The SPARC internalization pathway in Skm-PCs from young and old rats. (A) Skm-PCs derived from young and old rats were incubated with Alexa-SPARC (green) and immunostained with anti-MyoD (red). Representative photographs of MyoD-positive and MyoD-negative cells in Skm-PCs from young and old rats are shown in the left panel. Black arrowheads indicate MyoD-positive and white arrowheads indicate MyoD-negative cells, respectively. Scale bar = 50 μm. SPARC fluorescence per cell was quantified by ImageJ and graphed (right). Error bars represent means ± SEM (n = 10 cells). **, P < 0.01. The expression of Cltc (B) and Rab7 (C) was compared in Skm-PCs from young and old rats at 2 days after isolation. Graphed data are expressed as means ± SEM (n = 3). *, P < 0.05. Co-immunostaining of MyoD and Cltc (D) or Rab7 (E) and quantification of Skm-PCs from young and old rats at 2 days after isolation. Representative photographs of MyoD and Cltc or Rab7 in Skm-PCs from young and old rats are shown above. Black arrowheads indicate MyoD-positive and white arrowheads indicate MyoD-negative cells, respectively. Scale bar = 50 μm. Fluorescence intensity was quantified by ImageJ and graphed (below). Error bars represent means ± SEM (n = 10 cells). *, P < 0.05, **, P < 0.01. Skm-PCs, skeletal muscle progenitor cells; SPARC, secreted protein acidic and rich in cysteine.