川芎嗪对单侧输尿管梗阻大鼠肾脏的保护作用及缺氧诱导因子1α表达的影响

Reno-protective effects of Ligustrazine and its action on expression of hypoxia-inducible factor-1α in rats with unilateral ureteral obstruction

目的 探讨川芎嗪对单侧输尿管梗阻大鼠肾功能的保护作用及肾组织中缺氧诱导因子1α（HIF-1α）表达的影响.方法 6周龄健康雄性SD大鼠60只,随机分为治疗组、模型组和假手术组,每组各20只.治疗组建立单侧输尿管梗阻模型,并给予川芎嗪注射处理;模型组则仅进行模型建立,给予同样剂量的生理盐水输注;假手术组仅在手术中分离输尿管,不进行结扎.术后第7、14天各组随机选取10只大鼠进行血肌酐、尿素氮、尿β2微球蛋白的检测.3组于术后第7、14天各处死10只大鼠,模型组、治疗组留取解除梗阻侧肾脏,假手术组留取手术侧肾脏,用于肾病理形态学分析,并且采用免疫组化及Western免疫印迹行肾组织HIF-1α表达的检测.结果 川芎嗪治疗7d后,模型组与治疗组大鼠血肌酐、尿素氮、尿β2微球蛋白含量差异无统计学意义,但均较假手术组升高（均P＜0.05）.川芎嗪治疗14d后,模型组与治疗组大鼠血肌酐、尿素氮及尿β2微球蛋白含量仍较假手术组升高（均P＜0,05）,但治疗组较模型组已明显下降（均P＜0.05）.川芎嗪治疗7d和14d假手术组肾脏无明显病理变化,上皮细胞无变性,间质未见炎症细胞;模型组川芎嗪治疗7d时肾间质可见肾小管扩张,淋巴细胞浸润,纤维组织增生,14d肾小管上皮细胞萎缩或坏死,管腔扩张,大量淋巴细胞、单核巨噬细胞浸润,肾间质纤维组织明显增生;治疗组川芎嗪治疗7d时肾间质可见淋巴细胞浸润,纤维组织增生,散在局灶出血,14d可见间质纤维组织增生及炎症细胞浸润,较川芎嗪治疗7d及模型组治疗7d和14 d明显好转,间质内出血灶减少.川芎嗪治疗7d和14d模型组与治疗组大鼠的肾组织中HIF-1α表达较假手术组增高,治疗7d治疗组与模型组肾组织中HIF-1α表达差异无统计学意义,而14d较模型组明显下降（免疫组化7 d：11.12±3.10、19.70±4.10比2.66±0.88,14 d：7.64±3.11比35.44±5.01比2.82±0.73;Westen免疫印迹7 d：13.12±3.23、20.70±3.14比2.82±1.18,14 d：7.66±3.67比31.41±2.11比2.52±0.13;均P＜0.05）.结论 川芎嗪可改善单侧输尿管梗阻大鼠的肾功能,并对其肾脏HIF-1α的表达起下调作用,可作为梗阻性肾病的辅助治疗.

Objective To investigate the reno-protective effects of Ligustrazine and its action on expression of hypoxia-inducible factor-1 α （HIF-1 α） in rats with unilateral ureteral obstruction.Methods Sixty healthy,6-week-old male SD rats were randomly divided into the treatment group,model group,and sham operation group （n=20 each）.The treatment group was processed with modeling of unilateral ureteral obstruction,and was given the Ligustrazine by injection;the model group was processed with modeling alone,and was given the same dose of normal saline by infusion;the sham operation group only underwent surgical isolation of ureter,without ligation.At 7 and 14 d after the operation,10 rats in each group were randomly selected to be determined for serum creatinine,urea nitrogen,and urine β2-microglobulin.At 7 and 14 d after the operation,10 rats in each group were executed.The rats in model group and the treatment group were harvested for the kidneys on the obstructed side,and those in sham operation group were harvested for the kidneys on the operated side.The harvested kidneys were subjected to pathomorphological analysis.The expression of HIF-1α in renal tissues was determined by immunohistochemistry and Western blotting.Results After the Ligustrazine treatment at 7 d,there were no statistically significant differences in the serum creatinine,urea nitrogen,and urinary β2-microglobulin content of rats between the model group and the treatment group,but these were higher compared with those in the sham operation group （all P〈0.05）.After the Ligustrazine treatment at 14 d,the serum creatinine,urea nitrogen and urinary β2-microglobulin content of rats continued to be higher in the model group and the treatment group compared with those in the sham operation group （all P〈0.05）,but these levels were significantly lower in the treatment group compared with those in the model group （all P〈0.05）.After the Ligustrazine treatment at 7 and 14 d,no significant pathological changes,degeneration of epithelial cells,and interstitial inflammatory cells were found in the kidneys of the sham operation group.After the Ligustrazine treatment in the model group,renal tubular ectasia,lymphocyte infiltration,and hyperplasia of fibrous tissues were found at 7 d,and renal tubular epithelial cell atrophy or necrosis,lumen ectasia,massive lymphocyte and mononuclear macrophage infiltration,and significant hyperplasia of fibrous tissues were found at 14 d in renal interstitium.After the Ligustrazine treatment in the treatment group at 7 d,lymphocyte infiltration,hyperplasia of fibrous tissues,and focal bleeding were found.After the Ligustrazine treatment in the treatment group at 14 d,hyperplasia of interstitial fibrous tissues and inflammatory cell infiltration were significantly improved and the interstitial bleeding lesions were decreased,compared with those in the treatment group at 7 d and the model group at 7 and 14 d.The expression of HIF-1α in renal tissues of rats in the model group and the treatment group after the Ligustrazine treatment at 7 d and 14 d was significantly increased compared with that in the sham operation group.There was no statistically significant difference in the HIF-1 αt expression in renal tissues of rats between the treatment group and the model group after the Ligustrazine treatment at 7 d,whereas the HIF-1 α expression in the treatment group was significantly decreased compared with that in the model group at 14 d （immunohistochemistry 7 d：11.12±3.10,19.70±4.10 vs 2.66±0.88,14 d：7.64±3.11 vs 35.44±5.01 vs 2.82±0.73;Western blotting 7 d：13.12±3.23,20.70±3.14 vs 2.82±1.18,14 d：7.66±3.67 vs 31.41±2.11 vs 2.52±0.13;all P〈0.05）.Conclusion Ligustrazine can improve the renal function in rats with unilateral ureteral obstruction,and down-regulate the expression of renal HIF-1α,which can be used as an adjuvant therapy for obstructive nephropathy.