摘要
目的 探讨胡黄连苷Ⅱ(PicrosideⅡ,PⅡ)对脊髓损伤(spinal cord injury,SCI)大鼠神经功能恢复的影响及对环磷酸腺苷(cAMP)-蛋白激酶A(PKA)信号通路的调节作用.方法 建立SCI大鼠模型,大鼠分为正常组(CT组)、SCI模型组(SCI组)、PⅡ低剂量组(PⅡ L组,5 mg·kg-1·d-1)、PⅡ高剂量组(PⅡ H组,20 mg·kg-1·d-1)和PⅡ高剂量+PKA抑制剂组(PⅡ H+H-89 组,20 mg·kg-1·d-1 PⅡ+5 mg·kg-1·d-1),每组 18 只.Basso-Beattie-Bresnahan(BBB)评分评价SCI大鼠运动功能,HE染色评价脊髓组织病理学特征,劳克坚牢蓝(LFB)染色观察脱髓鞘情况,免疫荧光检测胶质纤维酸性蛋白(GFAP)、离子钙结合适配器分子-1(IBA-1)表达,ELISA检测丙二醛(MDA)、超氧化物歧化酶(SOD)、环腺苷酸(cAMP)的含量,Western blot 检测磷酸化(p)-PKA、PKA、p-环磷酸腺苷反应成分结合蛋白(CREB)、CREB蛋白表达.结果 与CT组比较,SCI组大鼠脊髓组织缺损、空腔,大量炎性细胞浸润,脱髓鞘以及GFAP、IBA-1、MDA表达增加,BBB评分以及SOD、cAMP、p-PKA/PKA、p-CREB/CREB表达减少(P<0.05);与SCI组比较,PⅡ L组、PⅡ H组组织损伤改善,空腔及炎性细胞减少,脱髓鞘以及GFAP、IBA-1、MDA表达降低,BBB评分以及SOD、cAMP、p-PKA/PKA、p-CREB/CREB表达增加(P<0.05);与PⅡ H组比较,PⅡ H+H-89 组组织损伤严重,脱髓鞘以及GFAP、IBA-1、MDA表达增加,BBB评分以及SOD、cAMP、p-PKA/PKA、p-CREB/CREB表达减少(P<0.05).结论 PⅡ可能通过激活cAMP/PKA信号轴促进SCI大鼠神经功能恢复.
Abstract
Objective To investigate the impact of picroside Ⅱ(PⅡ)on the recovery of neural function in spinal cord injury(SCI)rats and its regulatory effect on the cyclic adenosine monophosphate(cAMP)-protein kinase A(PKA)signaling pathway.Methods The SCI rat model was established.Rats were randomly divided into control group(CT group),SCI group,low-dose PⅡ group(PⅡ L group,5 mg/kg/d),high-dose PⅡ group(PⅡ H group,20 mg/kg/d),and high-dose PⅡ+PKA inhibitor group(PⅡ H+H-89 group,20 mg/kg/d PⅡ+5 mg/kg/d H-89).The Basso,Beattie and Bresnahan(BBB)scoring was applied to evaluate the motor function of SCI rats.Hematoxylin and eosin(H&E)and Luxol fast blue(LFB)staining were performed to evaluate the pathological characteristics of spinal cord tissue and demyelination,respectively.Immunofluorescence was applied to detect the expressions of glial fibrillary acidic protein(GFAP)and ion calcium binding adapter molecule-1(IBA-1).Enzyme-linked immunosorbent assay(ELISA)was applied to measure relative levels of malondialdehyde(MDA),superoxide dismutase(SOD)and cyclic adenosine monophosphate(cAMP).Western blot was applied to detect the protein expressions of phosphorylated PKA(p-PKA),PKA,phosphorylated cyclic adenosine phosphate responsive component binding protein(p-CREB)and CREB.Results Compared with those the CT group,rats in SCI group had spinal cord tissue defects,cavities,infiltration of massive inflammatory cells,demyelination,significantly higher levels of GFAP,IBA-1 and MDA,but lower BBB,relative levels of SOD and cAMP,p-PKA/PKA and p-CREB/CREB(P<0.05).Compared with rats of SCI group,rats in the PⅡ L group and PⅡ H group had alleviated histological injuries,reduced cavities,inflammatory cells and demyelination,significantly lower levels of GFAP,IBA-1 and MDA,but higher BBB,relative levels of SOD and cAMP,p-PKA/PKA and p-CREB/CREB(P<0.05).Compared with rats of PⅡ H group,rats in PⅡ H+H-89 group had severer histological injuries and demyelination,significantly higher levels of GFAP,IBA-1 and MDA,but lower BBB,relative levels of SOD and cAMP,p-PKA/PKA and p-CREB/CREB(P<0.05).Conclusion PⅡ may promote the recovery of neurological function in SCI rats by activating the cAMP/PKA signaling pathway.
基金项目
新疆维吾尔自治区自然科学基金青年基金(2019D01C323)
NETL
NSTL
万方数据