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Virally expressed connexin26 restores gap junction function in the cochlea of conditional Gjb2 knockout mice.

ABSTRACT: Mutations in GJB2, which codes for the gap junction (GJ) protein connexin26 (Cx26), are the most common causes of human nonsyndromic hereditary deafness. We inoculated modified adeno-associated viral (AAV) vectors into the scala media of early postnatal conditional Gjb2 knockout mice to drive exogenous Cx26 expression. We found extensive virally expressed Cx26 in cells lining the scala media, and intercellular GJ network was re-established in the organ of Corti of mutant mouse cochlea. Widespread ectopic Cx26 expression neither formed ectopic GJs nor affected normal hearing thresholds in wild-type (WT) mice, suggesting that autonomous cellular mechanisms regulate proper membrane trafficking of exogenously expressed Cx26 and govern the functional manifestation of them. Functional recovery of GJ-mediated coupling among the supporting cells was observed. We found that both cell death in the organ of Corti and degeneration of spiral ganglion neurons in the cochlea of mutant mice were substantially reduced, although auditory brainstem responses did not show significant hearing improvement. This is the first report demonstrating that virally mediated gene therapy restored extensive GJ intercellular network among cochlear non-sensory cells in vivo. Such a treatment performed at early postnatal stages resulted in a partial rescue of disease phenotypes in the cochlea of the mutant mice.

SUBMITTER: Yu Q

PROVIDER: S-EPMC3881370 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature

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Virally expressed connexin26 restores gap junction function in the cochlea of conditional Gjb2 knockout mice.

Yu Q Q Wang Y Y Chang Q Q Wang J J Gong S S Li H H Lin X X

Gene therapy 20131114 1

Mutations in GJB2, which codes for the gap junction (GJ) protein connexin26 (Cx26), are the most common causes of human nonsyndromic hereditary deafness. We inoculated modified adeno-associated viral (AAV) vectors into the scala media of early postnatal conditional Gjb2 knockout mice to drive exogenous Cx26 expression. We found extensive virally expressed Cx26 in cells lining the scala media, and intercellular GJ network was re-established in the organ of Corti of mutant mouse cochlea. Widesprea ...[more]

PMID: 24225640

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Project description:Connexin26 (Cx26) and connexin30 (Cx30) are two major protein subunits that co-assemble to form gap junctions (GJs) in the cochlea. Mutations in either one of them are the major cause of non-syndromic prelingual deafness in humans. Because the mechanisms of cochlear pathogenesis caused by Cx mutations are unclear, we investigated effects of Cx30 null mutation on GJ-mediated ionic and metabolic coupling in the cochlea of mice. A novel flattened cochlear preparation was used to directly assess intercellular coupling in the sensory epithelium of the cochlea. Double-electrode patch clamp recordings revealed that the absence of Cx30 did not significantly change GJ conductance among the cochlear supporting cells. The preserved electrical coupling is consistent with immunolabeling data showing extensive Cx26 GJs in the cochlea of the mutant mice. In contrast, dye diffusion assays showed that the rate and extent of intercellular transfer of multiple fluorescent dyes (including a non-metabolizable D-glucose analogue, 2-NBDG) among cochlear supporting cells were severely reduced in Cx30 null mice. Since the sensory epithelium in the cochlea is an avascular organ, GJ-facilitated intercellular transfer of nutrient and signaling molecules may play essential roles in cellular homeostasis. To test this possibility, NBDG was used as a tracer to study the contribution of GJs in transporting glucose into the cochlear sensory epithelium when delivered systemically. NBDG uptake in cochlear supporting cells was significantly reduced in Cx30 null mice. The decrease was also observed with GJ blockers or glucose competition, supporting the specificity of our tests. These data indicate that GJs facilitate efficient uptake of glucose in the supporting cells. This study provides the first direct experimental evidence showing that the transfer of metabolically-important molecules in cochlear supporting cells is dependent on the normal function of GJs, thereby suggesting a novel pathogenesis process in the cochlea for Cx-mutation-linked deafness.

Dataset Information

Virally expressed connexin26 restores gap junction function in the cochlea of conditional Gjb2 knockout mice.

Publications

Virally expressed connexin26 restores gap junction function in the cochlea of conditional Gjb2 knockout mice.

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