中文摘要：

目的:检测卡介苗(BCG)胞壁蛋白对人β-防御素-1(hBD-1)基因表达的影响,并初步分析该基凶5′-端旁侧区中的BCG作用元件。方法:经Sephadex G-150层析柱分离得到BCG细胞壁蛋白组份,用逆转录多聚酶链反应(RT-PCR)和Northern杂交分析法检测hBD-1 mRNA在肺腺上皮细胞的表达,系列逐渐缩短的hBD-1基因5′-端序列被连接入无启动子的pCAT Basic质粒,构建了CAT报告质粒,ELISA法检测报告基因表达。结果:热灭活的BCG全菌(50mg/L)及分子量在18-30 kDa的胞壁蛋白组份(3mg/L)刺激SPC-A-1细胞8h后,hBD-1 mRNA表达明显增强,hBD-1基因-314到+54区域具有BCG诱导的转录活性,其中含有转录因子C/EBPβ、AP-1、CP2结合位点。结论:BCG胞壁蛋白组份(18-30 kDa)能够增强SPC-A-1细胞hBD-1 mRNA表达,其基因上游序列(-314/+54)含有C/EBPβ、AP-1、CP2结合位点,与BCG的诱导作用有关。

英文摘要：

AIM: To examine the stimulatory effect of bacille Calmette-Guérin (BCG) cell wall components on human βdefensin-1 (hBD-1) gene expression and analyze the response element in the 5’-flanking region of the gene. METHODS: BCG cell wall proteins were fractionated by Sephadex G-150 chromatography. Using reverse-transcription polymerase chain reaction (RT-PCR) and Northern hybridization analysis, hBD-1 mRNA expression was detected in a human pulmonary gland epithelial cell line SPC-A-1 cells. Progressive deletions of 5′-flanking region of hBD-1 gene were produced by PCR and ligated into promoterless chloramphenicol acetyltransferase (CAT) expression plasmid to construct pCAT reporter plasmids. Reporter gene expression was determined by ELISA. RESULTS: There was an obvious enhancement of hBD-1 mRNA expression after stimulation with heat-inactivated BCG whole cells (50mg/L), or the cell wall components with a molecular weight of 18-30 kDa (3 rng/L) for 8 h. The upstream sequence between-314 bp and+54 bp had the inducible activity by BCG, which contained CCAAT/enhancer binding protein-β (C/EBPβ), activator protein-1 (AP-1), and CP2 cis element. CONCLUSION: BCG cell wall components (18-30 kDa) can stimulate hBD-1 mRNA expression in pulmonary gland epithelial cells. The sequence (-314/+54) containing C/EBPβ, AP-1, and CP2 binding sites in the upstream of hBD-1 is involved in this induction.