司美格鲁肽对脑缺血大鼠的治疗作用及机制

Therapeutic effects and mechanism of semaglutide on cerebral ischemia rats

目的观察胰高血糖素样肽1(GLP-1)受体激动剂司美格鲁肽对脑缺血大鼠的治疗作用,并探讨其作用机制。方法将72只大鼠随机分为假手术组、模型组、司美格鲁肽组、司美格鲁肽+LY294002组,每组18只。模型组、司美格鲁肽组、司美格鲁肽+LY294002组用线栓法建立大脑中动脉闭塞模型,假手术组分离结扎动脉但不插入线栓。司美格鲁肽+LY294002组造模前30 min经侧脑室注射10μL 10 mmol/L PI3K抑制剂LY294002,其余组给予同等剂量生理盐水。司美格鲁肽组及司美格鲁肽+LY294002组造模后即刻经腹腔注射司美格鲁肽溶液(10 nmol/kg),之后隔日空腹经腹腔注射司美格鲁肽溶液(10 nmol/kg),其余组注射等量生理盐水。分别于造模前及造模后1、3、5、7 d测量血糖;用Longa 5分制评分法对各组神经行为进行评价;TTC染色法观察脑梗死情况,计算脑梗死体积;TUNEL染色法检测缺血脑组织细胞凋亡情况;Western blotting法检测缺血脑组织中磷脂酰肌醇-3-激酶(PI3K)/蛋白激酶B(AKT)信号通路蛋白(PI3K、p-PI3K、AKT、p-AKT)及凋亡相关蛋白(Bcl-2、Bax、cleaved-Caspase-3)表达,并计算p-PI3K/PI3K、p-AKT/AKT;用实时荧光定量PCR法检测缺血脑组织p53 mRNA表达。结果造模前后各组血糖水平比较差异均无统计学意义(P均>0.05)。与假手术组比较,模型组神经行为学评分、脑梗死体积比、凋亡细胞百分比及缺血脑组织中Bax、cleaved-Caspase-3蛋白表达和p53 mRNA表达高(P均<0.05),p-PI3K/PI3K、p-AKT/AKT及Bcl-2蛋白表达低(P均<0.05);与模型组比较,司美格鲁肽组神经行为学评分、脑梗死体积比、凋亡细胞百分比及缺血脑组织中Bax、cleaved-Caspase-3蛋白表达和p53 mRNA表达低(P均<0.05),p-PI3K/PI3K、p-AKT/AKT及Bcl-2蛋白表达高(P均<0.05);与司美格鲁肽组比较,司美格鲁肽+LY294002组神经行为学评分、脑梗死体积比、凋亡细胞百分比及缺血脑组织中Bax、cleaved-Caspase-3蛋白表达和p53 mRNA表达高(P均<0.05),p-PI3K/PI3K、p-AKT/AKT及Bcl-2蛋白表达低(P均<0.05)。结论司美格鲁肽可改善脑缺血大鼠神经功能障碍,抑制神经细胞凋亡,减轻脑组织损伤;其作用机制可能与激活PI3K/AKT通路有关。

Objective To observe the effect of glucagon-like peptide 1(GLP-1)receptor agonist semaglutide on ce-rebral ischaemia rats and to investigate its mechanism of action.Methods Seventy-two rats were randomly divided into the sham-operated group,model group,semaglutide group,and semaglutide+LY294002 group,with 18 rats in each group.The middle cerebral artery occlusion model was established with a wire plug in the model group,the semaglutide group and the semaglutide+LY294002 group,and the rats in the sham-operated group were dissected and ligated in the same way but without inserting a wire plug.TenμL of 10 mmol/L PI3K inhibitor LY294002 was injected into the lateral ventricle at 30 min prior to modeling in the semaglutide+LY294002 group,and the rest of the group were given the same dose of normal saline.Rats in the semaglutide group and semaglutide+LY294002 group were injected with semaglutide so-lution(10 nmol/kg)intraperitoneally after modeling,followed by Semaglutide injection(10 nmol/kg)intraperitoneally on the next day,and equal volume of normal saline was injected in the rest of the groups.Blood glucose was measured before and 1,3,5 and 7 days after modeling.We evaluated the neurobehavior in each group using the Longa 5-point scale.Cere-bral infarction was observed by TTC staining,and infarct volume was calculated;we detected the apoptosis in ischemic brain tissues by TUNEL staining.PI3K/AKT pathway proteins PI3K,p-PI3K,AKT,and p-AKT expression and apopto-sis-related proteins Bcl-2,Bax,cleaved-Caspase-3 expression were detected by Western blotting,and p-PI3K/PI3K and p-AKT/AKT were calculated.P53 mRNA expression in the ischemic brain tissues was detected by real-time fluorescent quantitative PCR.Results There was no statistically significant difference in blood glucose levels between the pre-and post-modelling groups(all P>0.05).Compared with the sham-operated group,the model group showed higher neurobe-havioral scores,cerebral infarct volume ratio,percentage of apoptotic cells and expression of Bax,cleaved-Caspase-3 pro-tein and p53 mRNA in the ischemic brain tissues(all P<0.05),and lower expression levels of p-PI3K/PI3K,p-AKT/AKT and Bcl-2 protein(all P<0.05).In comparison with the model group,the smeaglutide group showed lower neurobe-havioral scores,cerebral infarct volume ratio,percentage of apoptotic cells and expression levels of Bax,cleaved-Caspase-3 protein and p53 mRNA in ischemic brain tissue(all P<0.05),and higher expression levels of p-PI3K/PI3K,p-AKT/AKT and Bcl-2 protein(all P<0.05).Compared with the semaglutide group,the semaglutide+LY294002 group exhibit-ed higher neurobehavioral scores,cerebral infarct volume ratio,percentage of apoptotic cells and expression levels of Bax,cleaved-Caspase-3 protein and p53 mRNA in the ischemic brain tissues(all P<0.05)and lower expression levels of p-PI3K/PI3K,p-AKT/AKT,and Bcl-2 protein(all P<0.05).Conclusion Semeglutide improves neurological deficits,inhibits the neuronal apoptosis,and reduces brain tissue damage in cerebral ischemia rats,and the mechanism of action may be related to the activation of the PI3K/AKT pathway.