Huang Qiang, Sun Li-Jun, Dong Jun, Li Xiao-Nan, Wang Ai-Dong, Lan Qing
Neurosurgery Department and Brain Tumor Research Laboratory,The Second Affiliated Hospital, Suzhou University, Suzhou, Jiangsu, 215004, PR China.
Ai Zheng. 2003 Jul;22(7):673-9.
BACKGROUND & OBJECTIVE: Glioma cells can be induced in vitro from low differentiation state to high differentiation state with differentiation-inducing agents, as our previous studies have showed. However, the relevant molecular mechanisms remained unclear. This paper was to establish differentiation-inducing gene expression pedigree of glioma cells and clone new relevant genes, for the purpose of offering genomic information on the molecular mechanisms of differentiation-inducing effects of tumor cells.
A low differentiated human brain glioma cell line (SHG-44-9) was treated with differentiation-inducing agents. The differentiation-inducing effects on glioma cells were determined using MTT assay, colony-forming efficiency in double-layer soft agar, cell morphological changes, flow cytometry, and fluoroimmunoassay of glial fibrillary acidic protein expression. Gene expression differences of tumor cells before and after inducing were assayed with cDNA microarray techniques. The relevant genes were cloned with differential display polymerase chain reaction (DD-PCR), and verified with Northern blot analysis, then the genomic information about these genes were analyzed with bioinformatics technologies.
At dose of 1.75 mmol x L(-1), sodium butyrate caused a marked proliferation inhibitory effect on the SHG-44-9 cells, and drove them to a more mature phenotype. The expression of 98 genes changed obviously when were screened with gene chips immobilized with 18,000 human cDNAs, which were further proved by Northern blot analysis. Twelve differentiation-associated genes were cloned, including MIBP1 gene, which can regulate the c-myc gene expression and may be relevant with glioma cell differentiation.
Ninety-eight genes associated with differentiation and 12 genes were cloned. These data may help to further study the molecular mechanisms of glioma cells differentiation, and to find new targets for gene therapy against glioma.
如我们之前的研究所示，胶质瘤细胞可在体外被分化诱导剂从低分化状态诱导至高分化状态。然而，相关分子机制仍不清楚。本文旨在建立胶质瘤细胞分化诱导基因表达谱系并克隆新的相关基因，以提供关于肿瘤细胞分化诱导作用分子机制的基因组信息。
用分化诱导剂处理低分化人脑胶质瘤细胞系（SHG-44-9）。采用MTT法、双层软琼脂集落形成率、细胞形态变化、流式细胞术以及胶质纤维酸性蛋白表达的荧光免疫测定法来确定对胶质瘤细胞的分化诱导作用。用cDNA微阵列技术检测诱导前后肿瘤细胞的基因表达差异。用差异显示聚合酶链反应（DD-PCR）克隆相关基因，并用Northern印迹分析进行验证，然后用生物信息学技术分析这些基因的基因组信息。
丁酸钠在1.75 mmol·L⁻¹剂量时对SHG-44-9细胞有明显的增殖抑制作用，并使其向更成熟的表型发展。用固定有18,000个人类cDNA的基因芯片筛选时，98个基因的表达明显改变，Northern印迹分析进一步证实了这一点。克隆了12个与分化相关的基因，包括可调节c-myc基因表达且可能与胶质瘤细胞分化相关的MIBP1基因。
克隆了98个与分化相关的基因和12个基因。这些数据可能有助于进一步研究胶质瘤细胞分化的分子机制，并找到针对胶质瘤基因治疗的新靶点。
