异泽兰黄素改善蛛网膜下腔出血模型大鼠学习记忆能力的机制

Mechanism by which eupatilin improves learning and memory abilities in rats with subarachnoid hemorrhage

背景:异泽兰黄素是一种源自青蒿属的黄酮类活性成分,可缓解蛛网膜下腔出血大鼠的炎症反应,改善神经学评分,但其在学习记忆方面的作用和机制仍不清楚。目的:探究异泽兰黄素对蛛网膜下腔出血模型大鼠学习记忆能力及P38有丝分裂素激活蛋白激酶(p38 MAPK)/信号传导和转录活化因子3(STAT3)通路蛋白的影响。方法:采用随机数字表法将50只SD大鼠随机分为假手术组、模型组、异泽兰黄素组、橙皮素组、异泽兰黄素+橙皮素组,每组10只。除假手术组外,其余4组通过血管内穿孔构建蛛网膜下腔出血模型,造模成功2 h后,异泽兰黄素组尾静脉注射10 mg/kg异泽兰黄素,橙皮素组尾静脉注射50 mg/kg橙皮素(p38 MAPK/STAT3信号通路激活剂),异泽兰黄素+橙皮素组尾静脉注射10 mg/kg异泽兰黄素+50 mg/kg橙皮素,假手术组与模型组尾静脉注射10 mL/kg生理盐水。药物治疗24 h后,采用神经功能评分及Morris水迷宫实验检测大鼠神经功能及学习记忆能力,苏木精-伊红染色检测海马组织病理学变化,TUNEL法检测神经细胞凋亡,免疫荧光染色检测海马组织双皮质素阳性细胞数,Western blot检测海马组织p38 MAPK/STAT3蛋白表达。结果与结论:①与假手术组比较,模型组大鼠神经功能评分、学习记忆能力、双皮质素阳性细胞数均降低(P<0.05),神经细胞凋亡率及p-p38 MAPK/p38 MAPK、p-STAT3/STAT3蛋白表达均升高(P<0.05);②与模型组比较,异泽兰黄素组大鼠神经功能评分、学习记忆能力、双皮质素阳性细胞数均升高(P<0.05),神经细胞凋亡率及p-p38 MAPK/p38 MAPK、p-STAT3/STAT3蛋白表达均降低(P<0.05);橙皮素组大鼠神经功能评分、学习记忆能力、双皮质素阳性细胞数均降低(P<0.05),神经细胞凋亡率及p-p38 MAPK/p38 MAPK、p-STAT3/STAT3蛋白表达均升高(P<0.05);③与异泽兰黄素组比较,异泽兰黄素+橙皮素组大鼠神经功能评分、学习记忆能力、双皮质素阳性细胞数均降低(P<0.05),神经细胞凋亡率及p-p38 MAPK/p38 MAPK、p-STAT3/STAT3蛋白表达均升高(P<0.05);④苏木精-伊红染色显示,相较于模型组,异泽兰黄素组神经细胞排列较为整齐,橙皮素组神经细胞排列紊乱,异泽兰黄素+橙皮素组神经细胞排列与模型组相似;⑤结果表明,异泽兰黄素可能通过抑制p38 MAPK/STAT3信号通路改善蛛网膜下腔出血模型大鼠的学习记忆能力。

BACKGROUND:Eupatilin,a flavonoid active component derived from Artemisia sinensis,has been reported to relieve inflammation and improve neurological scores in rats with subarachnoid hemorrhage,but its role and mechanism in learning and memory remain unclear.OBJECTIVE:To investigate the effects of eupatilin on learning and memory abilities and P38 mitogen activated protein kinase(p38 MAPK)/signal transducer and activator of transcription-3(STAT3)pathway proteins in rats with subarachnoid hemorrhage.METHODS:A total of 50 Sprague-Dawley rats were randomly divided into sham surgery group,model group,eupatilin group,hesperetin group,eupatilin+hesperetin group,with 10 rats in each group.Except for the sham surgery group,the rats in the other groups were used to construct a subarachnoid hemorrhage model through intravascular perforation.Two hours after successful modeling,the eupatilin group was injected with 10 mg/kg eupatilin via the tail vein,the hesperetin group was injected with 50 mg/kg hesperetin(p38 MAPK/STAT3 signaling pathway activator)via the tail vein,the eupatilin+hesperetin group was injected with 10 mg/kg eupatilin and 50 mg/kg hesperetin via the tail vein,and the sham surgery group and the model group were injected with 10 mL/kg saline via the tail vein.The drug treatment lasted for 24 hours.The neurologic deficit score and Morris water maze experiment were applied to detect the neurological function and learning and memory abilities of rats.Hematoxylin-eosin staining was performed to detect the histopathological changes of the hippocampus.TUNEL method was used to detect neuronal apoptosis.Immunohistochemical staining was conducted to detect the number of doublecortinpositive cells in hippocampal tissue.Western blot was applied to detect the expression of p38 MAPK/STAT3 protein in hippocampal tissue.RESULTS AND CONCLUSION:Compared with the sham surgery group,rats in the model group had lower neurological deficit scores,learning and memory abilities,and number of doublecortin-positive cells(P<0.05),and higher neuronal apoptosis rate and protein expression of p-p38 MAPK/p38 MAPK and p-STAT3/STAT3(P<0.05).Compared with the model group,rats in the eupatilin group showed higher neurological deficit scores,learning and memory abilities,and number of doublecortin-positive cells(P<0.05),and lower neuronal apoptosis rate and protein expression of p-p38 MAPK/p38 MAPK and p-STAT3/STAT3(P<0.05),while those in the nerolone group showed lower neurological deficit scores,learning and memory abilities,and number of doublecortin-positive cells(P<0.05),and higher neuronal apoptosis rate and protein expression of p-p38 MAPK/p38 MAPK and p-STAT3/STAT3(P<0.05).Compared with the eupatilin group,rats in the eupatilin+hesperetin group had lower neurological deficit scores,learning memory abilities,and number of doublecortin-positive cells(P<0.05),and higher neuronal apoptosis rate and protein expression of p-p38 MAPK/p38 MAPK and p-STAT3/STAT3(P<0.05).Hematoxylin-eosin staining showed that compared with the model group,the nerve cells were more neatly arranged in the eupatilin group,disorganized in the hesperetin group,and arranged in a similar way to the model group in the eupatilin+hesperetin group.To conclude,eupatilin may improve learning and memory abilities of rats with subarachnoid hemorrhage by inhibiting the p38 MAPK/STAT3 signaling pathway.