CAJ | 학술논문

目的:分析B16-F10黑色素移植瘤及其肺转移瘤的差异表达蛋白,以筛选黑色素瘤转移相关的分子标志.方法:应用荧光差异凝胶电泳(two-dimensional differential gel electrophoresis,2D-DIGE)结合基质辅助激光解析电离飞行时间质谱技术(matrix assisted laser desorption ionisation time-of-flight mass spectrometry,MALDI-TOF-MS)分离鉴定B16-F10黑色素移植瘤及其肺转移瘤的差异表达蛋白,部分差异蛋白经Real-time PCR进行mRNA表达水平验证.结果:Decyder6.0软件分析结果显示2D-DIGE图谱分辨率高、重复性好,30个蛋白点在实验组和对照组间存在表达差异(Iratiol≥2,P<0.01),经质谱分析和数据库查询鉴定出9个蛋白在实验组表迭上调.包括肌红蛋白(myoglobin,MB)、波形蛋白(vimentin,VIM)、磷酸甘油激酶1(phosphoglycerate kinasel,PGK1)、磷酸丙糖异构酶(Triosephosphate isomerase,TPI或TIM)、重链结合蛋白(heavy-chain binding protein,BiP)、α-烯醇化酶(α-enolase或enolasel)、β-肌动蛋白(β-actin)、γ-肌动蛋白(γ-actin)、层连蛋白结合蛋白(laminin-binding protein),这些蛋白主要参与了细胞骨架构成、糖酵解等生物学过程.Real-time PCR结果显示糖酵解酶PGK1及TPI mRNA表达水平在实验组显著高于对照组(P=0.001,0.003),变化趋势与蛋白质组学相一致.结论:小鼠黑色素瘤转移过程与多种蛋白的异常表达有关,糖酵解酶PGK1、TPI可能参与了黑色素瘤的转移过程.
목적:분석B16-F10흑색소이식류급기폐전이류적차이표체단백,이사선흑색소류전이상관적분자표지.방법:응용형광차이응효전영(two-dimensional differential gel electrophoresis,2D-DIGE)결합기질보조격광해석전리비행시간질보기술(matrix assisted laser desorption ionisation time-of-flight mass spectrometry,MALDI-TOF-MS)분리감정B16-F10흑색소이식류급기폐전이류적차이표체단백,부분차이단백경Real-time PCR진행mRNA표체수평험증.결과:Decyder6.0연건분석결과현시2D-DIGE도보분변솔고、중복성호,30개단백점재실험조화대조조간존재표체차이(Iratiol≥2,P<0.01),경질보분석화수거고사순감정출9개단백재실험조표질상조.포괄기홍단백(myoglobin,MB)、파형단백(vimentin,VIM)、린산감유격매1(phosphoglycerate kinasel,PGK1)、린산병당이구매(Triosephosphate isomerase,TPI혹TIM)、중련결합단백(heavy-chain binding protein,BiP)、α-희순화매(α-enolase혹enolasel)、β-기동단백(β-actin)、γ-기동단백(γ-actin)、층련단백결합단백(laminin-binding protein),저사단백주요삼여료세포골가구성、당효해등생물학과정.Real-time PCR결과현시당효해매PGK1급TPI mRNA표체수평재실험조현저고우대조조(P=0.001,0.003),변화추세여단백질조학상일치.결론:소서흑색소류전이과정여다충단백적이상표체유관,당효해매PGK1、TPI가능삼여료흑색소류적전이과정.
Objective: To investigate differentially expressed protein profiles in B16-F10 grafted melanoma and its metastasis in the lung in order to identify molecular markers of melanoma metastasis. Methods: Differentially expressed proteins in B16-F10 grafted melanoma and its metastatic lesion in the lung were isolated and identified by fluorescence two-dimensional differential gel electrophoresis(2D-DIGE)coupled with matrix assisted laser desorption ionisation time-of-flight mass spectrometry(MALDI-TOF-MS).Some of identified proteins were further confirmed by Real-time PCR analysis. Results: High resolutional images of differential gel electrophoresis were obtained and 9 of 30 differentially expressed proteins (IRatiol≥2,P<0.01)were identified by MALDI-TOF-MS.The expression of Myoglobin(MB),vimentin(VIM),phosphoglycerate kinase 1(PGK1),Triosephosphate isomerase(TPI or TIM),heavy-chain binding protein(BiP),α-enolase,β-actin,γ-actin,and laminin-binding protein were up-regulated in the experimental group compared with the control group.These proteins were involved in the cytoskeletal formation,glycolysis and so on.Real-time PCR analysis showed up-regulation of mRNA expression of PGK1 and TPI in the experimental group(P=0.001 and 0.003),which was in consistent with the resuits of proteomic analysis. Conclusion: A variety of abnormally expressed proteins contribute to the metastasis of mice melanoma.Glycolytic enzymes PGK1 and TPI may be involved in this process.