The new prognosis assessment method which the Interaction between NGFR and GM1 is applied to, and NGF sensitive regulatory differentiation derivation therapy for neuroblastma patients

本研究的目的是探讨神经生长因子受体(NGFR，TrkA)与GM1的相互作用在神经母细胞瘤临床诊断和治疗中的应用。我们研究了GM1的检测方法，结合NGFR(TrkA)对临床标本进行检测。尝试了提高免疫印迹法和薄层色谱法的灵敏度。因为不存在抗GM1的高敏抗体。用霍乱毒素B亚单位(CTB)检测。虽然用Trk特异性抗体的免疫沉淀法是可行的，但结合Trk的GM1的检测在敏感性方面掌握困难。对谈话状态不同的临床样本的分析不能适应这一过程。因此，我们利用细胞系集中了NGFR(TrkA)和GM1相互作用的详细机制。利用激光共聚焦显微镜荧光抗体技术，发现神经生长因子受体(NGFR，TrkA)和GM1在分化为神经元的PC12细胞和神经母细胞瘤细胞株NB-1中的细胞表面分布情况有明显不同。这一结果提示，神经母细胞瘤在NGF刺激下不能分化为神经元，这是由于Trk和GM1相互作用系统的破坏所致。但是，如果这种机制对分化调控起主导作用，我们就不能解释在NGF高浓度培养条件下高表达Trk的神经母细胞瘤分化的机制。我们比较了PC12细胞和可被NGF诱导分化的PCTrk(Trk高表达PC12)细胞系，假设在低浓度NGF作用下，高Trk表达细胞存在竞争抑制机制。本研究为神经母细胞瘤的新诊断和治疗提供了重要依据。

our purpose was the application of the interaction with GM1 of NGFR (TrkA) to the clinical diagnosis and the cure of neuroblastoma. We studied the detection method of GM1 which combined in NGFR (TrkA) was examined to cover clinical samples. Increase in sensitivity of the immunoblotting method and the TLC (thin layer chromatography) was tried. Because an anti-GM1 high sensitive antibody didn't exist. It was detected by using CTB (cholera toxin B subunit). Though the immunoprecipitation method with a Trk specific antibody was possible the detection of GM1 which combined in Trk mastered distress in the point of the sensitivity. The analysis of the clinical samples which conversation states varied couldn't adapt this procedure. So, we concentrated the detailed mechanism of NGFR (TrkA) and the GM1 interaction by using cell lines. We found the cell surface distribution situation of NGFR (TrkA) and GM1 was remarkably different in the PC12 cell which can differentiate to neuron, and the neuroblastoma cell line NB-1 by the fluorescent antibody technique which a confocal laser microscope was used for. This result suggested that neuroblastma cannot differentiate to neuron with NGF stimulation because of the destruction of the interaction system between Trk and GM1. But, if this mechanism is dominant for the differentiation regulation, we cannot explain the mechanism which a hyper Trk expression neuroblastma differentiates under the NGF high concentration culture condition. We compared PC12 cell line with the PCTrk (Trk hyper expression PC12) cell line that can differentiate by NGF on the assumption that the competitive inhibition mechanism occurred in the hyper Trk expression cells under the low concentration of NGF. We could get essential findings for new diagnosis and the cure of neuroblastoma in this study.