To evaluate the neurotrophic growth functions of SKP-SCs and primary SCs, PC12 cells were separately cocultured with either SKP-SCs or primary SCs. The percentage of neurite outgrowth and the number of neurites originating from the soma were then observed [26 (link)]. The lower chambers of an 8-μm membrane-based culture insert (PIEP12R48, Millipore) contained PC12 cells that were pre-treated with 50 ng/μL β-nerve growth factor (Peprotech, 450–01-20) for 24 h to stop their proliferation and induce neurite outgrowth [26 (link)]. The upper chambers were filled with SKP-SCs or primary SCs (1 × 106 cells in 300 μL per well) in a serum-free medium. After co-culturing for 48 h, neuritogenesis was measured as previously described [27 (link)]. Briefly, neurite length was measured by the ratio of neurite length to the size of soma, and the measurements were defined as follows. ‘L0’ for cells with no neurites, ‘L1’ for cells whose neurite length was shorter than the size of the soma, ‘L2’ for cells with neurite length between the original size of the soma and twice the size of the soma, and ‘L3’ for cells whose neurite length was longer than twice the size of the soma.
β nerve growth factor
Manufactured by Thermo Fisher Scientific
β-nerve growth factor is a recombinant protein that is a member of the nerve growth factor family. It supports the survival and differentiation of specific target neurons.
Automatically generated - may contain errors
2 protocols using β nerve growth factor
1
Evaluating Neurotrophic Effects of Stem Cells
2
Differentiation of iPSCs to Sensory Neurons
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Differentiation of the iPSCs into sensory neurons was adapted from a previous study.10 (link) Briefly, Essential 8TM Flex medium was replaced by knockout serum replacement medium (KSR) consisting of KnockOutTM DMEM (10829018, Thermo Fisher), KnockOutTM Serum Replacement (10828028, Thermo Fisher), GlutaMAXTM, non-essential amino acids (11140050, Thermo Fisher) and β-mercaptoethanol. RevitaCellTM was used to enhance cell survival. SB431542 and LDN-193189 were added to the KSR medium to promote anterior neuro-ectoderm specification. On Day 2, CHIR99021, DAPT and potent and selective vascular endothelial growth factor receptor and fibroblast growth factor receptor inhibitor SU5402 (SU, 3300, Tocris) were supplemented to the KSR medium. The medium was then progressively transitioned into neuronal medium consisting of neurobasal medium, N2 supplement, B-27TM supplement (without vitamin A), GlutaMAXTM and β-mercaptoethanol. Additional compounds used to promote neural crest cells and sensory neuron differentiation and maturation were: ascorbic acid, β-nerve growth factor (450-34, Peprotech), neurotrophin-3 (450-03, Peprotech), BDNF and GDNF. Sensory neurons were replated on Matrigel®-coated plates at a density of 2.5 × 105 cells/ml. Additionally, as in our motor neuron protocol, a single treatment with 1 µM Ara-C was performed on Day 14.
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