tBHQ对膜性肾病大鼠肾组织损伤、氧化应激抑制及肾皮质和细胞核Nrf2/NF-κB信号通路调控作用

Inhibitory effects of tBHQ on renal injury and oxidative stress and its regulatory effect on Nrf2/NF-κB signaling pathway in renal cortex and nucleus of rats with membranous nephropathy

目的观察特丁基对苯二酚(tBHQ)对膜性肾病(MN)大鼠肾组织损伤、氧化应激的抑制及肾皮质和细胞核Nrf2/NF-κB信号通路的调控作用,以探讨tBHQ对MN的潜在治疗作用及机制。方法采用随机数字表法将32只大鼠均分为正常对照组、tBHQ对照组、模型组及tBHQ干预组。正常对照组腹腔注射3 mL正常兔血清;tBHQ对照组同法注射正常兔血清后予tBHQ 50 mg/kg灌胃,每天1次,共15次;模型组腹腔注射3 mL兔Fx1A抗血清建立被动Heymann肾炎(PHN)模型;tBHQ干预组建立PHN模型后tBHQ 50 mg/kg灌胃,每天1次,共15次。结束实验当天检测各组大鼠肾病综合征相关指标[24 h尿蛋白(24 h-UPro)、血清白蛋白(ALB)、总胆固醇(T-CHOL)、甘油三酯(TG)、高密度脂蛋白(HDL)、低密度脂蛋白(LDL)]、肾功能相关指标[sCr、尿素氮]、肾脏足细胞损伤情况(足细胞足突融合、肾小球Podocin/Desmin表达及分布)]、肾皮质氧化应激标志物[丙二醛(MDA)、8-异前列腺素(8-iso-PG)、8-羟基脱氧鸟苷(8-OHDG)]、肾皮质抗氧化酶[超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、γ-谷氨酰半胱氨酸合成酶(γ-GCS)]活力及肾皮质和细胞核Nrf2/NF-κB信号通路相关蛋白。结果与正常对照组相比,模型组24 h-UPro、T-CHOL、TG、HDL、LDL、sCr、尿素氮均升高,ALB降低(P均<0.05);肾小球上皮下电子致密物沉积,足细胞足突弥漫性融合,足细胞足突宽度增加,Podocin表达量减少、分布异常,Desmin表达量上调;肾皮质MDA、8-iso-PG、8-OHDG均升高(P均<0.05),SOD、CAT、γ-GCS活力均降低(P均<0.05);肾皮质及细胞核Nrf2均下调,NF-κB p65、p-NF-κB p65(Ser536)均上调(P均<0.05)。与模型组相比,tBHQ干预组24 h-UPro、T-CHOL、TG、HDL、LDL、sCr均下降,ALB上升(P均<0.05);足细胞足突弥漫性融合减轻,足细胞足突宽度下降,Podocin/Desmin表达及分布趋向正常;肾皮质MDA、8-iso-PG、8-OHDG均下降(P均<0.05),SOD、CAT、γ-GCS活力均上升(P均<0.05);肾皮质及细胞核Nrf2均上调,NF-κB p65、p-NF-κB p65(Ser536)均下调(P均<0.05)。结论tBHQ对MN大鼠肾组织损伤及氧化应激均有抑制作用,可调控肾皮质及细胞核Nrf2/NF-κB信号通路。tBHQ可能通过激动Nrf2、抑制NF-κB活化,从而抑制肾组织氧化应激反应,进而减轻MN大鼠肾组织损伤。

Objective To observe the inhibitory effects of tert-butylhydroquinone(tBHQ)on renal injury and oxidative stress and its regulatory effect on the nuclear factor erythroid 2-related factor 2(Nrf2)/nuclear factor-κB(NF-κB)signaling pathway in renal cortex and nucleus of rats with membranous nephropathy(MN),so as to illustrate the potential therapeutic effect and mechanism of tBHQ on MN.Methods Thirty-two rats were randomly divided into the normal control group,tBHQ control group,model group,and tBHQ intervention group,respectively.Rats in the normal control group were injected intraperitoneally with 3 mL of normal rabbit serum,rats in the tBHQ control group were injected with normal rabbit serum in the same way and then were gavaged with 50 mg/kg tBHQ once a day for a total of 15 injections,rats in the model group were injected intraperitoneally with 3 mL of rabbit Fx1A antiserum to establish passive Heymann ne‐phritis(PHN)models,and rats in the tBHQ intervention group were injected with rabbit Fx1A antiserum in the same way and then were gavaged with 50 mg/kg tBHQ once a day for a total of 15 injections.At the end of the experiment,indicators related to nephrotic syndrome[24 h urinary protein(24 h-UPro),serum albumin(ALB),total cholesterol(T-CHOL),tri‐glyceride(TG),high-density lipoprotein(HDL),and low-density lipoprotein(LDL)],renal function-related indicators[serum creatinine(sCr),urea nitrogen],renal podocyte injury(podocyte foot process fusion,glomerular Podocin/Des‐min expression and distribution),renal cortex oxidative stress markers[malondialdehyde(MDA),8-isoprostaglandin(8-iso-PG),8-hydroxydeoxyguanosine(8-OHDG)],renal cortex antioxidant enzymes[superoxide dismutase(SOD),cata‐lase(CAT),γ-glutamylcysteine synthase(γ-GCS)]activities and the Nrf2/NF-κB signaling pathway in the renal cortex and nucleus of the rats in each group were detected.Results Compared with the normal control group,the 24 h-UPro,T-CHOL,TG,HDL,LDL,sCr,and urea nitrogen increased,and ALB decreased(all P<0.05);subepithelial electron dense deposits in the glomeruli,diffuse fusion of the podocyte foot processes,and the width of the podocyte foot processes increased;the expression level and abnormal distribution of Podocin decreased,the expression level of Desmin was up-reg‐ulated;the MDA,8-iso-PG,and 8-OHDG levels all increased in the renal cortex(all P<0.05),and the activities of SOD,CAT,andγ-GCS decreased(all P<0.05);Nrf2 was down-regulated,whereas NF-κB p65 and p-NF-κB p65(Ser536)were up-regulated in the renal cortex and nucleus in the model group(all P<0.05).Compared with the model group,the 24 h-UPro,T-CHOL,TG,HDL,LDL,and sCr levels all decreased,and the ALB increased(all P<0.05);the diffuse fusion of the podocyte foot processes was relieved,the podocyte foot processes width decreased,and the expres‐sion and distribution of Podocin/Desmin tended to be normal;the MDA,8-iso-PG,and 8-OHDG levels decreased in the renal cortex(all P<0.05),and the activities of SOD,CAT,andγ-GCS increased(all P<0.05);Nrf2 was up-regulated,whereas NF-κB p65 and p-NF-κB p65(Ser536)were down-regulated in the renal cortex and nucleus of the tBHQ interven‐tion group(all P<0.05).Conclusions The tBHQ inhibits renal injury and oxidative stress of MN rats,and can regulate the Nrf2/NF-κB signaling pathway in the renal cortex and nucleus.The tBHQ may activate Nrf2 and inhibit NF-κB activa‐tion,thereby alleviating oxidative stress and renal injury in MN rats.