Analyses of Molecular Mechanisms underlying Medulloblastoma Oncogenesis

虽然髓母细胞瘤（MB）的分子发病机制仍然知之甚少，Wnt和Hedgehog发育信号通路的重要性最近变得明显。为了进一步了解正常小脑细胞转化为MB的分子事件，我们利用抑制性消减杂交（SSH）技术来鉴定失调的分子，因此可能在MB肿瘤发生中起重要作用。在相应的正常小脑组织的cDNA的消减杂交后，从人和Ptch杂合小鼠MBs的SSH文库被产生。通过对文库的差异筛选，分离并测序了100多个在肿瘤中上调的cDNA片段。使用NCBI BLAST程序鉴定这些序列。我们选择了参与致癌重要过程的基因，如细胞增殖、凋亡调节和小脑分化，以进一步分析。在人MB文库中鉴定的基因包括Unc 33样蛋白（ULIP）、SOX 4、神经元蛋白、果蝇BarH样1的哺乳动物同源物（BARHL 1）、核基质蛋白NRP/B（ENC 1）和同源框OTX 2基因。在小鼠MB中发现上调的基因包括细胞周期蛋白D2、胸腺生成素、Musashi-1、蛋白磷酸酶2A抑制剂-2（I-2 PP 2A）和Unc 5 H4（D）。使用半定量RT-PCR，这些基因的mRNA表达水平显着高于人类MB比正常小脑。此外，一些基因主要在MB中表达。这些基因的过度表达在正常细胞转化中所起的作用仍有待充分确定。

Although the molecular pathogenesis of medulloblastoma (MB) remains poorly understood, the importance of the Wnt and Hedgehog developmental signaling pathways has recently become apparent. In order to further the understanding of molecular events that transform a normal cerebellar cell into a MB, we utilized the technique of suppression subtractive hybridization (SSH) to identify molecules that are dysregulated and therefore may be important in MB oncogenesis. After the subtractive hybridization of cDNA from corresponding normal cerebellar tissue, SSH libraries from both human and Ptch heterozygous murine MBs were generated. Through differential screening of the libraries, over 100 cDNA fragments up-regulated in the tumor were isolated and sequenced. These sequences were identified using the NCBI BLAST program. We selected genes involved in oncogenically important processes such as cellular proliferation, apoptosis regulation, and differentiation of the cerebellum to analyze further. Genes identified in the human MB library included Unc33-like protein (ULIP), SOX4, neuronatin, the mammalian homologue of Drosophila BarH-like 1(BARHL1), the nuclear matix protein NRP/B (ENC1), and the homeobox OTX2 gene. Genes found to be up-regulated in the murine MB included cyclin D2, thymopoietin, Musashi-1, protein phosphatase 2A inhibitor-2 (I-2PP2A), and Unc5H4 (D). Using semi-quantitative RT-PCR, the mRNA expression levels for these genes are markedly higher in human MB than in the normal cerebellum. Furthermore some genes were expressed predominantly in MB. The role played by the over-expression of these genes in the transformation of normal cells remains to be fully determined.