Thick transverse cross-sections (8 µm) of tissues were obtained using a cryostat (2800 FrigocutN, Reichert Jung, Heidelberg, Germany) equilibrated at -30 and -20°C for adipose tissue and muscle, respectively. Sections were mounted on silane-coated slides for enhanced adhesion. Immunocyto-localization of PDGFR marker was carried out using a standard procedure. Briefly, after incubation in 2% bovine serum albumin (BSA; Sigma, Saint-Quentin Fallavier, France) to prevent non-specific binding, sections were incubated in PBS/0.1% BSA for 45 min at room temperature with the following primary mouse antibodies: anti-PDGFR and anti-laminin complex (Interchim, Montluçon, France). Nuclei were counterstained with 4'-6-diamidino-2-phenylindole (DAPI; Dako, Trappes, France). No significant staining was detected in slides incubated with control serum and/or goat IgG in the absence of the primary antibody. A Nikon DS-Ri1 epifluorescence microscope was used to acquire digital images using an Eclipse E400 digital camera and NIS-Elements software version 3.0 (Nikon).
Qualitative and quantitative analyses of stained cells were performed using a self-developed plugin for ImageJ (ImageJ 1.43, National Institutes of Health, Bethesda, MD, USA).
Qualitative and quantitative analyses of stained cells were performed using a self-developed plugin for ImageJ (ImageJ 1.43, National Institutes of Health, Bethesda, MD, USA).