Analysis of gene expression associated cancer invasion and metastasis using in situ RT-PCR method

1.检测尿激酶型纤溶酶原激活物(UPA)、尿激酶型纤溶酶原激活剂受体(UPAR)及纤溶酶原激活物抑制物-1、2(PAI-1、PAI-2)在膀胱癌组织中的表达，探讨其在膀胱癌发生发展中的作用。UPA、uPAR、PAI-1和PAI-2mRNA在浸润性肿瘤中的表达水平明显升高，并与组织学分级有关。我们还分析了SPARC基因在膀胱癌中的表达及其与临床病理表现的关系。浸润性肿瘤的SPARC表达水平明显高于浅表性肿瘤。这些结果表明uPA、uPAR、PAI-1、PAI-2和SPARC在膀胱癌的发生发展中起重要作用。为了明确其在尿路上皮癌中的作用，本研究采用明胶酶谱技术研究了活化形式的基质金属蛋白酶-2的表达。侵袭性肿瘤组织中活化形式的基质金属蛋白酶-2的表达水平明显升高。高水平的激活形式的基质金属蛋白酶-2与病程特定的生存期缩短密切相关。提示基质金属蛋白酶-2的活化形式在尿路上皮癌的侵袭中起重要作用。我们建立了非侵袭性和侵袭性尿路上皮癌细胞系，并将侵袭性尿路上皮癌细胞系移植到SCID小鼠的膀胱内。我们分析了侵袭性尿路上皮癌细胞系向SCID小鼠膀胱形成的膀胱癌组织中MT1-MMP1和MMP2mRNA的表达定位。MT1-MMPmRNA表达位于肿瘤晚期，间质细胞表达MMP2mRNA。此外，我们现在正在分析手术获得的组织中mRNAs的定位。

1. we investigated the expression of urokinase-type plasminogen activator (uPA), urokinase-type plasminogen activator receptor (uPAR), and plasminogen activator inhibitor-1, 2 (PAI-1, PAI-2) in bladder cancer to determine the role of their gene expression in invasion and progression of bladder cancer. Expression levels of uPA, uPAR, PAI-1 and PAI-2 mRNA was significantly higher in invasive tumor, and was correlated with histological grade. We also analyzed the gene expression of SPARC in bladder cancer and its relationship with clinical-histopathological manifestation. Invasive tumors expressed significantly higher level of SPARC than superficial tumors. These results showed that uPA, uPAR, PAI-1, PAI-2, and SPARC played an important role in the tumor progression of bladder cancer.2. The present study investigated the expression of the activated form of MMP-2 using zelatin zymography in order to define its role in urothelial cancer. Expression levels of activated forms of MMP-2 were significantly higher in invasive tumor tissue. High levels of activated forms of MMP-2 were strongly associated with shortened course-specific survival. These findings suggest that the activated form of MMP-2 plays a significant role in invasion of urothelial cancer.3. We established non invasive and invasive urothelial cancer cell lines, and transplanted invasive urothelial cancer cell line to the bladder of SCID mice. We analyzed the mRNA localization of MT1-MMP and MMP-2 mRNA in bladder tumor tissues formed from invasive urothelial cancer cell line to the bladder of SCID mice. MT1-MMP mRNA was observed in the advanced part of tumor, and MMP-2 mRNA was observed in interstitial cells . Furthermore we are now analysing mRNAs localization in tissues obtained from surgery.