支持细胞外泌体调控OHC caveolin信号通路参与Cx26基因迟发性耳聋

Recent studies showed that the Cx26-caused delayed hearing loss may result from the active cochlear amplification reducing, but not impairing K+ recycling in the cochlea. We hypothesized that the suppression to caveolin expressing in OHC by the exosomes secreted from the supporting cells is reduced. Following, the functions of the caveolin signal pathway（caveolin/prestin and/or caveolin/Ca2+/CaM/prestin）in OHC are changed and the OHC electromotility and the active cochlear amplification is impaired, which induce delayed hearing loss, when some types of Cx26 deficiency occur. This project will use a time-controlled, inducible Cx26 KO mice to make the mouse model with Cx26 deficiency-inducing delayed hearing loss. Then, exosomes and exosomal miRNAs from the Corti’s Organ will be identified, the expressions of caveolin and CaM and prestin in OHC of the Cx26 deficiency cochlea be observed, and Ca2+ concentration in OHCs and the nonline ccapacity of the plasma membrane of OHC be measured by the methods of single cell RT-PCR, western blot and confocal microscopy and whole-cell voltage-clamp. Finally, the interfering by the exosome miRNA to the caveolin expressing, and the signal pathway caveolin/prestin or caveolin/Ca2+/CaM/prestin in OHCs will be identified and confirmed in Corti’s Organs and in HEK293 cell and HEI-OC1 cell in vitro. The results will help to illuminate the functions of the exosome miRNA from the supporting cells and the caveolin signal pathway in OHCs in the pathogenesis of the Cx26-induced delayed hearing loss.

近年发现耳蜗K+循环破坏不能完全解释Cx26耳聋，Cx26迟发性聋主要因耳蜗主动放大受损。我们科学假说：Cx26某些缺陷，支持细胞外泌体对OHC caveolin表达抑制下降，引起OHC内caveolin信号通路（caveolin/prestin和（或）caveolin/Ca2+/CaM/prestin）功能异常，导致OHC电运动暨耳蜗主动放大受损，致迟发性聋。项目首先建立Cx26敲除迟发性聋小鼠模型，鉴定Corti器外泌体miRNA，通过单细胞RT-PCR、共聚焦显微镜和膜片钳等方法观察OHC膜caveolin、prestin、CaM表达、OHC Ca2+浓度及电运动；再用miRNA干预、caveolin过表达等方法在体外Corti器和细胞干预caveolin信号通路，分析其功能，进一步鉴别和证实该信号通路。有助于探明耳蜗外泌体和 caveolin信号通路在Cx26迟发性耳聋的致聋机理。

近年发现耳蜗K+循环破坏不能完全解释Cx26耳聋，Cx26迟发性聋主要因耳蜗主动放大受损。我们提出科学假说：Cx26某些缺陷，支持细胞外泌体对OHC caveolin表达抑制下降，引起OHC内caveolin信号通路（caveolin/prestin和（或）caveolin/Ca2+/CaM/prestin）功能异常，导致OHC电运动暨耳蜗主动放大受损，致迟发性聋。项目首先建立Cx26敲除迟发性聋小鼠模型，鉴定Corti器外泌体miRNA，通过单细胞RT-PCR、共聚焦显微镜和膜片钳等方法观察OHC膜caveolin、prestin、CaM表达、OHC Ca2+浓度及电运动；再用miRNA干预、caveolin过表达等方法在体外Corti器和细胞干预caveolin信号通路，分析其功能，进一步鉴别和证实该信号通路。有助于探明耳蜗外泌体和 caveolin信号通路在Cx26迟发性耳聋的致聋机理。