γ干扰素和神经生长因子诱导神经母细胞瘤细胞分化及TrkA的表达

目的　在人神经母细胞瘤 (neuroblastoma ,NB)SMS KCNR细胞系中分别应用γ干扰素(IFN γ)、神经生长因子 (NGF)及联合应用IFN γ和NGF作为诱导剂诱导NB细胞分化 ,观察细胞形态学的变化、细胞增殖情况及TrkAmRNA的表达。方法　常规方法培养人SMS KCNR细胞系细胞 ,收取第 10天培养的细胞 ,提取总RNA ,应用逆转录 聚合酶链反应 (RT PCR)方法检测TrkAmRNA的表达水平 ,应用锥虫兰染色方法计数活细胞数并用倒置相差显微镜观察细胞形态学的改变 ,计数细胞分化率。结果　在KCNR细胞中同时负荷IFN γ和NGF的细胞表现出比单独负荷IFN γ或NGF更加显著的形态学分化 (P <0 0 1)。前者细胞增殖抑制更加明显 (P <0 0 1)。在IFN γ组 ,TrkAmRNA表达增加 (P <0 0 1) ,NGF组TrkAmRNA的表达未见明显改变 (P >0 0 5 ) ,联合用药组TrkAmRNA的表达有明显降低 (P <0 0 1)。结论　在人SMS KCNR细胞中 ,联合应用IFN γ和NGF比单独应用二者有更加显著诱导分化的作用。与IFN γ诱导TrkAmRNA表达增加不同 ,联合用药导致TrkAmRNA表达下调 ,可能存在某种负调节机制。研究结果表明 ,联合应用IFN γ和NGF可能是治疗NB甚至是晚期NB的合理方案

Neuronal differentiation and TrkA expression of neuroblastoma induced by the combination treatment of interferon-gamma and NGF

OBJECTIVE: Neuroblastoma (NB) is one of the most common solid tumors of early childhood. A small percentage of NB has been found to undergo spontaneous or chemotherapeutically induced regression or differentiation into benign ganglioneuromas. The differentiation therapy is a respective method in the clinical treatment of NB. OBJECTIVE: Interferon-gamma (IFN-gamma), nerve growth factor (NGF), and combination of IFN-gamma and NGF were used as induction drugs respectively to induce the differentiation of human SMS-KCNR cells. The cells were observed for morphological differences, growth arrest, and the TrkA mRNA expression of the 10th day-cells. METHODS: The human SMS-KCNR cell lines were cultured. The TrkA mRNA expression was detected by RT-PCR. Cell counts were measured by using a hemocytometer and the viability was assessed with the trypan blue exclusion. The morphological differentiation was observed under the phase-contrast microscope. RESULTS: The simultaneous loading of NGF with IFN-gamma caused more prominent neurite outgrowth than independent treatment with either IFN-gamma or NGF in SMS-KCNR cells (P < 0.01). The cell proliferation was inhibited significantly in the former (P < 0.01). The TrkA mRNA level increased in the IFN-gamma group (P < 0.01). While there was no significant change of TrkA mRNA level in the NGF group (P > 0.05). The TrkA mRNA level was obvious decreased in the combination group (P < 0.01). CONCLUSION: In human SMS-KCNR cells, the combination of IFN-gamma and NGF showed prominent effect on the cell differentiation. In contrast with the induction of TrkA mRNA expression by IFN-gamma, the combination treatment could decrease TrkA mRNA expression, which might indicate that a kind of negative feedback mechanism existed. These data suggest that using IFN-gamma and NGF together may be an appropriate strategy in the treatment of children with advanced neuroblastoma.