表皮生长因子受体在苯并（a）芘致肺肿瘤中的表达变化

Study on the expression of epidermal growth factor receptor protein during benzo（a）pyrene induced carcinogenesis

目的 探讨表皮生长因子受体（EGFR）在苯并（a）芘（BaP）致肺肿瘤中的表达变化.方法 利用课题组前期构建的BaP诱导的人支气管上皮细胞恶性转化模型（BTC）作为研究对象,采用细胞免疫荧光和Western blot法分析比较恶性转化细胞（BTC细胞）和同期未转化对照组细胞（16HBE细胞）中的EGFR蛋白表达变化.将健康雌性SPF级SD大鼠36只,将其以单纯随机法分为2组,每组18只.染毒组以肺穿刺法注射10 mg/ml玉米油-BaP,每只浓度0.2 ml,构建SD雌性大鼠肺肿瘤模型,收集肺脏组织,利用组织免疫荧光和Western blot法分析肺脏组织中EGFR蛋白的表达变化.采用t检验对EGFR蛋白相对灰度值进行分析.结果 细胞免疫荧光结果显示,与正常16HBE细胞比较,BTC细胞中EGFR蛋白的荧光强度较高;进一步利用Western blot检测EGFR的蛋白表达发现,对照组相对灰度值为1.04±0.13,BTC细胞的相对灰度值为2.32±0.12,BTC细胞的EGFR蛋白的表达水平较高,差异有统计学意义（t=12.39,P〈0.001）.SD大鼠肺肿瘤模型检测发现,染毒组大鼠肺组织存在弥漫性的肺泡间隔增厚、肺泡壁破坏以及肺泡融合等,肺组织内可见较大肿块,病理学分析为未分化细胞癌,提示肺肿瘤模型建立成功.免疫荧光结果显示,与对照组相比,BaP诱导的动物肺癌模型的肺组织中EGFR的荧光强度明显增强;Western blot分析结果证实,对照组和染毒组EGFR蛋白的相对灰度值分别为0.21±0.03、1.30±0.07,染毒组动物肺肿瘤组织中EGFR蛋白的表达水平高于对照组,差异有统计学意义（t=12.84,P〈0.001）.结论 在BaP致癌模型中,EGFR蛋白表达上调,EGFR在BaP致癌中可能发挥重要作用.

Objective To explore the expression of epidermal growth factor receptor（EGFR） protein during benzo（a）pyrene（BaP） induced carcinogenesis. Methods This study, we firstly utilized immunofluorescence assay and Western-blot to examine EGFR expression of the BaP which was constructed previously by project team induced malignant transformation human bronchial epithelial cell （BTC） and the control （16HBE cell）. Then, we selected 36 healthy SD rats, divided into two groups according to simple random method, 18 rats each group. The constructed rat lung neoplasm model induced by pulmonary injection of BaP （10 mg/ml of BaP solution in 0.2 ml corn oil）, contrast group use 0.2 ml corn oil, lung tissue was collected and the EGFR expression of lung tissue was detected by immunofluorescence assay and Western blot. T analysis was used to test the different of EGFR between two groups. Results Immunofluorescence analysis showed that the EGFR expression in BTC was significantly higher than 16HBE cell. Meanwhile, Western blot also was used to confirmed this result, the relative expression of EGFR protein in the rats of the model group the control group were 1.04±0.13 and 2.32±0.12, respectively, and the difference was statistically significance （t=12.39, P〈0.001）. In vivo, well-defined tumor was found in the rat with pulmonary injection of BaP, and the lung showed diffuse alveolar septal thickening, alveolar wall destruction and pulmonary alveoli fusion, which suggested that the rat lung neoplasm model was constructive successfully. Furthermore, we found the EGFR expression of lung was increased dramatically in the rat lung neoplasm model by immunofluorescence detection and Western blot. The relative expression of EGFR protein in the rats of the model group the control group were 0.21±0.03 and 1.30±0.07,respectively, and the difference was statistically significance （t=12.84, P〈0.001）. Conclusion Expression of EGFR protein was increased during BaP carcinogenesis, and EGFR may play an important role in the carcinogenesis of BaP.