CAJ | 학술논문

目的观察丙酮酸乙酯(EP)对小鼠缺血/再灌注(I/R)损伤肾脏的保护作用,研究其对肾组织中炎症因子及丝裂素活化蛋白激酶(MAPK)信号通路相关蛋白表达的影响. 方法将50只雄性BABL/c小鼠按随机数字表法分为假手术组(n=8)、模型组(n=10)和EP治疗组(n=32);EP治疗组再分为EP预处理组和EP 4、6、12 h处理组,每组8只,分别于制模前30 min及制模后4、6、12 h腹腔注射EP 40 mg/kg.采用夹闭双肾动脉30 min制备肾I/R损伤模型.于I/R 24 h取肾组织,采用实时聚合酶链反应(PCR)检测白细胞介素(IL-1β、IL-6)、肿瘤坏死因子-α(TNF-α)、细胞间黏附分子-1(ICAM-1)、高迁移率族蛋白B1(HMGB1)的mRNA表达;采用蛋白质免疫印迹法(Western blotting)检测MAPK信号转导通路中细胞外信号调节激酶1/2(ERK1/2)、c-Jun氨基末端蛋白激酶(JNK)、p38MAPK等的蛋白表达. 结果实时PCR结果显示,与假手术组比较,模型组小鼠肾组织IL-1β、IL-6、TNF-α、ICAM-1及HMGB1的mRNA表达均显著增高(IL-1β:12.05±8.08比3.18±1.13;IL-6:10.26±6.85比0.81±0.34;TNF-α:5.83±3.85比0.67±0.34;ICAM-1:3.87±2.02比0.29±0.13;HMGB1:652.82±78.50比112.31±32.50,均P＜0.05);而EP各处理组能显著抑制上述炎症因子的表达,尤其以12 h处理组最为显著,分别为0.45±0.26、 0.66±0.13、 0.21±0.11、 0.05±0.02、 212.26±3.20(均P＜0.05).Western blotting结果显示,与假手术组比较,模型组小鼠肾组织磷酸化的ERK1/2、JNK、p38MAPK蛋白表达均显著升高(p-ERK1/2:1.13±0.38比0.48±0.34;p-JNK:1.40±0.15比0.36±0.15;p-p38MAPK:0.47±0.15比0.21±0.17,均P＜0.05);与模型组比较,EP各处理组在不同时间点均能显著抑制ERK1/2、JNK、p38MAPK的活化(均P＜0.05). 结论 EP能有效防治小鼠I/R肾脏损伤,可能与其调控炎症相关因子及MAPK信号转导的表达有关.
목적 관찰병동산을지(EP)대소서결혈/재관주(I/R)손상신장적보호작용,연구기대신조직중염증인자급사렬소활화단백격매(MAPK)신호통로상관단백표체적영향. 방법 장50지웅성BABL/c소서안수궤수자표법분위가수술조(n=8)、모형조(n=10)화EP치료조(n=32);EP치료조재분위EP예처리조화EP 4、6、12 h처리조,매조8지,분별우제모전30 min급제모후4、6、12 h복강주사EP 40 mg/kg.채용협폐쌍신동맥30 min제비신I/R손상모형.우I/R 24 h취신조직,채용실시취합매련반응(PCR)검측백세포개소(IL-1β、IL-6)、종류배사인자-α(TNF-α)、세포간점부분자-1(ICAM-1)、고천이솔족단백B1(HMGB1)적mRNA표체;채용단백질면역인적법(Western blotting)검측MAPK신호전도통로중세포외신호조절격매1/2(ERK1/2)、c-Jun안기말단단백격매(JNK)、p38MAPK등적단백표체. 결과 실시PCR결과현시,여가수술조비교,모형조소서신조직IL-1β、IL-6、TNF-α、ICAM-1급HMGB1적mRNA표체균현저증고(IL-1β:12.05±8.08비3.18±1.13;IL-6:10.26±6.85비0.81±0.34;TNF-α:5.83±3.85비0.67±0.34;ICAM-1:3.87±2.02비0.29±0.13;HMGB1:652.82±78.50비112.31±32.50,균P＜0.05);이EP각처리조능현저억제상술염증인자적표체,우기이12 h처리조최위현저,분별위0.45±0.26、 0.66±0.13、 0.21±0.11、 0.05±0.02、 212.26±3.20(균P＜0.05).Western blotting결과현시,여가수술조비교,모형조소서신조직린산화적ERK1/2、JNK、p38MAPK단백표체균현저승고(p-ERK1/2:1.13±0.38비0.48±0.34;p-JNK:1.40±0.15비0.36±0.15;p-p38MAPK:0.47±0.15비0.21±0.17,균P＜0.05);여모형조비교,EP각처리조재불동시간점균능현저억제ERK1/2、JNK、p38MAPK적활화(균P＜0.05). 결론 EP능유효방치소서I/R신장손상,가능여기조공염증상관인자급MAPK신호전도적표체유관.
Objective To investigate the effects of ethyl pyruvate(EP) on expression of proinflammatory related gene and proteins of mitogen-activated protein kinase(MAPK) in renal tissues in ischemic/reperfusion(I/R) injury in mice. Methods Fifty male BABL/c mice were randomly divided into sham operation group(n=8),model group(n=10),and EP treatment group(n=32).EP treatment group was subdivided into EP pretreatment group(administration of 40 mg/kg EP 30 minutes before reproduction of model,n=8),and 4,6 and 12 hours treatment groups(administration of 40 mg/kg EP 4,6 and 12 hours after reproduction of model,respectively,n=8 in each group).Bilateral renal artery was occluded with a microvascular clamp for 30 minutes to reproduce kidney I/R injury model,and the kidney was harvested at 24 hours after I/R.The mRNA expressions of interleukins(IL-1β,IL-6),tumor necrosis factor-α(TNF-α),intercellular adhesion molecule-1(ICAM-1) and high-mobility group box 1(HMGB1) were determined by real time reverse transcription-polymerase chain reaction(RT-PCR).The changes in protein levels of MAPKs[extracellular regulated protein kinase 1/2(ERK1/2),c-Jun N-terminal kinase(JNK),p38MAPK] were determined by Western blotting analysis. Results Real-time PCR assay showed that the mRNA expressions of IL-1β,IL-6,TNF-α,ICAM-1,HMGB1 in renal tissue were much higher than those in sham operation group(IL-1β:12.05±8.08 vs.3.18±1.13,IL-6:10.26±6.85 vs.0.81±0.34,TNF-α:5.83±3.85 vs.0.67±0.34,ICAM-1:3.87±2.02 vs.0.29±0.13,HMGB1:652.82±78.50 vs.112.31±32.50,all P<0.05);and the expression in EP treatment groups was markedly down-regulated than that in model group,especially in 12-hour treatment group(0.45±0.26,0.66±0.13,0.21±0.11,0.05±0.02,212.26±3.20,respectively,all P<0.05).Western blotting analysis revealed that the expression of the phosphorylated forms of ERK1/2,JNK,p38MAPK proteins was significantly higher than in sham operation group(p-ERK1/2:1.13±0.38 vs.0.48±0.34,p-JNK:1.40±0.15 vs.0.36±0.15,p-p38MAPK:0.47±0.15 vs.0.21±0.17,all P<0.05);the expression of the phosphorylated forms of ERK1/2,JNK,p38MAPK in each EP treatment group was significantly down-regulated compared with that in model group(all P<0.05). Conclusion EP can effectively protect kidney from acute injury produced by I/R,which may be related to the regulation of proinflammatory genes and the MAPKs in renal tissue.