Bierlein De la Rosa Metzere, Sharma Anup D, Mallapragada Surya K, Sakaguchi Donald S
Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA 50011, USA.
Department of Chemical and Biological Engineering, Iowa State University, Ames, IA 50011, USA; Neuroscience Program, Iowa State University, Ames, IA 50011, USA.
J Biosci Bioeng. 2017 Nov;124(5):572-582. doi: 10.1016/j.jbiosc.2017.05.014. Epub 2017 Jul 8.
The use of genetically modified mesenchymal stem cells (MSCs) is a rapidly growing area of research targeting delivery of therapeutic factors for neuro-repair. Cells can be programmed to hypersecrete various growth/trophic factors such as brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and nerve growth factor (NGF) to promote regenerative neurite outgrowth. In addition to genetic modifications, MSCs can be subjected to transdifferentiation protocols to generate neural cell types to physically and biologically support nerve regeneration. In this study, we have taken a novel approach by combining these two unique strategies and evaluated the impact of transdifferentiating genetically modified MSCs into a Schwann cell-like phenotype. After 8 days in transdifferentiation media, approximately 30-50% of transdifferentiated BDNF-secreting cells immunolabeled for Schwann cell markers such as S100β, S100, and p75. An enhancement was observed 20 days after inducing transdifferentiation with minimal decreases in expression levels. BDNF production was quantified by ELISA, and its biological activity tested via the PC12-TrkB cell assay. Importantly, the bioactivity of secreted BDNF was verified by the increased neurite outgrowth of PC12-TrkB cells. These findings demonstrate that not only is BDNF actively secreted by the transdifferentiated BDNF-MSCs, but also that it has the capacity to promote neurite sprouting and regeneration. Given the fact that BDNF production remained stable for over 20 days, we believe that these cells have the capacity to produce sustainable, effective, BDNF concentrations over prolonged time periods and should be tested within an in vivo system for future experiments.
基因改造间充质干细胞（MSCs）的应用是一个快速发展的研究领域，旨在为神经修复传递治疗因子。细胞可被编程以超分泌各种生长/营养因子，如脑源性神经营养因子（BDNF）、胶质细胞系源性神经营养因子（GDNF）和神经生长因子（NGF），以促进再生性神经突生长。除了基因改造外，MSCs还可进行转分化方案以生成神经细胞类型，从物理和生物学上支持神经再生。在本研究中，我们采用了一种新颖的方法，将这两种独特策略相结合，并评估了将基因改造的MSCs转分化为雪旺细胞样表型的影响。在转分化培养基中培养8天后，约30-50%的转分化分泌BDNF的细胞对雪旺细胞标志物如S100β、S100和p75进行免疫标记。诱导转分化20天后观察到增强，表达水平下降最小。通过ELISA对BDNF产量进行定量，并通过PC12-TrkB细胞测定法测试其生物学活性。重要的是，通过PC12-TrkB细胞神经突生长增加验证了分泌的BDNF的生物活性。这些发现表明，转分化的BDNF-MSCs不仅能主动分泌BDNF，而且具有促进神经突萌发和再生的能力。鉴于BDNF产量在20多天内保持稳定，我们认为这些细胞有能力在较长时间内产生可持续、有效的BDNF浓度，应在体内系统中进行测试以用于未来的实验。
