STUDY ON HAIR CELL REGENERATION IN THE MAMMALIAN COCHLEA

1)检测了腺病毒载体可转染腺病毒的细胞类型。我们将Adex1CAlacZ注射到豚鼠的耳蜗骨或前庭组织中。我们发现LacZ在包括螺旋神经节、血管纹、外毛细胞和支持细胞在内的多种耳蜗组织中都有表达。在前庭迷路中，淋巴管周围间隙的间皮细胞，椭圆体和壶腹的感觉细胞和支持细胞，壶腹的过渡细胞和暗细胞被转染组。在体内，我们发现含有苯酚的局麻液对豚鼠内耳的损伤允许LacZ基因通过圆窗膜导入豚鼠内耳。2)我们研究了地塞米松诱导的免疫抑制对腺病毒在豚鼠内耳腺病毒基因转移结果的影响，发现免疫抑制治疗减少了重复腺病毒介导的基因治疗的负面后果。3)我们证明了将Math 1基因插入豚鼠成熟耳蜗腺病毒的体内接种导致Math 1在无感觉耳蜗细胞中过表达。Math1的过度表达导致在Corti器中出现未成熟的毛细胞，并在牙间细胞、内沟和Hensen细胞区出现与Corti器相邻的新毛细胞。轴突从听神经束向一些新的毛细胞延伸，表明新的细胞吸引听神经元。4)我们评估了给予BrdU长达10天的耳聋豚鼠整个成熟耳蜗听感觉上皮的有丝分裂活性。耳聋后3~30天，在Corti器内可见BrdU阳性细胞。免疫组织化学证实BrdU阳性细胞为Deiters细胞，这些结果表明，成熟哺乳动物耳蜗中的Deiters细胞虽然有限，但仍具有有丝分裂活性，并可在有丝分裂后存活。

1)We examined the types of cells that can be tranfected with adenovirus vector. We injected an Adex1CAlacZ into the guinea pig cochlear or vestibular explants. We found expression of lacZ in a variety of cochlear tissues including the spiral ganglion, stria vascularis, and the outer hair cells and supporting cells. In the vestibular labyrinth, the mesothelial cells in the perilymphatic space, the sensory and supporting cells in the utricle and ampulla and the transitional and dark cells in the ampulla were transfected. In vivo, we found that damage from a local anesthetic solution containing phenol permitted LacZ gene delivery to the guinea pig inner ear via the round window membrane.2)We investigated the effects of dexamethasone-induced immunosuppression on the outcome of adenovirus gene transfer in guinea pig inner ears and found that immunosuppressive treatment reduced the negative consequences of repeated adenovirus-mediated gene therapy.3) We demonstrate that in vivo inoculation of adenovirus with the Math 1 gene insert into the mature guinea pig cochlea results in Math 1 overexpression in nonsensory cochlear cells. Math 1 overexpression leads to the appearance of immature hair cells in the organ of Corti and new hair cells adjacent to the organ of Corti in the interdental cell, inner sulcus, and Hensen cell regions. Axons are extended from the bundle of auditory nerve toward some of the new hair cells, suggesting that the new cells attract auditory neurons.4)We assessed mitotic activity in the auditory sensory epithelium throughout the mature cochlea of deafened guinea pigs given BrdU for up to 10 days. We found BrdU-positive cells within the organ of Corti from 3 to 30 days following deafening. Immunohistochemistry confirmed that the BrdU-positive cells were the Deiters cells, These findings indicate that, albeit limited, the Deiters cells in the mature mammalian cochlea possess a potential of mitotic activity and that they could survive after mitosis.