基于PI3K-Akt-eNOS通路探讨松果菊苷对脊髓损伤大鼠的神经保护作用

Neuroprotective Effect of Echinacein on Spinal Cord Injury in Rats Based on PI3K-Akt-eNOS Pathway

目的基于PI3K-Akt-eNOS通路探讨松果菊苷(ECH)对脊髓损伤(SCI)大鼠的神经保护作用并探讨其机制。方法大鼠随机分为假手术组、脊髓损伤组、松果菊苷干预组、PI3K抑制剂LY294002组和松果菊苷干预+PI3K抑制剂LY294002组。采用改良的Allen法建立SCI大鼠模型。通过脊髓运动功能(BBB)评分评估大鼠运动功能,HE染色观察脊髓组织病理变化,TUNEL检测脊髓组织细胞凋亡率,试剂盒检测血清一氧化氮(NO)、内皮型一氧化氮合酶(eNOS)、诱导型一氧化氮合酶(iNOS)含量,qRT-PCR法检测各组脊髓组织eNOS和iNOS mRNA表达水平,Western blotting法检测脊髓组织Bax、Bcl-2、磷脂酰肌醇3激酶(PI3K)、磷酸化PI3K(p-PI3K)、蛋白激酶B(Akt)和磷酸化蛋白激酶B(p-Akt)蛋白表达。结果与假手术比较,脊髓损伤组大鼠BBB评分、血清NO和eNOS水平、脊髓组织eNOS m RNA以及Bcl-2、p-PI3K和p-Akt蛋白表达显著降低(P <0.05),血清iNOS水平、脊髓组织细胞凋亡数、iNOS m RNA、Bax蛋白表达均显著升高(P <0.05)。与脊髓损伤组比较,松果菊苷干预组大鼠BBB评分、血清NO和eNOS水平、脊髓组织e NOS mRNA以及Bcl-2、p-PI3K和p-Akt蛋白表达显著升高(P <0.05),血清iNOS水平、脊髓组织细胞凋亡数、iNOS mRNA、Bax蛋白表达均显著降低(P <0.05)。PI3K抑制剂LY294002组和松果菊苷干预+PI3K抑制剂LY294002组两组大鼠各检测指标差异均无统计学意义(P> 0.05)。结论 ECH可能通过激活PI3K-Akt-eNOS信号通路对SCI大鼠发挥神经保护作用。

Objective:To investigate the neuroprotective effect of echinarin(ECH)on spinal cord injury(SCI) rats based on PI3K-Akt-eNOS pathway. Methods:Rats were randomly divided into the sham group,SCI group,ECH group,LY group and ECH + LY group. SCI rat model was established with modified Allen method. The motor function of rats was evaluated by spinal cord motor function(BBB)score;histopathological changes of spinal cord tissues were observed by HE staining;the apoptotic rate of spinal cord tissues was detected by TUNEL;serum NO,eNOS and iNOS contents were detected by the kit;the expression levels of eNOS and iNOS m RNAs in spinal cord tissues were detected by qRT-PCR;Western blot was used to detect the expressions of Bax,Bcl-2,PI3K,pPI3K,AKT and p-AKT in spinal cord. Results:Compared with the sham group,BBB score,serum NO and e NOS levels,expressions of eNOS m RNA and Bcl-2,p-PI3K and p-Akt proteins in spinal cord in SCI group were significantly decreased(P < 0.05);the level of serum iNOS,the number of apoptotic cells in spinal cord tissue and the expressions of iNOS m RNA and Bax protein were significantly increased(P < 0.05). Compared with SCI group,BBB score,serum NO and eNOS levels,expressions of e NOS m RNA and Bcl-2,p-PI3K and p-Akt proteins in spinal cord in ECH group were significantly increased(P < 0.05);the level of serum iNOS,the number of apoptotic cells in spinal cord tissue and the expressions of iNOS m RNA and Bax protein were significantly decreased(P <0.05). There was no significant difference in each index between LY group and ECH + LY group(P > 0.05). Conclusion:ECH may play a neuroprotective role in SCI rats by activating PI3K-Akt-eNOS signaling pathway.