远志皂苷元对脂多糖诱发神经细胞炎症损伤的保护作用

Senegenin protects against lipopolysaccharide-induced neurite toxicity in a nerve cell model

目的研究具有神经营养作用的远志皂苷元对神经细胞炎症因子的影响,探讨远志皂苷元是否具有降低神经细胞炎症反应的作用,从而为缓解抗肿瘤药物的毒副作用寻找新的突破口。方法体外培养小胶质细胞BV2细胞,采用脂多糖(lipopolysaccharide,LPS)制备细胞炎症模型,观察不同浓度的远志皂苷元对LPS诱导的细胞炎症因子—一氧化氮(nitric oxide,NO)的影响,从而发现较佳的远志皂苷元浓度;远志皂苷元2μmol/L、2.5μmol/L及3μmol/L和阳性对照组地塞米松10μmol/L,主要通过Griess Reagent法检测炎症介质NO浓度;定量聚合酶链反应(quantitative polymerase chain reaction,QPCR)法检测环氧化酶-2(cyclooxygenase-2,COX-2)mRNA表达量;Western blot法检测COX-2蛋白表达量;并用LPS诱导后的BV2炎症上清刺激SH-SY5Y细胞,再进行其细胞活力和NO浓度检测。结果不同浓度远志皂苷元对LPS诱导的BV2细胞产生NO的影响不同。与LPS组相比较,(5μmol/L、25μmol/L、50μmol/L、100μmol/L)剂量组远志皂苷元抗炎作用不明显,且50μmol/L及100μmol/L远志皂苷元可提高炎症介质NO的浓度,具有明显的促炎作用;而(1μmol/L、2μmol/L、4μmol/L、8μmol/L)剂量组中发现1μmol/L、2μmol/L远志皂苷元具有明显抗炎症作用(P<0.05或P<0.01),综上结果发现低剂量远志皂苷元具抗炎作用;(1.5μmol/L、2μmol/L、2.5μmol/L、3μmol/L)剂量组远志皂苷元均具有明显的降低NO浓度的作用,但以2μmol/L尤为显著(P<0.001),据该结果后续实验中远志皂苷元分为低、中、高(2μmol/L、2.5μmol/L、3μmol/L)剂量组;各组COX-2 mRNA表达情况对比发现,与LPS组相比,远志皂苷元2μmol/L、2.5μmol/L组中COX-2 mRNA表达显著降低(P<0.001),仍稍弱于地塞米松组;另外相比LPS组,远志皂苷元2μmol/L、2.5μmol/L、3μmol/L剂量组均可降低COX-2蛋白表达水平,以2μmol/L效果最为显著(P<0.05)。结论远志皂苷元可抑制LPS诱导的细胞炎症因子的释放及表达,表明具有神经营养作用的远志皂苷元可能还具有保护神经细胞炎症损伤的作用。

Objective To study the neurotrophic and anti-inflammatory effects of senegenin on inflammatory factors in nerve cells to identify novel drugs that alleviate the toxic and side effects of anti-tumor drugs.Methods Microglial BV2 cells were cultured in vitro,and lipopolysaccharide(LPS)was used to prepare the cell inflammation model.The effects of different concentrations of senegenin(2,2.5,3μmol/L)on the LPS-induced cellular inflammatory factor nitric oxide(NO)were observed to determine the optimal dose of senegenin compared with the positive control group(dexamethasone 10μmol/L).The concentration of NO in inflammatory medium was detected by Griess Reagent.Results The effects of different concentrations of senegenin on the production of NO in LPS-induced BV2 cells were different.Compared with the LPS group,the anti-inflammatory effects of senegenin(5,25,50,100μmol/L)were not obvious.Moreover,the concentration of NO in the inflammatory medium was increased by 50 and 100,indicating a proinflammatory effect.In addition,in the senegenin 1,2,4,and 8μmol/L dose range,1,2,and 2 kept up with each other and had significant anti-inflammatory effects(P<0.05 or P<0.01).The low doses of senegenin had anti-inflammatory effects.All doses of senegenin,especially 2μmol/L(P<0.001),significantly reduced the concentration of NO.According to this result,senegenin was divided into low,medium and high dose groups in a follow-up test.Co X-2 mRNA expression in the senegenin 2 and 2.5μmol/L groups was significantly decreased(P<0.001)compared with the LPS group,but was slightly lower than that in the dexamethasone group.In addition,compared with the LPS group,2,2.5,and 3μmol/L senegenin reduced Co X-2 protein levels,with the most significant effect shown for 2μmol/L senegenin(P<0.05).Conclusions Senegenin reduces the release and expression of inflammatory factors in microglia induced by LPS,suggesting senegenin has neurotrophic effects that might protect nerve cells from inflammatory injury.