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目的探讨大鼠肾缺血再灌损伤后血清中TNF-α、IL-6的变化以及Fas、Bcl-蛋白在肾小管上皮细胞中表达的变化,并分析在AKI发病机制中的作用。方法用无损伤动脉夹钳夹大鼠双侧肾蒂40min制成IRI动物模型。观察缺血再灌注(IR)后2h、6h、12h、24h、48h血清中TNF-α、IL-6的变化(ELISA法)及SCr的变化(苦味酸法)和以上各灌注时间点肾脏的病理变化(HE染色)以及IR后12h Fas、Bcl-2蛋白的表达情况(免疫组化法)和与肾小管上皮细胞凋亡的关系(TUNEL法)。结果肾小管上皮细胞凋亡在IR后12h明显增加,以皮髓交界区明显,阳性细胞率(%)为77.74±5.00%,而假手术组(Sham组)阳性率(%)为2.03±0.82%,两组差异具有显著性(P<0.01);血清TNF-α、IL-6在IR后12h含量明显增高,分别为130.58±36.57、352.70±58.32(pg/ml),并与SCr的升高对应;Fas在IR后12h肾小管上皮细胞阳性率(%)为83.23±2.22%,Sham组阳性率(%)为3.02±0.82%(P<0.01);Bcl-2在IR12h组及Sham组中肾小管上皮细胞阳性率(%)分别为0.40±0.05%、2.00±0.90%,差异具有显著性(P<0.01)。结论 TNF-α、IL-6参与了AKI的病理生理过程;TNF-α可能通过上调Fas和下调Bcl-2参与对肾小管上皮细胞的凋亡调控。
Objective To investigate the changes of serum TNF-α, IL-6 and the expression of Fas and Bcl-2 in renal tubular epithelial cells after renal ischemia-reperfusion injury in rats and analyze the role in the pathogenesis of AKI. Methods The IRI animal model was established by clamping the bilateral renal pedicles with an artery without injury for 40 minutes. The changes of serum TNF-α and IL-6 (ELISA method) and changes of SCr (picric acid method) at 2h, 6h, 12h, 24h and 48h after ischemia reperfusion were observed. The pathological changes (HE staining) and the expression of Fas and Bcl-2 protein (immunohistochemistry) and the relationship with the apoptosis of renal tubular epithelial cells (TUNEL) were observed 12h after IR. Results The apoptosis of renal tubular epithelial cells was significantly increased at 12 h after IR, and the positive rate of the positive cells was 77.74 ± 5.00% in the border zone of the skin and the positive rate (%) was 2.03 ± 0.82 in the sham operation group (Sham group) %, There was significant difference between the two groups (P <0.01). The levels of TNF-α and IL-6 in serum increased significantly at 12h after IR, which were 130.58 ± 36.57,352.70 ± 58.32 (pg / ml) The positive rate of Fas in renal tubular epithelial cells was 83.23 ± 2.22% at 12h after IR and 3.02 ± 0.82% in Sham group (P <0.01). The expressions of Bcl-2 in IR12h group and Sham group The positive rates of renal tubular epithelial cells (%) were 0.40 ± 0.05% and 2.00 ± 0.90%, respectively. The difference was significant (P <0.01). Conclusions TNF-α and IL-6 are involved in the pathophysiological process of AKI. TNF-α may be involved in the regulation of apoptosis of renal tubular epithelial cells by up-regulating Fas and down-regulating Bcl-2.