中枢神经细胞瘤和少突胶质细胞瘤基因差异表达的研究

Studies with DDRT-PCR on the Differential Display of Genes in Central Neurocytoma and Oligoden Droglioma

目的应用差异显示反转录PCR（differential display reverse transcription PCR,DDRT-PCR）技术,研究中枢神经细胞瘤（central neurocytoma,CN）和少突胶质细胞瘤（oligedendrogliomas,OG）基因表达差异。方法用DDRT-PCR技术比较2例CN、4例OG和2例正常脑组织标本基因表达的差异,鉴别、分离差异表达的基因片段,进行再扩增、克隆、测序。测序结果经BLAST检索进行同源性分析。结果CN、OG和正常脑组织中存在明显的基因表达差异。在选取的8条差异表达条带（W1～8）中,1个（W8）在CN中高表达,3个（W3,4,7）在OG中高表达,4个（W1、2、5、6）主要在正常脑组织中表达。BLAST查询证实,其中有2个（W3、8）差异片段所代表的基因编码为已知蛋白,为FAC1基因和AGA基因;有2个（W1、2）与人类染色体序列具有同源性,但无相关基因及功能的报道。另4个（W4、5、6、7）与鼠具有同源性,功能不详。结论差异显示片段W1所代表的基因可能是一个新的含有锌指结构的转录调节因子,位于人类第16号染色体上;基因FAC1在正常脑组织中不表达,在OG中高表达,可能是OG的相关基因。

Objective Central neurocytoma （CN） is a rare, benign intraventricular tumor of central nervous system （CNS）. Histologically, CN closely resembles oligodendroglioma（OG） in terms of morpholog. According to the WHO new classification of CNS tumors, CN is classified as neuronal and mixed neuronal-lial tumors of neuroepithelial tissue, and OG as oligodendroglial tumors of neuroepithelial tissue. The former is characterized by its high radiosensitivity, but the latter is sensitive to chemotherapy partly. This study aims at investigating the molecular mechanisms underlying the differences between CN and OG, and their pathogeneses. For this purpose, we applied fluorescent DD-CR/DDRT-PCR to identify differentially expressed genes in CN, OG and normal brain tissues. Methods In this study, tissues including 2 case of CN, 4 case of OG, and 2 normal brain tissues obtained during decompression surgery for cerebral trauma as control group. The total RNAs from frozen specimens were compared by fluorescent DDRT-PCR. Differential display bands of interest were chosen. Eight cDNA fragments isolated from DD denaturing gel were extracted and reamplified, cloned into pMD18-T vector and sequenced. Then the sequences of the bands were analyzed for homology with known genes using the BLAST search. Results Significant differences of gene expression were detected among CN, OG and normal brain tissues. Eight bands were selected. In these bands, one（W8） was highly expressed in CN, three（W3, 4, 7） in OG, and 4（W1, 2, 5, 6） in normal brain tissue. BLAST analysis and a GenBank search of these 8 clones showed that 2（W3, 8 ） represented known genes, namely fetal Alz-50 reactive clone 1 （FACl） located in chromosome 17q24.3 and aspartylglucosaminidase（AGA） in 4q32-q33. The protein encoded by FAC1 is a novel member of the PHD/LAP zinc finger family. Two clones（W1, 2） showed homology to human without known gene sequence. Another four clones （W4, 5, 6, 7） shared sequence homology to Mus musculus or Rattus norvegicus. Conclusion Differentially displayed band W1, a sequence located in chromosome 16, is expressed highly in normal brain tissue but not in CN and OG. It might be a novel gene encoding a protein of transcription factors regulator which contains a zinc finger motif. FAC1 is expressed at low level or not in normal brain tissues but higher in OG. It is supposed that FAC1 may play a role in the development of OG depending on the regulation of gene expression.