PMC

A. Venn diagram showing the functional pathways affected by the addition of calcitriol in cultures of Schwann cells or in co-cultures of DRG/Schwann cells. Five of the fifteen metabolic calcitriol-regulated pathways are affected in both cell types. B. Validation by qPCR of four selected up-regulated genes (Prx, Tspan2, IgF1, Spp1) involved in axogenesis and myelination.

A. All animals (n = 6 per group) improved their hindlimb locomotion during the 12 weeks of experiment. B. A similar pattern was observed when 12 animals were included in the Control, Vehicle and D3–500 groups. Crosses (+) indicate that the response was significantly increased, when compared to the Vehicle group (+ p<0.05;++p<0.01).

Ventilatory response of the animals after muscle stimulation (A) and response of metabosensitive afferent fibre activity after active muscle electrical stimulation (B) or intramuscular capsaicin injection (C). The experiment was first assessed in the 6 initial groups (n = 6 per group) (right histograms) and then in the 3 final groups (n = 12 per group) (left histograms). Crosses (+) indicate that the response was significantly increased, when compared to the Vehicle group (+ p<0.05;++p<0.01;+++p<0.001).

Muscle contractions were obtained using peroneal nerve electrical stimulation. A. Twitches were analysed in terms of peak Amplitude (A) and Maximum Relaxation Rate (MRR), defined as the slope of a tangent drawn to the steepest portion of the relaxation curve. B. Electrical stimulation frequencies were used to reach the tetanus threshold. The experiment was first assessed in the 6 initial groups (n = 6 per group; right histograms) and then in the 3 final groups (n = 12 per group; left histograms). Crosses (+) indicate significant changes when compared to the Vehicle group (+ p<0.05;++p<0.01).

A. List of functions for the genes involved in “nervous system development and function” whose expression was altered after addition of calcitriol to Schwann cells (A) or Schwann cells and dorsal root ganglion cells (B). Red arrows indicate an over-expression; green arrows, an under-expression. C. Twenty-five nervous system-related genes were used to generate a network representation. The genes shaded red are upregulated and those that are green are downregulated. The intensity of the shading shows to what degree each gene was up- or downregulated. The genes in white colour were not significantly changed in the analysis and can be considered as “missing links”. Orange solid lines represent a known direct interaction between calcitriol and the genes present in the network.

Peroneal nerves from the Control, Vehicle and D3–500 groups were either fixed with paraformaldehyde and included in paraffin (n = 6 per group) for counting axon numbers (A–D) or fixed with glutaraldehyde and included in resin (n = 6 per group) for assessing myelination (E–H). A,B. Representative pictures of nerve sections immunostained with an anti-neurofilament antibody in Vehicle (A) and D3–500 (B) groups. Quantitative analysis of axon numbers indicated that vitamin D3–500 induced a statistically significant doubling of axons in the proximal end (C) but not in the distal end (D). E. Low magnification view of a nerve section stained with p-phenyl-n-diamine (D3–500 group). F. High magnification view of a nerve section with arrows indicating how the G-ratio (the ratio between the diameter of the axon and the outer diameter of the myelinated fibre) was calculated (D3–500 group). Quantitative analysis indicates that vitamin D3–500 triggered myelination in the proximal (G) and the distal (H) ends of the nerve. Crosses (+) indicate significant changes when compared to the Vehicle group (+ p<0.05;++p<0.01). Scale bar:?