Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 plasma levels rise after running and XCL1 treatment increases the number of neurospheres. ( a ) XCL1 plasma levels measured by an ELISA in standard-housed mice (STD, n = 5 mice) and mice housed for 4 days with a running wheel (RUN, n = 6 mice). * p < 0.05, Student’s t -test. ( b ) qPCR gene expression analysis of lymphotactin receptors reveals that neural precursor cells express Itga9 but not Xcr1 (left), although both Itga9 and Xcr1 are detected in splenic control tissue (right). Uncropped gels are presented in Supplementary Fig. . ( c ) Representative images of a SVZ neurosphere (top) and a DG neurosphere (bottom). Scale bars: 100 μm. ( d ) Neurosphere assays with DG-derived primary cells cultured with XCL1. n = 9 to 10 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( e ) Neurosphere assays with SVZ-derived primary cells cultured with XCL1. n = 6 to 9 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( f ) Neurosphere assays with XCL1-neutralizing antibodies. n = 3 to 6 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( g ) Size distribution of DG-derived neurospheres cultured with XCL1. n = 7 to 8 independent experiments. Dashed lines represent control cultures normalized to 100%. All data represent the mean ± SEM.

Article Snippet: Compounds, including recombinant mouse XCL1 protein (R & D Systems) and mouse XCL1 antibodies (5 μg/ml; R & D Systems) diluted in 0.1% bovine serum albumin (BSA) in phosphate-buffered saline (PBS), were added immediately before seeding.

Techniques: Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Gene Expression, Control, Derivative Assay, Cell Culture

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 promotes neuronal differentiation in adherent monolayer and neurosphere cultures. ( a ) Viability assay in adherent NPC cultures with XCL1. n = 5 to 6 independent experiments. ( b ) CFSE proliferation assay in adherent NPC cultures with XCL1. n = 3 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( c ) Motility of adherent monolayer-cultured NPCs determined by semi-automated tracking. Data are plotted as the 5 th /95 th percentile with outliers represented as circles. XCL1: n = 92 cells, Control: n = 104 cells, *** p < 0.001, Student’s t -test. ( d ) Quantifica t ion of β-tubulin + cells in proliferating NPC cultures two days after the addition of XCL1. n = 4 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( e ) Representative image of differentiated NPCs in adherent monolayer cultures showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( f ) Quantification of GFAP + and β-tubulin + cells in differentiated adherent monolayer cultures treated with XCL1. n = 4 to 5 independent experiments, * p < 0.05, *** p < 0.001, one-way ANOVA with Dunnett test. ( g ) Representative image of differentiated neurospheres showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( h ) Quantification of GFAP + and β-tubulin + cells in differentiated neurosphere cultures treated with XCL1. n = 5 independent experiments, *** p < 0.001, one-way ANOVA with Dunnett test. Dashed lines represent control cultures normalized to 100%. All data represent the mean ± SEM.

Article Snippet: Compounds, including recombinant mouse XCL1 protein (R & D Systems) and mouse XCL1 antibodies (5 μg/ml; R & D Systems) diluted in 0.1% bovine serum albumin (BSA) in phosphate-buffered saline (PBS), were added immediately before seeding.

Techniques: Viability Assay, Proliferation Assay, Cell Culture, Control

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 influences the cell cycle progression of NPCs in vitro . ( a ) Representative images of a dividing NPC followed by time-lapse microscopy. Images are 5 min apart. Yellow arrows mark the process of cell division. Scale bar: 10 μm. ( b ) Example of a generation tree of a re-dividing cell obtained from semi-automated cell tracking of NPCs to calculate the mean generation time. ( c ) Generation time of NPCs cultured with and without XCL1 determined by semi-automated cell tracking. Data are plotted as the 5 th /95 th percentile with outliers represented as circles. Control: n = 23 cells, XCL1: n = 26 cells. ( d ) Representative flow cytometry plots of the click-iT EdU proliferation assay. Viable cells were first defined using forward scatter and side scatter (left). Doublets were then excluded from single cell signals by plotting Hoechst-width against Hoechst-area (middle). Finally, to determine the cell cycle phase, the DNA content (Hoechst label) was plotted against the EdU signal (right). ( e ) Percentage of NPCs in S phase. n = 4 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( f ) Percentage of NPCs in G2/M phases. n = 4 independent experiments. ( g ) Percentage of NPCs in G1/G0 phases. n = 4 independent experiments. All data represent the mean ± SEM.

Article Snippet: Compounds, including recombinant mouse XCL1 protein (R & D Systems) and mouse XCL1 antibodies (5 μg/ml; R & D Systems) diluted in 0.1% bovine serum albumin (BSA) in phosphate-buffered saline (PBS), were added immediately before seeding.

Techniques: In Vitro, Time-lapse Microscopy, Cell Tracking Assay, Cell Culture, Control, Flow Cytometry, Proliferation Assay

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: Neurogenesis in XCL1 KO mice is reduced ex vivo . ( a ) Neurosphere assays with primary DG cells from XCL1 KO mice (−/−) and WT littermates (+/+). n = 6 mice per group, * p < 0.05, paired Student’s t -test. ( b ) Representative image of differentiated neurospheres showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( c ) Quantification of GFAP + and β-tubulin + cells in differentiated neurosphere cultures from XCL1 −/− and +/+ mice. n = 4 mice per group, * p < 0.05, Student’s t -test. ( d ) and ( e ) Neurosphere assays with primary DG cells from XCL1 −/− and +/+ mice in the presence of ( d ) potassium chloride (n = 4 to 5 independent experiments) and ( e ) norepinephrine (n = 6 independent experiments). *** p < 0.001, **** p < 0.0001, Student’s t -test.

Article Snippet: Compounds, including recombinant mouse XCL1 protein (R & D Systems) and mouse XCL1 antibodies (5 μg/ml; R & D Systems) diluted in 0.1% bovine serum albumin (BSA) in phosphate-buffered saline (PBS), were added immediately before seeding.

Techniques: Ex Vivo

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 1 | Expression of XCL1 and XCR1 in periprosthetic tissues. (A–C) Detection of XCR1 in human tissues around loosening hip-implant by the immunofluorescence test. Deparaffinized sections were stained to observe F4/80, CD68, iNOS (green), XCR1 (red), and cell nuclei (blue). Scale bars are 100 µm. Representative images exhibit the sectioned tissues from three patients. (D) Detection of XCL1 in synovial fluid from same patients by Western blotting analysis. (E) Gene expressions of XCL1 and XCR 1 in calvarial bone tissues in a murine osteolysis calvarial model. Results represent the means of relative expression values ± SEM of three mice. *indicates a significant difference as determined by the Student t-test (p ≤0.05).

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Expressing, Staining, Western Blot

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 2 | Administration of XCL1 exaggerates osteolytic lesions in a polyethylene-particles-induced osteolysis model. (A) Representative images for micro-CT of calvariae. The right panel shows quantification of the lytic area on the calvarial bone tissues of mice. Results represent the means ± SEM of seven mice. *represents the significance determined by one-way ANOVA, followed by a Tukey’s multiple-comparison procedure. Arrows indicate osteolytic lesions. (B) Representative images for histological analyses of bone sections stained by H&E and TRAP. Scale bar is 100 µm. Arrows indicate bone lesions. (C) Cell count of inflammatory cells in calvarial bone sections. (D) Quantification of TRAP-stained areas in calvarial bone sections. The results represent the means ± SEM for three mice. *indicates a significant difference, as determined by one-way ANOVA, followed by the Tukey’s multiple-comparison procedure (p ≤0.05).

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Micro-CT, Comparison, Staining, Cell Counting

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 3 | Blockade of XCL1/lymphotactin by neutralizing antibody ameliorates severity of osteolysis triggered by polyethylene-particles in murine model. (A) Quantification of the lytic area on the calvarial bone tissues of mice determined by micro-CT. Left panel shows representative images of calvariae. Results represent the means ± SEM of seven mice. *represents the significance determined by one-way ANOVA, followed by a Tukey’s multiple-comparison procedure. Arrows indicate osteolytic lesions. (B) Histological analyses of bone sections stained by H&E and TRAP. Scale bar is 100 µm. (C) Cell count of inflammatory cells in calvarial bone sections. (D) Quantification of TRAP-stained areas in calvarial bone sections. Arrows indicate bone lesions. The results represent the means ± SEM for four mice. *indicates a significant difference, as determined the Student t-test (p ≤0.05).

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Micro-CT, Comparison, Staining, Cell Counting

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 4 | Sponge-soaked protein-induced murine model. (A) Representative images for micro-CT of calvariae. Arrows indicate osteolytic lesions. (B) Quantification of the lytic area on the calvarial bone tissues of mice determined by micro-CT. Results represent the means ± SEM of six mice. *represents the significance determined by one-way ANOVA, followed by a Tukey’s multiple-comparison procedure. (C) Representative images for histological analyses of bone sections stained by H&E and TRAP. Scale bar is 100 µm. Arrows indicate bone lesions. (D) Quantification of TRAP-stained areas in calvarial bone sections. Results represent the means ± SEM of three mice. *indicates a significant difference, as determined by one-way ANOVA, followed by the Tukey’s multiple-comparison procedure (p ≤0.05). (E) Heat map for the gene expression of inflammatory and osteoclast marker genes in bone tissues. Calvarial bone tissues were harvested for the analysis of gene expressions after the implantation of XCL1-soaked sponges. Scale bar (Log2) represents the means of relative expression values of each target gene after normalizing with the GAPDH ± SEM of three mice. *indicates a significant difference, as determined by the t-test (p ≤0.05).

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Micro-CT, Comparison, Staining, Gene Expression, Marker, Expressing

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 5 | Effect of XCL1 on osteoclast differentiation and bone resorption. (A) Count of TRAP-positive cells in RANKL-stimulated monocytes in the presence or absence of XCL1. Left panel shows representative images for cells stained by TRAP. Results represent the means ± SEM of triplicates. *indicates a significant difference, as determined by the Tukey’s multiple comparisons test (p ≤0.05). (B) Actin ring staining assay for cells in RANKL-stimulated monocytes in the presence or absence of XCL1. (C) Quantification of the bone resorbed areas on dentin slices. Results represent the means ± SEM of values from three dentin slices. Left panel shows representative images for the resorbed areas. Scale bars are 200 µm.

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Staining

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 7 | The summary of the current study. XCL1 promotes cells infiltrate, inflammatory response, and osteoclast differentiation leading to aseptic loosening. Scale bars are indicated on histological images. Blocking of the XCL1 might be a potent therapeutic target for this clinical problem.

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Blocking Assay

Journal: Frontiers in immunology

Article Title: Blockade of XCL1/Lymphotactin Ameliorates Severity of Periprosthetic Osteolysis Triggered by Polyethylene-Particles.

doi: 10.3389/fimmu.2020.01720

Figure Lengend Snippet: FIGURE 6 | Effect of XCL1 on osteoblast activation and function. (A) Effect of XCL1 on the gene expression of inflammatory and osteoclastogenic factors in differentiated osteoblasts. Results represent the means ± SEM of triplicates and *indicates a significant difference, as determined by the Tukey’s multiple comparisons test (p ≤0.05). (B) Effects of stimulation by recombinant proteins on NFKB, SAPK/JNK, P42/p44-MAPK (Erk1/2), and P38- activities. Human osteoblasts were cultured in a differentiation medium supplemented with either XCL1 or TNFα (positive control) and harvested for the gene expression analysis by qRT-PCR or for protein analysis by western blotting.

Article Snippet: Recombinant mouse XCL1 (R&D system) at concentrations of 1 or 2 μg (dissolved in 100 μl PBS) was subcutaneously injected (single injection) onto calvariae to examine its effect on the progression of osteolysis (n = 7 for each group).

Techniques: Activation Assay, Gene Expression, Recombinant, Cell Culture, Positive Control, Quantitative RT-PCR, Western Blot

Journal: The Journal of Experimental Medicine

Article Title: Superior antigen cross-presentation and XCR1 expression define human CD11c + CD141 + cells as homologues of mouse CD8 + dendritic cells

doi: 10.1084/jem.20100348

Figure Lengend Snippet: XCL1 induces a [Ca 2+ ] i signal in CD141 + DCs. CD141 + DCs were flow sorted to a purity >98.7%, immobilized on poly– l -lysine–coated glass coverslips, and loaded with 2 µM fura-2/AM. Cells were imaged in a monochromator-assisted digital video imaging system and challenged with 1 µg/ml XCL1 as indicated (left arrow). Subsequently, the same cells were challenged again with a mixture of 100 ng/ml CCL2, 200 ng/ml CCL21, 200 ng/ml CXCL9, and 1 ng/ml CX3CL1 used as a positive control (right arrow). The data shown represent [Ca 2+ ] i concentrations of 300 single cells (gray lines) measured in two independent experiments. The mean [Ca 2+ ] i signal averaged over all cells responding to XCL1 is indicated (black line).

Article Snippet: The lower chamber was filled with chemotaxis medium containing recombinant human XCL1 (R&D Systems) or any of the chemokines CCL2 (100 ng/ml), CCL21 (200 ng/ml), CX3CL1 (1 ng/ml), CXCL12 (200 ng/ml for T cells, B cells, NK cells, and monocytes; 100 ng/ml for pDCs), and CXCL8 (100 ng/ml; all from R&D Systems), and the cells were incubated for 150 min at 37°C in 5% CO 2 .

Techniques: Imaging, Positive Control

Journal: The Journal of Experimental Medicine

Article Title: Superior antigen cross-presentation and XCR1 expression define human CD11c + CD141 + cells as homologues of mouse CD8 + dendritic cells

doi: 10.1084/jem.20100348

Figure Lengend Snippet: XCL1 selectively induces chemotaxis in CD141 + DCs. (A) A mixture of highly purified, flow-sorted DC subtypes (20% CD141 + , 40% CD16 + , and 40% CD1c + DCs; Input DC) was tested for migration in response to medium alone or to serial dilutions of XCL1 (10–5,000 ng/ml) in a Transwell system. A combination of the chemokines CCL2, CCL21, and CX3CL1 was used as a positive control for the DC subsets (Migrated DC). The absolute numbers of CD141 + , CD1c + , and CD16 + DCs in input and migrated cell populations are truly represented in the dot plots, because all cells within a defined volume were included in the analysis in each instance. (B) Proportion of migrated CD1c + , CD16 + , and CD141 + DCs in the experiment shown in A. (C) Proportion of migrated pDCs, monocytes, granulocytes, T cells, B cells, and NK cells in response to XCL1 (10–1,000 ng/ml) or the chemokines CXCL12 and CXCL8, which were used as positive controls. For migration assays of B cells, NK cells, and monocytes, PBMCs were magnetically depleted of T cells, and for T cell migration, PBMCs were used directly. For migration assays of granulocytes, whole blood cells were used after erythrocyte lysis with ACK buffer, and pDCs were magnetically enriched from PBMCs with the Plasmacytoid Dendritic Cell Isolation Kit (Miltenyi Biotec). All experiments with DCs were performed three times; all other populations were assayed twice. Error bars represent means ± SEM.

Article Snippet: The lower chamber was filled with chemotaxis medium containing recombinant human XCL1 (R&D Systems) or any of the chemokines CCL2 (100 ng/ml), CCL21 (200 ng/ml), CX3CL1 (1 ng/ml), CXCL12 (200 ng/ml for T cells, B cells, NK cells, and monocytes; 100 ng/ml for pDCs), and CXCL8 (100 ng/ml; all from R&D Systems), and the cells were incubated for 150 min at 37°C in 5% CO 2 .

Techniques: Chemotaxis Assay, Purification, Migration, Positive Control, Lysis, Cell Isolation

Journal: The Journal of Experimental Medicine

Article Title: Superior antigen cross-presentation and XCR1 expression define human CD11c + CD141 + cells as homologues of mouse CD8 + dendritic cells

doi: 10.1084/jem.20100348

Figure Lengend Snippet: Involvement of the XCL1–XCR1 communication axis in the innate and adaptive cytotoxic responses to cross-presented microbial and tumor antigens. Secretion of the chemokine XCL1 by activated NK cells specifically attracts XCR1-expressing DCs capable of antigen cross-presentation. This ensures an effective communication between these cells in the innate phase of the immune response. In the adaptive phase, secretion of XCL1 by activated CD8 + T cells optimizes the communication with antigen cross-presenting DCs and facilitates the differentiation of CD8 + T cells to cytotoxic cells.

Article Snippet: The lower chamber was filled with chemotaxis medium containing recombinant human XCL1 (R&D Systems) or any of the chemokines CCL2 (100 ng/ml), CCL21 (200 ng/ml), CX3CL1 (1 ng/ml), CXCL12 (200 ng/ml for T cells, B cells, NK cells, and monocytes; 100 ng/ml for pDCs), and CXCL8 (100 ng/ml; all from R&D Systems), and the cells were incubated for 150 min at 37°C in 5% CO 2 .

Techniques: Expressing

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 plasma levels rise after running and XCL1 treatment increases the number of neurospheres. ( a ) XCL1 plasma levels measured by an ELISA in standard-housed mice (STD, n = 5 mice) and mice housed for 4 days with a running wheel (RUN, n = 6 mice). * p < 0.05, Student’s t -test. ( b ) qPCR gene expression analysis of lymphotactin receptors reveals that neural precursor cells express Itga9 but not Xcr1 (left), although both Itga9 and Xcr1 are detected in splenic control tissue (right). Uncropped gels are presented in Supplementary Fig. . ( c ) Representative images of a SVZ neurosphere (top) and a DG neurosphere (bottom). Scale bars: 100 μm. ( d ) Neurosphere assays with DG-derived primary cells cultured with XCL1. n = 9 to 10 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( e ) Neurosphere assays with SVZ-derived primary cells cultured with XCL1. n = 6 to 9 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( f ) Neurosphere assays with XCL1-neutralizing antibodies. n = 3 to 6 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( g ) Size distribution of DG-derived neurospheres cultured with XCL1. n = 7 to 8 independent experiments. Dashed lines represent control cultures normalized to 100%. All data represent the mean ± SEM.

Article Snippet: Protein levels were also measured using the mouse XCL1 PicoKineTM ELISA Kit (Boster Biological Technology), according to the manufacturer’s instructions.

Techniques: Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Gene Expression, Control, Derivative Assay, Cell Culture

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 promotes neuronal differentiation in adherent monolayer and neurosphere cultures. ( a ) Viability assay in adherent NPC cultures with XCL1. n = 5 to 6 independent experiments. ( b ) CFSE proliferation assay in adherent NPC cultures with XCL1. n = 3 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( c ) Motility of adherent monolayer-cultured NPCs determined by semi-automated tracking. Data are plotted as the 5 th /95 th percentile with outliers represented as circles. XCL1: n = 92 cells, Control: n = 104 cells, *** p < 0.001, Student’s t -test. ( d ) Quantifica t ion of β-tubulin + cells in proliferating NPC cultures two days after the addition of XCL1. n = 4 independent experiments, * p < 0.05, one-way ANOVA with Dunnett test. ( e ) Representative image of differentiated NPCs in adherent monolayer cultures showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( f ) Quantification of GFAP + and β-tubulin + cells in differentiated adherent monolayer cultures treated with XCL1. n = 4 to 5 independent experiments, * p < 0.05, *** p < 0.001, one-way ANOVA with Dunnett test. ( g ) Representative image of differentiated neurospheres showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( h ) Quantification of GFAP + and β-tubulin + cells in differentiated neurosphere cultures treated with XCL1. n = 5 independent experiments, *** p < 0.001, one-way ANOVA with Dunnett test. Dashed lines represent control cultures normalized to 100%. All data represent the mean ± SEM.

Article Snippet: Protein levels were also measured using the mouse XCL1 PicoKineTM ELISA Kit (Boster Biological Technology), according to the manufacturer’s instructions.

Techniques: Viability Assay, Proliferation Assay, Cell Culture, Control

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: XCL1 influences the cell cycle progression of NPCs in vitro . ( a ) Representative images of a dividing NPC followed by time-lapse microscopy. Images are 5 min apart. Yellow arrows mark the process of cell division. Scale bar: 10 μm. ( b ) Example of a generation tree of a re-dividing cell obtained from semi-automated cell tracking of NPCs to calculate the mean generation time. ( c ) Generation time of NPCs cultured with and without XCL1 determined by semi-automated cell tracking. Data are plotted as the 5 th /95 th percentile with outliers represented as circles. Control: n = 23 cells, XCL1: n = 26 cells. ( d ) Representative flow cytometry plots of the click-iT EdU proliferation assay. Viable cells were first defined using forward scatter and side scatter (left). Doublets were then excluded from single cell signals by plotting Hoechst-width against Hoechst-area (middle). Finally, to determine the cell cycle phase, the DNA content (Hoechst label) was plotted against the EdU signal (right). ( e ) Percentage of NPCs in S phase. n = 4 independent experiments, ** p < 0.01, one-way ANOVA with Dunnett test. ( f ) Percentage of NPCs in G2/M phases. n = 4 independent experiments. ( g ) Percentage of NPCs in G1/G0 phases. n = 4 independent experiments. All data represent the mean ± SEM.

Article Snippet: Protein levels were also measured using the mouse XCL1 PicoKineTM ELISA Kit (Boster Biological Technology), according to the manufacturer’s instructions.

Techniques: In Vitro, Time-lapse Microscopy, Cell Tracking Assay, Cell Culture, Control, Flow Cytometry, Proliferation Assay

Journal: Scientific Reports

Article Title: The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

doi: 10.1038/s41598-019-48360-5

Figure Lengend Snippet: Neurogenesis in XCL1 KO mice is reduced ex vivo . ( a ) Neurosphere assays with primary DG cells from XCL1 KO mice (−/−) and WT littermates (+/+). n = 6 mice per group, * p < 0.05, paired Student’s t -test. ( b ) Representative image of differentiated neurospheres showing GFAP + astrocytes in green and β-tubulin + neurons in red. Scale bar: 50 μm. ( c ) Quantification of GFAP + and β-tubulin + cells in differentiated neurosphere cultures from XCL1 −/− and +/+ mice. n = 4 mice per group, * p < 0.05, Student’s t -test. ( d ) and ( e ) Neurosphere assays with primary DG cells from XCL1 −/− and +/+ mice in the presence of ( d ) potassium chloride (n = 4 to 5 independent experiments) and ( e ) norepinephrine (n = 6 independent experiments). *** p < 0.001, **** p < 0.0001, Student’s t -test.

Article Snippet: Protein levels were also measured using the mouse XCL1 PicoKineTM ELISA Kit (Boster Biological Technology), according to the manufacturer’s instructions.

Techniques: Ex Vivo

Journal: The Journal of rheumatology

Article Title: Expression and function of the C-class chemokine lymphotactin (XCL1) in Wegener's granulomatosis.

doi: 10.3899/jrheum.090244

Figure Lengend Snippet: Figure 2. Flow cytometric analysis results of XCL1 expression. PBMC isolated from WG patients and healthy controls were subjected to flow cytometric analysis after stimulation with PMA/ionomycin. XCL1 was expressed in a significantly greater proportion of CD4+ and CD8+ T cells in WG patients in comparison to controls (A). XCL1 was detected in WG patients in a significantly greater proportion of CD4+ T cells lacking the costimulatory molecule CD28 (p = 0.007) (B). In CD8+ T cells, XCL1 expression was found mainly within the CD8+CD28– T cell subpopulation in WG patients, but there was no statistically significant difference of frequencies of XCL1-expressing CD8+CD28– T cells between WG patients and controls (C). Comparing T cells from patients with active and inactive WG, a significant difference was detected between these patient groups in frequencies of XCL1-positive CD4+ T cells and CD8+ T cells (p < 0.001) (D).

Article Snippet: Cells were subsequently stimulated with human recombinant XCL1 (R&D Systems, Wiesbaden, Germany) at a concentration of 100 ng/ml for 24 h. To control the specificity of XCL1 action, anti-human XCL1 antibody (R&D Systems) was added to the control wells.

Techniques: Expressing, Isolation, Comparison