黄连解毒汤有效部位抑制NogoA/NgR表达对脑缺血大鼠白质损伤的影响

目的观察黄连解毒汤有效部位对脑缺血大鼠白质损伤的影响,探讨黄连解毒汤有效部位对轴突修复抑制信号勿动蛋白A(NogoA)和勿动蛋白受体(Nogo receptor,NgR)的调控作用。方法将雄性SD大鼠随机分为假手术组、模型组、总生物碱组、总黄酮组、总环烯醚萜组。除假手术组外,其余各组大鼠采用线栓法建立大鼠大脑中动脉阻塞模型。造模后2 h灌胃给药,总生物碱组大鼠灌胃44 mg/kg总生物碱;总黄酮组大鼠灌胃50 mg/kg总黄酮;总环烯醚萜组大鼠灌胃80 mg/kg总环烯醚萜,假手术组、模型组灌胃等体积的生理盐水,1次/d,连续给药7 d。采用HE染色观察大鼠白质病理形态学改变,采用罗克沙尔坚牢蓝(Luxol fast blue,LFB)染色观察大鼠髓鞘的病理形态学改变,采用免疫组织化学染色法检测大脑内囊和外囊区域淀粉样前体蛋白(amyloid precursor protein,APP)、NogoA、NgR表达,采用实时荧光定量PCR技术检测缺血梗死灶周围组织NogoA、NgR基因表达。结果与模型组比较,总生物碱组、总黄酮组、总环烯醚萜组大鼠内囊和外囊区域病理损伤评分降低(P<0.05或P<0.01),LFB染色积分光密度值增高(P<0.01),APP、NogoA表达降低(P<0.01);总生物碱组和总环烯醚萜组大鼠内囊和外囊NgR表达降低(P<0.01或P<0.05),总黄酮组大鼠内囊NgR表达降低(P<0.01);总生物碱组、总黄酮组和总环烯醚萜组大鼠梗死灶周围组织NogoA(1.20±0.17)、(1.55±0.30)、(1.19±0.38)比(2.22±0.58)]和NgR(1.98±0.55)、(1.48±0.31)、(1.58±0.27)比(3.36±0.41)]基因表达降低(P<0.01)。结论黄连解毒汤有效部位可减轻脑缺血大鼠白质损伤,其作用机制可能与抑制NogoA/NgR表达有关。

Effect of the active fraction of Huanglian-Jiedu Decoction on white matter injury in rats with cerebral ischemia by inhibiting NogoA/NgR expression

Objective To study the effect of Huanglian-Jiedu Decoction on the white matter lesion of rats with focal cerebral ischemia and to explore the regulative role of the active fraction of Huanglian-Jiedu Decoction on NogoA/NgR.Methods Male SD rats were randomly divided into sham operation group,model group,total alkaloid group,total flavonoid group,and total iridoid group.Except for the sham operation group,the rats in the other groups were used to establish the middle cerebral artery occlusion rat model by the suture method.2 hours after modeling,rats in the total alkaloid group were given intragastric administration with 44 mg/kg total alkaloids;rats in the total flavonoid group were given intragastric administration 50 mg/kg total flavonoids;rats in the total iridoid group were given intragastric administration 80 mg/kg total iridoids,the sham operation group and the model group were intragastrically given equal volume of normal saline,once a day,for 7 consecutive days.The pathological changes of rat white matter were observed by HE staining,the pathological changes of rat myelin sheath were observed by Luxol fast blue(LFB)staining,and the expression of Amyloid precursor protein(APP),NogoA,and NgR in the internal and external capsule areas of the brain was detected by immunohistochemical staining.Real-time fluorescence quantitative PCR was used to detect the expression of NogoA and NgR in the tissues surrounding the ischemic infarct.Results Compared with the model group,the total alkaloid group,total flavonoid group,and total iridoid group had lower pathological damage scores in the internal and external capsule areas of rats(P<0.05 or P<0.01),increased integral optical density value of LFB staining(P<0.01),decreased expression of APP and NogoA;the expression of NgR in the internal and external capsules of rats in the total alkaloid group and the total iridoid group decreased(P<0.05 or P<0.01),and the expression of NgR in the inner capsule of rats in the total flavonoid group decreased(P<0.01);the expression of NogoA(1.20±0.17,1.55±0.30,1.19±0.38 vs.2.22±0.58)and NgR(1.98±0.55,1.48±0.31,1.58±0.27 vs.3.36±0.41)genes in the tissues around the infarct focus of rats in the total alkaloid group,total flavonoid group and total iridoid group decreased(P<0.01).Conclusion The present study investigates the therapeutic effects of Huanglian-Jiedu Decoction,promoting white matter repair by decreasing the overexpression of NogoA and NgR in an experimental animal model of stroke.