青蒿素对脂多糖活化的小胶质细胞炎症介质释放的影响

目的探讨青蒿素(artemisinin,ART)对脂多糖(lipopolysaccharide,LPS)诱导的小胶质细胞炎症介质释放的影响,研究青蒿素对帕金森病等慢性神经退行性疾病神经细胞炎症反应的抑制作用及其机制。方法采用CCK8法检测ART对BV2小胶质细胞的作用浓度。使用一定浓度的ART来处理经过LPS诱导的小胶质细胞。定量PCR技术和ELISA检测ART对促炎因子白细胞介素1β(interleukin-1β,IL-1β)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的影响。采用Western blotting检测核因子-κB(nuclear factor kappa-B,NF-κB)蛋白(P65)、人核因子κB抑制蛋白α(NF-kappa-B inhibitor alpha,IKBα)的表达水平,以及ART对诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)和环氧合酶2(cyclooxygenase-2,COX-2)基因表达的影响。结果当ART的剂量为0.1~10μmol/L时,BV2细胞的活性无明显变化,因此选择浓度为1μmol/L的ART用于进一步的实验。定量PCR结果表明,ART预处理组中促炎因子IL-1β、TNF-αmRNA的表达显著降低。Western blotting结果显示,ART预处理组NF-κB、IKBα的蛋白表达水平显著升高,炎症诱导酶iNOS、COX-2的表达水平显著降低。结论 ART可调节LPS活化的BV2小胶质细胞中炎性因子及炎症诱导酶的表达,发挥抗炎作用,这一作用可能是通过调控NF-κB信号通路来实现的。

Effect of artemisinin on the release of inflammatory mediators of LPS-activated microglia

Objective To investigate the effect of artemisinin (ART) on the release of inflammatory mediators induced by lipopolysaccharide (LPS) in microglia, and to investigate the inhibitory effect of artemisinin on the inflammatory response of neurons in Parkinson' s disease and its mechanism. Methods The concentration of ART on BV2 microglia was detected by CCK8 method. LPS-induced microglia was treated with a given ART drug concentration. The effects of ART on the proinflammatory factors interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) were detected by quantitative PCR and ELISA. Western blot was used to detect the expression of NF-κB protein (P65) and IKBα, and the effect of ART on the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) genes. Results A dosage range of 0.1 to 10 μmol/L ART did not show a significant toxicity to BV2 cells, so 1 μmol/L ART was selected for further experiments. Quantitative PCR results showed that the expression of pro-inflammatory factors IL-1β and TNF-α mRNA was significantly decreased in the ART pretreatment group. The results of Western blotting showed that the expression levels of NF-κB and IKBα in the ART pretreatment group were significantly increased, and the expression levels of inflammation-inducing enzymes iNOS and COX-2 were significantly decreased. Conclusion ART can regulate the expression of inflammatory factors and inflammation-inducing enzymes in LPS-activated BV2 microglia and exert anti-inflammatory effects, which may be achieved by regulating NF-κB signaling pathway.