白藜芦醇对狼疮性肾炎小鼠的保护作用

Protective effect of resveratrol on mice with lupus nephritis

目的探讨白藜芦醇对MRL/lpr狼疮肾炎小鼠肾损伤的影响,并分析其与Toll样受体4(TLR4)/核因子κB(NF-κB)信号通路的相关性。方法选取6只野生型C57BL/6小鼠作为正常组,另将MRL/lpr小鼠随机分为模型组、阳性对照组和低、高剂量实验组,每组6只。低、高2个剂量实验组腹腔注射白藜芦醇20,80 mg·kg^-1·d^-1,阳性对照组腹腔注射醋酸泼尼松6 mg·kg^-1·d^-1,正常组和模型组注射等量0.9%NaCl,连续给药4周。采用考马斯亮蓝法检测尿蛋白含量;用酶联免疫吸附法检测血清中抗双链DNA抗体(anti-dsDNA)、抗核抗体(ANA)及白细胞介素-1β(IL-1β)、肿瘤坏死因子α(TNF-α)的含量;以蛋白质印迹法检测肾组织中TLR-4、MyD88、磷酸化核因子κB p65[p-NF-κB p65(Ser536)]蛋白表达水平。结果给药后,正常组、模型组、低剂量实验组、高剂量实验组和阳性对照组的尿蛋白含量分别为(0.57±0.06),(118.39±0.30),(107.23±4.49),(58.52±3.10)和(45.71±6.39)mg·dL^-1;anti-dsDNA水平分别为(150.39±15.36),(293.71±15.69),(273.24±39.66),(176.32±10.40)和(158.80±14.21)μg·mL^-1;ANA水平分别为(158.95±16.12),(519.67±33.65),(488.44±21.19),(206.99±21.52)和(177.11±14.57)μg·L-1;IL-1β水平分别为(10.78±1.15),(43.09±2.61),(39.01±1.23),(19.40±2.63)和(14.45±1.37)pg·mL^-1;TNF-α水平分别为(41.76±10.51),(397.18±25.27),(363.03±19.26),(134.84±23.13)和(91.10±18.43)pg·mL^-1;TLR4蛋白相对表达量分别为0.08±0.01,1.21±0.03,0.98±0.12,0.23±0.04和1.03±0.16;MyD88蛋白相对表达量分别为0.17±0.05,0.87±0.05,0.82±0.03,0.21±0.03和0.79±0.03;p-NF-κB p65(Ser536)蛋白相对表达量分别为0.06±0.02,0.86±0.06,0.81±0.07,0.09±0.02和0.82±0.08。模型组与正常组比较或高剂量实验组与模型组比较,上述指标的差异均有统计学意义(P<0.05)。结论白藜芦醇通过降低血清抗体含量、抑制炎症反应,对MRL/lpr狼疮肾炎小鼠肾起到保护作用,其机制可能与抑制TLR4/NF-κB信号通路活化有关。

Objective To investigate the effect of resveratrol on renal injury in MRL/lpr lupus nephritis mice,and to analyze its correlation with toll-like receptor 4(TLR4)/nuclear factorκB(NF-κB)signaling pathway.Methods Six wild-type C57BL/6 mice were selected as the blank group,and MRL/lpr mice were randomly divided into model group,Test-L group,Test-H group and positive control group,with 6 mice in each group.The low and high dose groups were given intraperitoneal injection of 20,80 mg·kg-1·d-1 resveratrol,the positive control group was given intraperitoneal injection of 6 mg·kg-1·d-1 prednisone acetate,and the normal group and model group were given the same amount of 0.9%NaCl for 4 weeks.Coomassie blue method was used to detect urine protein content.Serum levels of anti-double-stranded DNA antibody(anti-dsDNA),anti-nuclear antibody(ANA),interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)were detected by ELISA.The protein expression levels of TLR4,MyD88,p-NF-κB p65(Ser536)in renal tissues were detected by Western blotting.Results After administration,the contents of urine protein in normal group,model group,Test-L group,Test-H group and positive control group were(0.57±0.06),(118.39±0.30),(107.23±4.49),(58.52±3.10)and(45.71±6.39)mg·dL^-1,respectively;the contents of anti-dsDNA were(150.39±15.36),(293.71±15.69),(273.24±39.66),(176.32±10.40)and(158.80±14.21)μg·mL^-1,respectively;the contents of ANA were(158.95±16.12),(519.67±33.65),(488.44±21.19),(206.99±21.52)and(177.11±14.57)μg·L^-1,respectively;the contents of IL-1βwere(10.78±1.15),(43.09±2.61),(39.01±1.23),(19.40±2.63)and(14.45±1.37)pg·mL^-1,respectively;the contents of TNF-αwere(41.76±10.51),(397.18±25.27),(363.03±19.26),(134.84±23.13)and(91.10±18.43)pg·mL^-1,respectively;the relative expression of TLR4 protein were 0.08±0.01,1.21±0.03,0.98±0.12,0.23±0.04 and 1.03±0.16,respectively;the relative expression of MyD88 protein were 0.17±0.05,0.87±0.05,0.82±0.03,0.21±0.03 and 0.79±0.03,respectively;the relative expression of p-NF-κB p65(Ser536)protein were 0.06±0.02,0.86±0.06,0.81±0.07,0.09±0.02 and 0.82±0.08,respectively.The comparison between the model group and the normal group or the Test-H group and the model group showed statistically significant differences in the above indicators(P<0.05).Conclusion Resveratrol plays a protective role in the kidney of MRL/lpr mice by decreasing the content of serum antibody and inhibiting inflammatory reaction,and its mechanism may be related to the inhibition of TLR4/NF-κB signaling pathway activation.