Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: cDNA array profiling of genetic alterations and the expression of CCL26 in mono-cultured and co-cultured MG63 cells and hMSCs. ( A ) Changes in mRNA expression in hMSCs at 48 h after the co-culture with MG63 and in MG63 co-cultured with hMSCs. Co-culture condition increased the expression of CCL26 mRNA in both cell lines. ( B ) Relative expression of CCL26 mRNA in alone and co-cultured condition in each cell lines. Changes in the mRNA expression with high expression variability ( C ) and changes in the expression of housekeeping genes (control) ( D ).

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Expressing, Cell Culture, Co-Culture Assay, Control

Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: Changes in CCL26 expression and cell growth in MG63 and hMSCs induced by the co-culture condition and rCCL26 administration. ( A ) Changes in CCL26 mRNA expression in MG63 and hMSCs were assessed by qRT-PCR. The co-culture and rCCL26 addition significantly increased the expression of mRNA of CCL26. ( B ) rCCL26 was administered to mono-cultured MG63. There was a significant increase in cell growth with rCCL26 at 10 ng/ml. (*) p < 0.05, (**) p < 0.01. ( C ) Changes in cell proliferation induced by Co-cultured condition and recombinant CCL26 administration in MG63 and hMSCs. ( D ) Changes in the degree of cell proliferation in the untreated, co-culture, and recombinant groups were analyzed by performing cell proliferation assay via BrdU incorporation. ( E ) Changes inCCL26 protein expression of intra-cellular hMSCs and MG63 were assessed by western blot analysis. The co-culture and addition of rCCL26 increased the expression ofCCL26 protein in MG63 and hMSCs. ( F ) The quantification of western blot analysis. Data represents represent the mean ± SD of three independent experiments. p < 0.05 was considered to indicate significance: (*) p < 0.05, (**) p < 0.01.

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Expressing, Co-Culture Assay, Quantitative RT-PCR, Cell Culture, Recombinant, Proliferation Assay, BrdU Incorporation Assay, Western Blot

Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: Effects of neutralizing anti-CCL26 Ab on CCL26 expression in mono-cultured and co-cultured MG63 and hMSCs. ( A ) Changes inCCL26 expression in MG63 were assessed by qRT-PCR. The addition of antiCCL26 Ab to MG63 decreased the expression ofCCL26 mRNA. ( B ) Changes inCCL26 expression in hMSCs were assessed by qRT-PCR. The addition of anti-CCL26 Ab to hMSCs decreased the expression ofCCL26 mRNA. ( C ) Changes inCCL26 protein expression in hMSCs and MG63 were assessed by western blot analysis. The addition of anti-CCL26 Ab to MG63 and hMSCs decreased the expression ofCCL26 protein in these cells. ( D ) The quantification of western blot analysis. Data represents represent the mean ± SD of three independent experiments. p < 0.05 was considered to indicate significance: (*) p < 0.05, (**) p < 0.01.

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Expressing, Cell Culture, Quantitative RT-PCR, Western Blot

Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: Effects of co-culture condition and administration of rCCL26 or anti-CCL26 Ab on motility of MG63. ( A ) Changes in protein expression of factors related to cell motility. ( B ) The quantification of western blot analysis. Data represents represent the mean ± SD of three independent experiments. ( C ) The influence of co-culture and rCCL26 or anti-CCL26 Ab on actin fiber morphology was evaluated using immunofluorescent imaging. Original magnification, × 400; Scale bars: 50 μm. ( D ) The cell migration of MG63 was assessed in each group at 24 h after the challenge with or without rCCL26 and neutralizing anti-CCL26 Ab. ( E ) The amount of MG63 cells that crossed the membrane was measured. A significant decrease in motility was found in the group given anti-CCL26 Ab. ( F ) The cell invasion in MG63 was assessed in each group after 24 h. ( G ) The amount of the cells of which the membrane with Matrigel was crossed by MG63 was measured. Decreased migration ability was found in the group administered anti-CCL26 Ab. p < 0.05 was considered to indicate significance: (*) p < 0.05, (**) p < 0.01.

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Co-Culture Assay, Expressing, Western Blot, Imaging, Migration, Membrane

Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: Changes in expression of Src and its downstream factors related to invasive potential. ( A ) Changes in phosphorylation and the expression of protein factors relating to invasive potential were analyzed. Decreased phosphorylation of Src, FAK, MEK and ERK in MG63 cells was noted in the group administered anti-CCL26 Ab. ( B ) The quantification of western blot analysis. ( C ) Immunofluorescence staining of cultured MG63 cells showed decreased CCL26 and p-Src in the group administered anti-CCL26 Ab. Original magnification, × 400; Scale bars: 50 μm. ( D ) The number of Rac and p-Src positive cells per unit area. Data represents represent the mean ± SD of three independent experiments. p < 0.05 was considered to indicate significance: (*) p < 0.05, (**) p < 0.01.

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Expressing, Phospho-proteomics, Western Blot, Immunofluorescence, Staining, Cell Culture

Journal: Scientific Reports

Article Title: Analysis of the signal cross talk via CCL26 in the tumor microenvironment in osteosarcoma

doi: 10.1038/s41598-021-97153-2

Figure Lengend Snippet: Changes in the lung nodules and the expression of Rac and phosphorylated Src in pulmonary metastatic lesions. ( A ) The group given anti-CCL26 Ab showed a significant suppression of the size of the pulmonary metastatic lesion. ( B ) Immunostaining of the tissues collected from the pulmonary metastatic lesion. Decreased expression of Rac and p-Src was observed in the group administered anti-CCL26 Ab. Original magnification, × 400; Scale bars: 50 μm. ( C ) The number of Rac and p-Src positive cells per unit area. Data represents represent the mean ± SD of three independent experiments. p < 0.05 was considered to indicate significance: (*) p < 0.05, (**) p < 0.01.

Article Snippet: Recombinant human CCL26 (R&D system, USA) (10 ng/ml) was administered to the culture medium.

Techniques: Expressing, Immunostaining

Journal: Hepatology (Baltimore, Md.)

Article Title: Hypoxia induces myeloid-derived suppressor cell recruitment to hepatocellular carcinoma through chemokine (C-C motif) ligand 26.

doi: 10.1002/hep.28655

Figure Lengend Snippet: Fig. 2. Hypoxia promotes MDSC invasion and CCL26 expression in HCC. (A) Representative IHC images of MDSC marker (CD11b) and two well-established hypoxia markers (GLUT1 and CA9) on consecutive sections

Article Snippet: Sources of the recombinant proteins used in Transwell invasion assays are listed as follow: human CCL26 (R&D Systems), mouse GM-CSF (R&D Systems), and mouse IL-4 (R&D Systems).

Techniques: Expressing, Marker

Journal: Hepatology (Baltimore, Md.)

Article Title: Hypoxia induces myeloid-derived suppressor cell recruitment to hepatocellular carcinoma through chemokine (C-C motif) ligand 26.

doi: 10.1002/hep.28655

Figure Lengend Snippet: Fig. 3. CCL26 is a direct transcriptional target of HIFs. CCL26 mRNA (i) and protein (ii) expression in (A) MHCC-97L-EV, -shHIF-1α, -shHIF-2α clones, (B) MHCC-97L-HIF1α-wildtype (WT) and –HIF1α-knockout (KO)

Article Snippet: Sources of the recombinant proteins used in Transwell invasion assays are listed as follow: human CCL26 (R&D Systems), mouse GM-CSF (R&D Systems), and mouse IL-4 (R&D Systems).

Techniques: Expressing, Clone Assay, Knock-Out

Journal: Hepatology (Baltimore, Md.)

Article Title: Hypoxia induces myeloid-derived suppressor cell recruitment to hepatocellular carcinoma through chemokine (C-C motif) ligand 26.

doi: 10.1002/hep.28655

Figure Lengend Snippet: Fig. 4. Hypoxia induces MDSC invasion though CCL26/CX3CR1. (A) (i) Percentages and number of CX3CR1

Article Snippet: Sources of the recombinant proteins used in Transwell invasion assays are listed as follow: human CCL26 (R&D Systems), mouse GM-CSF (R&D Systems), and mouse IL-4 (R&D Systems).

Techniques:

Journal: Hepatology (Baltimore, Md.)

Article Title: Hypoxia induces myeloid-derived suppressor cell recruitment to hepatocellular carcinoma through chemokine (C-C motif) ligand 26.

doi: 10.1002/hep.28655

Figure Lengend Snippet: Fig. 5. CCL26 promotes MDSC recruitment and tumor growth. (A) BALB/c mice were orthotopically implanted with 1×106 MHCC-97L-NTC, -shCCL26-1 (sh26-1), and –shCCL26-2 (sh26-2) clones (n = 5 for

Article Snippet: Sources of the recombinant proteins used in Transwell invasion assays are listed as follow: human CCL26 (R&D Systems), mouse GM-CSF (R&D Systems), and mouse IL-4 (R&D Systems).

Techniques: Clone Assay