摘要
目的 研究细胞外基质蛋白对肿瘤细胞基质金属蛋白酶 (MMP)基因表达的影响及机理。方法 以纤粘连蛋白处理SKOV3卵巢癌细胞后 ,用明胶 酶谱法和反转录PCR观察MMP分泌及基因表达。以MMP 2启动子转染细胞 ,通过测定萤火虫酶活性 ,观察纤粘连蛋白对MMP 2启动子活性的影响。细胞p5 3含量采用Westernblot分析。结果 5 μg/mL纤粘连蛋白刺激MMP 2分泌 ,但不影响MMP 9分泌 ,这一作用随纤粘连蛋白浓度增加而增强。经 10 μg/mL纤粘连蛋白刺激 1h后 ,细胞内MMP 2mRNA量显著增加 ;作用时间延长 ,诱导作用减弱。纤粘连蛋白可诱导转染细胞中萤火虫酶活性增加 ,AP 1功能抑制剂姜黄素 (5 0 μmol/L)不能阻断这一作用。经纤粘连蛋白处理后 ,细胞内p5 3含量明显增加。结论 纤粘连蛋白可刺激卵巢癌细胞中MMP 2分泌 ,诱导MMP 2启动子活性增强及mRNA含量增加 ,这一作用可能与p5 3蛋白转录因子活性有关 ,而与AP 1通路无关。
Objective To investigate the effects of extracelluar matrix proteins on matrix metalloproteinases (MMPs) expression by cancer cells, and the underlying mechanisms.Methods Following stimulation of SKOV3 ovarian cancer cells by fibronectin, MMPs secretion and cellular mRNA contents were assayed by gelatin zymography and RT PCR, respectively. The activity of MMP 2 promoter was monitored by cellular luciferase activity after the cells were transfected with MMP 2 promoter luciferase construct. Cellular p53 contents were determined by Western blot analysis.Results Fibronectin (5 μg/ml) was found to stimulate the secretion of MMP 2 but not MMP 9 by SKOV3 cells. The stimulation was enhanced with the increase in fibronectin concentration. When SKOV3 cells were treated with 10 μg/ml fibronectin for 1hr, the cellular MMP 2 mRNA dramatically increased. However, with increase in stimulation time, MMP 2 mRNA content decreased. Fibronectin induced an increase in luciferase activity in cells transfected with MMP 2 promoter construct, whilst curcumin (50 μmol/L), a potent transcription factor AP 1 inhibitor, could not block the fibronectin induced increase in MMP 2 promoter activity. Fibronectin also induced an increase in p53 content of SKOV3 cells.Conclusion Fibronectin stimulates ovarian cancer cells to secret MMP 2 via its enhancing effect on MMP 2 promoter activity with resultant increase in MMP 2 transcription. The effect might involve a pathway associated with p53 but independent of AP 1. Subject
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
2000年第2期109-112,共4页
Chinese Journal of Oncology
关键词
纤粘连蛋白
基质金属蛋白酶
卵巢癌细胞
Ovarian neoplasms
Matrix metalloproteinase
Fibronectin
Gene expression
