Journal: Science Advances

Article Title: Pathogenic GM-CSF drives functional diversification of inflammatory macrophages in autoimmune arthritis

doi: 10.1126/sciadv.aec0986

Figure Lengend Snippet: UMAP visualization showing the expression patterns of ( A ) key proinflammatory cytokines and ( B ) indicated genes across synovial MNPs. ( C ) Flow cytometry analysis of Arg1 and C1q expression, along with surface markers EpCAM and CD11c, within CD11b + Ly6G − Ly6C lo CD26 − synovial MNPs from control (healthy) and arthritic SKG mice. Arthritic SKG mice were analyzed 4 weeks after mannan injection. ( D ) Total numbers of Arg1 + and EpCAM + macrophages in the synovium of control and arthritic SKG mice ( n = 6 each). ( E and F ) Intracellular staining of TNF-α, IL-6, and pro–IL-1β in CD11b + Ly6G − Ly6C lo CD26 − CD200 − synovial macrophages derived from CD45.1 + WT and CD45.2 + Csf2rb −/− cells in mixed BM chimeric mice, 4 weeks post–mannan injection. (F) Percentages and total numbers of TNF-α + , IL-6 + , and pro–IL-1β + macrophages, as shown in (E) ( n = 6). ( G ) UMAP displaying the expression levels of Ccl17 , Irf4 , and Pdcd1lg2 . ( H ) Gating strategy for sorting synovial cell subsets from arthritic SKG mice, including cDCs, Ly6C hi monocytes, EpCAM + macrophages, CD11c lo macrophages, and CD11c hi EpCAM − macrophages from arthritic SKG mice. ( I ) Relative mRNA expression of Ccl17 in the indicated sorted subsets ( n = 4). Expression levels were normalized to those of Hprt . ** P < 0.01. Statistical analyses were performed using Student’s t test (D), paired t test (F), and one-way ANOVA, followed by Tukey’s multiple-comparison test (I). Error bars denote the SD in panels. Data in (C), (E), (H), and (I) are the representatives of two independent experiments; data in (D) and (F) are pooled from two independent experiments.

Article Snippet: Gene expression levels of Hprt and Ccl17 were quantified using TaqMan probes ( Hprt : Mm01545399_m1 and Ccl17 : Mm01244826_g1) and THUNDERBIRD Probe qPCR Mix (TOYOBO) on a StepOnePlus Real-Time PCR System (Thermo Fisher Scientific).

Techniques: Expressing, Flow Cytometry, Control, Injection, Staining, Derivative Assay, Comparison

Journal: Experimental & Molecular Medicine

Article Title: H3K18 lactylation in cancer-associated fibroblasts drives malignant pleural effusion progression via TNFR2 + T reg recruitment

doi: 10.1038/s12276-025-01557-3

Figure Lengend Snippet: a KEGG pathway enrichment of DEGs between FACS-sorted TNFR2 + and TNFR2 − CD4 + T cells from MPE ( n = 3). b , c Volcano plot ( b ) and heatmap ( c ) of DEGs related to the chemokine signaling pathway. d , e Flow cytometry histograms ( d ) and comparisons of chemokine receptor expression (CXCR6, CCR4 and CCR6) ( e ) on TNFR2 + T reg cells and TNFR2 − T reg cells ( n = 13). f Schematic of Transwell chemotaxis assay testing TNFR2 − T reg chemotaxis toward MPE supernatant (by Figdraw). g , h Transwell chemotaxis assay comparing TNFR2 + T reg frequencies between freshly isolated cells and cells that migrated toward MPE supernatant after 4 h incubation. i Concentrations of CXCL16, CCL17, CCL22 and CCL20 in MPE and PB were quantified using ELISA ( n = 16). j Schematic of chemotaxis assay to investigate the chemotatic axis to attract TNFR2 + T reg cells in MPE. k , l Flow cytometry histograms and comparisons of chemotaxis of TNFR2 + T reg cells in response to MPE in the presence of anti-CXCL16, anti-CCL17, anti-CCL22 or anti-CCL20 mAbs. Data shown in d , e , g – i , k and l are representative of at least three independent experiments (mean ± s.d.). Statistical analysis was performed using paired two-tailed Student’s t -test ( e and h ), Wilcoxon test ( i ) or one-way ANOVA ( l ). * P < 0.05, ** P < 0.01, **** P < 0.0001. ns not significant, mAbs monoclonal antibodies.

Article Snippet: In specific experimental conditions, the following reagents were added to the lower chamber: anti-CCL17 antibody (50 ng/ml, #MAB364-SP, R&D Systems), anti-CCL20 antibody (50 ng/ml, #MAB360-SP, R&D Systems), anti-CCL22 antibody (50 ng/ml, #MAB336-SP, R&D Systems), anti-CXCL16 antibody (50 ng/ml, #MAB976, R&D Systems) or recombinant human CXCL16 (100 ng/ml, #976-CX-025/CF, R&D Systems).

Techniques: Flow Cytometry, Expressing, Chemotaxis Assay, Isolation, Incubation, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Bioprocessing