PDF(Pubmed)
Abstract：
Inherited hearing impairment is a remarkably heterogeneous monogenic condition, involving hundreds of genes, most of them with very small (< 1%) epidemiological contributions. The exception is GJB2, the gene encoding connexin-26 and underlying DFNB1, which is the most frequent type of autosomal recessive non-syndromic hearing impairment (ARNSHI) in most populations (up to 40% of ARNSHI cases). DFNB1 is caused by different types of pathogenic variants in GJB2, but also by large deletions that keep the gene intact but remove an upstream regulatory element that is essential for its expression. Such large deletions, found in most populations, behave as complete loss-of-function variants, usually associated with a profound hearing impairment. By using CRISPR-Cas9 genetic edition, we have generated a murine model (Dfnb1em274) that reproduces the most frequent of those deletions, del(GJB6-D13S1830). Dfnb1em274 homozygous mice are viable, bypassing the embryonic lethality of the Gjb2 knockout, and present a phenotype of profound hearing loss (> 90 dB SPL) that correlates with specific structural abnormalities in the cochlea. We show that Gjb2 expression is nearly abolished and its protein product, Cx26, is nearly absent all throughout the cochlea, unlike previous conditional knockouts in which Gjb2 ablation was not obtained in all cell types. The Dfnb1em274 model recapitulates the clinical presentation of patients harbouring the del(GJB6-D13S1830) variant and thus it is a valuable tool to study the pathological mechanisms of DFNB1 and to assay therapies for this most frequent type of human ARNSHI.
摘要：
遗传性听力障碍是一种非常异质的单基因疾病，涉及数百个基因，他们中的大多数与非常小（<1％）的流行病学贡献。例外的是GJB2，该基因编码连接蛋白-26和潜在的DFNB1，这是大多数人群（高达40％的ARNSHI病例）中最常见的常染色体隐性遗传性非综合征性听力障碍（ARNSHI）类型。DFNB1是由GJB2中不同类型的致病变体引起的，也是由大的缺失引起的，这些缺失使基因保持完整，但去除对其表达至关重要的上游调控元件。如此大的删除，在大多数人群中发现，表现为完全丧失功能的变体，通常与严重的听力损伤有关。通过使用CRISPR-Cas9基因版，我们已经产生了一个鼠类模型（Dfnb1em274），再现这些缺失中最常见的，德尔（GJB6-D13S1830）。Dfnb1em274纯合小鼠是可行的，绕过Gjb2基因敲除的胚胎致死性，并呈现与耳蜗特定结构异常相关的深度听力损失表型(>90dBSPL)。我们表明Gjb2表达几乎被废除，其蛋白质产物，Cx26，几乎在整个耳蜗中都不存在，与以前的条件性敲除不同，在以前的条件性敲除中，没有在所有细胞类型中获得Gjb2消融。Dfnb1em274模型概括了携带del（GJB6-D13S1830）变体的患者的临床表现，因此它是研究DFNB1的病理机制和测定这种最常见类型的人类ARNSHI疗法的有价值的工具。
