An abnormal response of retinoblastoma cells (Y-79) to neurotrophins.
An abnormal response of retinoblastoma cells (Y-79) to neurotrophins.
复制标题
视网膜母细胞瘤细胞 (Y-79) 对神经营养素的异常反应。
DOI：
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发表时间：
2000
影响因子：
4.4
通讯作者：
R. Grantyn
中科院分区：
文献类型：
--
作者：
N. Wagner;K. Wagner;M. Sefton;A. Rodríguez‐Tébar;R. Grantyn
目的
目的：明确神经营养因子及其受体在视网膜母细胞瘤（Rb）细胞中的表达，阐明其在神经外胚层肿瘤细胞增殖中的作用，并建立Rb细胞分化的条件。
方法
Rb衍生的细胞系Y-79在无血清悬浮或单层培养中生长。分离出增殖和分化的细胞，并进行半定量逆转录-聚合酶链反应（RT-PCR）分析、免疫染色和流式细胞术。通过5-溴-2 '-脱氧尿苷（BrdU）掺入来估计细胞的增殖率，并确定神经营养因子和层粘连蛋白对BrdU掺入、突起生长或免疫染色的影响。
结果
与先前研究的正常视网膜前体细胞相比，Y-79细胞不仅表达神经生长因子（NGF）、脑源性神经营养因子（BDNF）、神经营养因子-3（NT-3）和p75，而且还表达相应的高亲和力受体TrkA、TrkB和TrkC。外源性和内源性神经营养因子受体配体刺激增殖。用K252 a抑制蛋白激酶磷酸化阻断增殖并促进分化。K252 a对分化的作用通过添加可溶性层粘连蛋白而增强。经过9天的联合治疗，分化细胞的分数达30%，分化的特点是改善附着，神经突生长，NF-68的表达，和神经胶质细胞酸性蛋白（GFAP）和小清蛋白免疫反应性的损失。这些变化伴随着TrkB和TrkC的下调，而不是TrkA或p75。分离分化的细胞并在不存在K252 a的情况下进一步生长。然而，尽管在分化的细胞中TrkA表达水平很高，但添加NGF对其存活没有影响。
结论
神经营养因子的促有丝分裂作用可能有助于视网膜肿瘤的生长。
PURPOSE
To clarify the expression of neurotrophins and their receptors in retinoblastoma (Rb) cells, to elucidate their potential role in the proliferation of neuroectodermal tumor cells, and to establish conditions for Rb cell differentiation.
METHODS
The Rb-derived cell line Y-79 was grown in serum-free suspension or monolayer culture. Proliferating and differentiated cells were isolated and submitted to semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis, immunostaining, and flow cytometry. The proliferation rate of the cells was estimated by 5-bromo-2'-deoxyuridine (BrdU) incorporation, and the effects of neurotrophins and laminin on BrdU-incorporation, process outgrowth, or immunostaining were determined.
RESULTS
In contrast to previously studied normal retinal precursor cells, Y-79 cells not only express nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and p75, but also the corresponding high affinity receptors TrkA, TrkB, and TrkC. Proliferation was stimulated by exogenous and endogenous neurotrophin receptor ligands. Inhibition of protein kinase phosphorylation with K252a blocked proliferation and promoted differentiation. The effect of K252a on differentiation was enhanced by the addition of soluble laminin. After 9 days of combined treatment, the fraction of differentiated cells amounted to 30%, differentiation being characterized by improved attachment, neurite outgrowth, expression of NF-68, and a loss of glial fibrillary acidic protein (GFAP) and parvalbumin immunoreactivity. These changes were accompanied by a downregulation of TrkB and TrkC, but not TrkA or p75. Differentiated cells were isolated and further grown in the absence of K252a. However, despite the high level of TrkA expression in differentiated cells, the addition of NGF had no effect on their survival.
CONCLUSIONS
A mitogenic action of neurotrophins could contribute to retinal tumor growth.
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