人参皂苷Rg1通过抑制NLRP3炎症小体途径减轻氧糖剥夺/复供后小胶质细胞炎症反应

Ginsenoside Rg1 reduces the inflammatory response of microglia after oxygen glucose deprivation/resupply by inhibiting the NLRP3 inflammasome pathway

目的探讨人参皂苷Rg1通过NLRP3炎症小体途径对小胶质细胞氧糖剥夺/复供损伤后炎症反应的影响,并进一步研究其抗炎的作用机制。方法将生长状态良好的BV-2小胶质细胞随机分为6组:无处理组(Con),氧糖剥夺/复供组(OGD/R),人参皂苷Rg1低、中、高剂量组(剂量分别为0.1、0.2、0.4 mmol/L,简称Rg1L、Rg1M、Rg1H),MCC950对照组(0.05 mmol/L,MCC950)。采用CCK8法检测细胞增殖;免疫荧光和免疫印迹法检测经不同浓度人参皂苷Rg1和MCC950作用48 h后,BV-2小胶质细胞内NLRP3炎症小体相关蛋白的表达。ELISA检测BV-2小胶质细胞培养液中炎症因子IL-1β、IL-18的表达水平。结果与Con组比较,经缺氧缺糖/复供处理的BV-2小胶质细胞胞质内可见明显的Iba-1绿色荧光表达;OGD/R处理BV-2小胶质细胞2 h后,加入不同浓度人参皂苷Rg1和MCC950培养48 h后,细胞增殖率呈现明显的下降趋势。OGD/R组呈现明显的NLRP3、ASC、Caspase-1、IL-1β和IL-18阳性表达。Rg1L、Rg1M、Rg1H 3组NLRP3蛋白表达水平显著下降,并且随着药物浓度的升高,表达越低。结果表明,与OGD/R组相比,人参皂苷Rg1和MCC950可抑制活化的BV-2小胶质细胞表达NLRP3(P<0.01)。结论人参皂苷Rg1可能通过抑制NLRP3、ASC和Caspase-1蛋白的表达、干扰NLRP3炎症小体合成,进一步抑制相关炎症因子IL-1β和IL-18的表达水平,从而抑制炎症反应。

Objective To investigate the effect of ginsenoside Rg1 on the inflammatory response of microglia cells after oxygen-glucose deprivation/resupply injury through NLRP3 inflammasome pathway,and to further investigate its anti-inflammatory mechanism.Methods BV-2 microglia were randomly divided into six groups:No treatment group(Con),oxygen glucose deprivation/resupply group(OGD/R),ginsenoside Rg1 low,medium and high dose groups(0.1,0.2,0.4 mmol/L,Rg1L,Rg1M,Rg1H),MCC950 control group(0.05 mmol/L,MCC950).The cell proliferation was detected by CCK8.Immunofluorescence and western blotting were used to detect the expression of NLRP3 inflammasome-related protein in BV-2 microglia after treatment with different concentrations of ginsenoside Rg1 and MCC950 for 48 h.The expression levels of IL-1β and IL-18 in BV-2 microglia were detected by ELISA.Results Compared with Con group,Iba-1 green fluorescence was observed in the cytoplasm of BV-2 microglia cells treated with hypoxia and glucose deficiency/resupply.After OGD/R treatment of BV-2 microglia for 2h,the proliferation rate of BV-2 microglia cells showed an obvious downward trend after 48 h culture with different concentrations of ginsenoside Rg1 and MCC950.There were significant positive expressions of NLRP3,ASC,Caspase-1,IL-1β and IL-18 in the OGD/R group.The NLRP3 protein expression in Rg1L,Rg1M and Rg1H groups was significantly decreased,and the expression decreased with the increasing of drug concentration.The results showed that compared with OGD/R group,ginsenoside Rg1 and MCC950 inhibited the expression of NLRP3 in activated BV-2 microglia cells(P<0.01).Conclusions Ginsenoside Rg1 may inhibit the expression levels of IL-1β and IL-18,thus inhibiting the inflammatory response by inhibiting the expressions of NLRP3,ASC,Caspase-1 and interfering with NLRP3 inflammasome synthesis.