CAJ | 학술논문

目的分析甲基丙烯酸环氧丙酯(glycidyl methacrylate,GMA)致人支气管上皮(16HBE)细胞恶性转化过程中不同时点阿片样物质结合蛋白/细胞黏附分子样蛋白(opioid binding protein/cell adhesion molecule-like,OPCML)基因甲基化状态,探讨其在GMA诱导16HBE细胞恶性转化过程中OPCML基因甲基化状态的变化及其意义.方法收获GMA第10、20、30代16HBE细胞,应用甲基化芯片和甲基化特异性PCR(methylation-specific-PCR,MSP)技术检测OPCML基因在GMA诱导16HBE细胞恶性转化不同时点甲基化状态,采用实时荧光定量PCR(qPCR)法检测该基因在不同转化时点基因表达量,并与同期DMSO溶剂对照组细胞比较.结果甲基化芯片结果显示,GMA染毒组16HBE细胞在第10、20、30代OPCML基因均发生甲基化,对照组均未发生甲基化,且MSP法检测结果与芯片结果一致.qPCR结果显示,与同期溶剂对照组相比,GMA染毒组16HBE细胞在第20、30代OPCML基因表达量均下调,经甲基化转移酶抑制剂(5-氮杂-2'-脱氧胞苷(5-Aza-2'-deoxycytidine,5-Aza-cdr)处理后,该基因与同代龄GMA组细胞相比,表达量水平上升,差异有统计学意义(P值分别为0.004、0.001、0.001).结论 OPCML基因可能作为GMA诱导16HBE细胞发生恶性转化的一个特异指标.
목적 분석갑기병희산배양병지(glycidyl methacrylate,GMA)치인지기관상피(16HBE)세포악성전화과정중불동시점아편양물질결합단백/세포점부분자양단백(opioid binding protein/cell adhesion molecule-like,OPCML)기인갑기화상태,탐토기재GMA유도16HBE세포악성전화과정중OPCML기인갑기화상태적변화급기의의.방법 수획GMA제10、20、30대16HBE세포,응용갑기화심편화갑기화특이성PCR(methylation-specific-PCR,MSP)기술검측OPCML기인재GMA유도16HBE세포악성전화불동시점갑기화상태,채용실시형광정량PCR(qPCR)법검측해기인재불동전화시점기인표체량,병여동기DMSO용제대조조세포비교.결과 갑기화심편결과현시,GMA염독조16HBE세포재제10、20、30대OPCML기인균발생갑기화,대조조균미발생갑기화,차MSP법검측결과여심편결과일치.qPCR결과현시,여동기용제대조조상비,GMA염독조16HBE세포재제20、30대OPCML기인표체량균하조,경갑기화전이매억제제(5-담잡-2'-탈양포감(5-Aza-2'-deoxycytidine,5-Aza-cdr)처리후,해기인여동대령GMA조세포상비,표체량수평상승,차이유통계학의의(P치분별위0.004、0.001、0.001).결론 OPCML기인가능작위GMA유도16HBE세포발생악성전화적일개특이지표.
Objective To analyze the opioid binding protein/cell adhesion molecule-like (OPCML) methylation status at different stages of malignant transformation of human bronchial epithelial (16HBE) cells induced by Glycidyl Methacrylate (GMA) and to explore the effect of OPCML methylation in the process of malignant transformation.Methods Cells were harvested at different stages (the 10th generation,the 20th generation and the 30th generation).To verify the Methylation chip result of OPCML methylation status in the process of malignant transformation,we detected it by methylation-specific-PCR (MSP);Real-time fluorescence Quantitative PCR (qPCR) were applied to measure the gene expression levels of OPCML at different transformed stage,and compared with the control groups (treated with DMSO).Results Based on the result of methylation chip,the gene of OPCML methylation occurred in all stages,which was consistent to the result of MSP;qPCR showed that the levels of gene expression decreased in the 20th generation and 30th generation.Conclusion Methylation status of OPCML gene promoter could be considered as a stable and specific biomarker in the transformation process.