TNF-α拮抗剂对大鼠缺血再灌注肺组织中致炎性细胞因子及HIF-1α表达的影响

摘要

目的 观察肿瘤坏死因子-α(TNF-α)拮抗剂TNFRI-Fc融合蛋白对大鼠肺缺血一再灌注损伤过程中肺内致炎性细胞因子释放与缺氧诱导因子-1α(HIF-lα)表达的影响.方法 选择体质量250～ 350 g的雄性SD大鼠54只,随机分为3组,假手术组(C组)6只仅行开胸,不行缺血及再灌注处理；Ⅰ组与Ⅱ组各24只,均行左肺缺血和再灌注处理,Ⅱ组在肺门夹闭前及肺门开放前5 min经颈静脉留置针推注溶于生理盐水中的可溶性TNFRl-Fc融合蛋白(3μg/kg).Ⅰ组与Ⅱ组于再灌注后1、3、6、12h各处死6只动物,C组于Ⅰ组、Ⅱ组再灌注后1h同时处死动物,均取左肺上2/3制成匀浆ELISA法检测TNF-α、白细胞介素(IL)-8、IL-6含量,下1/3通过免疫组化分析HIF-1α蛋白表达水平.结果 Ⅰ组TNF-α、IL-8、IL-6含量在1h即较C组明显升高；Ⅱ组各时间点肺组织匀浆内TNF-α、IL-8、IL-6含量较Ⅰ组明显降低,且TNF-α达峰时间明显错后；HIF-1α蛋白主要表达于肺泡上皮细胞的胞核或胞浆,再灌注后6h达高峰,表达水平Ⅱ组较Ⅰ组明显降低(P＜0.01),C组HIF-1α极少量表达.Ⅰ组和Ⅱ组的HIF-1α蛋白表达与TNF-α水平成正比.结论 TNF-α拮抗剂通过有效中和TNF-α并阻断其生物学活性,抑制致炎性细胞因子释放、减弱HIF-1α蛋白表达,起到减轻缺血再灌注损伤的作用.%Objective To investigate the effects of tumor necrosis factor-α (TNF-α) antagonist (TNFRI-Fc) on the release of proinflammatory cytokines and the expression of hypoxia-inducible factor-la (HIF-lα) in lung ischemia-reperfu-sion injury (I/R) in rats. Methods Fifty-four male SD rats (250 ~ 350 g) were randomly divided into sham operation group (group C,n=6),ischemia-reperfusion (I/R) group (group Ⅰ,n=24) and TNFRI-Fc group (group Ⅱ,n=2A). Rats were killed 1,3,6 and 12 h after reperfusion (6 rats in each time point) respectively in group Ⅰ and group Ⅱ. Values of TNF-a,IL-8 and IL-6 were detected by ELISA method using pulmonary homogenate of the left upper 2/3 lung tissue of rats. The HIF-la expression was detected by immunohistochemistry using left lower 1/3 pulmonary homogenate. Results The levels of TNF-a,IL-6 and IL-8 were significantly higher in group Ⅰ than those of group C and group Ⅱ (P < 0.05). But levels of TNF-α,IL-6 and IL-8 were significantly lower in group Ⅱ than those of group Ⅰ,and the peak level of TNF-a was significantly pushed back. The expressions of HIF-lα were mainly in the nucleus or cytoplasm of alveolar epithelial cells,and reached the peak 6 h after reperfusion. The expression levels of HIF-lα were significantly lower in group Ⅱ than those of group Ⅰ (P < 0.01). The very lower expression of HIF-lα was found in group C. The expressions of HIF-lα were proportional to the level of TNF-a in group I and group Ⅱ. Conclusion The TNF-a antagonist can effective neutralize TNF-a to block the biological activity,inhibit the release of proinflammatory cytokines and attenuate the expression of HIF-lα to protect the lung function against ischemia-reperfusion injury.