In vivo outer hair cell gene editing ameliorates progressive hearing loss in dominant-negative Kcnq4 murine model.
In vivo outer hair cell gene editing ameliorates progressive hearing loss in dominant-negative Kcnq4 murine model.
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作者：
Noh B;Rim JH;Gopalappa R;Lin H;Kim KM;Kang MJ;Gee HY;Choi JY;Kim HH;Jung J
外毛细胞（OHC）变性是导致进行性听力损失和老年性耳聋的主要原因。尽管这些疾病的发病率很高，但目前还没有靶向治疗。研究方法：我们产生了一种携带Kcnq 4 W276 S/+的小鼠模型，以重现DFNA 2，DFNA 2是一种伴随OHC变性的进行性听力损失的常见遗传形式。在对指导RNA、Cas9、载体和递送途径进行全面优化后，我们应用了体内基因编辑策略来破坏Kcnq 4中的显性负等位基因并预防进行性听力损失。结果如下：使用靶向OHC的双重腺相关病毒包进行体内基因编辑显著改善了听觉脑干反应和畸变产物耳声发射的听觉阈值。此外，我们开发了一种新的活细胞成像技术，使用铊离子来研究OHC的膜电位，并成功地证明了突变等位基因破坏导致更多的超极化OHC，表明KCNQ 4通道活性升高。结论：这些发现可以促进DFNA 2靶向疗法的开发，并支持使用基于CRISPR的基因疗法来纠正OHC中的缺陷。
Outer hair cell (OHC) degeneration is a major cause of progressive hearing loss and presbycusis. Despite the high prevalence of these disorders, targeted therapy is currently not available. Methods: We generated a mouse model harboring Kcnq4W276S/+ to recapitulate DFNA2, a common genetic form of progressive hearing loss accompanied by OHC degeneration. After comprehensive optimization of guide RNAs, Cas9s, vehicles, and delivery routes, we applied in vivo gene editing strategy to disrupt the dominant-negative allele in Kcnq4 and prevent progressive hearing loss. Results: In vivo gene editing using a dual adeno-associated virus package targeting OHCs significantly improved auditory thresholds in auditory brainstem response and distortion-product otoacoustic emission. In addition, we developed a new live-cell imaging technique using thallium ions to investigate the membrane potential of OHCs and successfully demonstrated that mutant allele disruption resulted in more hyperpolarized OHCs, indicating elevated KCNQ4 channel activity. Conclusion: These findings can facilitate the development of targeted therapies for DFNA2 and support the use of CRISPR-based gene therapy to rectify defects in OHCs.
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影响因子：
12.8
作者：
Jung, Jinsei;Lin, Haiyue;Gee, Heon Yung
通讯作者：
Gee, Heon Yung
DOI：
10.1056/nejmra1616601
发表时间：
2017-12-21
期刊：
The New England journal of medicine
影响因子：
--
作者：
Cunningham LL;Tucci DL
通讯作者：
Tucci DL
影响因子：
3.5
作者：
Akita, J;Abe, S;Usami, S
通讯作者：
Usami, S
影响因子：
5.3
作者：
Beisel, KW;Rocha-Sanchez, SM;Fritzsch, B
通讯作者：
Fritzsch, B
影响因子：
11.4
作者：
Kharkovets, T;Dedek, K;Jentsch, TJ
通讯作者：
Jentsch, TJ