Hedgehog信号通路通过趋化因子调控成肌细胞迁移的分子机理研究

Craniofacial abnormalities account for most of congenital malformations. The normal development of tongue is critical for the development of palate. It is well known that Hedgehog signaling in neural crest cells (NCCs) is essential for normal patterning and growth of the mandible and the tongue. Whereas underlying molecular mechanisms need to be studied further. The defect of tongue development was found in Hedgehog signaling conditional knockout mice or overactivation mice.The muscle precursors fail to migrate into the tongue anlage. The abnormal expression of Chemokine SDF1/CXCL12 was detected in the tongue anlage of mutant mice. Recombinant Sonic Hedgehog/Shh could induce the expression of SDF1 in organ culture experiments. Furthermore the preliminary data showed that SDF-1α could induce the migration of myogenic progenitors towards and away from a chemokine concentration gradient. The results of experiments suggested that SDF1 mediate shh induced migration of myogenic progenitors and the expression level of SDF1 was regulated by Hedgehog signaling. Conditinal knockout mice, Dual-Luciferase Reporter Assay, in vitro organ culture, Chip assay and Flow cytometery will be used to address the mechanism of tongue development. The project will contribute to the treatment of the congenital craniofacial malformation.

新生儿颌面部发育畸形发病率位于先天性发育畸形的首位，其中舌发育异常会导致舌裂、巨舌、无舌以及腭裂等发育缺陷疾病。Hedgehog信号通路对于颌面部发育至关重要，而目前对其调控舌发育的分子机理少有报道。我们前期研究结果显示在神经嵴细胞中敲除或过度激活Hedgehog信号通路都导致舌不能正常发育，而且舌成肌细胞也没有正常迁移到舌原基部位。突变型小鼠舌原基部位趋化因子SDF1表达异常，体外试验进一步证实Shh可以调控SDF1的表达。我们提出以下假设：Hedgehog信号通路通过调控舌原基部位基质细胞中趋化因子SDF1的表达水平从而诱导舌肌肉细胞向舌原基迁移。本项目拟通过基因条件敲除小鼠、体外双荧光报告基因、染色质免疫沉淀分析、体外器官培养、流式细胞分选等方法，进一步深入研究Hedgehog信号通路对于舌发育过程中肌肉细胞迁移的调控机制，为寻找治疗颌面部发育畸形疾病有效途径和新药靶点提供理论依据。

来自枕骨体节的成肌细胞的迁移是舌形态发生的必要条件。然而，目前对调控成肌细胞迁移的分子机理少有报道。我们的研究结果显示在神经嵴细胞中敲除Hedgehog信号通路会导致舌不能正常发育，而且舌成肌细胞也没有正常迁移到舌原基部位。Hedgehog信号通路敲除（SmoWnt1Cre）及Isl1突变型小鼠（Isl1ShhCre）舌原基部位趋化因子Sdf1/Cxcl12表达异常，试验进一步证实Isl1通过Shh/Wnt5a级联调节第一鳃弓中趋化因子Cxcl12的表达模式。Isl1ShhCre胚胎中的Cxcl12+间充质细胞不能迁移到下颌远端区域，而是聚集在下颌的近端区域。CXCL12以浓度依赖性方式调控表达其受体Cxcr4的成肌细胞的迁移，CXCL12在低浓度下吸引Cxcr4+成肌细胞的迁移，但在高浓度下排斥Cxcr4+成肌细胞的迁移。Cxcl12+间充质细胞的聚集导致CXCL12的局部浓度升高，从而阻止了Cxcr4+成肌细胞的迁移。Ihh基因过表达挽救了突变小鼠舌发育的缺陷及成肌细胞的迁移，证实了Hedgehog信号在舌发育中的重要功能。本研究首次证实ISL1/SHH/WNT5A/CXCL12轴在舌发育过程中调节成肌细胞的迁移，有助于深入了解舌发育的分子机制，为寻找治疗颌面部发育畸形疾病有效途径和新药靶点提供理论依据。