Fig 1. Effect of SPARC on the modulation of ECM proteins in proliferating C2C12 cells.

Representative western blots for collagen 1a1 and fibronectin expressions in C2C12 myoblasts after induction/inhibition of SPARC. C2C12 myoblasts were cultured in GM. One day before adding anti-SPARC (40 μg/ml) or rSPARC (8 μg/ml), cells were trypsinezed and plated at density of 4×10⁵/well in 12-well plates. Cells were cultured in four different conditions for 48 h. Proteins extraction was performed as described above and collagen 1a1 and fibronectin protein expression levels were measured by western blot. The experiments were repeated three times (three different passages of C2C12 cells) and a representative western blot image was shown. Data were expressed as a ratio to the positive control (pooled samples). One-way ANOVA with repeated measurements was used to adjust the validations of the different passages and a contrast analysis was performed. P value was set at < 0.05 after the Bonfferoni adjustments. (a) Thirty μg of whole cell lysate proteins were loaded to measure collagen 1a1 levels in C2C12 proliferating myoblasts. Addition of SPARC induced collagen 1a1 expression, however, SPARC inhibition decreased it. (b) Five μg of the proteins from proliferating cells were loaded to measure fibronectin levels. Fibronectin expression trended to decrease after rSPARC addition and anti-SPARC increased it. Abbreviations: PBS: phosphate buffered saline, rSPARC: recombinant SPARC protein, and anti-SPARC: anti-SPARC antibody. *Significant differences between experimental conditions.