Journal: Genes

Article Title: Differentially Expressed Genes of Natural Killer Cells Can Distinguish Rheumatoid Arthritis Patients from Healthy Controls

doi: 10.3390/genes11050492

Figure Lengend Snippet: ELISA Quantification of ( A ) CXCL16 and ( B ) IL-1β and ( C ) IFN-γ in the plasma of RA patients and healthy controls.

Article Snippet: The kits used were human CXCL16 (DY1164), IFN-γ (DY285) and IL-1β (DY201) duosets (R&D systems, Minneapolis, MN, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Clinical Proteomics

Journal: International immunopharmacology

Article Title: MiR-625-5p is a potential therapeutic target in sepsis by regulating CXCL16/CXCR6 axis and endothelial barrier.

doi: 10.1016/j.intimp.2024.112508

Figure Lengend Snippet: Fig. 1. Predictive value of biomarkers and clinical variables on cumulative ICU survival in patients with sepsis or septic shock patients. Blood samples were collected from patients withsepsis or septic shock and healthy subjects (control) for qRT-PCR analysis of miR-625-5p. miR-625-5p was normalized with the level of h-SNORD44 (internal control) to determine the ratios, and the ratios of the control were arbitrarily set at 1. The relative levels of miR-625-5p (a) and CXCL16 (b) were detected by qRT-PCR. The levels of CXCL16 (c), SDC-1 (d), HS (e), and VE-cadherin (f) were detected by using ELISA. The red rulers in Figures a and d represent mean values with SDs, quantitative data was compared with one-way ANOVA between three groups. The blue rulers in Figures b, c, e, and f represent the medians with ranges, quantitative data was compared with Kruskal–Wallis analysis between three groups. ROC curve analysis of APACHE II score, SOFA score, miR-625-5p, CXCL16, SDC-1, PCT, and lactate at admission for the prediction of 28-day mortality (g). Kaplan–Meier survival estimates for all patients with sepsis or septic shock according to the respective level of miR-625-5p (miR-625-5p, cut-of: 25) (h). Correlations between biomarker levels and various clinical parameters (i–n). *P < 0.05, **P < 0.01.

Article Snippet: Serum levels of syndecan-1 (EK1339, BOSTER, China), heparan sulfate (356350, Usbiological, USA), and VE-cadherin (DCADV0, R&D Systems, USA), and CXCL16 (EK0741, BOSTER, China) levels in the serum and cell supernatant were determined with an enzyme-linked immunosorbent assay (ELISA) kit following the manufacturer’s instructions.

Techniques: Control, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Biomarker Discovery

Journal: International immunopharmacology

Article Title: MiR-625-5p is a potential therapeutic target in sepsis by regulating CXCL16/CXCR6 axis and endothelial barrier.

doi: 10.1016/j.intimp.2024.112508

Figure Lengend Snippet: Fig. 3. miR-625-5p regulated CXCL16 transcription and expression. EA.hy926 cells were transfected with miR-625-5p mimic (100 nM) or negative control (miR- 625-5p mimic NC) for 48 h. qRT-PCR analysis of miRNA expression levels for miR-625-5p by transfection of miR-625-5p mimics (a). The relative level of CXCL16 was detected by qRT-PCR (b), CXCL16 supernatant levels were quantified by ELISA (c), whereas the expression levels in EA.hy926 cells were detected by Western blotting (d). Quantification of CXCL16 is illustrated in (e). *P < 0.05, **P < 0.01.

Article Snippet: Serum levels of syndecan-1 (EK1339, BOSTER, China), heparan sulfate (356350, Usbiological, USA), and VE-cadherin (DCADV0, R&D Systems, USA), and CXCL16 (EK0741, BOSTER, China) levels in the serum and cell supernatant were determined with an enzyme-linked immunosorbent assay (ELISA) kit following the manufacturer’s instructions.

Techniques: Expressing, Transfection, Negative Control, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: International immunopharmacology

Article Title: MiR-625-5p is a potential therapeutic target in sepsis by regulating CXCL16/CXCR6 axis and endothelial barrier.

doi: 10.1016/j.intimp.2024.112508

Figure Lengend Snippet: Fig. 4. Inhibitor of miR-625-5p attenuated LPS-induced EA.hy926 cell barrier injury. EA.hy926 cells were transfected with the miR-625-5p inhibitor (50 nM) or negative control (micrOFF inhibitor NC) for 48 h and then exposed to LPS (5 and 10 µg/mL) for 6 h. The relative level of CXCL16 was detected by qRT-PCR (a), the expression levels of CXCL16 in the EA.hy926 cells were detected by Western blotting (b), and CXCL16 supernatant levels were quantified by ELISA (c). Quantification of CXCL16 is illustrated in (d). The effect of LPS and miR-625-5p mimic on the permeability of EA.hy926 cell were assessed using FITC-dextran and TEER methods (e, f). *P < 0.05, **P < 0.01.

Article Snippet: Serum levels of syndecan-1 (EK1339, BOSTER, China), heparan sulfate (356350, Usbiological, USA), and VE-cadherin (DCADV0, R&D Systems, USA), and CXCL16 (EK0741, BOSTER, China) levels in the serum and cell supernatant were determined with an enzyme-linked immunosorbent assay (ELISA) kit following the manufacturer’s instructions.

Techniques: Transfection, Negative Control, Quantitative RT-PCR, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Permeability

Journal: International immunopharmacology

Article Title: MiR-625-5p is a potential therapeutic target in sepsis by regulating CXCL16/CXCR6 axis and endothelial barrier.

doi: 10.1016/j.intimp.2024.112508

Figure Lengend Snippet: Fig. 6. CXCL16 knockdown inhibited LPS-induced endothelial cell injury in vitro. EA.hy926 cells were transfected with CXCL16 siRNA (siCXCL16-1, 2, or 3) or negative control (siRNA NC) and CXCL16 expression was determined after 24 h. The relative level of CXCL16 were detected by qRT-PCR (a). EA.hy926 cells were divided into four groups: Group 1, control; Group 2, LPS; Group 3, LPS + siCXCL16; Group 4, siRNA NC. The cells in Group 3 were transfected with CXCL16 siRNA (siCXCL16), whereas cells in the other groups were transfected with negative control siRNA (siRNA NC). At 24 h post-transfection, the cells in Groups 2 and 3 were treated with 10 µg/mL LPS. Endothelial permeability was measured by using FITC- dextran and TEER 6 h after LPS treatment (b, c). The levels of HS, SDC-1, claudin5, occludin, and VE-cadherin were detected by Western blotting (d). Quantification of HS, SDC-1, claudin5, occludin, and VE-cadherin are shown in (e): *P < 0.05, vs. Control group; & P < 0.05, vs. LPS group. Immunofluorescence images of HS and SDC-1 in EA.hy926 cells (f, h; magnification, ×200; scale bar, 150 µm). Fluo rescence intensity analysis of g and i. *P < 0.05, **P < 0.01.

Article Snippet: Serum levels of syndecan-1 (EK1339, BOSTER, China), heparan sulfate (356350, Usbiological, USA), and VE-cadherin (DCADV0, R&D Systems, USA), and CXCL16 (EK0741, BOSTER, China) levels in the serum and cell supernatant were determined with an enzyme-linked immunosorbent assay (ELISA) kit following the manufacturer’s instructions.

Techniques: Knockdown, In Vitro, Transfection, Negative Control, Expressing, Quantitative RT-PCR, Control, Permeability, Western Blot, Immunofluorescence

Journal: International immunopharmacology

Article Title: MiR-625-5p is a potential therapeutic target in sepsis by regulating CXCL16/CXCR6 axis and endothelial barrier.

doi: 10.1016/j.intimp.2024.112508

Figure Lengend Snippet: Fig. 9. Mechanisms by which miR-625-5p disrupts lung endothelial barrier integrity. miR-625-5p level may be an effective biomarker for predicting 28-day mortality in patients with sepsis or septic shock. Furthermore, LPS-induced vascular endothelial hyper-permeability by regulating miR-625-5p/CXCL16/CXCR6 axis in sepsis. ① LPS increased miR‑625-5p expression in EA.hy926 cells; ② miR-625-5p positively regulated CXCL16 in EA.hy926 cells; ③ Treatment of EA.hy926 cells with LPS significantly increased CXCL16 release; ④ CXCL16 combines with CXCR6; ⑤ CXCR6 mediated the effects of CXCL16 on endothelial barrier dysfunction.

Article Snippet: Serum levels of syndecan-1 (EK1339, BOSTER, China), heparan sulfate (356350, Usbiological, USA), and VE-cadherin (DCADV0, R&D Systems, USA), and CXCL16 (EK0741, BOSTER, China) levels in the serum and cell supernatant were determined with an enzyme-linked immunosorbent assay (ELISA) kit following the manufacturer’s instructions.

Techniques: Biomarker Discovery, Permeability, Expressing

Journal: Genome Medicine

Article Title: Single-cell RNA transcriptome analysis of CNS immune cells reveals CXCL16/CXCR6 as maintenance factors for tissue-resident T cells that drive synapse elimination

doi: 10.1186/s13073-022-01111-0

Figure Lengend Snippet: CXCL16 levels increase on IBA1 + cells during acute infection and return to baseline levels during recovery. a Representative microscopy images of RNA in situ/immunohistochemistry (ISH/IHC) for Cxcl16 and IBA1 in the cortex of mock- or WNV-infected mice at 7, 25, ad 52 DPI. The middle panel is the inset of the white box in 7 DPI. b – d Quantification of the ISH/IHC for Cxcl16 + area ( b ); IBA1 + Cxcl16 + area normalized to the total IBA1 + area, represented as fold change over mock ( c ); or IBA1 + Cxcl16 + area normalized to the total Cxcl16 + area ( d ) in the cortex or hippocampus of mock- or WNV-infected mice at 7, 25, and 52 DPI. e ELISA for CXCL16 in the cortex, hippocampus, cervical lymph nodes, meninges, and blood at 7 and 25 DPI. f Representative immunostaining of the cortex for CXCR6-GFP, CD3, IBA1, and DAPI. Scale bars, 50 μm. Data represent the mean±s.e.m. and were analyzed by one-way ANOVA or unpaired Student’s t -test. * P <0.05, ** P <0.005, *** P < 0.001, **** P < 0.0001

Article Snippet: The level of CXCL16 was detected using a commercial ELISA kit (DY503, R&D Systems, Inc., MN, USA), according to the manufacturer’s guidelines.

Techniques: Infection, Microscopy, In Situ, Immunohistochemistry, Enzyme-linked Immunosorbent Assay, Immunostaining

Journal: Advanced Science

Article Title: CXCL16/CXCR6/TGF‐β Feedback Loop Between M‐MDSCs and Treg Inhibits Anti‐Bacterial Immunity During Biofilm Infection

doi: 10.1002/advs.202409537

Figure Lengend Snippet: The interaction of MDSCs and Treg by CXCL16‐CXCR6 signaling was obvious in PJI. a) The interaction plot showed the cell communications between myeloid cells and lymphocytes by ligand‐receptor pair analysis. The thicker the line represented, the stronger the interaction weights/strength, and the more the number of interactions among the cell types. b) The bar chart showed the interaction strength and the number of interactions among PJI, AF and OA. c) The bar chart showed the strength comparison of the interaction pathways among PJI, AF and OA. d) The heatmap showed relative strength of all enriched signals (outgoing and incoming) across myeloid cells and lymphocytes. e) The circle plot showed the inferred intercellular communication network for CXCL signaling pathway. f) The heatmap showed the CXCL signaling pathway among myeloid cells and lymphocytes. g) The bar chart showed the ligand‐receptor pair of CXCL signaling pathway among myeloid cells and lymphocytes. h) The bar chart showed the relative contribution of ligand‐receptor pair to the overall communication network of CXCL signaling pathway. i) The dot plot showed the incoming communication patterns of CXCL signaling pathway from M‐MDSCs among PJI, AF and OA. j) Multiplex immunofluorescence image showed the co‐location of CXCR6+ Treg and CD68+ myeloid cells with excretive CXCL16. Yellow arrow highlighted the CD68+ myeloid cells expressing CXCL16, and red arrow highlighted the CD4+FOXP3+ Treg expressing CXCR6. Scale bar, 20 mm (PJI, n = 3; AF, n = 3; OA, n = 3). Unpaired t test; ** p < 0.01, *** p < 0.001.

Article Snippet: CXCL16 in human synovial fluid samples was also measured by ELISA (70‐EK1254‐96, Hangzhou, China, MultiSciences).

Techniques: Comparison, Multiplex Assay, Immunofluorescence, Expressing

Journal: Advanced Science

Article Title: CXCL16/CXCR6/TGF‐β Feedback Loop Between M‐MDSCs and Treg Inhibits Anti‐Bacterial Immunity During Biofilm Infection

doi: 10.1002/advs.202409537

Figure Lengend Snippet: PJI patients with high CXCR6 showed a higher recurrence rate. a) The Volcano plot showed differentially expressed genes in PJI versus AF patients. The x and y axes represented the log2 fold change and p‐value, respectively, based on the Mann‐Whitney U test. Red dots: upregulated genes, blue dots: downregulated genes. b) The heatmap showed the distribution of 28 types of immune cells in PJI and AF. c) The bar chart showed the difference in proportion of M‐MDSCs and Treg between PJI and AF (PJI, n = 18; AF, n = 18). Unpaired t test; *** p < 0.001. d) The heatmap showed the characteristic genes of chemokines, PJI markers, cell markers and immune checkpoint between PJI and AF. e–l) The violin plots showed part of the differential genes (CXCL16, CXCR6, S100A8, S100A9, FOXP3, CTLA4, TIGIT and LAG3) between PJI and AF as shown in (d). Unpaired t test; ns p > 0.05, *** p < 0.001, **** p < 0.0001. m,n) Quantitative analysis of immunohistochemical staining for CXCL16 and CXCR6 (PJI, n = 80; AF, n = 25; OA = 25). 40x magnification. Scale bar, 20 mm. Unpaired t test; ** p < 0.01, *** p < 0.001. o) The Kaplan‐Meier curve showed the recurrence curves of PJI patients characterized by either low or high expression of CXCR6 (CXCR6‐high PJI, n = 30; CXCR6‐low PJI, n = 30). Significance was calculated using the log‐rank test. p,q) Quantitative analysis of Harris Hip score and HSS knee score in CXCR6‐high PJI and CXCR6‐low PJI group (CXCR6‐high PJI, n = 30; CXCR6‐low PJI, n = 30). Unpaired t test; * p < 0.05.

Article Snippet: CXCL16 in human synovial fluid samples was also measured by ELISA (70‐EK1254‐96, Hangzhou, China, MultiSciences).

Techniques: MANN-WHITNEY, Immunohistochemical staining, Staining, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: High metastatic tumor-derived CXCL16 mediates liver colonization metastasis by inducing Kupffer cell polarization via the PI3K/AKT/FOXO3a pathway

doi: 10.1016/j.neo.2025.101174

Figure Lengend Snippet: CXCL16 mediate the intercommunication between CRC cells and KCs. (A)RNA-seq gene expression heatmap in CT26 and CT26-LM group. (B)RNA-seq gene expression heatmap in CT26 and co-CT26 group. (C)Venn diagram of genes positively correlated with liver metastasis analyzed through transcriptome microarrays between parent CT26 cells and CT26-LM or CT26 and co-CT26 cells. (D)Heatmap of top 20 changed genes among parent CT26 cells, LM and co-cultured CT26 cells. (E)Heatmap of top 10 changed cytokine genes among parent CT26 cells, LM and co-cultured CT26 cells. (F)qRT-PCR analysis of the mRNA expression of CXCL14, IL24, CXCL16 and IL1A in CT26 and CT26-LM cells. (G)qRT-PCR analysis of the mRNA expression of CXCL14, IL24, CXCL16 and IL1A in CT26 and co-CT26 cells. (H)ELISA assay revealed the CXCL16 upregulated in culture supernatants from CT26-LM cells. (I)Western blot analysis showed the changes of CXCL16 protein level in CT26, CT26-LM and co-CT26 cells.

Article Snippet: CXCL16 levels in cell culture media and mouse serum were quantified using a Mouse CXCL16 ELISA kit (Elabscience, China) according to the manufacturer's protocol.

Techniques: RNA Sequencing, Gene Expression, Cell Culture, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Neoplasia (New York, N.Y.)

Article Title: High metastatic tumor-derived CXCL16 mediates liver colonization metastasis by inducing Kupffer cell polarization via the PI3K/AKT/FOXO3a pathway

doi: 10.1016/j.neo.2025.101174

Figure Lengend Snippet: CXCL16/PI3K/FOXO3a signaling pathway mediates M2 polarization of KCs. (A)Western blot showed the protein expression of M2 markers in different concentration of CXCL16. (B)Western blot showed the protein expression of M2 markers in different concentration of ML339. (C)Western blot analysis of M2 markers (CD206, ARG1) and PI3K/AKT1/FOXO3a pathway in ImKC when treated with CXCL16 (200ng/ml), LM-CM or ML339. (D)Western blot analysis of M2 markers (CD206, ARG1) and PI3K/AKT1/FOXO3a pathway in ImKC when treated with CXCL16 (200ng/ml), LM-CM or LY294002. (E)Identification of transcription factor FOXO3a binding site. (F)Prediction of FOXO3a binding sites in the CD206 promoter region using JASPAR. (G)The promoter sequence of CD206 contains 3 putative FOXO3a transcriptional factor binding sites. (H)Luciferase reporter assays for the activity of wild-type and mutant CD206 promoters. (I)Quantitative analysis of ChIP experiments performed on DNA samples precipitated with antibodies against FOXO3a and IgG using primers detecting FOXO3a binding sites on CD206 promoter.

Article Snippet: CXCL16 levels in cell culture media and mouse serum were quantified using a Mouse CXCL16 ELISA kit (Elabscience, China) according to the manufacturer's protocol.

Techniques: Western Blot, Expressing, Concentration Assay, Binding Assay, Sequencing, Luciferase, Activity Assay, Mutagenesis

Journal: Neoplasia (New York, N.Y.)

Article Title: High metastatic tumor-derived CXCL16 mediates liver colonization metastasis by inducing Kupffer cell polarization via the PI3K/AKT/FOXO3a pathway

doi: 10.1016/j.neo.2025.101174

Figure Lengend Snippet: Intervention of CXCL16/CXCR6 for suppression of CT26-LM metastasis in vivo. (A)Schematic representation of BALB/c mice were treated with ML339 once every two days after spleen injection CT26. (B)Liver metastasis nodules between control(DMSO) group and ML339 group (metastatic tumor nodules shown by arrows). (C)Dotplot showed the number of metastatic tumors between control(DMSO) group and ML339 group. (D)Barplot showed the whole liver weight between control(DMSO) group and ML339 group. (E)F4/80 IHC staining and quantitative analysis of AOD in tumor; scale bar 50μm. (F)Relative mRNA expression of M2 markers and M1 markers in control(DMSO) and ML339 group liver tissue. (G-H)IF staining of ARG1 and CD206 in control(DMSO) and ML339 group liver tissue; scale bar 50μm.

Article Snippet: CXCL16 levels in cell culture media and mouse serum were quantified using a Mouse CXCL16 ELISA kit (Elabscience, China) according to the manufacturer's protocol.

Techniques: In Vivo, Injection, Control, Immunohistochemistry, Expressing, Staining

Journal: Neoplasia (New York, N.Y.)

Article Title: High metastatic tumor-derived CXCL16 mediates liver colonization metastasis by inducing Kupffer cell polarization via the PI3K/AKT/FOXO3a pathway

doi: 10.1016/j.neo.2025.101174

Figure Lengend Snippet: Diagrammatic sketch for CXCL16/PI3K/AKT1/FOXO3a activation in M2-KCs to promote MET of liver-metastatic cells.

Article Snippet: CXCL16 levels in cell culture media and mouse serum were quantified using a Mouse CXCL16 ELISA kit (Elabscience, China) according to the manufacturer's protocol.

Techniques: Activation Assay