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

摘要:

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、40 μg/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40 μg/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、40 μg/mL时抑制率达到37.28%、41.05%、54.09%，在浓度为40 μg/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40 μg/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。

Abstract:

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-α and 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 40 μg/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 40 μg/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 40 μg/mL. The inhibition rates of iNOS, COX-2, p-p65, and p-IκBα were 29.27%, 95.04%, 27.18%, and 28.45% at 40 μg/mL, which was able to promote the expression of Nrf2 and HO-1 proteins. At 40 μg/mL, mRNA expression levels of IL-6 and TNF-α were 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.