Scala S, Wosikowski K, Giannakakou P, Valle P, Biedler J L, Spengler B A, Lucarelli E, Bates S E, Thiele C J
Medicine Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.
Cancer Res. 1996 Aug 15;56(16):3737-42.
Brain-derived neurotrophic factor (BDNF) and its receptors are necessary for the survival and development of many neuronal cells. Because BDNF and TrkB are expressed in many poor-prognosis neuroblastoma (NB) tumors, we evaluated the role of BDNF in affecting sensitivity to chemotherapeutic agents. We investigated the effects of activation of the BDNF-TrkB signal transduction pathway in two NB cell lines, 15N and SY5Y. 15N cells lack the high-affinity receptor p145TrkB and express BDNF; 15N cells were used along with 15N-TrkB cells, a subline transfected with a TrkB expression vector. In cytotoxicity assays, 15N-TrkB cells were consistently 1.4-2 fold more resistant to vinblastine than 15N cells. Drug accumulation assays showed a 50% reduction in[3H]vinblastine accumulation in 15N-TrkB cells compared with control 15N cells. Addition of 30 ng/ml BDNF resulted in a reduction to 46% of control in 15N cells and a reduction to 28% of control in 15N-TrkB cells. SY5Y cells were chosen as a second model because they lack both endogenous BDNF and TrkB expression. p145TrkB expression is induced by 1 nM retinoic acid. Vinblastine accumulation was not significantly affected by 1 nM retinoic acid in SY5Y cells. Addition of 30 ng/ml BDNF decreased [3H]vinblastine accumulation to 58% of control in SY5Y cells and decreased [3H]vinblastine accumulation to 62% of control in TrkB-expressing SY5Y cells. Although an increase in BDNF expression in seen in multidrug-resistant sublines of SY5Y and BE(2)-C NB cells, the protective effect of BDNF in vinblastine toxicity may be unrelated to mdr-1, because the activity of other agents transported by P-glycoprotein was not affected. There was no increase in mdr-1 expression in 1 nM RA SY5Y cells and 15N-TrkB cells, as assessed by Northern blot analysis. In addition to the effects of BDNF on vinblastine cytotoxicity and accumulation, there was an inhibition in the ability of vinblastine to depolymerize tubulin in BDNF-treated cells. Thus, BDNF and TrkB may partially rescue NB cells from vinblastine toxicity and thereby may contribute to a more chemoresistant phenotype.
脑源性神经营养因子（BDNF）及其受体对许多神经元细胞的存活和发育至关重要。由于BDNF和TrkB在许多预后不良的神经母细胞瘤（NB）肿瘤中表达，我们评估了BDNF在影响化疗药物敏感性方面的作用。我们研究了BDNF-TrkB信号转导通路激活在两种NB细胞系15N和SY5Y中的作用。15N细胞缺乏高亲和力受体p145TrkB并表达BDNF；15N细胞与15N-TrkB细胞一起使用，15N-TrkB细胞是用TrkB表达载体转染的亚系。在细胞毒性试验中，15N-TrkB细胞对长春碱的耐药性始终比15N细胞高1.4至2倍。药物蓄积试验显示，与对照15N细胞相比，15N-TrkB细胞中[3H]长春碱的蓄积减少了50%。添加30 ng/ml BDNF导致15N细胞中[3H]长春碱的蓄积降至对照的46%，15N-TrkB细胞中降至对照的28%。选择SY5Y细胞作为第二个模型，因为它们既缺乏内源性BDNF表达，也缺乏TrkB表达。1 nM视黄酸可诱导p145TrkB表达。在SY5Y细胞中，1 nM视黄酸对长春碱的蓄积没有显著影响。添加30 ng/ml BDNF可使SY5Y细胞中[3H]长春碱的蓄积降至对照的58%，在表达TrkB的SY5Y细胞中降至对照的62%。尽管在SY5Y和BE(2)-C NB细胞的多药耐药亚系中观察到BDNF表达增加，但BDNF对长春碱毒性的保护作用可能与mdr-1无关，因为P-糖蛋白转运的其他药物的活性未受影响。通过Northern印迹分析评估，在1 nM RA SY5Y细胞和15N-TrkB细胞中mdr-1表达没有增加。除了BDNF对长春碱细胞毒性和蓄积的影响外，BDNF处理的细胞中长春碱使微管蛋白解聚的能力也受到抑制。因此，BDNF和TrkB可能部分挽救NB细胞免受长春碱毒性，从而可能导致更具化疗耐药性的表型。
