Time-Dependent Changes in Muscle IGF1-IGFBP5-PAPP System after Sciatic Denervation

Abstract

Denervation-induced muscle atrophy is a frequent cause of skeletal muscle diseases. However, the role of the most important muscle growth factor, insulin-like growth factor (IGF-1), in this process is poorly understood. IGF-1 activity is controlled by six IGF-1 binding proteins (IGFBPs). In skeletal muscle, IGFBP-5 seems to have an important role in atrophic processes. Furthermore, pappalysins (PAPP-A) modulate muscle growth by increasing IGF-1 bioavailability through IGFBP cleavage. We aimed to study the time-dependent changes in the IGF1-IGFBP5-PAPP system and its regulators in gastrocnemius muscle after sciatic denervation. Gastrocnemius atrophy and overexpression of IGF-1 was observed from day 3 post-denervation. The proteolytic factors measured were elevated from day 1 post-denervation onwards. Expression of both IGFBP-5 and pappalysins were increased on days 1 and 3. Subsequently, on days 7 to 14 pappalysins returned to control levels while IGFBP-5 remained elevated. The ratio IGFBP-5/PAPP-A was correlated with the main proteolytic markers. All data suggest that the initial increase of pappalysins could facilitate the IGF-1 action on muscle growth, whereas their subsequent decrease could lead to further muscle wasting.

Keywords: IGF-1; IGFBP-5; pappalysins; skeletal muscle atrophy.

Figure 1

The effect of sciatic denervation on relative gastrocnemius weight (A), cross-sectional fiber area (B), and mRNA levels of myosin heavy chain isoforms (MHC) I and IIb (C,D), at different time points post-denervation (DNX). Values are represented as mean ± standard error of the mean (SEM) of left (SHAM operated) and right (denervated) paws in comparison with control animals (SHAM operated in both hind limbs). Statistics: *** p < 0.001, ** p < 0.01 and * p < 0.05 in the comparisons between right vs. left paw with Student’s t test; a ≠ b ≠ c ≠ d in the comparisons between different time-points groups of the right paw by least post hoc analysis after significant one-way ANOVA.

Figure 2

Effect of sciatic denervation on gastrocnemius mRNA levels of insulin-like growth factor 1 (IGF-1) (A), IGF-1 receptor (IGF-1R) (B), IGF-1 binding protein 5 (IGFBP-5) (C), eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) (D), pregnancy-associated plasma protein-A (PAPP-A) (E) and PAPP-A2 (F), stanniocalcin 1 (STC-1) (G) and STC-2 (H) at different days after denervation (DNX). Values are represented as mean ± standard error of the mean (SEM) of left (SHAM operated) and right (denervated) paws in comparison with control animals (SHAM operated in both hind limbs). Statistics: *** p < 0.001 and * p < 0.05 in the comparisons between right vs. left paw with Student’s t test; a ≠ b ≠ c in the comparisons between different time-points groups of the right paw with least post hoc analysis after significant one-way ANOVA.

Figure 3

Effect of sciatic denervation on gastrocnemius mRNA levels of histone deacetylase 4 (HDAC-4) (A), myogenin (B), atrogin 1 (C), muscle RING-finger protein-1 (MuRF-1) (D) at different time points post-denervation (DNX). Values are represented as mean ± standard error of the mean (SEM) of left (SHAM operated) and right (denervated) paws in comparison with control animals (SHAM operated in both hind limbs). Statistics: *** p < 0.001, ** p < 0.01 and * p < 0.05 in the comparisons between right vs. left paw with Student’s t test; a ≠ b ≠ c in the comparisons between different time-points groups of the right paw with least post hoc analysis after significant one-way ANOVA.

Figure 4

Effect of sciatic denervation on the IGFBP-5/PAPP-A expression levels ratio (A), total protein concentration (B) and the protein levels of HDAC-4 (C), myogenin (D), cleaved alpha II spectrin (E) and light chain 3b II (LC3b-II) (F) at different time points post-denervation (DNX). Values are represented as mean ± standard error of the mean (SEM) of right (denervated) paws in comparison with control animals (SHAM operated in both hind limbs). Representative Western blots are shown in (G). Statistics: a ≠ b ≠ c in the comparisons between different time-point groups of the right paw by least post hoc analysis after significant one-way ANOVA.

Figure 5

Linear correlation between IGFBP-5/PAPP-A expression levels ratio and myogenin mRNA (A) and protein levels (B), MuRF-1 expression levels (C) and LC3b-II protein levels (D). Statistics: Pearson’s correlations.