Journal: Cell Communication and Signaling : CCS

Article Title: PD-L1 mediates M1 polarization of macrophages via the AIM2/IL-18/STAT1 signaling axis during sepsis

doi: 10.1186/s12964-025-02578-1

Figure Lengend Snippet: PD-L1 expression positively correlates with proinflammatory markers in M1-polarized macrophages. (A-C) PD-L1-overexpressing (PD-L1 HI ) and control (PD-L1 NC ) THP-1-M cells were stimulated with LPS (100 ng/ml) + IFN-γ (20 ng/ml) for 12 h followed by RNA sequencing. A Heatmap of inflammatory gene expression in PD-L1 HI vs. PD-L1 NC cells. B MA plot showing differential gene expression between PD-L1 HI and PD-L1 NC cells. C TPM expression values of IL-6, IL-27 and NOS2. D - E PD-L1 HI and PD-L1 NC cells were stimulated with LPS + IFN-γ for 24 h. ( n = 3 independent biological replicates) ( D ) qRT-PCR analysis of IL-6, IL-27 and NOS2 mRNA levels ( n = 3 independent biological replicates). E ELISA for IL-6/IL-27 and Griess reagent assay for NO in supernatants ( n = 3 independent biological replicates). (F-G) PD-L1-knockout THP-1-M (PD-L1 KO /THP-1-M) and PD-L1 NC /THP-1-M were stimulated with LPS + IFN-γ for 24 h. F qRT-PCR analysis of IL-6, IL-27 and NOS2 mRNA levels ( n = 3 independent biological replicates). G ELISA and Griess reagent detection of IL-6/IL-27 proteins and NO in supernatants ( n = 3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for C, D, E, F and G was performed by Student’s t-test. * p < 0.05

Article Snippet: IL-27, IL-6, IL-18, and IFN-γ concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm.

Techniques: Expressing, Control, RNA Sequencing, Gene Expression, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Knock-Out

Journal: Cell Communication and Signaling : CCS

Article Title: PD-L1 mediates M1 polarization of macrophages via the AIM2/IL-18/STAT1 signaling axis during sepsis

doi: 10.1186/s12964-025-02578-1

Figure Lengend Snippet: Triple-verification strategy (gene silencing/antibody neutralization/site-directed mutagenesis) confirms regulatory hierarchy of PD-L1/AIM2/IL-18/STAT1 signaling axis in macrophage M1 polarization. A PD-L1 HI /THP-1-M and PD-L1 NC /THP-1-M cells were pretreated with AIM2-targeting siRNA or scramble siRNA (scr siRNA) prior to 24 h LPS (100 ng/ml) + IFN-γ (20 ng/ml) co-stimulation, with subsequent Western blotting analysis of STAT1 phosphorylation. B - C Anti-IL-18 neutralizing antibody (1 µg/ml) and isotype control (1 µg/ml) were pretreated for 24 h followed by LPS + IFN-γ 24 h co-stimulation. B qRT-PCR quantification of IFN-γ mRNA ( n = 3 independent biological replicates). C STAT1 phosphorylation profiling by Western blotting. D Truncation mutant validation: LPS + IFN-γ-stimulated (24 h) PD-L1 Δ35–90 cells analyzed for STAT1 activation vs. full length type. E Following AIM2 silencing in PD-L1 HI /THP-1-M and PD-L1 NC /THP-1-M cells, qRT-PCR quantification of IL-6, IL-27 and NOS2 mRNA after 24 h LPS + IFN-γ stimulation ( n = 3 independent biological replicates). F Post IL-18 neutralization, mRNA levels of IL-6, IL-27 and NOS2 were assessed by qRT-PCR under LPS + IFN-γ 24 h stimulation ( n = 3 independent biological replicates). G STAT1-silenced cells were analyzed for IL-6/IL-27/NOS2 transcriptional changes post 24 h LPS + IFN-γ exposure ( n = 3 independent biological replicates). H Truncated PD-L1 Δ35–90 mutants were evaluated for IL-6, IL-27 and NOS2 mRNA expression following LPS + IFN-γ 24 h challenge ( n = 3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for B, E, F, G and H was performed by Student’s t-test. * p < 0.05

Article Snippet: IL-27, IL-6, IL-18, and IFN-γ concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm.

Techniques: Neutralization, Mutagenesis, Western Blot, Phospho-proteomics, Control, Quantitative RT-PCR, Biomarker Discovery, Activation Assay, Expressing

Journal: Cell Communication and Signaling : CCS

Article Title: PD-L1 mediates M1 polarization of macrophages via the AIM2/IL-18/STAT1 signaling axis during sepsis

doi: 10.1186/s12964-025-02578-1

Figure Lengend Snippet: PD-L1 knockout reduces septic mortality dampens systemic inflammation and alleviates vital organ (cardiac/hepatic/renal) injury in murine models. A PD-L1 wild-type (WT) and knockout (KO) mice were subjected to CLP followed by survival monitoring, inflammatory marker assessment, and multi-organ functional evaluation. B Survival rates of PD-L1 WT and PD-L1 KO mice at indicated time points post-CLP ( n = 15 mice per group). Results were compared by log-rank test. C mRNA levels of IL-6, IL-27, and NOS2 in monocytes analyzed by qRT-PCR at 24 h post-CLP ( n = 3 mice per group). D Serum concentrations of IL-6 and IL-27 proteins measured by ELISA, and NO levels determined by Griess assay at 24 h post-CLP ( n = 3 mice per group). E Serum biomarkers of organ function including ALT and AST for hepatic injury, BUN for renal dysfunction, and cTnI for cardiac damage at 24 h post-CLP ( n = 3 mice per group). The data are presented as the mean ± SEM. Statistical analysis for C, D and E was performed by Student’s t-test. * p < 0.05

Article Snippet: IL-27, IL-6, IL-18, and IFN-γ concentrations in cell supernatants or plasma were quantified using ELISA kits (Boster Biological Technology; Wuhan, China) with absorbance measurements at 450 nm.

Techniques: Knock-Out, Marker, Functional Assay, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Griess Assay