Fig. 1: SCIMP can be secreted in exosomes by macrophages stimulated with bacteria.

a The chemotactic activity of the SCIMP-containing supernatant (n = 5) from SCIMP-overexpressing HEK293 cells (SCIMP-HEK293) toward peripheral neutrophils was measured using a chemotaxis assay, with Interleukin 8 (IL-8, 10 ng/mL) used as the positive control (n = 5). b The exogenous SCIMP-6×his recombinant protein in the HEK293 cells could be detected by the anti-6×his antibody both in the cell culture supernatant and cell lysate via western blot. c The exosomes present in the cell culture supernatant of SCIMP-HEK293 were purified using the ultracentrifugation method, and the presence of SCIMP-6×his recombinant protein was confirmed by western blot analysis, along with the exosomal markers CD9, CD63, CD81 and TSG101. d The exosomes from SCIMP-HEK293 or the control HEK293 were analyzed by NTF, and the diameter of the particles was indicated by the V-SSC channel, and PE-anti-CD63 antibody and FITC-anti-SCIMP antibody were used as detection antibodies. e The purified exosomes from SCIMP-HEK293 were observed under the TEM, stained with gold-conjugated anti-SCIMP antibody (scale bar = 200 nm). f The endogenous SCIMP protein secreted in exosomes by RAW264.7 cells with the stimulation of PBS, LPS (1 μg/mL), or heat-denatured E. coli (MOI = 100) for 0.5, 1, 2, and 6 h, was detected by western blot using anti-SCIMP antibody. g The percentage of SCIMP-positive particles in the total exosomes (CD63 positive), which were purified from the supernatant of RAW264.7 cells treated with PBS (n = 8) or E. coli (MOI = 100, n = 10) for 2 h, were stained and measured by NTF. h The endogenous SCIMP protein in the pulmonary tissue of the mice with E. coli-induced pneumonia was detected by IHC using anti-SCIMP antibody (scale bar = 200 μm/50 μm). i The colocalization of SCIMP (exogenously expressed SCIMP-GFP recombinant protein, green), EEA1 (stained with anti-EEA1 antibody, blue), and LAMP2A (stained with anti-LAMP2A antibody, red) was observed by confocal microscopy (IF, Immunofluorescence) in RAW264.7 cells stimulated with heat-denatured E. coli (MOI = 1000) for 0, 1, and 4 h (scale bar = 20 μm). The raw data is available in the “Source Data”.