Department of Otolaryngology-Head and Neck Surgery, Wayne State University School of Medicine, Detroit, Michigan.
Stem Cells Dev. 2019 Jul 1;28(13):860-870. doi: 10.1089/scd.2018.0247. Epub 2019 May 29.
Auditory signals are processed in multiple central nervous system structures, including the auditory cortex (AC). Development of stem cell biology provides the opportunity to identify neural stem cells (NSCs) in the central nervous system. However, it is unclear whether NSCs exist in the AC. The aim of this study is to determine the existence of NSCs in the postnatal mouse AC. To accomplish this aim, postnatal mouse AC tissues were dissected and dissociated into singular cells and small cell clumps, which were suspended in the culture medium to observe neurosphere formation. The spheres were examined by quantitative real-time polymerase chain reaction and immunofluorescence to determine expression of NSC genes and proteins. In addition, AC-spheres were cultured in the presence or absence of astrocyte-conditioned medium (ACM) to study neural differentiation. The results show that AC-derived cells were able to proliferate to form neurospheres, which expressed multiple NSC genes and proteins, including SOX2 and NESTIN. AC-derived NSCs (AC-NSCs) differentiated into cells expressing neuronal and glial cell markers. However, the neuronal generation rate is low in the culture medium containing nerve growth factor, ∼8%. To stimulate neuronal generation, AC-NSCs were cultured in the culture medium containing ACM. In the presence of ACM, ∼29% AC-NSCs differentiated into cells expressing neuronal marker class III β-tubulin (TUJ1). It was observed that the length of neurites of AC-NSC-derived neurons in the ACM group was significantly longer than that of the control group. In addition, synaptic protein immunostaining showed significantly higher expression of synaptic proteins in the ACM group. These results suggest that ACM is able to stimulate neuronal differentiation, extension of neurites, and expression of synaptic proteins. Identifying AC-NSCs and determining effects of ACM on NSC differentiation will be important for the auditory research and other neural systems.
听觉信号在多个中枢神经系统结构中进行处理，包括听觉皮层 (AC)。干细胞生物学的发展为识别中枢神经系统中的神经干细胞 (NSC) 提供了机会。然而，目前尚不清楚 AC 中是否存在 NSCs。本研究旨在确定出生后小鼠 AC 中是否存在 NSCs。为了实现这一目标，分离出生后小鼠的 AC 组织并解离成单个细胞和小细胞团，将其悬浮在培养基中以观察神经球的形成。通过定量实时聚合酶链反应和免疫荧光检查来检查球体，以确定 NSC 基因和蛋白质的表达。此外，在存在或不存在星形胶质细胞条件培养基 (ACM) 的情况下培养 AC 球体，以研究神经分化。结果表明，AC 衍生细胞能够增殖形成表达多种 NSC 基因和蛋白质的神经球，包括 SOX2 和 NESTIN。AC 衍生的 NSCs (AC-NSCs) 分化为表达神经元和神经胶质细胞标志物的细胞。然而，在含有神经生长因子的培养基中培养时，神经元的生成率约为 8%。为了刺激神经元生成，将 AC-NSCs 在含有 ACM 的培养基中培养。在 ACM 的存在下，约 29%的 AC-NSCs 分化为表达神经元标志物 III 型 β-微管蛋白 (TUJ1) 的细胞。观察到 ACM 组中 AC-NSC 衍生神经元的突起长度明显长于对照组。此外，突触蛋白免疫染色显示 ACM 组中突触蛋白的表达明显更高。这些结果表明，ACM 能够刺激神经元分化、突起延伸和突触蛋白的表达。鉴定 AC-NSCs 并确定 ACM 对 NSC 分化的影响，对于听觉研究和其他神经系统的研究都非常重要。
