TXNIP/NLRP3信号通路在糖尿病大鼠肾缺血再灌注损伤中的作用

Role of TXNIP/NLRP3 signaling pathway in renal ischemia-reperfusion injury in diabetic rats

目的评价硫氧还蛋白相互作用蛋白/NOD样受体热蛋白结构域相关蛋白3（TXNIP/NLRP3）信号通路在糖尿病大鼠肾缺血再灌注损伤中的作用。 方法清洁级健康雄性SD大鼠，8～12周龄，体重200～220 g，采用腹腔注射1%链脲佐菌素65 mg/kg的方法建立糖尿病模型。取糖尿病模型制备成功的大鼠24只，采用随机数字表法分为3组（n＝8）：假手术组（S组）、肾缺血再灌注组（I/R组）和TXNIP抑制剂白藜芦醇组（R组）。R组于糖尿病模型制备成功后第3周每天腹腔注射白藜芦醇10 mg/kg，连续7 d。糖尿病模型制备成功后第4周，采用双侧肾蒂夹闭25 min再灌注的方法制备肾缺血再灌注损伤模型。再灌注48 h时处死大鼠取肾组织，观察病理学结果，采用比色法测定MDA含量、SOD活性和组织超氧阴离子清除力，采用ELISA法检测IL-1β和IL-18含量，采用TUNEL法检测细胞凋亡情况。采用Western blot法测定肾组织TXNIP、NLRP3和caspase-1的表达。左心室取血标本，测定血清BUN和Cr浓度。 结果与S组比较，I/R组和R组血清Cr浓度和细胞凋亡指数升高，肾组织超氧阴离子清除力降低，TXNIP、NLRP3和caspase-1表达上调，I/R组血清BUN浓度、肾组织MDA、IL-1β和IL-18含量升高，SOD活性降低（P〈0.05），肾组织病理学损伤加重；与I/R组比较，R组血清BUN和Cr浓度降低，肾组织MDA、IL-1β、IL-18含量和细胞凋亡指数降低，SOD活性和组织超氧阴离子清除力升高，TXNIP、NLRP3和caspase-1表达下调（P〈0.05），肾组织病理学损伤减轻。 结论糖尿病大鼠肾缺血再灌注损伤的病理生理机制与TXNIP/NLRP3信号通路激活有关。

Objective To evaluate the role of thioredoxin-interacting protein （TXNIP）/oligomer- ization domain-like receptor family pyrin domain-containing 3 （ NLRP3 ） signaling pathway in renal ische- mia-reperfusion （I/R） injury in diabetic rats. Methods Pathogen-free healthy male Sprague-Dawley rats, aged 8-12 weeks, weighing 200-220 g, were used in the study. Diabetes mellitus was induced by intrap- eritoneal injection of 1% streptozotocin 65 mg/kg and confirmed by blood glucose ≥ 16. 7 mmol/L 3 days lat- er. Twenty-four diabetic rats were divided into 3 groups （n = 8 each） using a random number table： sham operation group （group S）, renal I/R group （group I/R） and resveratrol （TXNIP inhibitor） group （group R）. Resveratrol 10 mg/kg was intraperitoneally injected every day for 7 consecutive days starting from 3rd week after successful establishment of the model in group R. At 4th week after successful establish- ment of the model, renal I/R was produced by occlusion of bilateral renal pedicles for 25 min followed byreperfusion in anesthetized rats in group R. The animals were sacrificed at 48 h of reperfusion, and renal specimens were obtained for microscopic examination of pathologic changes and for measurement of malondi- aldehyde （MDA） content, superoxide dismutase （SOD） activity and superoxide anion scavenging capa- bility （ using eolorimetric method）, interleukin-lbeta （IL-1β） and IL-18 contents （ by enzyme-linked immunosorbent assay）, cell apoptosis （using TUNEL） and expression of TXNIP, NLRP3 and caspase-1 in renal tissues （ using Western blot）. Blood samples were obtained from the left ventricle for determination of serum urea nitrogen （BUN） and creatinine （Cr） concentrations. Results Compared with group S, the serum Cr concentration and apoptosis index were significantly increased, superoxide anion scavenging capability in renal tissues was decreased, and the expression of TXNIP, NLRP3 and caspase-1 was up-reg- ulated in I/R and R groups, and the serum BUN concentration and contents of MDA, IL-1β and IL-18 in renal tissues were increased, the SOD activity was decreased （P〈0.05） , and the pathological changes of renal tissues were aggravated in group I/R. Compared with group I/R, the serum BUN and Cr concentra- tions were significantly decreased, the contents of MDA, IL-1β and IL-18 and apoptosis index were de- creased, the SOD activity and superoxide anion scavenging capability were increased, the expression of TXNIP, NLRP3 and caspase-1 was down-regulated （P〈0.05）, and the pathological changes of renal tis- sues were significantly attenuated in group R. Conclusion The pathophysiological mechanism of renal I/R injury is associated with the activation of TXNIP/NLRP3 signaling pathway in diabetic rats.