Schomann Timo, Mezzanotte Laura, De Groot John C M J, Rivolta Marcelo N, Hendriks Sanne H, Frijns Johan H M, Huisman Margriet A
Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands.
Optical Molecular Imaging Group, Department of Radiology, Erasmus Medical Center, Rotterdam, South Holland, the Netherlands.
PLoS One. 2017 Oct 30;12(10):e0187183. doi: 10.1371/journal.pone.0187183. eCollection 2017.
Stem-cell-based repair of auditory neurons may represent an attractive therapeutic option to restore sensorineural hearing loss. Hair-follicle-bulge-derived stem cells (HFBSCs) are promising candidates for this type of therapy, because they (1) have migratory properties, enabling migration after transplantation, (2) can differentiate into sensory neurons and glial cells, and (3) can easily be harvested in relatively high numbers. However, HFBSCs have never been used for this purpose. We hypothesized that HFBSCs can be used for cell-based repair of the auditory nerve and we have examined their migration and incorporation into cochlear modiolus explants and their subsequent differentiation. Modiolus explants obtained from adult wild-type mice were cultured in the presence of EF1α-copGFP-transduced HFBSCs, constitutively expressing copepod green fluorescent protein (copGFP). Also, modiolus explants without hair cells were co-cultured with DCX-copGFP-transduced HFBSCs, which demonstrate copGFP upon doublecortin expression during neuronal differentiation. Velocity of HFBSC migration towards modiolus explants was calculated, and after two weeks, co-cultures were fixed and processed for immunohistochemical staining. EF1α-copGFP HFBSC migration velocity was fast: 80.5 ± 6.1 μm/h. After arrival in the explant, the cells formed a fascicular pattern and changed their phenotype into an ATOH1-positive neuronal cell type. DCX-copGFP HFBSCs became green-fluorescent after integration into the explants, confirming neuronal differentiation of the cells. These results show that HFBSC-derived neuronal progenitors are migratory and can integrate into cochlear modiolus explants, while adapting their phenotype depending on this micro-environment. Thus, HFBSCs show potential to be employed in cell-based therapies for auditory nerve repair.
基于干细胞修复听觉神经元可能是恢复感音神经性听力损失的一种有吸引力的治疗选择。毛囊隆突来源的干细胞（HFBSCs）是这类治疗的有前景的候选者，因为它们（1）具有迁移特性，移植后能够迁移，（2）可以分化为感觉神经元和神经胶质细胞，以及（3）可以相对容易地大量获取。然而，HFBSCs从未被用于此目的。我们假设HFBSCs可用于基于细胞的听神经修复，并且我们已经研究了它们向耳蜗蜗轴外植体的迁移和整合以及随后的分化。从成年野生型小鼠获得的蜗轴外植体在组成型表达桡足类绿色荧光蛋白（copGFP）的EF1α-copGFP转导的HFBSCs存在下培养。此外，将没有毛细胞的蜗轴外植体与DCX-copGFP转导的HFBSCs共培养，后者在神经元分化过程中双皮质素表达时显示copGFP。计算HFBSCs向蜗轴外植体的迁移速度，两周后，将共培养物固定并进行免疫组织化学染色处理。EF1α-copGFP HFBSCs的迁移速度很快：80.5±6.1μm/h。到达外植体后，细胞形成束状模式并将其表型转变为ATOH1阳性神经元细胞类型。DCX-copGFP HFBSCs整合到外植体后变为绿色荧光，证实了细胞的神经元分化。这些结果表明，源自HFBSCs的神经祖细胞具有迁移能力，能够整合到耳蜗蜗轴外植体中，同时根据这种微环境改变其表型。因此，HFBSCs显示出可用于基于细胞的听神经修复治疗的潜力。
