基于基因芯片技术的肝癌糖类相关基因表达谱研究
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摘要
研究背景:糖链及其缀合物不仅参与生命活动的基本生理生化过程,而且在肿瘤的发生和发展中发挥着重要作用。肿瘤细胞中的糖链与正常细胞相比有明显变化,包括表达增多,减少或出现新的结构等。糖链的合成没有模板,而是通过一系列定位有序的糖基转移酶,糖苷酶以及磺基转移酶的作用完成的,这些酶的表达差异将直接导致糖链结构的变化。因此,从这些糖类相关基因入手可以研究它们如何调控糖链的合成,寻找癌症相关糖基因,为筛选抗癌药物提供靶分子,为癌症的早期诊断和防治提供有效方法。
目的:运用基因芯片技术为检测手段,筛选在肝癌中差异表达的糖类相关基因,作为肝癌检测的标记物,为肝癌早期诊断和抗肝癌药物提供靶分子,并为肝癌及其他肿瘤的研究建立平台。
方法:本实验以正常肝细胞系CL(chang's liver)和肝癌细胞系7721,以及正常人血液和肝癌病人血液为研究对象,利用本实验室自主研发的糖类相关基因芯片(其中包括糖基转移酶130个,糖苷酶12个,磺基转移酶48个,质控点20个),系统地研究肝癌样本及正常样本中糖类相关基因发生的变化,筛选肝癌中异常表达的糖类相关基因,并用荧光定量PCR技术加以验证。
结果:用糖类相关基因芯片对正常肝细胞系CL(chang's liver)和肝癌细胞系7721进行比较研究,筛选出了差异表达的糖类相关基因38个,其中上调基因19个,下调基因19个。又对肝癌病人血液和正常人血液样本进行了研究,筛选出了上调基因15个,下调基因13个。细胞样本和血液样本中上调基因重合的有7个,下调基因重合的有5个。从中挑选出了四个表达差异较大的基因,上调基因两个Alg10和Galnt11,下调基因两个Hmcs,Galnt12。用荧光定量PCR技术对这四个基因的表达水平进行验证,其结果与芯片结果呈现出一致性。
结论:利用糖类相关基因芯片与荧光定量PCR等技术系统的研究了肝癌糖类相关基因表达谱,筛选出12个差异表达的基因,为今后肝癌的深入研究以及临床诊断和基因治疗创造了条件。该试验中成功设计和制备出的糖类相关基因芯片,为其它肿瘤与相关糖类基因的研究建立了平台。
Background: A growing body of evidence supported oligosaccharides in tumors modulate their function with respect to controlling tumor cell adhesion, motility, and invasiveness. Oligosaccharide structures in tumors are synthesized by a series of glycosyltransferases, glycosidase and sulfotransferase. So it is indispensable to monitor the expression profile of the glycosylation-related gene expression for the investigation of tumor.
Purpose: The objective of this contribution was to investigate the glycosylation-related gene between liver cancer and normal liver cell or blood sample.Selected the differential expressed glycosylation-related gene as the biomarker of liver cancer and establish a platform for the investigation of tumor.
Method:Using the cell line (7721 ,CL) and the blood sample (liver cancer patient, health adult) as the research object.Hybridization with the oligonucleotide arrays which desigened and prepared by ourselves(including 130 glycosyltransferases,12 glycosidase,48 sulfotransferase and 20 contral gene). Select the differential expressed glycosylation-related gene . RT-PCR confirmed the result of oligonucleotide arrays.
Result: Using the cell line as the research object we found 38 differential expressed glycosylation-related gene.19 high expression level gene and 19 low expression level gene. Then we use the blood sample as the research object found 15 high expression level gene, 13 low expression level gene.There are 7 common up regulated glycosylation-related gene in cell line and blood sample,and 5 common down regulated glycosylation-related gene.pick up four of them: up regulated gene Alg10 ,Galnt11; down regulated gene Hmcs and Galnt12 . The RT-PCR confirmed the result of oligonucleotide arrays.
Conclusion: The investigation of liver cancer using the way of oligonucleotide arrays found out 12 differential expressed glycosylation-related gene compared with nomal sample.Four of them have confirmed by RT-PCR.Results of this study are expected to be developed into new approaches for the prevention, diagnosis, prognosis and analysis of liver cancer. successfully desigened and prepared glycosylation-related gene array, establish a platform for the investigation of tumor.
目的:运用基因芯片技术为检测手段,筛选在肝癌中差异表达的糖类相关基因,作为肝癌检测的标记物,为肝癌早期诊断和抗肝癌药物提供靶分子,并为肝癌及其他肿瘤的研究建立平台。
方法:本实验以正常肝细胞系CL(chang's liver)和肝癌细胞系7721,以及正常人血液和肝癌病人血液为研究对象,利用本实验室自主研发的糖类相关基因芯片(其中包括糖基转移酶130个,糖苷酶12个,磺基转移酶48个,质控点20个),系统地研究肝癌样本及正常样本中糖类相关基因发生的变化,筛选肝癌中异常表达的糖类相关基因,并用荧光定量PCR技术加以验证。
结果:用糖类相关基因芯片对正常肝细胞系CL(chang's liver)和肝癌细胞系7721进行比较研究,筛选出了差异表达的糖类相关基因38个,其中上调基因19个,下调基因19个。又对肝癌病人血液和正常人血液样本进行了研究,筛选出了上调基因15个,下调基因13个。细胞样本和血液样本中上调基因重合的有7个,下调基因重合的有5个。从中挑选出了四个表达差异较大的基因,上调基因两个Alg10和Galnt11,下调基因两个Hmcs,Galnt12。用荧光定量PCR技术对这四个基因的表达水平进行验证,其结果与芯片结果呈现出一致性。
结论:利用糖类相关基因芯片与荧光定量PCR等技术系统的研究了肝癌糖类相关基因表达谱,筛选出12个差异表达的基因,为今后肝癌的深入研究以及临床诊断和基因治疗创造了条件。该试验中成功设计和制备出的糖类相关基因芯片,为其它肿瘤与相关糖类基因的研究建立了平台。
Background: A growing body of evidence supported oligosaccharides in tumors modulate their function with respect to controlling tumor cell adhesion, motility, and invasiveness. Oligosaccharide structures in tumors are synthesized by a series of glycosyltransferases, glycosidase and sulfotransferase. So it is indispensable to monitor the expression profile of the glycosylation-related gene expression for the investigation of tumor.
Purpose: The objective of this contribution was to investigate the glycosylation-related gene between liver cancer and normal liver cell or blood sample.Selected the differential expressed glycosylation-related gene as the biomarker of liver cancer and establish a platform for the investigation of tumor.
Method:Using the cell line (7721 ,CL) and the blood sample (liver cancer patient, health adult) as the research object.Hybridization with the oligonucleotide arrays which desigened and prepared by ourselves(including 130 glycosyltransferases,12 glycosidase,48 sulfotransferase and 20 contral gene). Select the differential expressed glycosylation-related gene . RT-PCR confirmed the result of oligonucleotide arrays.
Result: Using the cell line as the research object we found 38 differential expressed glycosylation-related gene.19 high expression level gene and 19 low expression level gene. Then we use the blood sample as the research object found 15 high expression level gene, 13 low expression level gene.There are 7 common up regulated glycosylation-related gene in cell line and blood sample,and 5 common down regulated glycosylation-related gene.pick up four of them: up regulated gene Alg10 ,Galnt11; down regulated gene Hmcs and Galnt12 . The RT-PCR confirmed the result of oligonucleotide arrays.
Conclusion: The investigation of liver cancer using the way of oligonucleotide arrays found out 12 differential expressed glycosylation-related gene compared with nomal sample.Four of them have confirmed by RT-PCR.Results of this study are expected to be developed into new approaches for the prevention, diagnosis, prognosis and analysis of liver cancer. successfully desigened and prepared glycosylation-related gene array, establish a platform for the investigation of tumor.
引文
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