PCP信号通路介导肠神经嵴细胞分化异常在先天性巨结肠发生中的作用及其调控机制研究

Hirschsprung disease (HSCR) is a congenital neuro-developmental disorder implicating the enteric nervous system (ENS).Research find that the alterations of intestinal micro-environment in primarily embryogenic period can affect migration and differentiation of neural crest cells．Planar cell polarity pathway(PCP)was well known to have key roles in axon guidance. Recently,research has revealed that Feedback regulation of apical progenitor fate by immature neurons through PCP signaling pathway. Our previous study of Fzd3 and Celsr3 gene sequencing identified that related to HSCR. Based on the previously confirmed effects of 4 newly-detected loss-of-function SNP mutations among HD patients, we plan to first explore their influence on regulation the developmental features of the enteric neural crest cell in vitro. Secondly, we try to study how the mutants may affect the PCP-JNK-Notch signaling activit,and examine the pathway mechanism by inhibition the inactivity or rescue of core-enzyme. Lastly, we shall analyze the pathological characteristics of neurogenesis and gliogenesis in the colon of HD patients. Our proposed study will disclose the biological and cellular mechanisms and their pathological roles in predisposing people to HD.

先天性巨结肠（HD）是一种典型的肠神经嵴细胞（ENCC）发育异常性疾病，胚胎早期肠内微环境改变是其发生的重要病因。既往认为平面细胞极性信号通路（PCP）主要参与调控细胞定向迁移和极性分布，最近研究发现该通路在神经前体细胞有序分化过程中同样发挥重要作用。申请人前期针对HD患者PCP信号通路候选基因重测序、核心蛋白表达及神经节细胞形态研究，发现该通路核心基因FZD3和CELSR3与疾病发生密切相关。本课题拟运用离体培养的PC12细胞系和ENCC模型，以新发现的罕见变异为切入点，首先研究PCP-JNK-Notch通路的活性效应分子和下游机制，进而明确其对ENCC发育特性的影响并通过抑制核心酶活化、补充下游分子证实其特异性，最后在肠组织标本中进一步验证相应分子标志物及相关病理改变。阐明FZD3、CELSR3基因突变干扰肠神经嵴细胞有序分化的调控机制，为揭示HD的复杂发病机制提供新线索和理论依据。

先天性巨结肠（HD）是一种典型的肠神经嵴细胞（ENCC）发育异常性疾病，既往认为平面细胞极性信号通路（PCP）主要参与调控细胞定向迁移和极性分布，最近研究发现该通路在中枢神经前体细胞同样发挥重要作用。本研究分析了Celsr3在胎鼠后肠表达的时空特意性与ENCC迁移与定植时空相一致，验证CELSR3在小鼠ENCC中高表达，验证c1961t位点突变可导致Celsr3表达异常；探讨HD患者CELSR3基因突变所致的PCP信号通路功能失常引起ENS发育过程中增殖/分化失平衡的可能机制。