Department of Analytical Neurobiology, Faculty of Pharmacy, Meijo University, Tempaku, Nagoya 4688503, Japan.
Cell Biol Int. 2011 Apr;35(4):325-34. doi: 10.1042/CBI20090344.
We established a clonal adrenal medullary cell line, named tsAM5NE, from transgenic mice harbouring the temperature-sensitive Simian virus 40 large T-antigen gene, under the control of the tyrosine hydroxylase promoter. tsAM5NE cells conditionally grew at a permissive temperature of 33°C and exhibited the noradrenergic chromaffin cell phenotype. To understand the characteristics of tsAM5NE cells, we first examined the responsiveness of the cells to ligands of the GDNF (glial cell line-derived neurotrophic factor) family. tsAM5NE cells proliferated at the permissive temperature of 33°C in response to either GDNF or neurturin, but not artemin or persephin. At the non-permissive temperature of 39°C, GDNF or neurturin caused tsAM5NE cells to differentiate into neuron-like cells; however, the differentiated cells died in a time-dependent manner. Interestingly, LIF (leukaemia inhibitory factor) did not affect the GDNF-mediated cell proliferation at 33°C, but promoted the survival and differentiation of GDNF-treated cells at 39°C. In the presence of GDNF plus LIF, the morphological change induced by the temperature shift was associated with up-regulated expression of neuronal markers, indicating that the cells had indeed undergone neuronal differentiation. Thus, we demonstrated that tsAM5NE cells had the capacity to terminally differentiate into neuron-like cells in response to GDNF plus LIF when the oncogene was inactivated by the temperature shift. Thus, this cell line provides a useful model system for studying the mechanisms regulating neuronal differentiation.
我们从携带受温度敏感型猿猴病毒 40 大 T 抗原基因控制的酪氨酸羟化酶启动子的转基因小鼠中建立了一个克隆肾上腺髓质细胞系，命名为 tsAM5NE。tsAM5NE 细胞在允许温度 33°C 下条件性生长，并表现出去甲肾上腺素能嗜铬细胞表型。为了了解 tsAM5NE 细胞的特征，我们首先检查了细胞对 GDNF（胶质细胞系衍生的神经营养因子）家族配体的反应性。tsAM5NE 细胞在允许温度 33°C 下增殖，对 GDNF 或神经调节素有反应，但对 artemin 或 persephin 没有反应。在非允许温度 39°C 下，GDNF 或神经调节素使 tsAM5NE 细胞分化为神经元样细胞；然而，分化的细胞以时间依赖性方式死亡。有趣的是，LIF（白血病抑制因子）在 33°C 下不影响 GDNF 介导的细胞增殖，但促进了 39°C 下 GDNF 处理细胞的存活和分化。在 GDNF 加 LIF 的存在下，温度转换引起的形态变化与神经元标记物的上调表达相关，表明细胞确实经历了神经元分化。因此，我们证明了当温度转换使癌基因失活时，tsAM5NE 细胞有能力对 GDNF 加 LIF 作出反应，终末分化为神经元样细胞。因此，该细胞系为研究调节神经元分化的机制提供了一个有用的模型系统。
