Centre for Stem Cell Biology, Department of Biomedical Science, University of Sheffield, Sheffield, UK.
School of Clinical Dentistry, University of Sheffield, Sheffield, UK.
Stem Cells Transl Med. 2020 Nov;9(11):1462-1476. doi: 10.1002/sctm.20-0037. Epub 2020 Jul 7.
The potential of obtaining cell cultures with neural crest resemblance (neural crest-derived stem cells [NCSCs]) from dental-related tissues, including human dental pulp cells (hDPCs), has been discussed in the literature. However, most reports include the use of serum-rich conditions and do not describe the potential for neural differentiation, slowing translation to the clinic. Therefore, we aimed to culture and characterize NCSCs from the human dental pulp in vitro and evaluate their ability to differentiate into neurons; we also investigated the effectiveness of the addition of BMP4 to enhance this potential. Cultures were established from a varied cohort of patient samples and grown, as monolayers, in serum, serum-free, and also under sphere-aggregation conditions to induce and identify a NCSC phenotype. hDPC cultures were characterized by immunocytochemistry and reverse transcription quantitative polymerase chain reaction. Monolayer cultures expressed stem cell, neural progenitor and neural crest-related markers. Culturing hDPCs as neurospheres (hDPC-NCSCs) resulted in an increased expression of neural crest-related genes, while the addition of BMP4 appeared to produce better NCSC characteristics and neural differentiation. The neural-like phenotype was evidenced by the expression of TUJ1, peripherin, NFH, TAU, SYN1, and GAP43. Our results describe the establishment of hDPC cultures from a large variety of patients in serum-free medium, as NCSC that differentiate into neural-like cells, as well as an important effect of BMP4 in enhancing the neural crest phenotype and differentiation of hDPCs.
从牙齿相关组织（包括人牙髓细胞 [hDPC]）中获得具有神经嵴相似性的细胞培养物（神经嵴衍生干细胞 [NCSC]）的潜力在文献中已有讨论。然而，大多数报告都包含使用富含血清的条件，并且没有描述神经分化的潜力，从而减缓了向临床的转化。因此，我们旨在体外培养和表征人牙髓中的 NCSC，并评估其分化为神经元的能力；我们还研究了添加 BMP4 以增强这种潜力的效果。从不同患者样本中建立培养物，并在血清、无血清以及球体聚集条件下单层培养，以诱导和鉴定 NCSC 表型。hDPC 培养物通过免疫细胞化学和逆转录定量聚合酶链反应进行表征。单层培养物表达干细胞、神经祖细胞和神经嵴相关标志物。培养 hDPC 作为神经球（hDPC-NCSC）会导致神经嵴相关基因的表达增加，而添加 BMP4 似乎会产生更好的 NCSC 特征和神经分化。神经样表型通过 TUJ1、 peripherin、NFH、TAU、SYN1 和 GAP43 的表达得到证实。我们的结果描述了在无血清培养基中从大量不同患者中建立 hDPC 培养物作为 NCSC 的过程，这些 NCSC 分化为神经样细胞，以及 BMP4 对增强 hDPC 的神经嵴表型和分化的重要作用。
