棱术排石颗粒对肾草酸钙结石作用机制的研究

Study of the mechanism whereby lengzhupaishi(rixolipite)granules prevent renal calcium oxalate stones

目的探讨棱术排石颗粒对肾草酸钙结石作用及对蛋白激酶C(PKC)-核因子E2相关因子2(Nrf2)/抗氧化反应元件(ARE)信号通路的影响。方法将32只SPF级SD大鼠随机分为4组(n=8):对照组、模型组、治疗组和抑制剂组。除对照组外,所有大鼠28 d内连续给予乙二醇及氯化铵诱导大鼠泌尿系草酸钙结石形成,造模同时,治疗组灌胃给予10.0 g/kg的棱术排石颗粒,抑制剂组在此基础上腹腔注射30 mg/kg R031-8220,每天1次,连续28 d。实验结束后,检测各组大鼠24 h尿量、尿pH值、尿草酸(Ox)分泌量、血和24 h尿中Ca^(2+)、Mg^(2+)含量、血液中血尿素氮(BUN)、肌酐(Cr)水平。Vonkossa染色、Western blot检测各组大鼠肾组织病理改变和PKC、Keap1、Nrf2、ARE蛋白的表。结果与对照组相比,模型组大鼠24 h尿量、尿pH及血、尿中Mg^(2+)明显降低,肾中SOD酶活性明显下降,Ox、血、尿中Ca^(2+)、肾MDA水平、ROS荧光强度水平与PKC、Keap1、Nrf2、ARE蛋白表达量明显升高(P<0.05);经棱术排石颗粒治疗后,上述指标均明显逆转,差异均具有统计学意义(P<0.05);抑制剂组与模型组差异无统计学意义(P>0.05)。结论棱术排石颗粒可有效改善肾草酸钙结石大鼠的病理状态,可能与激活PKC-Nrf2/ARE抗氧化信号通路降低氧化应激水有关。

Objective To determine the effects of rixolipite granules on renal calcium oxalate calculi and protein kinase C(PKC)-nuclear factor erythroid-2-related factor 2(Nrf2)signaling to antioxidant response elements(AREs).Methods Thirty-two specific pathogen-free Sprague Dawley rats were randomly allocated to four groups(n=8 each):a control group,a model group,a treatment group,and an inhibitor group.All of the rats except the controls were administered ethylene glycol and ammonium chloride in their diet to induce urinary calcium oxalate calculus formation within 28 days.The treatment group was also administered 10.0 g/kg Lengzhu Paishi granules,and the inhibitor group was administered 30 mg/kg r031-8220 once daily by gavage for 28 days.Afterward,24 h urine volume,urine pH,urinary oxalic acid(Ox)concentration,the circulating Ca^(2+)and Mg^(2+)concentrations and the 24 h urinary output of each,blood urea nitrogen(BUN),and circulating creatinine(Cr)concentrations were measured.The protein expression of PKC,Keap1,Nrf2 was characterized using von Kossa staining and Western blot.Results The model group had significantly lower 24 h urine volume,urine pH,circulating Mg^(2+),urinary Mg^(2+),and renal superoxide dismutase activity than the control group;and the Ox,circulating and urinary Ca^(2+),renal malondialdehyde concentration,reactive oxygen species fluorescence intensity,and PKC,Keap1,and Nrf2 protein expression were significantly higher(P<0.05).Lengzhu Paishi granule treatment significantly ameliorated all these effects(P<0.05).There was no significant difference between the two groups(P>0.05).Conclusions Lengzhu Paishi granules prevent the pathological changes associated with renal calcium oxalate calculus formation in rats,which may be mediated through the activation of the antioxidant signaling pathway PKCNrf2/ARE and a reduction in oxidative stress.