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Enhanced axonal outgrowth and nerve cell network formation of mouse dorsal root ganglion neurons in vitro supplemented with vitamins B1, B6 and B12
Abstract
Biochemical functions of the neurotropic B vitamins such as thiamine (B1), pyridoxine (B6), and cobalamin (B12) in the nervous system are well described in bibliographic literature. However, it is still not well understood how depletion of neurotropic B vitamins affects neuronal cell health. In order to further study the important role of vitamins B1, B6 and B12 in maintaining nerve cell viability, the following experiment was carried out. In this current study we investigated the effects of vitamins B1, B6 and B12 compared to neurotropic B vitamin depletion on axonal outgrowth and nerve cell network formation of mouse dorsal root ganglion (mDRG) neurons in vitro over a 6-day period and quantified axonal length as well as cell body area automatically. Axonal outgrowth (neurite length [Phase] (mm)/cell body cluster) was imaged every 6 hours in live cells in real time during 6 days using the IncuCyte® S3 Live-Cell Analysis System with 20x magnification (9 fields/well). Resulting values were statistically analysed using One-way ANOVA (analysis of variance). In addition, we performed immunocytochemistry using beta-III tubulin staining in order to assess the health of the underlying beta-tubulin network necessary for supporting axonal outgrowth. Our results demonstrated that the absence of neurotropic vitamins B1, B6, and B12 leads to progressive neurodegeneration of primary cultures of mDRG neurons. While the time of onset of the degeneration response varied slightly between cultures, it generally appeared to begin 3–4 days after B vitamin removal. Therefore, we conclude that neurotropic vitamins B1, B6, and B12 support cell viability, neurite growth as well as nerve cell network formation of healthy mouse DRG neurons and are, thus, essential for nerve health and the maintenance of a healthy nerve function.
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References
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