淮山药灌胃预处理对大鼠肾脏缺血再灌注损伤保护作用的实验研究

Protective Effect of Pretreatment with Rhizome Dioscoreae Lavage on Renal Ischemia-reperfusion Injury in Rats

目的探讨淮山药灌胃预处理对大鼠肾脏缺血再灌注损伤的可能保护作用。方法 48只3-4周龄雄性SD大鼠随机分为缺血再灌注淮山药灌胃预处理组（淮山药组,n=24）和缺血再灌注无菌生理盐水灌胃对照组（生理盐水组,n=24）。淮山药组、生理盐水组在缺血再灌注损伤前连续灌胃5天。分别于再灌注损伤后2、12、24、48h四个时间点随机选淮山药组、生理盐水组各6只大鼠再次手术行下腔静脉采血和收获肾脏标本。测定血清肌酐（Scr）、尿素氮（BUN）以及血清丙二醛（MDA）含量。肾组织常规病理切片H-E染色光镜下参照Paller评分标准进行评分,TUNEL法检测细胞凋亡。结果再灌注后12、24、48h,同时间点淮山药组与生理盐水组比较,其血清Scr、BUN含量显著下降[BUN 12h：（15.30±2.60）mmol/L比（22.07±4.57）mmol/L,24h：（19.56±3.89）mmol/L比（34.75±6.32）mmol/L,48h：（14.72±2.99）mmol/L比（20.16±2.40）mmol/L;Scr 12h：（56.38±8.70）μmol/L比（98.72±12.37）μmol/L,24h：（81.53±10.17）μmol/L比（128.52±17.30）μmol/L;48h：（55.01±9.71）μmol/L比（80.72±11.13）μmol/L;P〈0.05,P〈0.01]。再灌注后2、12、24h,淮山药组血清MDA含量较同时间点生理盐水组显著下降[2h：（4.74±0.50）nmol/L比（7.45±1.03）nmol/L,12h：（6.19±0.68）nmol/L比（10.52±1.57）nmol/L,24h：（5.16±0.58）nmol/L比（6.93±1.32）nmol/L;P〈0.05,P〈0.01]。肾组织学评分于再灌注后12、24、48h,同时间点淮山药组较生理盐水组有显著减低[12h：（7.50±0.55）分比（8.50±0.55）分,24h：（6.17±0.75）分比（7.50±1.05）分,48h：（4.17±0.75）分比（5.50±1.05）分;P〈0.05]。再灌注后2h就有凋亡细胞出现,12h时可见大量凋亡细胞脱落到肾小管腔中,此时间点淮山药组凋亡细胞数较生理盐水组明显减少[（10.17±2.14）个/HPF比（13.67±2.94）个/HPF,P〈0.05]。结论淮山药灌胃预处理可以减轻缺血再灌注模型大鼠肾组织氧化损伤和减少肾小管上皮细胞凋亡的发生,降低血清Scr、BUN和MDA水平,有效保护肾功能。

Objective To investigate the protective effect of pretreatment with Rhizome Dioscoreae on alleviating ischemia-reperfusion injury in rats.Methods Fifty-four Spargue-Dawley rats were randomly divided into 3 groups：sham operation group（n =6）,ischemia-reperfusion group with pretreatment of Phizoma Dioscoreae via intragastric administration（Rhizoma dioscoreae group,n=24）,and ischemia-reperfusion group with aseptic normal saline lavege（aseptic saline group,n=24）.The rats in Rhizoma Dioscoreae group and aseptic saline group were pretreated with rhizoma Dioscoreae and placebo aseptic normal saline for 5d,respectively before being subjected to bilateral renal vascular occlusion for 45 min.At 2,12,24 and 48 h after reperfusion,every 6 rats in each group were randomly euthanized for blood sampling and kidney harvesting.The blood urea nitrogen（BUN） and serum creatinine（Scr）and serum malondialdehyde（MDA） were measured.Renal morphologic changes were scored with Paller＇s criterion on hematoxylin and eosin（HE） stained sections.Renal tubular cell apoptosis were detected with terminal deoxynucleotidyl transferase-mediated d UTP-biotin nick end labeling procedure（TUNEL）.Results After 12,24,and 48 h of reperfusion,the Scr and BUN value in Rhizoma Dioscoreae group were significantly lower than those in aseptic saline group（BUN 12h：15.3±2.60mmol/L vs 22.07±4.57mmol/L,24h：19.56±3.89mmol/L vs 34.75±6.32mmol/L,48h：14.72 ±2.99mmol/L vs 20.16 ±2.40mmol/L; Scr 12h：56.38 ±8.72mmol/L vs 98.72 ±12.37mmol/L,24h：81.53 ±10.17mmol/L vs 128.50±17.30mmol/L,48h：55.01±9.71mmol/L vs 80.72±11.13mmol/L; P〈0.05 or P〈0.01）.At 2,12,and 24 h,the serum content of MDA in Rhizoma Dioscoreae group were significantly lower than those in aseptic saline group（2h：4.74 ±0.50nmol/L vs 7.45 ±1.03nmol/L,12h：6.19 ±0.68nmol/L vs 10.52 ±1.5nmol/L,24h：5.16 ±0.58nmol/L vs 6.93 ±1.32nmol/L;P〈0.05 orP〈0.01）.Renal apoptosis score at 12,24,and 48 h of reperfusion in Rhizoma Dioscoreae group were less than that in aseptic saline group（12h：7.50±0.55 vs 8.50±0.55,24h：6.17±0.75 vs 7.00±1.05; 48h：4.17±0.75 vs 5.50±1.05; P〈0.05）.The tubular cell apoptosis was detected in the kidney at 2h after ischemia-reperfusion injury and reached the highest level at 12h（Rhizoma Dioscoreae group 10.17±2.14/HPF vs aseptic saline group 13.67±2.94/HPF,P〈0.05）.Conclusion The pretreatment of Rhizoma Dioscoreae lavage in rats results in lessening the oxidative damage and apoptosis,lowering the level of Scr,BUN and MDA,thereby protecting kidney from ischemia-reperfusion injury.