Percp cy5.5 cd80

Raw 264.7 macrophages were treated with IL-33 (1–100 ng) or deferoxamine mesylate (0.1–1.5 mM) for 24 h before being trypsinized and counted. Cells were then washed with 500 µL cell sorting buffer (CSB; 1× PBS, 2% newborn calf serum, 0.1% NaN₃, 5 mM EDTA, pH 8.0) and blocked for 15 min with an Fc block derived from a 2.4G2 hybridoma cell (ATCC, HB-197) supernatant. PerCP/Cy5.5 CD80 (BioLegend, cat# 104722, San Diego, CA, USA), PE-CD86 (BioLegend, cat# 159204), APC-CD40 (BioLegend, cat# 124612), PE-Cy7-IA/IE (BioLegend, cat# 107630), FITC IL-33R (MD Bioproducts, cat# 101001F, St. Paul, MN, USA), and aqua fluorescent live/dead stain (Invitrogen, cat# L34966) were added directly to cells following Fc block incubation for 30 min at 4 °C protected from light. Then, the cells were washed with CSB and fixed for 20 min at room temperature in 2% paraformaldehyde protected from light. The cells were washed again with CSB and resuspended in 500 µL fresh CSB for analysis via flow cytometry on a BD LSR Fortessa.

Pg strain ATCC33277 was obtained from BeNa Culture Collection Center (Beijing, China). Anti‐CD63 & anti‐TSG101 were bought from Umibio (Shanghai, China). Exosome‐depleted FBS was provided by HAKATA (Shanghai, China). Exo‐spin mini standard and exo‐spin buffer were bought from TheWell Bioscience (North Brunswick, USA). LPS, hemin, and transferrin were purchased from Sigma‐Aldrich (St. Louis, USA). Mouse IL‐4 was purchased from PeproTech (RockyHill, USA). Firefly luciferase reporter gene lentiviral vector was bought from Genechem (Shanghai, China). DiR and FITC were bought from Beyotime Biotechnology (Shanghai, China). DCFH‐DA and Rosup were bought from Solarbio (Beijing, China). HO‐1 Assay Kit, Ferrous Iron Colorimetric Assay Kit, and Cell Total Iron Colorimetric Assay Kit were provided by Elabscience (Wuhan, China). n situ Apoptosis Detection Kit was bought from TaKaRa (Beijing, China). APC‐CD11b, FITC‐CD206, Percp/Cy5.5‐CD80, PE‐CD86, FITC‐CD3, PE‐CD4, Percp/Cy5.5‐CD8, FITC‐PD1 was bought from BioLegend (San Diego, USA). Mouse IL‐4 and Mouse IL‐1 ELISA Kits were bought from Boster (Pleasanton, USA).