2.3. Polarization of M1 and M2 macrophages in vitro and identification

For inflammatory macrophage (M1) differentiation, MФ were stimulated with 100 ng/mL LPS and 100 ng/mL recombinant IFN‐γ (R&D Systems, Minneapolis, MN, USA) for 24 hours. To generate anti‐inflammatory macrophages (M2), MФ were stimulated with 100 ng/mL recombinant IL‐4 (R&D Systems), 20ng/mL recombinant IL‐10 (R&D Systems) and 100 ng/mL recombinant IL‐13 (R&D Systems) for 24 hours. Polarized cells were identified by flow cytometry with myeloid and lymphoid immunophenotyping panels. Primary antibodies used in the flow cytometry analysis are as follows: CD11b (BD Biosciences, Franklin Lakes, NJ, USA), CD206 (BD Biosciences) and F4/80 (Thermo Fisher Scientific, Waltham, MA, USA). Data were collected using a BD LSRFortessa analyser and analysed using FlowJo 10.0 software.