Activation of HMGI-C gene and its significance in well-differentiated liposarcomas

为阐明高分化脂肪肉瘤发生发展的分子机制，本研究从HMGI-C、HMGI-C、HMGI-C基因激活的分子机制及HMGI-C基因在成纤维细胞转化中的作用研究如下：1.HMGI-C基因在蛋白质和信息水平的表达：(1)免疫组织化学和蛋白质印迹分析(2)原位杂交(ISH)分析；(2)HMGI-C基因激活的分子机制：(1)巢式RT-PCR和DNA测序(2)Southern印迹分析用于基因扩增；(3)3‘端快速扩增(3’RACE)嵌合体基因形成。HMGI-C-/-或HMGI-C+/+小鼠成纤维细胞体外转化研究高分化脂肪肉瘤细胞HMGI-C基因高表达(21/21例)。相比之下，去分化脂肪肉瘤发生在分化良好的脂肪肉瘤…中肿瘤多为高度恶性，HMGI-C表达缺失。高分化脂肪肉瘤中HMGI-C蛋白表达的相对分子质量和HMGI-C基因蛋白编码区的cDNA巢式RT-PCR结果均未见异常。Southern印迹分析未见HMGI-C基因在高分化脂肪肉瘤中的扩增。3‘RACE分析尚未证实HMGI-C在高分化脂肪肉瘤中形成新的嵌合体基因。体外研究表明，HMGI-C-/-小鼠来源的成纤维细胞比HMGI-C-/-小鼠来源的成纤维细胞转化更频繁，并且HMGI-C-/-成纤维细胞的转化被HMGI-C-/-小鼠成纤维细胞的转化抑制，这些结果表明HMGI-C基因的激活参与了脂肪肉瘤的发生发展。体内和体外研究表明，HMGI-C在分化良好的脂肪肉瘤中过表达可能抑制其向高度恶性的肿瘤发展，即去分化脂肪肉瘤。本研究结果提示，在分化程度较高的脂肪肉瘤中，3‘UTRAUUA基序和/或启动子活性的异常可能与HMGI-C的激活有关。较少

In order to elucidate the molecular mechanism involved in the development and progression of well-differentiated liposarcoma, expression of HMGI-C, molecular mechanism of the activation of HMGI-C gene and functions of HMGI-C in the transformation of fibroblasts were investigated as followings :1. Expressions of HMGI-C gene at both protein and message levels :(1) Immunohistochemical and Western blot analyses(2) Analysis by in situ hybridization (ISH)2. Molecular mechanism of the activation of HMGI-C gene :(1) nested reverse transcription polymerase chain reaction (nested RT-PCR)and DNA sequencing(2) Southern blot analysis for gene amplification(3) 3' rapid amplifiction of cDNA ends (3'RACE) for chimera gene formation3. In vitro transformation studies using fibroblasts derived from HMGI-C-/- or HMGI-C +/+ mouseTumor cells of well differentiated liposarcoma overexpressed HMGI-C gene (21/21 cases). In contrast, dedifferentiated liposarcomas, which developed in well-differentiated liposarco … More ma and are highly malignant, showed loss of HMGI-C expressions. No abnormality was disclosed in the moleular weight of HMGI-C protein expressed in well-differentiated liposarcomas or in cDNA of protein coding region of HMGI-C gene demonstrated by nested RT-PCR.Southern blot analysis did not show any amplification of HMGI-C gene in well differentiated liposarcomas. 3'RACE analysis has not demonstrated any newly formed chimera gene of HMGI-C in well-differentiated liposarcoma so far. In vitro studies showed that fibroblasts derived from HMGI-C -/- mouse transformed more frequently than those derived from HMGI-C -/- mouse, and the transformation in HMGI-C -/- fibroblasts was inhibited by transfection of HMGI-C cDNA.These data clearly demonstrated that activation of HMGI-C gene is involved in the development and progression of well-differentitated liposarcoma. Both in vivo and in vitro data indicated that HMGI-C overexpressed in well-differentiated liposarcomas may inhibit progression to highly malignant tumor, i.e. dedifferentiated liposarcoma. The present data suggested that abnormalies in 3' UTR AUUUA motifs and / or promotor resion might contribute to the activation of HMGI-C in well- differentiated liposarcomas. Less