| ObjectiveUsing MCAO model to observe protective effect of bezoar active components on brain damage and the influence of endoplasmic reticulum stress (endoplasmic reticulin stress, ERS) related pathway.From the endoplasmic reticulum stress intervention angle discuss its anti-cerebral ischemia mechanism.MethodsIThe rats were stratified randomly divided sham group,model group,qingkailing group,taurine group,hyodeoxycholic group,ursodeoxycholic acid group,hyodeoxycholic acid group,taurine-conjugated ursodeoxycholic acid group.Through establish MCAO model in rats,to observe the scores of the neurologic impairment,to measure infarct volume by TTC,the morphological changes of glial cells were observed by HE staining,and the specific conditions of the nissl were observed by the method of nissl staining.2The rats were stratified randomly divided sham group,model group,qingkailing group,taurine group,ursodeoxycholic acid group,hyodeoxycholic acid group,taurine-conjugated ursodeoxycholic acid group.The ultrastructure of endoplasmic reticulum was observed by transmission electron microscope.Detecting ischemic brain tissue and endoplasmic reticulum stress-associated antibodies (P-PERK,P-EIF2α,ATF4,CHOP,caspase-12,GRP78) expression,from the endoplasmic reticulum stress intervention angle reveal bezoar active ingredient in anti-cerebral ischemia in rats.ResultsNeurological functionThe sham group had no neurologic impairment.After cerebral ischemia 24 hours,the majority of rats showed when walking on the left side hemiplegia,rotated to the left circle or dumping.When hanging tail,forelimb symmetry,left forelimb flexion,adduction, close to the chest wall,ipsilateral no response,contralateral reduced reactivity.Compared with sham group,the model group significantly reduced the score(P<0.01).Compared with the model group,the Qingkailing group and TUDCA group had significantly improved neurological function in rats(P<0.01),UDCA group also improve neurological function(P<0.05).At the same time,taurine group and hyodeoxycholic acid group also can improve neurological score,but there was no statistical difference(P>0.05).TTC stainingIn the sham-operated group,the TTC staining of the brain tissue showed uniform red color. Compared with the sham group,the model group rats formed obvious focal ischemia(P<0.01).Compared with the model group,the treatment group could reduce the volume of cerebral infarction(P<0.01).Compared with the TUDCA group, taurine group and UDCA group can significantly enlarged brain infarct volume(P<0.01). The area of infarction:model group>HDCA group>UDCA group>Taurine group>TUDC A group>Qingkailing group,and HDCA group,the mortality rate is high,so the next experiment,we mainly studied the positive medicine Qingkailing, taurine,UDCA,TUDCA on brain MCAO rats.HE stainingIn the sham group,the neuronal cells and glial cells were arranged in order,the chromatin was distributed evenly,the nucleolus was clear, and the model group saw a large area of infarction,which was a liquefaction necrosis,a screen mesh structure,a large number of nerve cell disappeared and small glial cells hperplasia.Necrosis margin was narrow,and there was a lot neutrophil infiltration.In the taurine group,the area of the infarct area was moderately reduced,and there was a number of small glial cells in thesoftening range.The edge of the group was widened,Nerve cell degeneration, shrinkage,triangular,nuclear stained.UDCA group,infarction area with moderate reduction, showing extensive neuronal degeneration,cell shrinkage,small mild gliosis,marginal zone broadening,visible red neurons cells degeneration,vascular dilation.TUDCA significantly reduced infarct area,marginal zone widened,reducing the number of neurons is not obvious,morecellular degeneration,reduce the degree of degeneration,cell shrinkage, microglia mild hyperplasia. Qingkailing group significantly reduced infarct area, marginal zone was widened,slightly reduced the number of neurons,showed mild degeneration of neurons,microglia proliferation was not obvious.Nissl stainingIn the sham group,nerve cells arranged in order,no edema,soma,nucleolus clear nissl common.In the model group,nissl significantly reduced,and some of the nerve cells were contracted and stained. After drug intervention,reducing infarct volume.The number of neurons in the structure is more,and the range of degeneration and necrosis is relatively small.The effect of Qingkailing and TUDCA is the most obvious,the injury area has preserved more complete structure of neurons.Endoplasmic reticulum ultrastructureIn the model group,endoplasmic reticulum flat vesicular structures disappear,swelling significantly,showing a small blister-like,disorganized,and reduce the number of ribosomes.Endoplasmic reticulum stress-related antibody expressionIn the model of cerebral ischemia reperfusion injury,it is proved that cerebral ischemia reperfusion injury can induce endoplasmic reticulum stress and activate PERK /EIF2a/ATF4 pathway by immunohistochemistry and western detection.On the one han d,the high expression of GRP78 can promote the repair of the endoplasmic reticulum,o n the other hand,the high expression of CHOP,caspase-12 can initiate cell damage pathway.After the intervention of qingkailing,taurine,ursodeoxycholic acid and taurine-conjugated ursodeoxycholic acid can increase the expression of GRP78,and decrease the expression of caspase-12 and CHOP to promote the recovery of endoplasmic reticulum,reduce the damage to the cells.Conclusions1 Qingkailing,taurine,ursodeoxycholic acid,hyodeoxycholic acid and taurine-conjugated ursodeoxycholic acid can reduce the volume of cerebral infarction,improve neurological function score and improve the pathological damage of brain tissue, including Qingkailing,taurine-conjugated ursodeoxycholic acid effect is the most obvious and taurine-conjugated ursodeoxycholic acid effect is superior to that of taurine and ursodeoxycholic acid,hyodeoxycholic acid.2 In MCAO rats,I/R induced ERS and activated the unfolded protein response PERK/EIF2a/ATF4 pathways, suggesting bezoar active ingredients of has a protective effects.3 Cerebral ischemia-reperfusion injury after 24h,significantly increased expression of GRP78,initiated protection mechanisms of ERS,after administration of the intervention to further promote the expression of GRP78,reducing the ERS,which plays a neuroprotective role, especially with Qingkailing group and TUDCA group effect is most obvious.4 Cerebral ischemia-reperfusion injury after 24h,CHOP,caspase-12 expression increased, indicating a long ERS initiated apoptotic pathway cells,whereas positive drug group and TUDCA Qingkailing group decreased hippocampal cortex in rats CHOP the expression of caspase-12,significantly inhibited cell apoptosis,and plays a role in the protection of brain injury. |
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