摘要
背景与目的:我们先前的研究已经证明,在诱导分化剂作用下,胶质瘤胞由分化不良向分化良好方向演进,但该效应的分子机制不明。本研究旨在建立质瘤细胞诱导分化的相关基因表达谱并克隆新基因,为进一步阐明产生该效应的子机制提供基因信息。方法:采用MTT法、软琼脂克隆形成率检测、细胞形态学观察、流式细胞术、胶质纤维酸性蛋白免疫荧光检测等方法,确定诱导分化剂对低分人脑胶质瘤细胞株SHG-44-9的分化效应;用cDNA微阵列技术检测诱导分化前后的基因表达变化;用随机引物差异显示PCR克隆相关基因,并以反Northern杂交验证,再与GenBank比对分析相关基因信息。结果:1.75mmol/L的苯丁酸钠能显著抑制SHG-44-9胶质瘤细胞的生长,并促其向良性方向分化。在含18000个人cDNA的芯片中发现、并经反Northern杂交验证,有98个基因的表达发生显著变化,并克隆12个与分化有关的基因,其中调控c-myc表达的MIBP1基因可能为调控胶质瘤细胞分化的基因。结论:本研究发现的98个与分化相关的基因和克隆的12个基因,有助于进一步研究胶质瘤细胞分化的分子机制和发现新的基因治疗靶点。
BACKGROUND &OBJECTIVE:Glioma cells can be induced in vitro from low differentiation stat e to hig h differentiation state with differentiation-inducing ag ents,as our previous studies have showed.However,the relevant molecular mechanisms r emained unclear.This paper was to establish differentiation-induci ng g ene expression pedig ree of g liom a cells and clone new relevant g enes ,for the pur pose of offering g enomic informatio n on the molecular mechanisms of differentiation-inducing effects of tumor cel ls.METHODS :A low differentiated human brain g li oma cell line(SHG-44-9)was treated with differentiation-inducing ag ents.The differentiation-inducing effects on g lioma cells were determined usin g MTT assay,colony-forming efficie ncy in double-layer soft ag ar,cell morpho log ical chang es,flow cytometry,and fluoroimmunoassay of g lial fibrill ary acidic protein expression.Gene expression differences of tumor cells before an d after inducing were assayed with cD NA microarray techniques.The relevan t g enes were cloned with differentia l display polymerase chain reaction(DD-PCR),and verified with Northern blot ana lysis ,then the g enomic information about t hese g enes were analyzed with bioinformatics technolog ies.RESULTS :At dose of 1.75mmol ·L -1 ,sodium butyrate caused a marked proliferat ion inhibitory effect on the SHG-44-9cells ,and drove them to a more mature phenot ype.The expression of 98g enes chang ed obviously when were screene d with g ene chips immobilized with 18,000human cDNAs,which were further proved by Northern blot analysis.Twe lve differentiation-associated g enes were cloned,including MIBP1g ene,w hich can reg ulate the c-myc g ene expression a nd may be relevant with g lioma cell differentiation.CONCLUSION:Ninety-eig ht g enes associated with differentiation and 12g enes were cloned.These data may help to further study the molecula r mechanisms of g lioma cells differen tiation,and to find new targ ets for g ene therapy ag ainst g lioma.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2003年第7期673-679,共7页
Chinese Journal of Cancer
基金
国家自然科学基金资助项目(No.39870826)
关键词
脑胶质瘤细胞株
体外
诱导分化
基因
细胞形态学
流式细胞术
Glioma
Inducing differentiation
Gene expression
Complementary deoxyribonucleic ac id array
