Morgan Claire, Alazawi William, Sirieix Pierre, Freeman Tom, Coleman Nicholas, Fitzgerald Rebecca
MRC Cancer Cell Unit, Hutchison-MRC Research Centre, Hills Road, Cambridge CB2 2XZ, UK.
Am J Gastroenterol. 2004 Feb;99(2):218-24. doi: 10.1111/j.1572-0241.2004.04054.x.
Acid, a principal component of refluxate, may contribute to the neoplastic progression of Barrett's esophagus. Brief acid exposure in vivo and in vitro has been shown to increase cell proliferation. The mechanisms underlying the hyperproliferative response are not well elucidated but may include alterations in Na(+)-H+ exchanger activity and MAPK signaling pathways.
To ascertain the effects of pulsatile acid exposure on gene expression in a Barrett's adenocarcinoma cell line (SEG-1).
SEG-1 cells were exposed to either acidified DMEM at pH 3.5 (0.1 M hydrochloric acid) or pH 7.4 (control) for 20 min followed by neutralization of the medium. Total RNA was extracted before acid exposure and over a 10-h time course (0.5, 2, 4, 6, 8, and 10 hours) and hybridized to Affymetrix human U133A oligonucleotide arrays. Data were analyzed using the Affymetrix statistical expression algorithms. Only alterations in gene expression that were > or = 2 and < or = -2 fold were studied further and a subset was further investigated by reverse transcription polymerase chain reaction (RT-PCR) and densitometry. Apoptosis was assayed in SEG-1 cells by western blot for cleaved caspase 3 and an apoptosis ELISA assay.
Changes in expression were identified for 138 genes. Analysis of gene function identified immediate downregulation of genes associated with apoptosis and early upregulation of genes associated with proliferation. The gene expression profiles suggest that MAPK pathways may be involved and suppression of apoptosis may occur via p53-dependent mechanisms.
Microarray analysis of gene expression changes in a Barrett's adenocarcinoma cell line has identified cellular pathways that may be disrupted following acid exposure.
酸是反流物的主要成分，可能促使巴雷特食管发生肿瘤进展。体内和体外的短暂酸暴露已显示可增加细胞增殖。这种过度增殖反应的潜在机制尚未完全阐明，但可能包括钠氢交换体活性和丝裂原活化蛋白激酶（MAPK）信号通路的改变。
确定脉动酸暴露对巴雷特腺癌细胞系（SEG-1）基因表达的影响。
将SEG-1细胞暴露于pH 3.5（0.1 M盐酸）或pH 7.4（对照）的酸化杜氏改良 Eagle培养基（DMEM）中20分钟，随后中和培养基。在酸暴露前以及10小时的时间进程（0.5、2、4、6、8和10小时）中提取总RNA，并与Affymetrix人类U133A寡核苷酸阵列杂交。使用Affymetrix统计表达算法分析数据。仅对基因表达变化大于或等于2倍且小于或等于 -2倍的情况进行进一步研究，并通过逆转录聚合酶链反应（RT-PCR）和光密度测定法对一个子集进行进一步研究。通过蛋白质免疫印迹法检测裂解的半胱天冬酶3并采用凋亡酶联免疫吸附测定法检测SEG-1细胞中的凋亡情况。
鉴定出138个基因的表达发生变化。基因功能分析确定与凋亡相关的基因立即下调，与增殖相关的基因早期上调。基因表达谱表明MAPK通路可能参与其中，并且凋亡抑制可能通过p53依赖性机制发生。
对巴雷特腺癌细胞系基因表达变化进行的微阵列分析确定了酸暴露后可能被破坏的细胞通路。
