Isolation and culture of hair cell progenitors from postnatal rat cochleae.
Cochlear hair cells are a terminally differentiated cell population that is crucial for hearing. Although recent work suggests that there are hair cell progenitors in postnatal mammalian cochleae, isolation and culture of pure hair cell progenitors from a well-defined cochlear area have not been reported. Here we present an experimental method that allows isolation and culture of hair cell progenitors from postnatal rat cochleae. These progenitor cells are isolated from the lesser epithelial ridge (LER, or outer spiral sulcus cell) area of pre-plated neonatal rat cochlear segments. They express the same markers as LER cells in vivo, including ZO1, Islet1, Hes1, and Hes5. When these cells are induced to express Hath1, they show the potential to differentiate into hair cell-like cells. Interestingly, these cells can be lifted from monolayer cultures and maintained in aggregate cultures in which spheres can be formed. Hair cell progenitors in the spheres display their proliferating capability and express only epithelial markers. Furthermore, when these spheres are mixed with dissociated mesenchymal cells prepared from postnatal rat utricular whole mounts, and replated onto a collagen substratum, the epithelial progenitor cells are able to differentiate into cells expressing markers of hair cells and supporting cells in epithelial islands, which mirrors the inner ear sensory epithelium in vivo. Successful isolation and culture of hair cell progenitors from the mammalian cochlea will facilitate studies on gene expression profiling and mechanism of differentiation/regeneration of hair cells, which are crucial for repairing hearing loss.
Zhai
Suoqiang
S
Institute of Otolaryngology, General Hospital of PLA, Beijing 100853, China.
Shi
Li
L
Wang
Bu-er
BE
Zheng
Guiliang
G
Song
Wei
W
Hu
Yinyan
Y
Gao
Wei-Qiang
WQ
D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D051792:Basic Helix-Loop-Helix Proteins / Q000378:metabolism; D001973:Bromodeoxyuridine / Q000378:metabolism; D002452:Cell Count / Q000379:methods; D049109:Cell Proliferation; D002469:Cell Separation; D002478:Cells, Cultured; D003051:Cochlea / Q000166:cytology*; D018920:Coculture Techniques / Q000379:methods; D015870:Gene Expression / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006198:Hair Cells, Auditory / Q000166:cytology* / Q000502:physiology; D018398:Homeodomain Proteins / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods; D007633:Keratins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D024507:Nonmuscle Myosin Type IIA / Q000378:metabolism; D009924:Organ Culture Techniques; D010750:Phosphoproteins / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D051381:Rats; D012097:Repressor Proteins / Q000378:metabolism; D013234:Stem Cells / Q000502:physiology*; D000072056:Transcription Factor HES-1; D014162:Transfection / Q000379:methods; D014746:Vimentin / Q000378:metabolism; D062826:Zonula Occludens-1 Protein
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D051792:Basic Helix-Loop-Helix Proteins; D001973:Bromodeoxyuridine; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins; D007633:Keratins; D008565:Membrane Proteins; D024507:Nonmuscle Myosin Type IIA; D010750:Phosphoproteins; D018809:Proliferating Cell Nuclear Antigen; D012097:Repressor Proteins; D000072056:Transcription Factor HES-1; D014746:Vimentin; D062826:Zonula Occludens-1 Protein; C489735:Hes1 protein, rat; C489686:Hes5 protein, rat; C110528:Mapk8ip1 protein, rat; C569044:Tjp1 protein, rat; C485184:enhanced green fluorescent protein; C413374:p27 antigen