赣南野生黄花倒水莲抗炎作用及其作用机制的初步研究

Preliminary study on the anti-inflammatory effect and mechanism of wild polygala fallax in Gannan

目的 分析赣南野生黄花倒水莲抗炎作用及其作用机制。方法 提取黄花倒水莲总黄酮后取对数生长期的RAW264.7细胞,分为空白对照组(以无血清DMEM孵育)、模型组(加入脂多糖100μg/L孵育24 h)及低、中、高剂量总黄酮组,ELISA方法测定一氧化氮(NO)、肿瘤坏死因子-α(TNF-α)、白介素(IL)-1α、IL-1β、IL-6、IL-10含量,Western blot法检测细胞中诱导型一氧化氮合酶(iNOS)、环氧合酶-2(COX-2)蛋白相对表达。应用对角叉菜胶致大鼠足肿胀模型,分为模型组(灌胃蒸馏水)、阳性对照组(灌胃给予阿司匹林)、低、中、高剂量总黄酮组,各组分别以0.2、2、20μmol/L的黄花倒水莲总黄酮提取物灌胃,连续给药5 d,观察各组大鼠足跖肿胀度及大鼠血清iNOS、COX-2、TNF-α、IL-1β水平。应用冰醋酸致大鼠腹腔毛细血管通透性模型,观察各组大鼠伊文斯蓝含量及毛细血管抑制通透性率。结果 与空白对照组比较,模型组RAW264.7细胞NO、TNF-α、IL-1α、IL-1β、IL-6水平升高,IL-10水平降低(P<0.01);与模型组比较,低剂量总黄酮组、中剂量总黄酮组及高剂量总黄酮组NO、TNF-α、IL-1α、IL-1β、IL-6水平降低,IL-10水平依次升高(P<0.05)。与空白对照组比较,模型组iNOS、COX-2蛋白相对表达量升高(P<0.05);与模型组比较,低剂量总黄酮组、中剂量总黄酮组及高剂量总黄酮组iNOS、COX-2蛋白相对表达量依次降低(P<0.05)。与模型组比较,阳性对照组、中剂量总黄酮组致炎成功后1、3 h及高剂量总黄酮组致炎成功后1、2、3 h的大鼠足跖肿胀度均降低(P<0.05或P<0.01)。与模型组比较,阳性对照组、低剂量总黄酮组、中剂量总黄酮组及高剂量总黄酮组血清iNOS、COX-2、TNF-α、IL-1β水平均依次降低(P<0.01)。与模型组比较,阳性对照组、低剂量总黄酮组、中剂量总黄酮组及高剂量总黄酮组腹膜毛细血管伊文斯蓝含量均降低(P<0.05或P<0.01);阳性对照组、低剂量总黄酮组、中剂量总黄酮组及高剂量总黄酮组腹膜毛细血管抑制率均增加(P<0.05或P<0.01)。结论 赣南野生黄花倒水莲总黄酮可有效发挥抗炎作用,抑制大鼠角叉菜胶所致足趾肿胀程度,降低大鼠腹腔毛细血管通透性,具有良好的市场发展前景和社会效益。

Objective Analysis of the anti-inflammatory effect and mechanism of wild polygala fallax in Gannan.Methods RAW264.7 cells in logarithmic growth phase were obtained after extraction of total flavonoids from polygala fallax.They were divided into blank control group(incubated with serum-free DMEM),model group(incubated with lipopolysaccharide 100 μg/L for 24 hours) and low,medium and high dose total flavonoids groups.The contents of NO,TNF-α,IL-1α,IL-1β,IL-6,IL-10 in cells were determined by ELISA.The protein relative expression of iNOS,COX-2 in cells were detected by Western blot.The rat foot swelling model induced by carrageenan were applied and divided into model group(gavage with distilled water),positive control group(gavage with aspirin),low,medium and high dose total flavonoids groups.Each group was gavaged with 0.2,2 and 20 μmol/L polygala fallax total flavonoids extract,respectively,for 5 days.The degree of rat foot swelling and the levels of serum iNOS,COX-2,TNF-α,and IL-1β in each group were observed.Used a rat peritoneal capillary permeability model induced by acetic acid,the content of Evans blue and the inhibitory permeability of capillaries in each group were observed.Results Compared with the blank control group,the levels of NO,TNF-α,IL-1α,IL-1β,IL-6 in RAW264.7 cells in the model group increased,while the level of IL-10 decreased(P<0.01).Compared with the model group,the levels of NO,TNF-α,IL-1α,IL-1β and IL-6 in the low-dose total flavone group,the medium-dose total flavone group and the high-dose total flavone group decreased,while the level of IL-10 increased(P<0.05).Compared with the blank control group,the relative expression of iNOS and COX-2 protein in the model group increased(P<0.05).Compared with the model group,the relative expression of iNOS and COX-2 protein in low-dose total flavonoids group,medium-dose total flavonoids group and high-dose total flavonoids group decreased(P<0.05).Compared with the model group,the swelling degree of rat foot in the positive control group,the medium-dose total flavonoids group and the high-dose total flavonoids group decreased 1,3 hours after successful inflammation and 1,2,3 hours after successful inflammation(P<0.05 or P<0.01).Compared with the model group,the serum levels of COX-2 and TNF-α in the positive control group,as well as the serum levels of iNOS,COX-2,TNF-α,and IL-1β in the low-dose total flavonoids group,medium-dose total flavonoids group and high-dose total flavonoids group were all reduced(P<0.01).Compared with the model group,the Evans blue content in the peritoneal capillaries of the positive control group,low-dose total flavonoids group,medium-dose total flavonoids group and high-dose total flavonoids group decreased(P<0.05 or P<0.01).The inhibition rate of peritoneal capillaries in the positive control group,low-dose total flavonoids group,medium-dose total flavonoids group and high-dose total flavonoids group all increased(P<0.05 or P<0.01).Conclusion The total flavonoids of wild polygala fallax in Gannan can effectively exert anti-inflammatory effects,inhibit the degree of foot swelling caused by carrageenan in rats,reduce the permeability of abdominal capillaries in rats,and have good market development prospects and social benefits.