刺猬蛋白促进内耳Math1表达的实验研究

The Expression of Math1 Induced by Sonic Hedgehog in the Inner Ear

目的研究刺猬蛋白(Shh)对体外小鼠耳蜗多潜能祖代细胞(CNPs)的Math1的调节作用,探讨治疗感音神经性耳聋的新方法。方法体外分离培养出生后第1天小鼠的CNPs细胞,在DMEM/F12+Shh培养基培养,用RT-PCR、荧光素酶检测系统、免疫荧光细胞化学技术和流式细胞仪等方法检测诱导CNPs细胞早期分化的重要转录因子Math1的表达。结果Shh诱导分化组的Math1mRNA表达、Math1启动子的转录水平及Math1蛋白表达阳性率均高于未诱导细胞(P<0.05);抑制剂LY294002具有显著下调Shh对Math1启动子转录水平和Math1蛋白表达的作用(P<0.05)。结论出生后第1天小鼠的CNPs细胞能够在Shh的诱导下,促进Math1的转录和表达,诱导耳蜗毛细胞的早期分化。

Objective To study the influence of sonic hedgehog（Shh） on the expression of Mathl for cochlear neural progenitors（CNPs） in mice in vitro. It is a new method for the therapy of sensorineural hearing loss. Methods CNPs were isolated from the postnatal day 1 organ of Corti in mice in vitro, cultured in DMEM/F12 ＋ Shh. Mammalian atonal homologl （Math1 or Atoh1 ） is a transcription factor for the initial differentiation of auditory hair cells. Shh was used to examine the expression of Math1 by RT-PCR, luciferase assays, immunohistochemistry and flow cytometry. Results Shh increased the expression of Math1 mRNA transcripts by RT-PCR, the promoter activity of Math1 by luciferase assays, and the expression of Math1 protein by immunohistochemisty and flow cytometry（P〈0.05）. LY294002 attenuated the promoter activity of Math1 and the expression of Math1 protein induced by Shh（P〈0.05）. Conclusions Addition of Shh increases the expression of Math1 for CNPs from the mouse postnatal day 1 organ of Corti in vitro and, therefore, induces the initiation of auditory hair cells.