Molecular regulation of constitutive expression of interleukin-8 in human pancreatic adenocarcinoma
Molecular regulation of constitutive expression of interleukin-8 in human pancreatic adenocarcinoma
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DOI：
10.1089/10799900050198372
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发表时间：
2000-11-01
影响因子：
2.3
通讯作者：
Xie, KP
中科院分区：
文献类型：
--
作者：
Le, XD;Shi, Q;Xie, KP
最近的研究表明，白介素8（IL-8）在人类胰腺癌的生长和转移中发挥着重要作用。在本研究中，我们确定了人胰腺癌细胞中组成型 IL-8 表达的分子调控。在体外培养各种人胰腺癌细胞系。使用酶联免疫吸附测定法测定，67% 的细胞系持续分泌高水平的 IL-8。一致地，根据 Northern 印迹分析确定，这些细胞持续表达高水平的 IL-8 mRNA。为了确定 IL-8 mRNA 高稳态水平的机制，测量了 IL-8 半衰期和转录率。表达高水平和低水平IL-8的细胞之间IL-8半衰期没有显着差异。然而，在组成型表达高水平 IL-8 的细胞中观察到更高的转录率和增加的 IL-8 启动子活性。使用缺失突变的详细 IL-8 启动子分析表明，-85 至 -133 bp 的区域对于组成型 IL-8 启动子活性至关重要。此外，点突变分析表明 NF-κB、AP-1 或 NF-IL-6 结合位点的突变显着降低或消除了组成型 IL-8 启动子活性。与组成型 IL-8 转录活性一致，在过表达 IL-8 的细胞中检测到高水平的组成型 NF-κB 和 AP-1 活性（使用电泳迁移率变动测定法测定）。此外，显性失活 I-kappaB α 表达载体 (I-kappaB αM) 的转染可抑制胰腺癌细胞中的组成型 NF-kappaB 活性和 IL-8 表达。总的来说，我们的数据表明，组成型 NF-kappaB 和 AP-1 激活有助于 IL-8 的过度表达，而 IL-8 反过来在肿瘤血管生成中发挥重要作用，并有助于人类胰腺癌的侵袭性生物学。
Recent studies have shown that interleukin-8 (IL-8) plays an important role in the growth and metastasis of human pancreatic cancer. In the present study, we determined the molecular regulation of constitutive IL-8 expression in human pancreatic cancer cells. Various human pancreatic cancer cell lines were incubated in vitro. Sixty-seven percent of the cell lines constitutively secreted high levels of IL-8, as determined using enzyme-linked immunosorbent assay. Consistently, these cells constitutively expressed high levels of IL-8 mRNA, as determined using Northern blot analysis. To determine the mechanisms of the high steady-state levels of IL-8 mRNA, the IL-8 half-life and transcription rate were measured. There was no significant difference in IL-8 half-life between cells expressing high and low levels of IL-8. However, higher transcription rates and increased IL-8 promoter activity were observed in the cells constitutively expressing high levels of IL-8. Detailed IL-8 promoter analysis using deletion mutation revealed that the region from -85 to -133 bp was essential for the constitutive IL-8 promoter activity. Also, point-mutation analysis indicated that mutation of NF-kappaB, AP-1, or NF-IL-6 binding sites significantly reduced or eliminated the constitutive IL-8 promoter activity. Consistent with the constitutive IL-8 transcription activity, high levels of constitutive NF-kappaB and AP-1 activity were detected in the cells overexpressing IL-8, as determined using electrophoretic mobility shift assay. In addition, transfection of a dominant-negative I-kappaB alpha expression vector (I-kappaB alphaM) inhibited constitutive NF-kappaB activity and IL-8 expression in pancreatic cancer cells. Collectively, our data demonstrated that constitutive NF-kappaB and AP-1 activation contributes to the overexpression of IL-8, which in turn plays an important role in tumor angiogenesis and contributes to the aggressive biology of human pancreatic cancer.