门静脉高压症脾亢脾巨噬细胞基因表达谱研究

Study on the macrophage in portal hypertensive spleen by cDNA microarray

目的检测门静脉高压症(PHT)脾亢脾和正常脾巨噬细胞的差异表达基因,为从基因水平观察巨噬细胞在门静脉高压症脾亢发生中的作用奠定基础。方法提取门静脉高压症脾亢脾巨噬细胞和正常脾巨噬细胞的总RNA,分别用标有荧光素的dCTP反转录制备cDNA探针,将探针与含有14112点cDNA的Biostar-H140s cDNA表达谱芯片杂交后扫描荧光强度,上述方法重复3次,从而筛选出恒定的差异表达基因。结果3张芯片分别检测到896、1 330和898个差异表达基因。恒定的差异表达基因共有121个,占总基因数的0．86%。其中表达上调的已知基因有21个,表达下调的已知基因有73个。差异表达基因涉及离子通道和运输蛋白、细胞周期蛋白类、细胞骨架和运动、细胞受体、细胞信号和传递蛋白、代谢、免疫相关等多个方面。结论筛选出的差异表达基因,对了解脾脏巨噬细胞在门静脉高压症脾亢发生中的作用有重要意义。

Objective To detect the differently expressed genes in macrophages in normal spleen and portal hypertensive spleen by cDNA microarray,and establish the foundation for studying the role of macrophages associated with hypersplenism in portal hypertension at gene level. Methods The Biostar- H140s chip containing 14112 spots of cDNAs was used to investigate the difference of the gene expression. Both the total RNA extracted from macrophages isolated from normal spleen and portal hypertensive spleen was reversely transcribed to cDNA with the incorporation of fluorescent （Cy3 and Cy5 ） labeled dCTP to prepare the hybridization probes. After hybridization,the gene chip was scanned for the fluorescent intensity. The differently expressed genes were screened. That was repeated three times, and only the genes that had differential expression in all three chips were considered to be associated with hypersplenism in portal hypertension. Results 896,1 330 and 898 genes were identified to be differently expressed in three chips respectively. 121 genes （0.86%） were identified to be differently expressed in all three chips, including 21 up-regulated known genes and 73 down- regulated known genes. The differently expressed genes were related with ionic channel and transport protein, cyclin, cytoskeleton, cell receptor, cell signal conduct, metabolism, immune, and so on. These genes might be related with the hypersplenism in portal hypertension. Conclusion The investigations based on cDNA microarray can screen differently expressed genes of macrophages in normal spleen and portal hypertensive spleen,that was important for comprehending the function of macrophages associated with hypersplenism in portal hypertension.