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Virally mediated Kcnq1 gene replacement therapy in the immature scala media restores hearing in a mouse model of human Jervell and Lange-Nielsen deafness syndrome.

Chang Q, Wang J, Li Q, Kim Y, Zhou B, Wang Y, Li H, Lin X - EMBO Mol Med (2015)

Bottom Line: A modified adeno-associated virus construct carrying a Kcnq1 expression cassette was injected postnatally (P0-P2) into the endolymph, which resulted in Kcnq1 expression in most cochlear marginal cells where native Kcnq1 is exclusively expressed.We also found that extensive ectopic virally mediated Kcnq1 transgene expression did not affect normal cochlear functions.Our results demonstrate the first successful gene therapy treatment for gene defects specifically affecting the function of the stria vascularis, which is a major site affected by genetic mutations in inherited hearing loss.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology, Emory University School of Medicine, Atlanta, GA, USA.

No MeSH data available.


Related in: MedlinePlus

Quantification of Kcnq1-positive marginal cells and the effect of treatment on the cellular organization of the marginal cells in the SVThe membranes of marginal cells are labeled (red) by phalloidin conjugated with rhodamine.A Immunolabeling results (Kcnq1 labeled in green) in WT mice.B, C Immunolabeling results (Kcnq1 labeled in green) of treated Kcnq1−/− mice, middle (B) and apical (C) turns, respectively.D The percentage of marginal cells having positive Kcnq1 immunolabeling signal above a visually detectable level is shown for WT (gray bars, left), untreated Kcnq1−/− (middle), and treated Kcnq1−/− mice (black bars, right). Data are given as mean ± SD (n = 6).E, F The organization of marginal cells in the SV is outlined by labeling with phalloidin conjugated with rhodamine. Results from WT (E) and untreated Kcnq1−/− mice (F) are compared.Data information: Scale bars represent approximately 50 μm.
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fig03: Quantification of Kcnq1-positive marginal cells and the effect of treatment on the cellular organization of the marginal cells in the SVThe membranes of marginal cells are labeled (red) by phalloidin conjugated with rhodamine.A Immunolabeling results (Kcnq1 labeled in green) in WT mice.B, C Immunolabeling results (Kcnq1 labeled in green) of treated Kcnq1−/− mice, middle (B) and apical (C) turns, respectively.D The percentage of marginal cells having positive Kcnq1 immunolabeling signal above a visually detectable level is shown for WT (gray bars, left), untreated Kcnq1−/− (middle), and treated Kcnq1−/− mice (black bars, right). Data are given as mean ± SD (n = 6).E, F The organization of marginal cells in the SV is outlined by labeling with phalloidin conjugated with rhodamine. Results from WT (E) and untreated Kcnq1−/− mice (F) are compared.Data information: Scale bars represent approximately 50 μm.

Mentions: Immunolabeling using the flattened cochlear preparation (Chang et al, 2008) allowed us to quantify cellular Kcnq1 expression (Fig3). The hexagonal cell membrane of individual marginal cells was outlined by labeling with phalloidin (labeled in red in Fig3). In the untreated cochleae of Kcnq1−/− mice, we observed that the orderly hexagonal organization of the marginal cells (Fig3A–C,E) was damaged (Fig3F). We also found that, compared to the marginal cells in WT mice (Fig3E), the sizes of the marginal cells in untreated Kcnq1−/− mice vary greatly. Moreover, many cells in untreated Kcnq1−/− mice had missing nuclei (arrows in Fig3F), suggesting cellular degeneration or distress. Counting positively transduced marginal cells (Fig3B and C) yielded viral transduction efficiencies ranging from 75 ± 5% (n = 6) to 71 ± 8% (n = 6) to 61 ± 10% (n = 6) for marginal cells, respectively, located in the basal, middle, and apical turns (black bars on the right of Fig3D). These values were slightly lower than those in WT counterparts (Fig3D, gray bars), but were sufficient to prevent deafness (Fig 5B).


Virally mediated Kcnq1 gene replacement therapy in the immature scala media restores hearing in a mouse model of human Jervell and Lange-Nielsen deafness syndrome.

Chang Q, Wang J, Li Q, Kim Y, Zhou B, Wang Y, Li H, Lin X - EMBO Mol Med (2015)

Quantification of Kcnq1-positive marginal cells and the effect of treatment on the cellular organization of the marginal cells in the SVThe membranes of marginal cells are labeled (red) by phalloidin conjugated with rhodamine.A Immunolabeling results (Kcnq1 labeled in green) in WT mice.B, C Immunolabeling results (Kcnq1 labeled in green) of treated Kcnq1−/− mice, middle (B) and apical (C) turns, respectively.D The percentage of marginal cells having positive Kcnq1 immunolabeling signal above a visually detectable level is shown for WT (gray bars, left), untreated Kcnq1−/− (middle), and treated Kcnq1−/− mice (black bars, right). Data are given as mean ± SD (n = 6).E, F The organization of marginal cells in the SV is outlined by labeling with phalloidin conjugated with rhodamine. Results from WT (E) and untreated Kcnq1−/− mice (F) are compared.Data information: Scale bars represent approximately 50 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4551345&req=5

fig03: Quantification of Kcnq1-positive marginal cells and the effect of treatment on the cellular organization of the marginal cells in the SVThe membranes of marginal cells are labeled (red) by phalloidin conjugated with rhodamine.A Immunolabeling results (Kcnq1 labeled in green) in WT mice.B, C Immunolabeling results (Kcnq1 labeled in green) of treated Kcnq1−/− mice, middle (B) and apical (C) turns, respectively.D The percentage of marginal cells having positive Kcnq1 immunolabeling signal above a visually detectable level is shown for WT (gray bars, left), untreated Kcnq1−/− (middle), and treated Kcnq1−/− mice (black bars, right). Data are given as mean ± SD (n = 6).E, F The organization of marginal cells in the SV is outlined by labeling with phalloidin conjugated with rhodamine. Results from WT (E) and untreated Kcnq1−/− mice (F) are compared.Data information: Scale bars represent approximately 50 μm.
Mentions: Immunolabeling using the flattened cochlear preparation (Chang et al, 2008) allowed us to quantify cellular Kcnq1 expression (Fig3). The hexagonal cell membrane of individual marginal cells was outlined by labeling with phalloidin (labeled in red in Fig3). In the untreated cochleae of Kcnq1−/− mice, we observed that the orderly hexagonal organization of the marginal cells (Fig3A–C,E) was damaged (Fig3F). We also found that, compared to the marginal cells in WT mice (Fig3E), the sizes of the marginal cells in untreated Kcnq1−/− mice vary greatly. Moreover, many cells in untreated Kcnq1−/− mice had missing nuclei (arrows in Fig3F), suggesting cellular degeneration or distress. Counting positively transduced marginal cells (Fig3B and C) yielded viral transduction efficiencies ranging from 75 ± 5% (n = 6) to 71 ± 8% (n = 6) to 61 ± 10% (n = 6) for marginal cells, respectively, located in the basal, middle, and apical turns (black bars on the right of Fig3D). These values were slightly lower than those in WT counterparts (Fig3D, gray bars), but were sufficient to prevent deafness (Fig 5B).

Bottom Line: A modified adeno-associated virus construct carrying a Kcnq1 expression cassette was injected postnatally (P0-P2) into the endolymph, which resulted in Kcnq1 expression in most cochlear marginal cells where native Kcnq1 is exclusively expressed.We also found that extensive ectopic virally mediated Kcnq1 transgene expression did not affect normal cochlear functions.Our results demonstrate the first successful gene therapy treatment for gene defects specifically affecting the function of the stria vascularis, which is a major site affected by genetic mutations in inherited hearing loss.

View Article: PubMed Central - PubMed

Affiliation: Department of Otolaryngology, Emory University School of Medicine, Atlanta, GA, USA.

No MeSH data available.


Related in: MedlinePlus