基因芯片筛选ghrelin miRNA转染胰腺炎细胞模型后的差异表达基因

Screening for Differentially Expressed Genes in Pancreatitis Cell Model Transfected With Ghrelin MiRNA by Gene Chip

背景:前期研究提示ghrelin在急性胰腺炎发病过程中发挥重要作用,但其机制尚不明确。目的:应用基因芯片技术筛选出ghrelin miRNA转染胰腺炎胰腺腺泡细胞后的差异表达基因。方法:以雨蛙肽构建胰腺炎胰腺腺泡细胞模型。将AR42J细胞分为ghrelin miRNA+雨蛙肽组、阴性对照+雨蛙肽组。提取细胞RNA,合成为ds-DNA,与大鼠全基因组基因芯片杂交、扫描、分析。筛选与炎症和钙通路相关的差异表达基因,并采用RT-PCR法对其中3个基因(Bcl-2、caspase-8、caspase-12)进行验证。结果:给予AR42J细胞ghrelin miRNA+雨蛙肽干预后,共筛选出2938个差异表达基因,其中1435个基因表达上调,1503个基因表达下调。与CAMP/Ca^2+信号通路相关的差异表达基因有60个,与炎症通路相关的差异表达基因有199个。RT-PCR结果表明,ghrelin miRNA+雨蛙肽组Bcl-2、caspase-12表达下调,caspase-8表达上调,与基因芯片筛查结果一致。结论:基因芯片技术可筛选出ghrelin miRNA转染胰腺炎胰腺腺泡细胞过程中发挥关键作用的基因,从而为研究内源性ghrelin对胰腺炎胰腺腺泡细胞的作用机制提供依据和参考。

Background:Previous studies suggest that ghrelin plays an important role in the pathogenesis of acute pancreatitis,but the mechanism is still unclear.Aims:To screen the differentially expressed genes in pancreatitis pancreatic acinar cells transfected with ghrelin miRNA by gene chip.Methods:Pancreatitis was induced by caerulin.AR42J cells were divided into ghrelin miRNA+caerulin group and negative control+caerulin group.AR42J cells were collected to extract RNA and synthesize ds-DNA,and was then hybridized with rat whole genome microarray for scanning and analyzing.Differentially expressed genes related to inflammation and calcium pathways were screened,and expressions of Bcl-2,caspase-8,caspase-12 were verified by RT-PCR.Results:For AR42J cells after intervention with ghrelin miRNA+caerulin,2938 differentially expressed genes were screened,including 1435 up-regulated genes and 1503 down-regulated genes.There were 60 differentially expressed genes involved in CAMP/Ca^2+signaling pathway and 199 differentially expressed genes involved in inflammation pathway.RT-PCR results showed that expressions of Bcl-2,caspase-12 were down-regulated and expression of caspase-8 was up-regulated in ghrelin miRNA+caerulin group,consistent with the results of gene chip.Conclusions:Gene chip can be used to screen the genes that play a key role in the process of pancreatitis pancreatic acinar cells transfected with ghrelin miRNA,and provide a theoretical basis and reference for the study of mechanism of endogenous ghrelin on pancreatitis pancreatic acinar cells.