*2.4. Immunofluorescence*

Sections of the rat femur were fixed in 4% ( *w*/*v*) paraformaldehyde (PFA (P6148, Sigma, Lezennes, France) in PBS (phosphate-buffered saline without Ca and Mg, GAUPBS0001, Eurobio, Les Ulis, France). After three washes in PBS, the sections were permeabilized for 15 min with 0.5% (*v*/*v*) Triton X100 buffered with PBS. The non-specific binding sites were blocked with Emerald Antibody Diluent (936B-08, Sigma, Lezennes, France) for 1 h at room temperature. Then, sections were incubated overnight at 4 ◦C with the primary rabbit anti-CD68 (ab125212, Abcam, Amsterdam, The Netherlands) antibody at 1:1000 di lution; the primary goa<sup>t</sup> anti-CD206 (C20) (sc-34577, Santa Cruz Bio., Heidelberg, Germany) antibody at 1:1000 dilution; the primary rabbit anti-CD163 (ab182422, Abcam, Amsterdam, The Netherlands) antibody at 1:500 dilution; and the primary rabbit anti-Iba1 (ab178846, Abcam, Amsterdam, The Netherlands) antibody at 1:200 dilution. Then, the sections were washed in PBS and incubated with the secondary anti-rabbit Alexa Fluor 488 (A-21206, Thermo Scientific, Waltham, MA, USA) antibody at 1:500 dilution and the secondary anti-goat Alexa Fluor 568 (ab175704, Abcam, Amsterdam, The Netherlands) antibody at 1:500 dilution for 2 h at room temperature. Finally, sections were washed in PBS for 20 min and mounted using a Fluoroshield mounting medium with DAPI (ab104139, Abcam, Amsterdam, The Netherlands). The fluorescence was detected using an epifluorescence microscope DM6000 (Leica, Schönwaldeglien, Germany) equipped with monochrome and color digital cameras.
