【摘要】 目的探讨硝基油酸(OA-NO2)对肾急性缺血再灌注(I/R)模型小鼠肾的保护作用。方法 C57小鼠分为假手术、I/R模型对照、I/R+OA-NO2500μg·kg-1和I/R+油酸(OA)500μg·kg-1组。I/R小鼠使用乙醚麻醉后,开腹采用夹闭双侧肾动脉30 min,去除血管夹,再灌注24 h制备肾I/R模型。I/R+OANO2和I/R+OA组在去除血管夹后分别ip给予OA-NO2和OA 500μg·kg-1,每6 h注射1次。假手术和I/R模型组ip给予乙醇0.8 ml·kg-1。24 h后处死小鼠,取血和肾组织,用全自动生化检测仪检测小鼠血浆尿素氮(BUN)和肌酐(Cr)水平,HE染色检测肾组织病理改变,ELISA检测血浆肿瘤坏死因子α(TNF-α)浓度,实时PCR检测肾组织细胞黏附分子1(ICAM-1)、白细胞介素1β(IL-1β)、烟酰胺腺嘌呤二核苷酸磷酸氧化酶胞浆亚基(p47)和烟酰胺腺嘌呤二核苷酸磷酸氧化酶催化亚基(gp91)基因表达,Western蛋白质印迹法检测肾组织TNF-α和IL-1β蛋白表达,ELISA检测肾组织丙二醛(MDA)含量。结果 I/R模型组小鼠血浆BUN和Cr水平较假手术组明显增高(P<0.01);OA-NO2处理后,与I/R模型组比较,血浆BUN水平降低36%(P<0.01),Cr水平降低44%(P<0.01);I/R+OA组BUN和Cr水平无明显变化。I/R模型组小鼠肾组织出现肾小管上皮细胞坏死、细胞结构消失、肾小管管腔扩张和肾小管管腔管型堵塞等改变,血浆TNF-α浓度、肾组织ICAM-1,IL-1β,p47和gp91mRNA表达、TNF-α和IL-1β蛋白表达及MDA含量均较假手术组明显升高(P<0.01);应用OA-NO2处理后,与I/R模型组比较,肾组织病理改变减轻,肾小管管腔扩张明显改善,未发现明显的肾小管管腔管型堵塞;血浆TNF-α浓度由I/R模型组的(590±73)ng·L-1降低至(259±71)ng·L-1(P<0.01),肾组织ICAM-1,IL-1β,p47和gp91基因表达以及TNF-α和IL-1β蛋白表达降低(P<0.01),MDA含量由I/R模型组的(3.6±0.7)mol·g-1组织降低至(1.8±0.4)mol·g-1组织(P<0.01);应用OA处理后,上述指标与I/R模型组比较均无明显差异。结论 OA-NO2对肾I/R导致的急性肾损伤具有明显的治疗作用,其作用机制可能与抗炎通路有关。更多还原

【Abstract】 OBJECTIVE To evaluate the potential therapeutic effect of nitro-oleic acid(OA-NO2)on acute kidney injury.METHODS C57 mice were subjected to bilateral renal ischemia for 30min,followed by 24h of reperfusion.After ischemia,the mice were ip given OA-NO2500μg·kg-1 or oleic acid(OA)500 μg·kg-1 every 6 hduring the 24-h recovery period,while those of ischemia/reperfusion(I/R)model group were ip given 0.8ml·kg-1 ethanol every 6h.The blood and kidney tissue of mice were collected after 24h reperfusion.Plasma urea nitrogen(BUN)and creatinine(Cr)were tested by the automatic biochemical de-tector.Histopathological changes of the kidney were detected by HE staining.The plasma concentration of tumor necrosis factor-α(TNF-α)and renal tissue malondialdehyde(MDA)content was detected by ELISA,and the mRNA expression of renal tissue intercellular adhesion molecule-1(ICAM-1),interleu-kin-1β(IL-1β),nicotinamide adenine dinucleotide phosphate oxidase cytoplasm subunit(p47)and nicotin-amide adenine dinucleotide phosphate oxidase catalytic subunit(gp91)was examined by real-time PCR.The protein expression of renal tissue TNF-αand IL-1β was determined using Western blotting.RESULTS Compared with sham group,the plasma BUN and Cr level was elevated in the mice of I/R model group(P<0.01),but decreased after adminstration by OA-NO2 by 36% and 44%,respectively,compared with I/R model group(P<0.01).Morphology changes of the kidney in I/R model group,including renal tubu-lar epithelial cell necrosis,cell structure collapse,tubular expansion and tube cast jam,were observed.The concentration of TNF-αand MDA,the mRNA expression of ICAM-1,IL-1β,p47 and gp91 and the pro-tein expression of TNF-αand IL-1β were significantly increased in I/R model group compared with sham group(P<0.01).After treatment with OA-NO2,the pathological changes in renal tissue were attenua-ted,while renal tubular lumen expansion was decreased compared with the I/R model group,but without obvious renal tubular cast jam.The plasma TNF-αconcentration decreased from(590±73)ng·L-1 in I/R model group to(259±71)ng·L-1 in I/R+OA-NO2 group(P<0.01).The renal tissue MDA content was decreased from(3.6±0.7)mol·g-1 tissue in I/R model group to(1.8±0.4)mol·g-1 tissue in I/R+OA-NO2 group(P<0.01),while the level of BUN,Cr and MDA,the expression of ICAM-1,IL-1β,p47,gp91,TNF-αand IL-1β and histological damage were not significantly different between I/R model and I/R +OA groups.CONCLUSION OA-NO2 attenuates kidney I/R injury likely by inhibiting the inflammatory response.更多还原