monocytic wild type thp 1 cell line  (ATCC)

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 is upregulated in both sepsis and THP-1-derived macrophages upon LPS + INF-γ stimulation. A PD-L1 expression in septic patients was analyzed using datasets GES134347 and GSE65682 retrieved from the GEO database. Statistical significance was determined by the Wilcoxon test. * p < 0.05. B THP-1-derived macrophages (THP-1-M) were co-stimulated with LPS (100 ng/ml) and IFN-γ (20 ng/ml) for 12–24 h, followed by quantification of PD-L1 mRNA levels via qRT-PCR ( n = 3 independent biological replicates). C PD-L1 protein expression in THP-1-M after LPS + IFN-γ stimulation for 6, 12, or 24 h was assessed by Western blotting. D Flow cytometry analysis of PD-L1 protein expression in THP-1-M treated with LPS + IFN-γ for 24 h. E Subcellular localization of PD-L1 in THP-1-M following 12 h LPS + IFN-γ stimulation was visualized by confocal laser scanning microscopy. The relative fluorescence intensity and Pearson’s R value were analyzed by ImageJ ( n = 3 independent biological replicates). The data are presented as the mean ± SEM. Statistical analysis for B was performed by one-way ANOVA, and that for E was determined by Student’s t-test. * p < 0.05

Article Snippet: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Western Blot, Flow Cytometry, Confocal Laser Scanning Microscopy, Fluorescence

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: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

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: Bioinformatics analysis integrating RNA sequencing data and GEO datasets demonstrated PD-L1-mediated significant activation of the NOD-like receptor signaling pathway. A PD-L1 NC /THP-1-M cells were co-stimulated with LPS (100 ng/ml) + IFN-γ (20 ng/ml) for 12 h followed by RNA sequencing, with subsequent KEGG pathway analysis of activated signaling pathways. B - C PD-L1-centered single-gene GSEA across datasets GSE57065 , GSE28750 and GSE95233 . B Ridge plot displaying top 20 activated pathways. C GSEA enrichment plot illustrating NOD-like receptor signaling pathway activation. D Comparative PD-L1-specific GSEA of NOD-like receptor signaling pathway activation in PD-L1 HI /THP-1-M vs. PD-L1 NC /THP-1-M cells following 12 h LPS + IFN-γ co-stimulation with RNA sequencing verification

Article Snippet: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: RNA Sequencing, Activation Assay, Protein-Protein interactions

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: Multi-omics integration identified AIM2 as the critical mediator for PD-L1-regulated macrophage M1 polarization. A - D PD-L1 NC /THP-1-M cells were co-stimulated with LPS (100 ng/ml) + IFN-γ (20 ng/ml) for 12 h for PD-L1-targeted IP-MS analysis. A Global proteomic landscape showing peptide/protein distribution and Venn intersections between PBS control vs. LPS + IFN-γ groups. B Triple-dataset intersection analysis integrating IP-MS, RNA-seq and GEO datasets ( GSE57065 , GSE28750 , GSE95233 ) for core candidate screening. C GO enrichment of 208 unique interacting proteins in LPS + IFN-γ group with inflammatory pathway emphasis. D Representative tandem MS/MS spectra identifying AIM2. E Co-IP validation of PD-L1-AIM2 interaction post 12 h LPS + IFN-γ co-stimulation in PD-L1 NC /THP-1-M cells ( F ) Differential AIM2 expression in PD-L1 HI /THP-1-M vs. PD-L1 NC /THP-1-M cells by MA plot (left) and TPM expression profile (right) following 12 h LPS + IFN-γ stimulation ( n = 3 independent biological replicates). G Temporal AIM2 regulation: qRT-PCR analysis at 6/12/24 h (left, n = 3 independent biological replicates) with Western blot confirmation at 24 h (right). H - J PD-L1-AIM2 structural docking studies. H AlphaFold-predicted full-length molecular models of PD-L1 and AIM2. I Top 5 scoring complexes from GRAMM docking server simulations. J High-resolution interface visualization of optimal complex with potential interaction sites. The data are presented as the mean ± SEM. Statistical analysis for F was performed by Student’s t-test, and that for G was determined by one-way ANOVA. * p < 0.05

Article Snippet: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Biomarker Discovery, Protein-Protein interactions, Control, RNA Sequencing, Tandem Mass Spectroscopy, Co-Immunoprecipitation Assay, Expressing, Quantitative RT-PCR, Western Blot

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 overexpression potentiates AIM2-mediated downstream effector (IL-18/IFN-γ) production and amplifies STAT1 activation cascade. A PD-L1 HI /THP-1-M vs. PD-L1 NC /THP-1-M cells were co-stimulated with LPS (100 ng/ml) + IFN-γ (20 ng/ml) for 12 h. RNA-seq-derived TPM profiles of IL-1β (left) and IL-18 (middle), with MA plot visualization of IL-18 differential expression (right) ( n = 3 independent biological replicates). B Dual-modality verification of IL-18 regulation: qRT-PCR analysis (left) with paired ELISA quantification (right) post 12 h co-stimulation ( n = 3 independent biological replicates). C Temporal dynamics of IFN-γ regulation: mRNA/protein measurement by qRT-PCR (left) and ELISA (right) at 24 h endpoint ( n = 3 independent biological replicates). D Western blot detection of STAT1 phosphorylation status and basal expression following 24 h LPS + IFN-γ co-stimulation. E - G PD-L1-specific single-gene GSEA analysis across GEO datasets ( GSE57065 , GSE28750 , and GSE95233 ). E Identification of 111 molecules positively correlated with PD-L1 expression. F MCODE algorithm-based core module identification in Cytoscape (40 hub molecules). G GO enrichment analysis of the 40-molecule module revealing JAK-STAT pathway activation. The data are presented as the mean ± SEM. Statistical analysis for A, B and C was performed by Student’s t-test. * p < 0.05

Article Snippet: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Over Expression, Activation Assay, RNA Sequencing, Derivative Assay, Quantitative Proteomics, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Phospho-proteomics, 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: 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: The human monocytic wild-type THP-1 cell line (PD-L1 WT ) was purchased from American Type Culture Collection (ATCC; Manassas, VA, USA).

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