粉葛与葛根多糖对脂多糖诱导RAW264.7细胞的抗炎作用

Anti-inflammatory Effect of Pueraria lobata(Willd.)Ohwi and P.thomsonii Benth.Polysaccharides on LPS-Induced RAW264.7 Cells

该研究通过脂多糖诱导RAW264.7细胞炎症模型,对粉葛粗多糖(FGC)、葛根粗多糖(GGC)以及粉葛均一多糖(FGJ)的抗炎作用进行比较。用CCK8试剂盒检测FGC、GGC以及FGJ对细胞活力的影响;相关试剂盒检测细胞产生NO、TNF-α、IL-6的分泌情况;蛋白印迹法检测NF-κB、Nrf2/HO-1通路相关蛋白表达,QPCR检测TNF-α、IL-6的mRNA表达,通过流式检测活性氧的产生。结果显示,在浓度为10、20、40μg/mL时,FGC、GGC以及FGJ对细胞均没有毒性(p>0.05)。各浓度的FGC、FGJ不能抑制LPS诱导产生的的NO、TNF-α、IL-6(p>0.05),GGC在浓度为40μg/mL时,对NO、TNF-α、IL-6的抑制率达到38.46%、52.90%、55.87%,可以抑制细胞内ROS释放,在10、20、40μg/mL时抑制率达到37.28%、41.05%、54.09%,在浓度为40μg/mL时,对iNOS、COX-2、p-p65、p-IκBα的抑制率达到29.27%、95.04%、27.18%、28.45%,对Nrf2、HO-1蛋白的表达有促进作用,在浓度40μg/mL时显著地抑制IL-6与TNF-α的mRNA表达,抑制率达到57.70%、81.68%。结果证明,GGC与FGC、FGJ相比抗炎活性更加明显,并且其抗炎作用是通过NF-κB与Nrf2/HO-1两条通路来实现的。

The anti-inflammatory effects of Pueraria crude polysaccharide(FGC),Pueraria crude polysaccharide(GGC),and Pueraria homogenized polysaccharide(FGJ)were compared in an LPS-induced RAW264.7 cell model.The effects of FGC,GGC,and FGJ on cell viability were detected using a CCK8 kit.Secretion of nitric oxide,TNF-α,and IL-6 in cells was detected using their corresponding kits.The expression levels of NF-κB and Nrf2/HO-1 pathway related proteins were detected using western blot,mRNA expression levels of TNF-αand IL-6 were detected by qPCR,and ROS production was detected using flow cytometry.The results showed that FGC,GGC,and FGJ had no toxicity to cells at concentrations of 10,20,and 40μg/mL(p>0.05).FGC and FGJ did not inhibit LPS-induced nitric oxide,TNF-α,and IL-6 secretion(p>0.05)at any concentration.At 40μg/mL,GGC inhibited nitric oxide,TNF-α,and IL-6 levels by 38.46%,52.90%,and 55.87%,respectively,which was able to inhibit intracellular ROS release.Inhibition rates reached 37.28%,41.05%,and 54.09%at concentrations of 10,20,and 40μg/mL.The inhibition rates of iNOS,COX-2,p-p65,and p-IκBαwere 29.27%,95.04%,27.18%,and 28.45%at 40μg/mL,which was able to promote the expression of Nrf2 and HO-1 proteins.At 40μg/mL,mRNA expression levels of IL-6 and TNF-αwere significantly inhibited,with inhibition rates reaching 57.70%and 81.68%,respectively.The results showed that GGC had a more distinct anti-inflammatory activity than FGC and FGJ,and its anti-inflammatory effect was realized through the NF-κB and Nrf2/HO-1 pathways.