Institute of Pharmacology and Toxicology, Tzu Chi University, Hualien 970, Taiwan, R.O.C.
Int J Oncol. 2010 Jun;36(6):1469-76. doi: 10.3892/ijo_00000633.
Pathologically, >90% of bladder cancer is transitional cell carcinoma (TCC). Previously, brain-derived neurotrophic factor (BDNF) but not tropomyosin-related kinase B (TrkB) was found in normal urothelium. TrkB activation by BDNF has been shown to promote the progression of several cancers, however, the existence and functional roles of both BDNF and TrkB in TCC have not been elucidated. In this study, three human TCC cell lines, BFTC905, TSGH8301, and T24 were used for the investigation. Both BDNF and TrkB but not TrkA or TrkC identified by RT-PCR and Western blotting were found in these cell lines. Immunostaining demonstrated the cytosolic expression of BDNF and TrkB, as well as membranous expression of TrkB in these cells. BDNF released from three cell lines was also detected in culture medium by ELISA. The proliferation of BFTC905 cells was enhanced by recombinant human BDNF (rhBDNF) in vitro, which was associated with increased phospho-TrkB and phospho-ERK levels. In contrast, TrkB-Fc chimeric protein served as BDNF scavenger eliciting cytotoxicity. Addition of rhBDNF in these cell lines cultured in poly-HEMA [Poly(2-hydroxyethyl methacrylate)] coated dishes for 48 h did not confer resistance to anoikis. Increased phospho-Akt expression was observed transiently within an hour after rhBDNF administration but disappeared 24 h later. Weekly injections of 100 ng rhBDNF into the cancer cell-loading site for 6 weeks promoted BFTC905 xenograft growth in SCID mice. Daily injection of 5 microg TrkB-Fc chimeric protein into the tumor 2 weeks after tumor cell implantation delayed tumor growth concomitant with phospho-TrkB suppression in xenografts. These results indicate that BDNF binding to TrkB receptor is a survival signal for TCC cells. Drugs that block BDNF or TrkB may provide a new and potential approach for TCC therapy.
从病理学角度来看，膀胱癌 90%以上为移行细胞癌（TCC）。先前研究发现，脑源性神经营养因子（BDNF）而非原肌球蛋白相关激酶 B（TrkB）存在于正常尿路上皮中。BDNF 激活 TrkB 已被证实可促进多种癌症的进展，但 BDNF 和 TrkB 在 TCC 中的存在和功能作用尚未阐明。在本研究中，我们使用了三种人 TCC 细胞系 BFTC905、TSGH8301 和 T24 进行研究。RT-PCR 和 Western blot 检测均发现这三种细胞系中存在 BDNF 和 TrkB，但不存在 TrkA 或 TrkC。免疫组化显示 BDNF 和 TrkB 存在于细胞质中，TrkB 存在于细胞膜上。ELISA 检测也发现三种细胞系均有 BDNF 分泌到细胞培养上清中。体外实验中，重组人 BDNF（rhBDNF）可增强 BFTC905 细胞的增殖，该作用与磷酸化-TrkB 和磷酸化-ERK 水平升高相关。相反，TrkB-Fc 嵌合蛋白作为 BDNF 清除剂可引起细胞毒性。在聚-HEMA（聚 2-羟乙基甲基丙烯酸酯）包被的培养皿中培养这些细胞 48 小时后，加入 rhBDNF 并不能使其抵抗失巢凋亡。rhBDNF 给药后 1 小时内观察到磷酸化-Akt 表达短暂增加，24 小时后消失。每周向 BFTC905 荷瘤部位注射 100ng rhBDNF，连续 6 周可促进 SCID 小鼠的 BFTC905 异种移植瘤生长。在肿瘤细胞植入后 2 周，每天向肿瘤内注射 5μg TrkB-Fc 嵌合蛋白可延迟肿瘤生长，同时抑制异种移植瘤中磷酸化-TrkB 的表达。这些结果表明，BDNF 与 TrkB 受体结合是 TCC 细胞的存活信号。阻断 BDNF 或 TrkB 的药物可能为 TCC 治疗提供一种新的潜在方法。
