基因芯片法检测胆囊胆固醇结石病人肝脏羧肽酶E的表达

Expression of hepatic carboxypeptidase E in patients with cholesterol gallstone

目的:利用基因芯片和实时定量聚合酶链反应(real-time PCR)分析胆囊胆固醇结石病人肝脏中羧肽酶E(carboxypeptidase E,CPE)的mRNA表达,探讨CPE与胆囊胆固醇结石形成可能相关的分子机制。方法:检测病人的血清总胆固醇、三酰甘油以及胆石、胆汁成分测定。正常肝细胞株(HL-7702)为共同参照,用含10 000个cDNA克隆的芯片对8份肝脏标本样品(包括6例胆囊结石和2例无胆囊结石对照)进行基因表达谱对比分析;通过实时PCR验证阳性结果。结果:有效表达基因6 068个,与正常对照组比较,胆固醇结石病人的肝脏有67条差异表达基因,其中CPE基因呈下调表达,经实时PCR验证与芯片筛选结果一致。进一步比较另12例胆固醇结石病人和8例对照组的肝脏组织CPE表达,CPE在胆囊胆固醇结石病人肝脏中的表达也显著低于对照组(39.52±6.21比124.50±36.36,P<0.05)。结论:肝脏CPE表达下降可能与胆囊胆固醇结石的形成有关。

Objective To survey carboxypeptidase E （CPE） mRNA expression by cDNA microarray and real-time PCR in the liver of patients with cholesterol gallstone （CGS）, and to reveal the role of CPE in the formation of CGS. Methods The serum total cholesterol, triglycerol, and biliary composition were assayed. Normal liver cell line HL-7702 was used as control, and 8 liver samples from 2 patients without CGS and 6 patients with CGS were studied by cDNA microarray containing 10 000 genes. The levels of gene expression changes were further confirmed by real-time PCR assay. Results A total of 6 068 genes expressed both in the human liver cell line and in the liver samples. Sixty-seven genes including lipid metabolism genes, regulatory genes of biological process and signal conduction genes in the patients with CGS expressed differently from those in the cases without CGS. The CPE expression was down regulated in the patients with CGS. The results from cDNA microarray were confirmed by real-time PCR. Further results in larger samples showed that the CPE mRNA level was lower in the patients with CGS than that in the controls （39.52±6.21 vs 124.50±36.36, P〈 0.05）. Conclusions The decrease of hepatic CPE mRNA level in patients with CGS seems to be related to the formation of cholesterol gallstone.