Ex vivo expansion of cochlear stem cells and the investigation of neural stem cell secreted-factors mediated survival promotion of auditory cells

• Main Authors: Hsin-Chien, Chen, 陳信傑
• Other Authors: Chih-Hung, Wang
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/67876378132591648154

博士 === 國防醫學院 === 醫學科學研究所 === 99 === It is generally accepted that cochlear hair cells exit cell cycle before birth. Sensorineural hearing loss thus is a handicapped disease due to irreversible and permanent hearing impairment. Stem cell-based therapy can be used as a potential approach to restore lost hearing. However, unlike neural stem cells that maintain populations in the adult brains of both rodents and humans, cochlear stem/progenitor cells (SPCs) appear to diminish in number after birth and may become quiescent in adult mammalian cochleae. For cochlear SPC application, a novel strategy to expand sufficient SPCs is obligatory. Our data demonstrate that ex vivo hypoxic culturing significantly increased sphere formation and viability of cochlear SPCs compared with those cultured under normoxic conditions. Concurrent with these self-renewal and proliferation promotion effects are changes in the expression of multiple stemness and associated gene targets, including Abcg2, nestin, p27Kip1and Vegf. Knockdown of Hif-1α expression by small interfering RNA inhibited hypoxia-induced cochlear SPC expansion and resulted in downregulation of Vegf, Abcg2, and nestin and upregulation of p27Kip1 gene expression. These results suggest that Hif-1α plays an important role in the stimulation of proliferation and maintenance of stemness of cochlear SPCs, which confers a great benefit of expanding cochlear SPCs via hypoxic conditions. The capacity for perpetual self-renewal is one of the main characteristics of stem cells. Little is known about the effect of embryonic neural stem cell (NSC)-secreted factors on auditory cell proliferation in vitro. 2 auditory cell types were cultured in the presence of NSC-secreted molecules and were evaluated in vitro. Our results demonstrated that both cell viability and cell proliferation were significantly enhanced on treatment with NSC-conditioned medium, which contains significantly elevated levels of leukemia inhibitory factor (LIF) secreted by NSCs. The NSC-conditioned medium not only activated the expression of leukemia inhibitory factor receptor in House Ear Institute-organ of Corti 1 cells but also upregulated the LIF downstream signal transducers and activators of transcription (STAT) 1 and STAT3. Blocking either the LIF signaling pathway with neutralizing antibodies or the downstream Janus kinase (JAK)/STAT pathway with JAK2 inhibitor AG490 resulted in a dose-dependent inhibition of cell proliferation, suggesting that NSC-secreted molecules promote auditory cell survival via the regulatory LIF/JAK/STAT signaling pathway.