anti mouse cxcl9  (Bio X Cell)

Journal: Frontiers in Immunology

Article Title: Macrophage TRIM21 lactylation exacerbates infection-induced orchitis through enhancing STAT1-mediated CXCL9 and CXCL10 production

doi: 10.3389/fimmu.2025.1684836

Figure Lengend Snippet: CXCL9 and CXCL10 contribute to inflammatory cell infiltration and impair spermatogenesis during infection-induced orchitis. (A) Venn diagram showing that chemokines CXCL9, CXCL10, and CXCL11 exhibit chemotactic effect on monocytes/macrophages, T cells, and dendritic cells, while CCL17 and CCL22 only target T cells. (B) Relative mRNA levels of Cxcl9 , Cxcl10 , Cxcl11 , Ccl17 , and Ccl22 in the testes from LPS-induced orchitic and PBS control mice ( n = 3). (C) Relative mRNA levels of Cxcl9 , Cxcl10 , Cxcl11 , Ccl17 , and Ccl22 in the testes from UPEC-induced orchitic and vehicle control mice ( n = 3). (D) Gross morphology of the testes from LPS-induced orchitic mice, orchitic mice with isotype IgG injection, and orchitic mice with single or dual neutralization of CXCL9 and CXCL10. Scale bar, 1 cm. (E) Weight and relative weight of the testes from the sham group of mice and aforementioned mice in (B) ( n = 3). (F) Comparison of representative seminiferous tubules’ diameter and seminiferous epithelium thickness between the sham group and the above groups ( n = 3). (G) Quantified flow cytometry showing the number of macrophages (CD45 + CD11b + F4/80 + ) per 10 6 testicular cells in the sham group and the above groups ( n = 3–4). (H) Representative H&E staining images of seminiferous tubules, testicular interstitium, caput epididymis, and cauda epididymis from the indicated mice mentioned above. In the testicular interstitium, different arrows indicate different immune cells as shown. We rely on the characteristic nuclear morphology and localization of immune cells as a supportive, histological complement to the flow cytometric data in (G) . Scale bars, 100 μm at 20× magnification and 20 μm at 100× magnification. Two groups of data are statistically compared with Student’s t- test, while multiple groups of data are statistically compared with one-way ANOVA. Error bars represent mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001; ns, not significant.

Article Snippet: The recombinant neutralizing antibodies anti-CXCL9 (AF-492-NA, R&D Systems, Minneapolis, MN, USA) and anti-CXCL10 (AF-466-NA, R&D Systems, Minneapolis, MN, USA) were diluted in sterile PBS and stored at −20°C.

Techniques: Infection, Control, Injection, Neutralization, Comparison, Flow Cytometry, Staining

Journal: Frontiers in Immunology

Article Title: Macrophage TRIM21 lactylation exacerbates infection-induced orchitis through enhancing STAT1-mediated CXCL9 and CXCL10 production

doi: 10.3389/fimmu.2025.1684836

Figure Lengend Snippet: Macrophages are a predominant cellular source of CXCL9 and CXCL10 in the testis during LPS-induced orchitis. (A) Flow cytometry showing immune cells (gated on CD45 + ) expressing chemokines CXCL9 or CXCL10 in the testes from LPS-induced orchitic mice, with the proportion of macrophages identified. (B) Flow cytometry showing the proportion of testicular macrophages 3 days after in situ injection of clodronate liposomes in the testes. PBS liposomes were injected as control, while the sham group served as the negative control. (C) Quantified flow cytometry showing the number of macrophages (CD45 + CD11b + F4/80 + ) per 10 6 testicular cells in LPS-induced orchitic mice and sham/orchitic mice with in situ injection of clodronate liposomes/PBS liposomes. The sham group of mice served as the negative control. Sample sizes: sham = 7; LPS + PBS = 6; clodronate liposome + PBS = 4; PBS liposome + PBS = 4; clodronate liposome + LPS = 6; PBS liposome + LPS = 6. (D) Representative immunofluorescence images of the testes showing the co-localization of F4/80 + macrophages and chemokines CXCL9 or CXCL10 in mice with PBS/LPS/clodronate liposomes/PBS liposomes + LPS/clodronate liposomes + LPS intratesticular injection. Scale bar, 100 μm at 20× magnification. (E) Representative H&E staining images of seminiferous tubules, caput epididymis, and cauda epididymis from the indicated mice mentioned in (D) . Scale bar, 100 μm at 20× magnification. Data are statistically compared with one-way ANOVA. Error bars represent mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: The recombinant neutralizing antibodies anti-CXCL9 (AF-492-NA, R&D Systems, Minneapolis, MN, USA) and anti-CXCL10 (AF-466-NA, R&D Systems, Minneapolis, MN, USA) were diluted in sterile PBS and stored at −20°C.

Techniques: Flow Cytometry, Expressing, In Situ, Injection, Liposomes, Control, Negative Control, Immunofluorescence, Staining

Journal: Frontiers in Immunology

Article Title: Macrophage TRIM21 lactylation exacerbates infection-induced orchitis through enhancing STAT1-mediated CXCL9 and CXCL10 production

doi: 10.3389/fimmu.2025.1684836

Figure Lengend Snippet: Lactylation promotes the transcriptional expression of CXCL9 and CXCL10 in macrophages. (A) The lactate levels in the testes from LPS-induced orchitic mice, PBS control, and sham group ( n = 6). (B) Representative immunofluorescence images showing the general localization of lysine lactylation modification (pan-Kla) and F4/80 + macrophages in the testes from mice mentioned in (A) . Scale bar, 200 μm at 10× magnification. (C) More precise co-localization of pan-Kla-positive signals and F4/80 + macrophages in the testicular interstitium under an oil immersion microscope. White dotted lines indicate the edge of the seminiferous tubules, and three yellow boxes indicate typical F4/80 + macrophages in the testicular interstitium. Scale bar, 20 μm at 100× magnification. (D) Co-localization analysis of pan-Kla and F4/80 fluorescent signals in the testes from LPS-induced orchitic mice, PBS control, or sham group. (E) Intracellular lactate levels of RAW264.7 cells 24 h after LPS + IFN-γ stimulation with/without oxamate (Oxa) or lactate (Lac) treatment, or after incubating with Oxa or Lac alone. RAW264.7 cells without LPS + IFN-γ stimulation served as control (Ctl). Pound signs (#) indicate comparison with the LPS + IFN-γ stimulation group ( n = 5, normalized with intracellular protein levels). (F) Global lysine lactylation levels in RAW264.7 cells from the groups mentioned in (E) . (G) Relative mRNA levels of Cxcl9 and Cxcl10 in RAW264.7 cells from the groups mentioned in (E) ( n = 3). Pound signs (#) indicate comparison with the LPS + IFN-γ stimulation group. Data are statistically compared with one-way ANOVA. Error bars represent mean ± SEM. * p < 0.05, ## p < 0.01, ### p < 0.001; ns, not significant.

Article Snippet: The recombinant neutralizing antibodies anti-CXCL9 (AF-492-NA, R&D Systems, Minneapolis, MN, USA) and anti-CXCL10 (AF-466-NA, R&D Systems, Minneapolis, MN, USA) were diluted in sterile PBS and stored at −20°C.

Techniques: Expressing, Control, Immunofluorescence, Modification, Microscopy, Comparison

Journal: Nature Communications

Article Title: Targeting fibroblast derived thrombospondin 2 disrupts an immune-exclusionary environment at the tumor front in colorectal cancer

doi: 10.1038/s41467-025-66485-2

Figure Lengend Snippet: a Spatial maps of the indicated cell types in Xenium 5 K data at tumor (T)–non-tumor (NT) interfaces in CMS1/MSI and CMS4/MSS CRCs. F: tumor front. White dashed lines denote tumor borders. b Violin plots showing the nearest distances of macrophages or cDCs to CD8 + T cells. c Dot plots of indicated gene expression across cell types. d Violin plots showing the nearest distances of CXCL9/10 + macrophages (Mac) or cDCs to CXCR3 + CD8 + T cells. e Proportions of indicated cells in CMS1/MSI and CMS4/MSS CRCs. f Spatial expression patterns of CXCL9 (orange), CXCL10 (red), and CXCR3 (green). Arrows indicate the corresponding cell types. g, h CellChat analysis of cell-cell interactions in Xenium data. The heatmap ( g ) shows the differential number of pairwise interactions between CMS1/MSI and CMS4/MSS. Top colored bar indicates the total incoming signaling and the right colored bar indicates the total outgoing signaling for each cell cluster. Red and blue bars denote an increase or decrease, respectively, in CMS4/MSS compared to CMS1/MSI. The chord diagram illustrates ligand-receptor pairs upregulated in CMS1/MSI ( h , left) or CMS4/MSS ( h , right), with edge weights reflecting their contribution to signaling among indicated cell types. Scale bars, 100 μm ( a ), 50 μm ( f ). Šídák’s multiple comparison test, two-sided ( b ); Mann-Whitney U-test, two-sided ( d ). Mean ± SEM. Adjustments for multiple comparisons were not made in ( b ). Source data are provided as a file.

Article Snippet: For CXCL9/10 inhibition, mice were intraperitoneally injected with anti-CXCL9 antibody (300 μg; Bio X Cell, BE 0309) and anti-CXCL10 antibody (300 μg; Bio X Cell, BE 0440) twice per week.

Techniques: Gene Expression, Expressing, Comparison, MANN-WHITNEY

Journal: Nature Communications

Article Title: Targeting fibroblast derived thrombospondin 2 disrupts an immune-exclusionary environment at the tumor front in colorectal cancer

doi: 10.1038/s41467-025-66485-2

Figure Lengend Snippet: a–e Combined blockage of CXCL9 (αCXCL9 ab) and CXCL10 (αCXCL10 ab) in MTO-bearing WT or Thbs2 -/- mice (n: WT control = 8, WT αCXCL9 ab/CXCL10 ab = 9, Thbs2 -/- control = 6, Thbs2 -/- αCXCL9 ab/CXCL10 ab = 9). Schematic representation ( a ), macroscopic images ( b ), and volumes and weights ( c ) of tumors. Immunofluorescence for CD8 ( d ) and its quantification ( e ). Bottom images show magnified views of the yellow dashed boxes in the top panels ( d ). f–o Anti-CXCR3 antibody (αCXCR3 ab) treatment in MTO-bearing WT or Thbs2 -/- mice (n: WT control = 10, WT αCXCR3 ab = 10, Thbs2 -/- control = 7, Thbs2 -/- αCXCR3 ab = 8). Schematic representation ( f ) macroscopic images ( g ) and volumes and weights ( h ) of tumors. i Representative IMC images of orthotopic tumors. T: tumor, NT: non-tumor. j, k CD8 + T cell infiltration analysis from ( f ). Histograms ( j ) and scatter plots ( k ) show distances of CD8 + T cells from tumor borders. l–o , Proximity analyses between CD8 + T cells and F4/80 + macrophages or CD11c + DCs in ( f ). Histograms ( l ) and scatter plots ( m ) show the nearest distances between CD8 + T cells and CD11c + DCs. Histograms ( n ) and scatter plots ( o ) show the nearest distances between CD8 + T cells and F4/80 + macrophages. Red dashed lines indicate tumor regions in ( b,g ). White lines denote tumor borders in ( i ). Scale bars, 1 mm ( b, g ), 500 μm ( d , top), 50 μm ( d , bottom), 100 μm ( i ). Fisher’s LSD test ( c,e,h ), Mann-Whitney U-test, two-sided ( k,m,o ). Mean ± SEM. Source data are provided as a Source Data file.

Article Snippet: For CXCL9/10 inhibition, mice were intraperitoneally injected with anti-CXCL9 antibody (300 μg; Bio X Cell, BE 0309) and anti-CXCL10 antibody (300 μg; Bio X Cell, BE 0440) twice per week.

Techniques: Control, Immunofluorescence, MANN-WHITNEY

Journal: Frontiers in Immunology

Article Title: Reciprocal activation between M1 macrophages and trophoblasts through CXCL9/STAT1/ZEB1/CCL2 axis promotes recurrent spontaneous abortion

doi: 10.3389/fimmu.2025.1629370

Figure Lengend Snippet: M1-Mφ-derived CXCL9 impairs trophoblasts invasion and migration. (A) mRNA expression levels in HTR-8 cells cultured alone or co-cultured with M1-Mφ. (B) ELISA assays of CXCL9 in the supernatant of M1-Mφ alone or co-cultured with HTR-8 cells. (C) CXCL9 mRNA expression in HTR-8 and M1-Mφ with or without 48 h of co-culture. (D-F) The expression of EMT markers in HTR-8 cells alone, CXCL9-supplemented HTR-8 cells, M1-Mφ-co-cultured HTR-8 cells, and anti-CXCL9 M1-Mφ-co-cultured HTR-8 cells were analyzed by qPCR, western blot and immunofluorescence. Scale bar: 20 μm. (G, H) Migration and invasion of anti-CXCL9 M1-Mφ-co-cultured HTR-8 cells and its control were measured by wound-healing assay and transwell assays, respectively. n = 3, Scale bar: 50 μm; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: In the anti-CXCL9 group, anti-Mouse CXCL9/MIG Antibody (1 mg/kg; MCE, Shanghai) were injected into female C57BL/6 mice by intravenously administered at 8:00 am on E7.5, E10.5 and E13.5.

Techniques: Derivative Assay, Migration, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay, Co-Culture Assay, Western Blot, Immunofluorescence, Control, Wound Healing Assay

Journal: Frontiers in Immunology

Article Title: Reciprocal activation between M1 macrophages and trophoblasts through CXCL9/STAT1/ZEB1/CCL2 axis promotes recurrent spontaneous abortion

doi: 10.3389/fimmu.2025.1629370

Figure Lengend Snippet: Anti-CXCL9 treatment alleviates embryo resorption rate in mice. (A) Experimental protocol for LPS-induced abortion model with anti-CXCL9 treatment. (B) The embryo resorption rates in the control, LPS-induced abortion and anti-CXCL9 groups. (C, D) Placental interface expression of E-cadherin and Vimentin. (E) IHC analysis of CD86 at the placental interface of mice. (F-H) IHC of p-STAT1, IRF1 and ZEB1 at the placental interface of mice. Scale bar, 50 µm. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: In the anti-CXCL9 group, anti-Mouse CXCL9/MIG Antibody (1 mg/kg; MCE, Shanghai) were injected into female C57BL/6 mice by intravenously administered at 8:00 am on E7.5, E10.5 and E13.5.

Techniques: Control, Expressing, Paraffin-embedded Immunohistochemistry