摘要
目的:膜型丝氨酸蛋白酶(ST14/MT-SP1)和它的同源物在细胞迁移和肿瘤转移中起重要作用。本研究目的是评估ST14/MT-SP1过度表达如何影响结直肠癌细胞的侵袭能力。方法:全长人ST14/MT-SP1基因被瞬时转染到结直肠癌细胞系RKO。表达产物由Ni2+-亲和层析柱纯化并通过明胶酶谱法分析蛋白的明胶酶活性。用ECM体外侵润试验确定细胞的体外侵袭力。用cDNA微阵列法测定ST14/MT-SP1转染细胞中MMPs和TIMPs表达变化情况。用实时定量PCR来验证这些基因表达的变化。结果:人全长ST14/MT-SP1基因被转染到结直肠癌细胞系RKO后,纯化表达的蛋白具有明胶酶的活力。RKO细胞过度表达ST14/MT-SP1后其体外侵润转移能力显著增强(P<0.01),而ST14/MT-SP1蛋白被阻断后使SW480和SW620细胞的侵袭能力降低(P<0.01)。进一步发现,ST14/MT-SP1过度表达使RKO细胞的MT2-MMP(MMP-15)表达显著上调(约2.5倍)和TIMP2表达下调(约0.35倍)。结论:ST14/MT-SP1过度表达导致了结直肠癌细胞侵袭力增强,这种能力的增强是由于ST14/MT-SP1自身具有明胶酶的活性和ST14/MT-SP1能上调MT2-MMP与下调TIMP-2的表达。因此,ST14/MT-SP1过度表达可能增强结直肠癌细胞的侵袭能力。
AIM： To evaluate how ST14/MT- SPI overexpression alters invasiveness of colorectal cancer cells. METHODS： Human full length ST14/MT- SP1 gene was transiently transfected into colorectal cancer （RKO） cell lines. The expression products were purified by chromatography on Ni^2＋ - affinity resin column and analyzed for gelatinase activity by gelatin zymography. Cell invasion and migration were determined by ECM invasion assay in vitro. RNA was isolated from stable STI4 -transfected RKO cells and a cDNA microarray was utilized to detect the change in expression of MMPs and TIMPs. Real - time quantitative PCR was used to validate the change of expression. RESULTS： The full length ST14/MT - SP1 was transfected and expressed in RKO cells. The expressed protein showed a gelatinase activity. In addition, invasiveness of RKO was significantly enhanced by ST14/MT - SP1 overexpression （ P 〈 0. 01 ）. Blockage of ST14/MT- SP1 resulted in a decrease in the invasiveness of SW480 and SW620 （P 〈0.01 ）. Furthermore, MT2 -MMP （ MMP - 15 ） expression was significantly up - regulated （ 2.5 - fold change） and TIMP2 down - regulated （ 0.35 - fold change） by ST14/MT - SP1 overexpression in RKO. CONCLUSION： STI4/MT - SP1 overexpression results in an increase in invasiveness of coloreetal cancer RKO cells. Increased invasiveness is due to an increase in the gelatinase activity of ST14/MT - SP1 anti a change in up - regulated MT2 - MMP and down - regulated TIMP - 2 expression. Therefore, ST14/MT - SP1 overexpression enhances invasiveness of coloreetal cancer cells.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2008年第12期2356-2362,共7页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.30200325)
浙江省卫生厅课题资助项目(No.2007B098)
浙江大学青年教师基金资助项目(No.491020-542970)
教育部博士学科点专项科研基金资助项目(No.J20070738)
