轴突导向分子EphA2/ephrinA1介导腺样囊性癌嗜神经侵袭的分子机制

Perineural invasion(PNI) is emerging as an important pathologic feature of adenoid cystic carcinoma(ACC). It can be a source of diffuse invasion and distant metastasis for ACC. Despite increasing recognition of this metastatic process, there has been little progress in the understanding of molecular mechanisms behind PNI. Axon guidance molecules Eph/ephrin could regulate the formation of nervous and vascular system in embryogenesis and tumor angiogenesis in adults. However, relative research on Eph/ephrin mediating interaction between tumor and nerve has not been found. In our previous study, we found a high expression of EphA2/ephrinA1 in ACC tissues and the nerve tissues invaded by ACC. The expression of EphA2/ephrinA1 was significantly correlated to perineural invasion of ACC. We propose that EphA2/ephrinA1 might be involved in the process of PNI in ACC and mediate the PNI through a series of signaling cascade. We plan to design the in vitro and in vivo models through which to investigate the mechanisms of PNI mediated by EphA2/ephrinA1. This research includes three parts. The first one is to investigate the behavior and phenotype change of ACC cells during the interaction between ACC cell and nerve tissue. The second one is to detect the function of EphA2/ephrinA1 during the process of PNI in ACC. The third one is to investigate the molecular mechanisms of PNI mediated by EphA2/ephrinA1 in ACC, including the regulation of cell migration and survival. We expect to realize the complexity of axon guidance molecules EphA2/ephrinA1 and reveal the molecular mechanisms of PNI in some extent.

嗜神经侵袭是腺样囊性癌(ACC)重要的生物学行为特征之一，其机制目前仍不清楚。轴突导向分子Eph/ephrin在发育过程中可以调节神经及血管系统的形成，在成体则参与调节肿瘤血管生成，那么Eph/ephrin在成体是否也参与了肿瘤与神经的相互作用？我们前期研究发现EphA2/ephrinA1在ACC组织中的表达与ACC嗜神经侵袭相关。综合已有的研究结果，我们推测EphA2/ephrinA1信号可能参与调节ACC嗜神经侵袭过程。本课题拟探讨以下内容：⑴ACC与神经相互作用过程中的细胞行为及分子表型的变化；⑵通过体外体内模型检测EphA2/ephrinA1在ACC嗜神经侵袭过程中的作用；⑶EphA2/ephrinA1对ACC嗜神经侵袭过程中细胞迁移、存活能力的分子调控。有望揭示EphA2/ephrinA1调节ACC嗜神经侵袭的作用机制，为深入理解肿瘤嗜神经侵袭的发生机制及肿瘤的防治提供线索。

腺样囊性癌容易复发和远处转移，嗜神经侵袭是其重要生物学行为特征之一。本研究通过对轴突引导分子EphA2在腺样囊性癌中的表达和功能研究，分析其对腺样囊性癌细胞生长、浸润及转移的作用和机制，初步探讨EphA2对腺样囊性癌嗜神经侵袭的调节机制。我们通过将肿瘤细胞与背根神经节在基质胶中共培养，建立了腺样囊性癌嗜神经侵袭体外模型，通过该模型分析了腺样囊性癌细胞与神经相互作用过程中的基因表达变化，通过基因芯片筛选出369个表达上调的基因和920个表达下调的基因，其中包括神经系统调节相关因子的变化。通过RNAi沉默腺样囊性癌细胞系SACC-LM中的EphA2表达，通过迁移试验、浸润试验、粘附试验检测，发现抑制EphA2可以促进腺样囊性癌细胞在体外的迁移、浸润、嗜神经性侵袭以及粘附能力。通过裸鼠皮下成瘤试验，我们发现抑制EphA2可以显著增加裸鼠移植肿瘤生长体积，并通过检测Ki-67表达确定抑制EphA2可增加肿瘤的增殖率。通过裸鼠尾静脉注射腺样囊性癌细胞检测EphA2抑制对转移的影响，我们发现SACC-EphA2-RNAi细胞注射组具有显著的较大的肿瘤负荷及肺重量。我们进一步检测EphA2和其配体ephrinA1在人腺样囊性癌标本中的表达情况。结果显示，在腺样囊性癌肿瘤组织中EphA2及其配体ephrinA1均呈高表达，这显示了EphA2/ephrinA1信号在细胞调节中的复杂性，抑制EphA2对腺样囊性癌细胞所的促进作用可能是由于其配体所产生的相应变化。通过对腺样囊性癌组织标本的免疫组化研究，我们发现在肿瘤细胞浸润神经组织的浸润前缘，E-Cadherin表达减少，而Slug的表达与肿瘤细胞的嗜神经侵袭正相关。在标本研究中我们发现，E-Cadherin表达与EphA2表达负相关，而Slug表达与EphA2表达正相关。我们的研究表明EphA2可能通过对肿瘤上皮间质转化的调节影响腺样囊性癌细胞的增殖、转移和对神经组织的侵袭。