不同剂量丁苯酞对UUO小鼠肾组织Nrf-2表达的影响

The effects of different doses of butylphthalide on Nrf-2 expression in mouse model with unilateral ureteral obstruction

目的探讨不同剂量丁苯酞(NBP)对单侧输尿管梗阻(UUO)小鼠肾间质纤维化的作用,并探讨核转录因子-E2相关因子2(Nrf-2)表达水平与肾间质纤维化的关联性。方法清洁级CD-1雄性小鼠72只随机均分为低剂量丁苯酞(NBPL)组、高剂量丁苯酞(NBPH)组、假手术(Sham)组和模型(UUO)组。除Sham组外,均制作UUO模型,NBPL、NBPH组自术后第1天始分别给予NBP 150 mg/(kg·d)、220 mg/(kg·d)灌胃。Sham及UUO组亦自术后第1天始均给予等量生理盐水灌胃,每天每公斤体质量入量恒定,直至处死。各组分别于术后第3、7、14天处死6只小鼠,取梗阻侧肾组织做免疫组化染色检测Ⅰ型胶原蛋白的表达,Western Blot检测7 d和14 d时Nrf-2、Ⅰ型胶原蛋白的表达。结果免疫组化结果显示,UUO组Ⅰ型胶原蛋白的IOD值随时间延长呈增高趋势,Sham组、NBPL组和NBPH组呈降低趋势(P<0.05)。UUO组、NBPL组和NBPH组各时点IOD值均较Sham组增高,NBPL、NBPH组较UUO组降低,且NBPH组低于NBPL组(P<0.05)。Western Blot结果显示,7 d和14 d时,UUO组、NBPL组和NBPH组Nrf-2、Ⅰ型胶原蛋白IOD值均较Sham组增高;NBPL、NBPH组Nrf-2蛋白IOD值较UUO组升高,而Ⅰ型胶原蛋白降低;NBPL组Nrf-2的IOD值低于NBPH组,而Ⅰ型胶原蛋白高于NBPH组(均P<0.05)。结论 NBP对UUO小鼠肾间质纤维化有一定的改善作用,作用机制可能与上调肾组织Nrf-2的表达,下调Ⅰ型胶原蛋白的表达有关。

Objective To explore the impact of different doses of butylphthalide on renal interstitial fibrosis in mice with obstructive nephropathy, and to discuss the correlation between Nrf-2 expression level and renal interstitial fibrosis.Methods Totally 72 male CD-1 mice of clean grade were selected and randomly assigned into 4 groups: sham operation group (Sham), model group (UUO), low dose of butylphthalide group (NBPL) and high dose of butylphthalide group (NBPH).In addition to the control group, UUO model was established in other three groups. NBPL group was given NBP 150 mg/(kg·d) by gavage since postoperative day 1,NBPH group was given NBP 220 mg/(kg·d) by gavage. Sham group and UUO groupwere given equal volume of saline by gavage. Six mice were sacrificed at the third, 7th, 14th day, respectively. The obstructive renal tissue was selected for immunohistochemical staining and Western blot assay. The expression levels ofNrf - 2 and type Ⅰ collagen were detected by Western blot assay. Results The IOD value of type Ⅰ collagen wasincreased with time in UUO group, and which showed a gradual decreasing trend in Sham group, NBPL group and NBPHgroup (P < 0.05). The IOD values at different time points were significantly higher in UUO group, NBPL group and NBPHgroup compared to those of Sham group. The IOD values were significantly decreased in NBPL and NBPH groups than that ofUUO group, and IOD value was lower in NBPH group than that of NBPL group (P < 0.05). Western blot assay showed thatIOD values of Nrf-2 and type Ⅰ collagen at 7 d and 14 d were increased in UUO group, NBPL group and NBPH group compared with those of Sham group. The IOD values of Nrf-2 protein were increased, and IOD values of type Ⅰ collagenwere decreased, in NBPL group and NBPH group than those of UUO group. The IOD value of Nrf - 2 protein was decreased in NBPL group than that of NBPH group, but the IOD value of type Ⅰ collagen was increased in NBPL group than that of NBPH group (P<0.05). Conclusion NBP can improve renal interstitial fibrosis in UUO mice, which may be related with the increased expression of Nrf - 2 and the down-regulated expression of type I collagen.