Dataset Information

Mouse otocyst transuterine gene transfer restores hearing in mice with connexin 30 deletion-associated hearing loss.

ABSTRACT: Although numerous causative genes for hereditary hearing loss have been identified, there are no fundamental treatments for this condition. Herein, we describe a novel potential treatment for genetic hearing loss. Because mutations or deletions in the connexin (Cx) genes are common causes of profound congenital hearing loss in both humans and mice, we investigated whether gene supplementation therapy using the wild-type Cx gene could cure hearing loss. We first generated inner ear-specific connexin 30 (Cx30)-deficient mice via the transuterine transfer of Cx30-targeted short hairpin RNA (shRNA-Cx30) into otocysts. The inner ear-specific Cx30-deficient mice mimicked homozygous Cx30-deficient mice both histologically and physiologically. Subsequently, we cotransfected the shRNA-Cx30 and the wild-type Cx30 gene. The cotransfected mice exhibited Cx30 expression in the cochleae and displayed normal auditory functions. Next, we performed the transuterine transfer of the wild-type Cx30 gene into the otocysts of homozygous Cx30-deficient mice, thereby rescuing the lack of Cx30 expression in the cochleae and restoring auditory functioning. These results demonstrate that supplementation therapy with wild-type genes can restore postnatal auditory functioning. Moreover, this is the first report to show that Cx-related genetic hearing loss is treatable by in vivo gene therapy.

SUBMITTER: Miwa T

PROVIDER: S-EPMC3677317 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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Mouse otocyst transuterine gene transfer restores hearing in mice with connexin 30 deletion-associated hearing loss.

Miwa Toru T Minoda Ryosei R Ise Momoko M Yamada Takao T Yumoto Eiji E

Molecular therapy : the journal of the American Society of Gene Therapy 20130416 6

Although numerous causative genes for hereditary hearing loss have been identified, there are no fundamental treatments for this condition. Herein, we describe a novel potential treatment for genetic hearing loss. Because mutations or deletions in the connexin (Cx) genes are common causes of profound congenital hearing loss in both humans and mice, we investigated whether gene supplementation therapy using the wild-type Cx gene could cure hearing loss. We first generated inner ear-specific conne ...[more]

PMID: 23587925

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Project description:Objective: Connexins are known to compose gap junctions by a pair of hemichannels connected in a head to head configuration. Each hemichannel is composed by hexameric cluster of connexins. There are 20 members of connexins known as connexin family in mice, and are expressed on each cell surface in different fashion. Connexin 30 (Cx30) is mainly expressed on the surface of oligodendroglia and astrocyte in central nervous system (CNS), but little is known about the functional relevance in neuroinflammatory diseases. To elucidate the role of Cx30 in the pathogenesis of neuroinflammatory disease, we induced experimental autoimmune encephalomyelitis (EAE) on Cx30 knock-out (Cx30-KO) mice and analyzed the clinical and neuropathological findings with wild-type control. Methods: C57BL/6J and Cx30-KO mice > 12 weeks of age were used in this study (N > 3 in each group). EAE was induced by immunization of mice with MOG35-55 peptide emulsified in CFA at a dose of 200 μg per mouse, followed by the administration of pertussis toxin (500 ng per mouse) on days 0 and 2. Mice were sacrificed and brain, spinal cord, spleen, and optic nerve were harvested for immunohistochemical analyses at the acute and chronic phases of EAE. Mice with EAE were scored as follows: 0, no disease; 1, limp tail; 2, abnormal gait and hind limb weakness (shaking); 2.5, paralysis of one hind limb; 3, paralysis of two hind limb; 3.5, ascending paralysis (able to move around); 4, tetraplegia; and 5, moribund (death). At the onset and chronic stage of EAE, mononuclear cells were isolated and analyzed by flow cytometry to check the distinct characteristics of cellular populations in inflamed CNS lesions. Results: Initial screening of immunohistological difference revealed basic activation of microglial cells in naïve Cx30-KO mice without any behavioral phenotype. Clinical signs of EAE were ameliorated in the Cx30-KO mice than in the control group mainly during the chronic phase of disease course. Immunohistochemical analyses of the fourth lumbar segment, brain and optic nerve revealed increased number of microglia in the Cx30-KO mice. Flow cytometric analysis also confirmed the findings. In contrast, there were no significant change in astroglial or oligodendroglial phenotype. Conclusion: Microglial activation appears to be the key factor in the Cx30-KO mice EAE with alleviation of chronic disease scores. Unexpectedly, microglia were already activated in naïve CNS, indicating protective phenotypic change of microglia in Cx30-KO mice.

Dataset Information

Mouse otocyst transuterine gene transfer restores hearing in mice with connexin 30 deletion-associated hearing loss.

Publications

Mouse otocyst transuterine gene transfer restores hearing in mice with connexin 30 deletion-associated hearing loss.

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