CAJ | 학술논문

目的分析不同剂量照射后成纤维细胞中玻连蛋白及胶原蛋白表达改变及其时间变化规律,初步探讨玻连蛋白作为放射性肺纤维化指标的意义.方法采用137 Cs辐射源,对人胚肺成纤维细胞WI-38、IMR-90各照射0、4、6、8、10和12 Gy,并于照后6、12、24、36、48和60 h收集细胞及培养上清液,采用Western blot法检测玻连蛋白和胶原蛋白Ⅰ、Ⅲ含量,采用实时PCR法及ELISA法分别检测玻连蛋白的转录水平及其细胞培养上清中的蛋白表达量.结果 Western blot结果显示,受照射后两种成纤维细胞玻连蛋白及胶原蛋白Ⅰ、Ⅲ表达呈正相关性(r=0.40～0.79,P＜0.05),都显著高于未经照射的对照组(t=3.04 ～25.45,P＜O.05),且蛋白表达的剂量效应和时间效应均呈现先增高后降低的变化趋势.玻连蛋白与胶原蛋白在8 ～10 Gy照后48 h,表达量最高(t=2.92～18.86,P＜0.05).Real-time PCR结果与ELISA结果显示,玻连蛋白mRNA表达及其在上清中含量受照射剂量影响较显著(F=27.09 ～ 42.62,P＜0.05),而受时间影响较小.结论照射后成纤维细胞中玻连蛋白在细胞内、上清中及mRNA表达水平均可能作为照射后纤维化的指标.而8 Gy照射及照射后48 h是产生成纤维细胞放射性纤维化的较优实验条件.
목적 분석불동제량조사후성섬유세포중파련단백급효원단백표체개변급기시간변화규률,초보탐토파련단백작위방사성폐섬유화지표적의의.방법 채용137 Cs복사원,대인배폐성섬유세포WI-38、IMR-90각조사0、4、6、8、10화12 Gy,병우조후6、12、24、36、48화60 h수집세포급배양상청액,채용Western blot법검측파련단백화효원단백Ⅰ、Ⅲ함량,채용실시PCR법급ELISA법분별검측파련단백적전록수평급기세포배양상청중적단백표체량.결과 Western blot결과현시,수조사후량충성섬유세포파련단백급효원단백Ⅰ、Ⅲ표체정정상관성(r=0.40～0.79,P＜0.05),도현저고우미경조사적대조조(t=3.04 ～25.45,P＜O.05),차단백표체적제량효응화시간효응균정현선증고후강저적변화추세.파련단백여효원단백재8 ～10 Gy조후48 h,표체량최고(t=2.92～18.86,P＜0.05).Real-time PCR결과여ELISA결과현시,파련단백mRNA표체급기재상청중함량수조사제량영향교현저(F=27.09 ～ 42.62,P＜0.05),이수시간영향교소.결론 조사후성섬유세포중파련단백재세포내、상청중급mRNA표체수평균가능작위조사후섬유화적지표.이8 Gy조사급조사후48 h시산생성섬유세포방사성섬유화적교우실험조건.
Objective To analyze radiation induced alterations of vitronectin and collagen expressions in fibroblasts at different times post-irradiation,so as to evaluate the potential to apply vitronectin as a biomarker of radiation-induced lung fibrosis.Methods The human fibroblast cells WI-38 and IMR-90 were irradiated with 137Cs γ-rays at doses of 0 (control),4,6,8,10 and 12 Gy,respectively.The cells and its supernatant were collected at 6,12,24,36,48 and 60 h post-irradiation.The expressions of vitronectin and collagen Ⅰ and Ⅲ were analyzed by Western blot,PCR and ELISA.Results After irradiation,the expressions of vitronectin and collagen Ⅰ and Ⅲ were positively correlated (r=0.40-0.79,P＜0.05) and were all significantly higher than that in control group (t =3.04-25.45,P ＜0.05) and reached the highest expression levels at 48 h after 8-10 Gy of irradiation (t =2.92-18.86,P ＜ 0.05).Analyses of Real-time PCR and ELISA assay showed that expressions of vitronectin mRNA and its protein level in the cell lysis were significantly increased by radiation (F =27.09-42.62,P ＜ 0.05).Conclusions The expressions of vitronectin in cellular supernatant and its mRNA may be a potential biomarker of radiation-induced fibrosis,and 48 h after 8 Gy irradiation may be an optimum condition of measurement.