YKL-40在肺纤维化大鼠中的动态表达

Dynamic expression of pulmonary YKL-40 protein in rats with bleomycin-induced pulmonary fibrosis

目的观察YKL-40在博莱霉素诱导的肺间质纤维化大鼠肺组织中的表达。方法以博莱霉素(7.5mg/kg体质量)气管内注入复制大鼠肺纤维化模型,对照组气管内注入等体积生理盐水。分别于第7天、第14天、第21天分批处死大鼠。通过H-E染色、Masson染色,并依据Szapiel评分方法判断肺组织肺泡炎及肺纤维化程度,采用免疫组化、real-timeRT-PCR及蛋白质印迹分析测定肺组织YKL-40的表达。结果博莱霉素组大鼠肺组织第7天肺泡炎程度(2.8±0.45)与对照组(0.42±0.25)相比差异有统计学意义(P<0.01),之后肺泡炎症减轻,但至第21天(1.8±0.84)与对照组间差异仍有统计学意义(P<0.05);肺组织纤维化程度在第14天(1.7±0.73)即与对照组(0.2±0.45)相比差异有统计学意义(P<0.05),至第21天(2.9±0.56)与对照组间差异有统计学意义(P<0.01)。YKL-40mRNA相对表达量在第7天(3.71±0.25)即高于对照组(P<0.05);蛋白质印迹分析结果显示,蛋白水平第14天(0.56±0.24)高于对照组(0.23±0.07,P<0.05),第21天(1.15±0.19)与对照组相比差异有统计学意义(P<0.01)。免疫组化结果显示,博莱霉素诱导大鼠肺组织YKL-40的上调表达呈时间依赖性,且主要来源于平滑肌细胞、肺泡巨噬细胞、肺泡上皮细胞及成纤维细胞。结论随着大鼠肺纤维化进展,YKL-40表达增加。YKL-40可能参与肺纤维化过程。

Objective To observe the expression of YKL-40 protein in the pulmonary tissues of rats with bleomycininduced pulmonary fibrosis （PF）. Methods The PF model group was induced with intratracheal instillation of bleomycin solution （7.5 mg/kg） and the control group was treated with normal saline. On day 7, 14, 21 after bleomycin challenge, rats were sacrificed and the pulmonary tissues were harvested. H-E staining, Masson staining and Szapiel score were employed to determine alveolitis and pulmonary fibrosis. Expressions of YKL-40 in lung tissues were detected by real-time RT PCR, Western blotting analysis and immunohistochemistry method. Results Bleomycin instillation induced alveolitis in the lung of rats, with inflammation score being significantly higher on day 7 （2.8_--t-0.45, P〈0.01） and on day 21 （1.8_＋0.84, P〈0.05） compared with that of control group （0.42±0.25）. Pulmonary fibrosis degrees in model group was significantly higher on day 14 （1.7±0.73, P〈0.05） and on day 21 （2.9±0.56, P〈0.01） compared with that of control group （0.2±0.45）. YKL-40 mRNA （YKL-40/β-actin） expression was significantly increased on day 7 （3.71±0.25） after bleomycin treatment compared with the control group （P 〈 0. 05）. Western blotting analysis showed that YKL-40 protein expression was significantly increased on day 14（0.56±0.24,P〈0.05） and on day 21（1.15±0.19, P〈0.01） after bleomycin treatment compared with the control group （0.23±0.07）. The results of immunohistochemistry showed that bleomycin up-regulated YKL-40 expression in a time-dependent manner, and YKL-40 expression was mainly located in the smooth muscle cells, alveolar macrophages, alveolar epithelium and fibroblasts. Conclusion YKL-40 expression may contribute to the pulmonary fibrosis, and may participate in the pathogenesis of pulmonary fibrosis.