Katayama Akihiro, Ogino Takeshi, Bandoh Nobuyuki, Nonaka Satoshi, Harabuchi Yasuaki
Department of Otolaryngology-Head and Neck Surgery, Asahikawa Medical College, Asahikawa, Japan.
Clin Cancer Res. 2005 Apr 15;11(8):2937-46. doi: 10.1158/1078-0432.CCR-04-1470.
The functional expression of CXCR4, which plays roles in cell migration and proliferation in response to its unique ligand stromal cell-derived factor-1 (SDF-1), has been reported in variety of carcinomas. However, CXCR4 expression and its functional role in head and neck squamous cell carcinomas (HNSCC) remain unclear. In this study, we investigated CXCR4 expression and analyzed its functions in HNSCC cell lines. We also attempted to regulate CXCR4 expression using cytokines, such as interleukin-1beta, tumor necrosis factor-alpha, and IFN-gamma. Finally, we investigated correlation between CXCR4 expression and clinical features in patients with HNSCC.
Six HNSCC cell lines were used in this study. Reverse transcription-PCR and flow cytometry analysis were shown for CXCR4 expressions with or without stimulations of cytokines. SDF-1-mediated cell migration was assayed in Matrigel-coated chemotaxis chamber. The SDF-1-mediated cell proliferation was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The SDF-1-mediated signaling pathways were analyzed by Western blot analysis. Biopsy specimens from 56 patients with HNSCC were used for immunohistologic analysis.
The significant CXCR4 expression was found in HSQ-89, IMC-3, and Nakamura cells. The SDF-1-mediated cell migration and proliferation were observed in CXCR4-positive cells. SDF-1 also promoted rapid phosphorylation of extracellular signal-regulated kinase 1/2 and Akt signaling pathways in CXCR4-positive cells. The SDF-1-mediated cell migration and proliferation of CXCR4-positive cells were inhibited by neutralization of CXCR4. Among three cytokines tested, IFN-gamma significantly reduced CXCR4 expression and SDF-1-induced cell migration and proliferation of CXCR4-positive cells. Immunohistologic analysis revealed that patients with advanced neck status and patients who developed distant metastases showed significantly higher CXCR4 expression, and the cause-specific survival of patients with CXCR4-expression was significantly shorter. Furthermore, multivariate analysis confirmed that CXCR4 positive was the independent factor for cause-specific death.
Our results may provide an insight into future therapeutic agent that inhibits tumor metastasis and progression via down-regulating CXCR4 expression in patients with HNSCC.
CXCR4在细胞迁移和增殖过程中对其独特配体基质细胞衍生因子-1（SDF-1）起反应，其功能性表达已在多种癌症中被报道。然而，CXCR4在头颈部鳞状细胞癌（HNSCC）中的表达及其功能作用仍不清楚。在本研究中，我们调查了CXCR4在HNSCC细胞系中的表达，并分析了其功能。我们还尝试使用细胞因子，如白细胞介素-1β、肿瘤坏死因子-α和干扰素-γ来调节CXCR4的表达。最后，我们研究了HNSCC患者中CXCR4表达与临床特征之间的相关性。
本研究使用了六种HNSCC细胞系。通过逆转录-聚合酶链反应（RT-PCR）和流式细胞术分析在有或无细胞因子刺激的情况下CXCR4的表达。在基质胶包被的趋化小室中检测SDF-1介导的细胞迁移。通过3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）试验分析SDF-1介导的细胞增殖。通过蛋白质免疫印迹分析SDF-1介导的信号通路。来自56例HNSCC患者的活检标本用于免疫组织学分析。
在HSQ-89、IMC-3和中村细胞中发现了显著的CXCR4表达。在CXCR4阳性细胞中观察到SDF-1介导的细胞迁移和增殖。SDF-1还促进了CXCR4阳性细胞中细胞外信号调节激酶1/2和Akt信号通路的快速磷酸化。CXCR4的中和抑制了SDF-1介导的CXCR4阳性细胞的迁移和增殖。在所测试的三种细胞因子中，干扰素-γ显著降低了CXCR4的表达以及SDF-1诱导的CXCR4阳性细胞的迁移和增殖。免疫组织学分析显示，颈部状态晚期的患者和发生远处转移的患者CXCR4表达显著更高，且CXCR4表达阳性患者的病因特异性生存期显著更短。此外，多变量分析证实CXCR4阳性是病因特异性死亡的独立因素。
我们的结果可能为未来通过下调HNSCC患者CXCR4表达来抑制肿瘤转移和进展的治疗药物提供思路。
