough point to further interpretation of traditional Chinesemedicine compound cerebral vein through promoting effect ofmicrovascular generation and its possible mechanism.Objective1in SPF rats as subjects in the early success of the preparation ofsuture-based cerebral artery occlusion focal cerebral ischemia animalmodel, the observed aging and young rats with cerebral ischemia (I)3h usingtransmission electron microscopy reperfusion (I/R)24h,3d,6d,12d brain angiogenesis change; protective effect of Naomaitong on cerebral ischemicinjury.2uses to detect aging and young rats with cerebral ischemia (I)3hreperfusion(I/R)24h,3d,6d,12d brain microvessel p-Akt1and F Ⅷ R:Agimmunohistochemistry; protein expression; research Naomaitong aged rats I/R microvascular p-Akt1and FⅧ R:Ag expression of change.Method1Prepare a legal reference Longa improved cerebral artery occlusionanimal model of focal cerebral ischemia (MCAO). Selected260healthymale SD rats were5-6months old youth, aging20-21months of age, theyoung rats were divided into six youth group, young model group36(basedon ischemia/reperfusion divided into ischemic time3h, reperfusion24h,3d,6d,12d, at each time point6); aging aged control rats were divided into sixgroups, aging model group36(based on ischemia/reperfusion divided intoischemic time3h, reperfusion24h,3d,6d,12d, at each time point6).Depending on the medication, the aging model rats were divided intonimodipine group (time point and the number of animals ditto) andNaomaitong groups (each group time and number of animals ibid).Treatment group five days before modeling and modeling gavage duringnimodipine (6mg·kg-1·d-1) and Naomaitong (0.9g·kg-1·d-1).2The two groups of rats were anesthetized at predetermined time points,the thorax was opened, the temperature in0.9%saline,4%paraformaldehyde perfusion of the heart chamber, the rapid separation ofwhole brain after decapitation ice tray, remove the olfactory bulb,cerebellum and the brain stem, in prechiasmatic2.0mm office, coronal cutback,followed by2.0mm thickness of brain tissue4.1st cut slices of braintissue samples used for routine pathological observation.2,3for vascularendothelial cell apoptosis, microvessel density (MVD) and field areadetection.4slices with ice saline three times to remove blood clots and drywater, with4%paraformaldehyde, to be doing routine pathological andimmunohistochemical specimens. SEM specimens selected from the frontalpole4.0mm, the amount of the left cerebral cortex in the tissues at themidline4.0mm1mm3,2.5%glutaraldehyde fixed tissue, using phosphatebuffer (pH7.2), washed3times (each10min); fixed with1%osmium tetroxide for2h, and then rinsed with distilled water three times (10minuteseach); finally graded ethanol dehydration, epoxy four alkyl substitution,pure resin impregnated, embedded, and ultrathin sections made of lead-Ustaining by HITACHI-7500transmission electron microscope. Brain tissuequickly than cold storage in liquid nitrogen molecular biological indicatorsrelated spare tested.3Using the above method, from the p-Akt1and FⅧ R:Ag aspects ofangiogenesis after cerebral ischemia, based on the observation Naomaitongp-Akt1protein expression after cerebral ischemia in the role ofangiogenesis mechanisms.Result1A cerebral ischemic brain tissue in each group of rats electronmicroscopy young sham group showed no change, young model groupI3h,I/R1d,I/R6d visible light to moderate perivascular edema,edema andendothelial cell mitochondria have medullary bodies; perivascular I/R12dslightly swollen. Aging sham group had no significant change; aging modelgroup I3h, I/R1d capillaries surrounding edema and even more, thebasement membrane serious defect, microvilli and endocytosis bubblereduced endothelial cell mitochondria visible edema, and with see most ofthe ridge, film disappear; microvascular changes after I/R3d reach peakvalues, most basement membrane is dissolved, destroyed or disappeared;I/R6d will be gradually reduced to only slightly after I/R12d perivascularedema. Nimodipine group I3h normal microvilli, mitochondrial edema;I/R3d,I/R6d, the I/R1d dilated rough endoplasmic reticulum, andmitochondria height perivascular edema, crest disappeared, microvilli,endocytosis reduce the number of bubbles;perivascular to I/R12d after alittle swollen, mitochondria then see the degree of edema, crest mostlydisappeared.Naomaitong group I3h, I/R3d vascular no significantimprovement; mild perivascular edema I/R6d visible microvilli andendocytosis bubble decreased slightly basement membrane defects,mitochondrial swelling was significantly reduced; to I/R12d vascular ultra-microstructure returned to normal.2A small amount of p-Akt1weakly positive P-AKT1protein expressionin rat brain tissue seen in each group comparing sham-operated rats; model group I3h,I/R1d,3d expression in rat p-Akt1increased significantly(P<0.01), no significant difference between the model group6d p-Akt1expression in rats with sham group, with the first12d group significantlyincreased (P<0.01). And the aging model group compared with the sametime:the nimodipine group I3h,I/R1d,3d,6d and Naomaitong group of rats ateach time point p-Akt1expression levels were enhanced(P<0.01). Comparedwith the same time point nimodipine group:Naomaitong group I3h,I/Rp-Akt1expression level was significantly enhanced3d rats (P <0.01).Naomaitong group between groups:I/R1d, p-Akt1higher expression levelsin rat3d I3h,I/R3d reached its peak (P<0.01); I/R6d,p-Akt1expressionlevels of12d rats lower than the I/R1d,3d (P<0.05,P<0.01).3Each group rat brain F ⅧR:Ag protein expression in brain tissue of ratsin each group in immunohistochemical expression were seen in Ⅷ factor,but mainly expressed in microvascular endothelial cells.Sham group only afew FⅧ R:Ag expression; compared with the normal group, model group FⅧR:Ag I3h appears in decreased expression, I/R1d reached the lowest point,and then began to rise,to I/R3d,6d,12d large mouse p-Akt1expression wassignificantly higher (p<0.01); compared with the aging model group thesame point in time:nimodipine group and Naomaitong rats each time point FⅧ R:Ag expression were increased (P<0.01), there was significantdifference (P<0.05). Nimodipine group compared with the same timepoint:Naomaitong rats at each time point F Ⅷ R:Ag expression levelenhancement (P<0.01), the difference was significant(P<0.05). Naomaitonggroup between groups:I/R3d,6d rat FⅧ R:Ag expression levels higher thanI3h,I/R1d(P<0.01), the difference12d after cerebral ischemia withsignificant statistical significa-nce(P<0.01), and with prolonged treatmenttime pulse through the brain and its expression was significantly increased,and the effect was sustained.Conclusion1. With the increase of age, aging rats brain tissue ischemia reperfusioninjury in early, heavy, slow recovery, weakened microvascular generatedafter cerebral ischemia injury, brain microvascular number is relativelyyoung rats decreased significantly. This is likely to be senile ICVD heavierconditions, and the one of the pathophysiology of morbidity and mortality is higher.2. P-AKT1can be directly or indirectly regulate various substrate proteinfactor, promote angiogenesis. Model group p-Akt1protein expression werehigher than that of control group, ischemia reperfusion injury in rats can beinduced in brain tissue p-Akt1to express and its expression changes with aregularity reperfusion time.3. After cerebral ischemia, vascular endothelial cell damage is serious,leading to FⅧ R:the synthesis of Ag, accompanied by microvascular densityand the increase of the number of new blood vessels.And clinical usingimmunohistochemical method to detect displays FⅧ R:Ag only expressed onthe new blood vessels, and F Ⅷ R:Ag positive expression intensity isproportional to the number of angiogenesis.4. Access through the combination of traditional Chinese medicines basedon cerebral vein to p-Akt1,F Ⅷ R:Ag expression level adjustment, isconducive to the formation of new blood vessels and nerve function afterbrain ischemia repair; The cerebral vein group I/R1d,3d express clearly,I/R6d,12d expression is more significant. |
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