乌司他丁通过抑制高迁移率族蛋白B1信号转导通路对大鼠早期放射性肺损伤的防治作用

Prevention effects of ulinastatin on the early radiation-induced lung injury in rats by inhibiting high mobility group box 1 protein pathway

目的观察乌司他丁对大鼠早期放射性肺损伤的防治作用及其机制。方法选取45只健康雌性SD大鼠,根据随机数字表法将大鼠分为正常组、模型组和实验组,每组15只。正常组不做处理,其余2组均全胸接受6 MV-X射线单次照射20 Gy制作放射性肺损伤模型。实验组大鼠予照射后48 h起予以尾静脉注射乌司他丁1×105U·kg-1qd,其余2组予以尾静脉注射同等体积0.9%Na Cl溶液。2周后留取肺组织,苏木精-伊红(HE)染色观察肺组织形态,计数肺泡灌洗液白细胞和红细胞数,实时荧光定量聚合酶链式反应(qRT-PCR)和免疫印迹法(Western blot)测定高迁移率族蛋白B1(HMGB1)、晚期糖基化终末产物受体(RAGE)、核转录因子kappa B(NF-κB)、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)和环氧合酶2(COX-2)的表达水平。结果 HE染色结果显示,实验组大鼠肺泡炎症程度较模型组显著减轻,实验组肺泡灌洗液中白细胞和红细胞数分别(3.42±1.53)×109/L,(3.95±1.24)×109/L,模型组为(7.04±2.18)×109/L,(7.12±1.35)×109/L,实验组均较模型组明显减少(P<0.05),但与正常组细胞数无明显差异(P>0.05)。正常组大鼠肺组织中HMGB1、RAGE、NF-ΚB、TNF-α、IL-6和COX-2 mRNA的表达分别为1.01±0.16,1.01±0.18,1.03±0.27,1.10±0.50,1.01±0.17和1.01±0.16;模型组中上述基因mRNA的表达分别为4.66±1.43,4.18±0.82,4.31±1.03,3.41±0.91,4.22±0.79和4.41±0.99;实验组中上述基因mRNA的表达分别为1.82±0.73,1.55±0.62,1.70±1.39,1.77±0.95,1.45±0.55和1.85±1.48。与模型组比较,实验组肺组织HMGB1信号通路表达水平均下降,差异均有统计学意义(均P<0.05);实验组与正常组比较,各基因的表达水平差异无统计学意义(P>0.05)。结论乌司他丁可通过干扰HMGB1信号通路的表达抑制胸部照射后大鼠肺组织炎性改变。

Objective To observe the preventive and therapeutic effects of ulinastatin on early radiation-induced lung injury in rats and to explore its possible mechanism.Methods A total of 45 healthy female SD rats were randomly divided into normal group,model group and experimental group,with 15 rats in each group.Radiation-induced lung injury model were established in the model group and the experimental group（whole-thorax irradiation utilizing 6 MV-X-ray,20 Gy by X-ray）,while the normal group received no treatment.The experimental group were treated with ulinastatin（1 × 105 U·kg-1 qd） 48 h after irradiation,while the other two groups were given 0.9% Na Cl.After2 weeks,lung tissue was collected and HE staining was used to observe the morphology of lung tissue.The number of white blood cells and red blood cells in alveolar lavage fluid were calculated.The expression of high mobility group box-1（HMGB1）,receptor for advanced glycation end-products（RAGE）,nuclear factor kappa B（NF-κB）,tumor necrosis factor alpha（TNF-α）,interleukin 6（IL-6） and cyclooxygenase 2（COX-2） were measured by qRT-PCR and Western blot.Results The degree of alveolitis in experimental group was significantly alleviated when compared with the model group.The number of white blood cells and red blood cells in the alveolar lavage fluid of rats in the experimental group were（3.42 ± 1.53） × 109/L,（3.95 ± 1.24） × 109/L,while those in the model group were（7.04 ± 2.18） × 109/L,（7.12 ± 1.35） × 109/L,respectively.The number of white blood cells and red blood cells in the alveolar lavage fluid of rats in the experimental group were also significantly lower than that in the model group（P 〈 0.05）,but there was no significant difference when compared with the normal group（P 〉 0.05）.The expression of HMGB1,RAGE,NF-κB,TNF-α,IL-6 and COX-2 mRNA in lung tissue of normal group were 1.01 ± 0.16,1.01 ± 0.18,1.03 ± 0.27,1.10 ± 0.50,1.01 ± 0.17 and 1.01 ± 0.16.The expression of these mRNAs in the model group were 4.66 ± 1.43,4.18 ± 0.82,4.31 ± 1.03,4.41 ± 0.91,4.42 ± 0.79 and 4.41 ± 0.99,respectively.The expression of these mRNAs in the experimental group were 1.82 ± 0.73,1.55 ± 0.62,1.70 ± 1.39,1.77 ± 0.95,1.45 ± 0.55 and 1.85 ± 1.48,respectively.The expression of HMGB1 signaling pathway in the lung tissue of experimental group was significantly decreased when compared with the model group（P 〈 0.05）,while no significant difference was found between the experimental group and the normal group（P 〉 0.05）.Conclusion Ulinastatin could alleviate the inflammatory changes of rat lung tissue by regulating HMGB1 signaling pathway.