Basic study on the differentiation induction of human germ cell tumors and its clinical application

1. 10^<-2>M N， N′-六亚甲基双乙酰胺（HMBA）在体外诱导多效性人胚胎癌细胞（NEC14）分化。分化伴随着人主要组织相容性抗原（HLA-A、B、C）的显著诱导，以及阶段特异性胚胎抗原（从SSEA-1^-/SSEA-3^+到SSEA-1^+/SSEA-3^-）的变化。分化后的NEC14细胞表达间充质相关分子vimentin和tenascin；因此，HMBA可以诱导这些细胞产生胚胎中胚层的某些间充质成分。在未分化的NEC14细胞中，N-myc的表达水平较高，但在HMBA诱导后，N-myc的表达水平瞬间下降至不到原始水平的1/10。为了研究N-myc基因在NEC14细胞分化中的作用，我们建立了几个表达外源N-myc基因的NEC14转化子。它们具有较短的群体倍增时间、较高的电镀效率和致瘤潜力。血清中神经元特异性烯醇化酶（NSE）的水平被发现是未成熟畸胎瘤和生殖细胞异常瘤的一种新的诊断指标。从人畸胎癌细胞系PA-1中，我们建立了一个克隆细胞系，在体外多细胞球体中产生独特的神经莲座。该细胞系表达神经外胚层抗原A2B5、HNK-1、NC-1、NSE、S-100蛋白和vimentin。这些玫瑰花结显示出明显的极化和重叠沉积的细胞外基质成分，包括层粘连蛋白、IV型胶原、硫酸肝素蛋白多糖和腱素。

1. A plupribotent human embryonal carcinoma cell line (NEC14) could be induced to differentiate morphologically by treatment with 10^<-2>M N, N'-hexamethylene-bis-acetamide (HMBA) in vitro. The differentiation accompanied the marked induction of human major histocompatibility antigens (HLA-A, B, C), and the changes of stage specific embryonic antigens (from SSEA-1^-/SSEA-3^+ to SSEA-1^+/SSEA-3^-). Differentiated NEC14 cells expressed mesenchym-associated molecules, vimentin and tenascin ; thus HMBA can induce these cells to certain mesenchymalle elements of embryonal mesoderm.2. The level of N-myc expression was high in undifferentiated NEC14 cells, but decreased transiently to less than 1/10 of the original level after the HMBA induction. To investigate the role of N-myc gene in the NEC14 cell differentiation, we established several NEC14 transformants expressing exogenous N-myc genes. They showed shorter population doubling time, increased plating efficiency and tumorigenic potential.3. The level of neuron specific enolase (NSE) in serum was found to be a novel diagnostic marker for immature teratomas and dysgerminomas.4. From the human teratocarcinoma line PA-1, we established a clonal line that produced a distinct neural rosettes within in vitro multicellular spheroids. This line expressed neuroectodermal antigens such as A2B5, HNK-1 and NC-1, NSE, S-100 protein and vimentin. These rosette showed a strikingly polarized and overlapped deposition of extracellular matrix components including laminin, type IV collagen, heparan sulfate proteoglycan and tenascin.