Expression profiling of genes involved in tumor - host stroma cell interaction in the metastasis of human fibrosarcoma cell (HT1080)

1. 人纤维肉瘤细胞HT1080转移过程中肿瘤-宿主间质细胞相互作用相关基因的选择1)HT1080高转移克隆与亲本克隆间基因表达无显著差异，选择HT1080高转移克隆，利用human Cancer 1.2 Atlas expression Arrays （Clonetech）软件进行cDNA macroarrays分析基因表达谱。HT1080高转移克隆与亲本体外培养基因表达无显著差异。2)肺微转移HT1080细胞中纤连蛋白1 （fn1）基因过表达采用裸鼠尾静脉注射HT1080肺转移模型，研究HT1080细胞中参与肺微转移形成的基因表达。在肺微转移中发现了几个过表达和过表达的基因。其中，纤连蛋白1 （fn1）基因的表达量较体外培养HT1080细胞增加了50倍。实时RT-PCR证实FN 1基因转录明显上调，激光捕获显微解剖/实时RT-PCR在肺微转移的栓塞HT1080肿瘤细胞中定位过表达。3)原位接种肌肉内肿瘤细胞后，连接处血小板红蛋白基因下调，肿瘤细胞生长和肺微转移增强。采用裸鼠愈合垫接种（h组）和肌肉内接种（m组）模型，研究HT1080细胞侵袭转移过程中肿瘤-宿主间质细胞相互作用相关基因的表达。与h组比较，m组肿瘤细胞生长和肺微转移明显增强。使用人类肿瘤1.2 Atlas表达阵列和小鼠Atlas表达阵列分析M组和h组肿瘤细胞及宿主基质细胞的基因表达。阵列分析显示m组与h组相比，连接血小板红蛋白基因表达明显下调。在mRNA水平上采用实时RT-PCR检测，在蛋白水平上采用特异性血小板蛋白单克隆抗体免疫组化检测。H-组与m -组间质细胞间无明显基因表达差异。在人软组织肉瘤中参与连接蛋白的下调：临床应用在不同类型的人软组织肉瘤中，我们分别在蛋白和mRNA水平上评价了连接蛋白基因的表达。结血小板红蛋白在滑膜肉瘤细胞中过度表达，尤其是上皮型。在恶性纤维组织细胞瘤（MFH）中，有肺转移的肿瘤与无肺转移的肿瘤相比，连接血小板红蛋白的表达不仅在蛋白水平上，而且在mRNA水平上均显著下调。少

1. Selection of genes involved in the tumor-host stroma cell interaction in the metastasis of human fibrosarcoma cell (HT1080)1) No significant difference of gene expressions between parent and highly metastatic clones of HT1080Highly metastatic clones were selectively cloned from HT1080 and gene expression profilings were analysed by cDNA macroarrays using Human Cancer 1.2 Atlas Expression Arrays (Clonetech). No significant difference of gene expressions between parent and highly metastatic clones of HT1080 in vitro cultures was found out.2) Fibronectin 1 (FN 1) gene is overexpressed in HT1080 cells in pulmonary micrometastasesGene expressions involved in pulmonary micrometastasis-formation were investigated with a lung metastatic model of HT1080 using nude mice injected through tail vein. Several overexpressed and underexpressed genes were demonstrated in pulmonary micrometastasis. Of them, expression of fibronectin 1 (FN 1) gene was increased 50 times compared with that of in vitro- … More cultured HT1080 cells. The prominent upregulation of FN 1 gene transcription was verified by real time RT-PCR and the overexpression was localized in embolized HT1080 tumor cells of the pulmonary micrometastases by laser captured microdissection / real time RT-PCR.3) Junction plakoglobin gene is down-regulated in orthotopically inoculated intramuscle tumor cells showing enhanced growth and pulmonary micrometastasesGenes involved in the tumor - host stroma cell interaction in the invasion & metastasis of HT1080 cells were investigated using heal pad inoculation (H-group) and intramuscular inoculation (M-group) models in nude mice. M-group showed significantly enhanced tumor cell growth and pulmonary micro-metastases compared with H-group. Gene expressions of tumor cells as well as host stroma cells of M- and H-groups were profiled using both Human Cancer 1.2 Atlas Expression Arrays and Mouse Atlas Expression Arrays. Array-analyses disclosed that expression of junction plakoglobin gene was significantly down-regulated in M-group compared with that of H-group. The down-regulation of junction plakoglobin gene was confirmed by real time RT-PCR at mRNA level and by immunohistochemistry using specific monoclonal antibody to plakoglobin at protein level. No significant genes differently expressed between stroma cells of H- and M-groups were shown.2. Involvement of the down regulation of junction plakoglobin in human soft tissue sarcomas : Clinical applicationExpressions of junction plakoglobin gene were evaluated at both protein and mRNA levels in various kinds of human soft tissue sarcomas.Junction plakoglobin was overexpressed in synovial sarcoma cells, especially epithelial type. In malignant fibrous histiocytomas (MFH), expressions of junction plakoglobin was significantly down-regulated, at not only protein but also mRNA level, in tumors with pulmonary metastases compared with those without pulmonary metastases. Less