Fig 1: IL-27 was highly expressed in mice with CLP-induced acute lung injury.A The level of serum IL-27 of mice in the sham group and CLP group was detected using ELISA kit. B The mRNA expressions of EBI3 and P28 in mouse lung tissue were detected using RT-qPCR. C The protein levels of EBI3 and P28 in mouse lung tissue were detected using Western blot. D The mRNA expressions of IL-6, TNF-a, and IL-1ß in mouse lung tissue were detected using RT-qPCR. E The protein contents of IL-6, TNF-a and IL-1ß in lung tissues of mice in each group were detected using ELISA. F The lung injury was detected using HE staining, scale bar = 50 µm. G The wet/dry ratio was used to calculate the formation of pulmonary edema, N = 8. H: 24 h after CLP, Evans blue dye was injected to measure pulmonary vascular leakage, N = 8. Measurement data are depicted as mean ± SD. The t test was used for the comparisons between two groups. One-way ANOVA was employed for the comparisons among multiple groups, followed by Tukey’s multiple comparisons test. *p < 0.05 vs. Sham group; #p < 0.05 vs. CLP group.
Fig 2: Mechanism diagram.ADMSC-exosomes inhibited IL-27 secretion in macrophages, thereby delaying sepsis-induced lung injury in mice.
Fig 3: ADMSC-exosomes reduced the number of pulmonary macrophages and the release of IL-27 in sepsis mice.A ADMSC-exosomes were isolated, purified and labeled with Dil dye; flow chart of injection in CLP sepsis mice. B The co-localization of exosomes (Dil pre-labeled red) and macrophages (F4/80, green) in lung tissue was detected using immunofluorescence staining, scale bar = 25 µm. C The number of macrophages CD68+ F4/80 + in single-cell suspension of whole lung tissue was detected using flow cytometry. D The content of IL-27 in serum of CLP mice treated with ADMSC-exosomes was detected using ELISA. E The mRNA expressions of P28 and EB13 in lung tissue of CLP mice treated with ADMSC-exosomes were detected using RT-qPCR. F The protein levels of P28 and EB13 in lung tissue of CLP mice treated with ADMSC-exosomes were detected using Western blot. G The content of IL-27 in macrophages after the treatment of ADMSC-exosomes or not detected using immunofluorescence. Blue fluorescence indicated DAPI, green fluorescence indicated F4/80, and red fluorescence indicated IL-27. N = 8. Measurement data are depicted as mean ± SD. One-way ANOVA was employed for the comparisons among multiple groups, followed by Tukey’s multiple comparisons test. *p < 0.05 vs. Sham group; #p < 0.05 vs. CLP group.
Fig 4: ADMSC-exosomes inhibited the LPS-mediated release of IL-27 in macrophages.A F4/80+ macrophages in lung tissue were detected using immunofluorescence (green staining) (nuclei were labeled by DAPI and presented blue staining). B The level of IL-27 in the supernatant of BMDMs treated with different concentrations of LPS for 12 h or LPS (100 ng/mL) for different times was detected using ELISA. C PKH67 (green)-labeled ADMSC-exosomes were co-cultured with BMDMs for 24 h, and then the fluorescence intensity of PKH67 in BMDMs was observed (DAPI-labeled nuclei were blue). D The time process of IL-27 release from BMDMs was detected using ELISA after the incubation of LPS (100 ng/mL) and ADMSC-exosomes (10 µg/µL). E The dose response of ADMSC-exosomes at different concentrations to IL-27 released by LPS-activated BMDMs within 12 h was detected using ELISA; Control indicated that the BMDMs were at rest. F BMDMs after LPS incubation were detected, with the addition of ADMSC-exosomes for 12 h or not. G The red staining of IL-27 in BMDMs was detected after 12 h-incubation, and Isotype represented the negative control. The cell experiment was repeated 3 times independently. Measurement data are depicted as mean ± SD. One-way ANOVA was employed for the comparisons among multiple groups, followed by Tukey’s multiple comparisons test. *p < 0.05.
Supplier Page from Abcam for Anti-IL-27-A antibody