1、第 14 卷 第 4 期2023 年 7 月Vol.14 No.4Jul.2023器官移植Organ Transplantation论著丁苯酞通过下调NF-B信号通路抑制细胞焦亡减轻大鼠肾缺血-再灌注损伤张瑞波申开文王强袁强沈俊【摘要】目的 探讨丁苯酞对大鼠肾缺血-再灌注损伤（IRI）的作用机制。方法 将 40 只 SD 大鼠随机分为假手术组（Sham 组）、模型组（IRI 组）、NF-B 抑制剂吡咯烷二硫代氨基甲酸酯（PDTC）组、丁苯酞低剂量组（NBP-L 组）及丁苯酞高剂量组（NBP-H 组），每组 8 只。检测各组大鼠血清肌酐（Scr）、血清胱抑素 C（Cys-C）、血尿素氮（BUN）
2、和血清白细胞介素（IL）-1、IL-18 水平，苏木素-伊红（HE）染色观察各组肾组织病理损伤情况，采用蛋白质印迹法和免疫组织化学法检测肾组织中炎症因子、核因子（NF）-B 信号通路及细胞焦亡相关蛋白表达水平。结果 与 Sham 组比较，IRI 组肾组织损伤较为严重，Scr、Cys-C、BUN 和血清IL-1、IL-18 水平均升高，蛋白质印迹法结果显示 NOD 样受体蛋白（NLRP3）、Gasdermin D（GSDMD）、半胱氨酸天冬氨酸蛋白酶（Caspase）-1、IL-18、IL-1、NF-B p65、p-NF-B p65 蛋白相对表达量均增加，免疫组织化学染色结果显示 NF-B p6
3、5、p-NF-B p65、IL-1、IL-18 和 NLRP3 蛋白表达均增多。与 IRI 组比较，PDTC 组、NBP-L 组和 NBP-H 组肾组织的损伤程度均减轻，Scr、Cys-C、BUN 和血清 IL-18、IL-1 水平均下降，蛋白质印迹法结果显示 NLRP3、GSDMD、Caspase-1、IL-1、IL-18、NF-B p65、p-NF-B p65 蛋白表达DOI:10.3969/j.issn.1674-7445.2023.04.010基金项目：贵州省卫生健康委科学技术基金（gzwkj2021-220）；国家自然科学基金培育项目 gyfynsfc（2020）-30；贵州医科大学
4、附属医院 2022 年博士科研启动基金（gyfybsky-2022-31）作者单位：550004 贵阳，贵州医科大学附属医院泌尿外科作者简介：张瑞波（ORCID:0009-0009-3056-7298），硕士研究生，肾移植相关缺血-再灌注损伤与保护，Email:通信作者：沈俊（ORCID:0000-0003-1208-5833），博士，副主任医师，肾移植相关缺血-再灌注损伤与保护，Email:结构式摘要HE 染色观察肾组织病理学改变免疫组化染色检测炎症因子、NF-B 信号通路及细胞 焦亡相关蛋白IRISham 组IRI 组PDTC 组NBP-H 组丁苯酞或 PTDC 或 0.9%氯化钠注射液N
5、BP-L 组蛋白质印迹法检测炎症因子、NF-B 信号通路及细胞 焦亡相关蛋白Scr、BUN、Cys-C 水平检测IL-1、IL-18 水平检测540第 14 卷器官移植肾 脏 缺 血-再 灌 注 损 伤（ischemia-reperfusion injury，IRI）通常由器官移植、肾部分切除手术、败血症等引起，如不及时采取有效措施加以预防，很容易发展为急性肾损伤，导致病死率上升1-6。肾 IRI【Abstract】Objective To elucidate the mechanism of dl-3-N-butylphthalide(NBP)on renal ischemia-reperf
6、usion injury(IRI)in rat models.Methods Forty SD rats were randomly divided into the sham operation group(Sham group),model group(IRI group),NF-B inhibitor pyrrolidine dithiocarbamate group(PDTC group),low-dose NBP group(NBP-L group)and high-dose NBP group(NBP-H group),with 8 rats in each group.Serum
7、 creatinine(Scr),serum cystatin C(Cys-C),blood urea nitrogen(BUN)and serum interleukin(IL)-1 and IL-18 levels were detected in all groups.Pathological injury of renal tissues in each group was observed by Hematoxylin-eosin(HE)staining.The expression levels of inflammatory factors and nuclear factor(
8、NF)-B signaling pathway and cell pyroptosis-related proteins in renal tissues were measured by Western blot and immunohistochemical staining.Results Compared with the Sham group,renal tissue injury was more severe,and the levels of Scr,Cys-C,BUN and serum IL-1 and IL-18 were all up-regulated in the
9、IRI group.Western blot showed that the relative expression levels of NOD-like receptor protein(NLRP3),Gasdermin D(GSDMD),cysteinyl aspartate specific proteinase(Caspase)-1,IL-18,IL-1,NF-B p65 and p-NF-B p65 proteins were all up-regulated,and immunohistochemical staining revealed that the expression
10、levels of NF-B p65 and p-NF-B p65,IL-1,IL-18 and NLRP3 proteins were all up-regulated in the IRI group.Compared with the IRI group,renal tissue injury was alleviated,and the levels of Scr,Cys-C,BUN and serum IL-18 and IL-1 were down-regulated in the PDTC,NBP-L and NBP-H groups.Western blot showed th
11、at the expression levels of NLRP3,GSDMD,Caspase-1,IL-1,IL-18,NF-B p65 and p-NF-B p65 proteins were down-regulated,and immunohistochemical staining indicated that the expression levels of NF-B p65,p-NF-B p65,IL-1,IL-18 and NLRP3 proteins were down-regulated in the PDTC,NBP-L and NBP-H groups,respecti
12、vely.Compared with the NBP-L group,renal tissue injury was mitigated,and the levels of Scr,Cys-C,BUN,serum IL-18 and IL-1 were all down-regulated in the NBP-H group.Western blot showed the expression levels of NLRP3,GSDMD,Caspase-1,IL-1,IL-18,NF-B p65 and p-NF-B p65 proteins were down-regulated in t
13、he NBP-H group.Immunohistochemical staining indicated that the expression levels of NF-B p65,p-NF-B p65,IL-1,IL-18 and NLRP3 proteins were down-regulated in the NBP-H group.Conclusions NBP may down-regulate the activity of NF-B/NLRP3 signaling pathway and reduce the expression levels of cell pyropto
14、sis-related proteins and inflammatory factors after renal IRI,thereby suppressing cell pyroptosis and alleviating renal IRI.【Key words】Dl-3-N-butylphthalide;Ischemia-reperfusion injury;Acute renal injury;Cell pyroptosis;Inflammation;NOD-like receptor protein 3(NLRP 3);Serum creatinine;Blood urea nit
15、rogen;Nuclear factor(NF)-B signaling pathway;Serum cystatin C(Cys-C);Interleukin-1;Interleukin-18均减少，免疫组织化学染色结果显示 NF-B p65、p-NF-B p65、IL-1、IL-18 和 NLRP3 蛋白表达均下降。与NBP-L 组比较，NBP-H 组肾组织的损伤程度减轻，Scr、Cys-C、BUN 和血清 IL-18、IL-1 水平均下降，蛋白质印迹法结果显示 NLRP3、GSDMD、Caspase-1、IL-1、IL-18、NF-B p65、p-NF-B p65 蛋白表达均减少，免疫组
16、织化学染色结果显示 NF-B p65、p-NF-B p65、IL-1、IL-18 和 NLRP3 蛋白表达均下降。结论 丁苯酞可下调 NF-B/NLRP3 信号通路的活性，降低肾 IRI 后焦亡相关蛋白的表达水平及炎症因子水平，进而抑制细胞焦亡，减轻肾 IRI。【关键词】丁苯酞；缺血-再灌注损伤；急性肾损伤；细胞焦亡；炎症；NOD 样受体蛋白 3（NLRP3）；血清肌酐；血尿素氮；核因子（NF）-B 信号通路；血清胱抑素 C（Cys-C）；白细胞介素-1；白细胞介素-18【中图分类号】R617，R816.8【文献标志码】A【文章编号】1674-7445（2023）04-0010-08Dl-3-N-butylphthalide alleviates renal ischemia-reperfusion injury by down-regulating NF-B signaling pathway and inhibiting cell pyroptosis in rat models Zhang Ruibo,Shen Kaiwen,Wang Qiang,Yuan Qiang,Shen
