Journal: Virology

Article Title: Identification of a Novel Signaling Complex Containing Host Chemokine Receptor CXCR4, Interleukin-10 Receptor, and Human Cytomegalovirus US27

doi: 10.1016/j.virol.2020.06.006

Figure Lengend Snippet: HEK293 cells cultured on glass coverslips were stained with primary antibodies to CXCR4, IL-10R, or OSMR in pairwise combinations as indicated, followed by secondary antibodies with oligonucleotide PLA+ or − probes. Red punctae indicate protein colocalization. The far right panel shows staining with anti-OSMR only followed by FITC-conjugated secondary antibody. Maximum intensity projection from Z-stack images taken by confocal microscopy. Scale bar = 10 μm.

Article Snippet: After blocking, anti-CXCR4 goat polyclonal (1:100, Santa Cruz) and anti-IL-10Rα rabbit polyclonal antibody (1:100, Santa Cruz) were diluted in incubation buffer (1% BSA, 0.3% Triton X-100, and 0.01% sodium azide in PBS) and added onto slides with overnight incubation at 4°C.

Techniques: Cell Culture, Staining, Confocal Microscopy

Journal: Virology

Article Title: Identification of a Novel Signaling Complex Containing Host Chemokine Receptor CXCR4, Interleukin-10 Receptor, and Human Cytomegalovirus US27

doi: 10.1016/j.virol.2020.06.006

Figure Lengend Snippet: A) HEK293 cells cultured on glass coverslips were treated with 100 ng/ml chemokine or cytokine as indicated and stained with antibodies to CXCR4 and IL-10R followed by secondary antibodies with oligonucleotide PLA+ or − probes. Red punctae indicate protein colocalization. Maximum intensity projection from Z-stack images taken by confocal microscopy. Scale bar = 10 μm. B) Average number of spots per cell; 10 cells counted per treatment. Error bars, standard error of mean (SEM). *p<0.00001, paired Student’s t-test vs. untreated, cmvIL-10, and each other. C) Calcium mobilization in HEK293 cells labeled with Fluo-4. Arrow indicates addition of 100 ng/ml stimulus: CXCL12 (solid black line), cmvIL-10 (gray dashed line), or CXCL12 + cmvIL-10 (black dashed line).

Article Snippet: After blocking, anti-CXCR4 goat polyclonal (1:100, Santa Cruz) and anti-IL-10Rα rabbit polyclonal antibody (1:100, Santa Cruz) were diluted in incubation buffer (1% BSA, 0.3% Triton X-100, and 0.01% sodium azide in PBS) and added onto slides with overnight incubation at 4°C.

Techniques: Cell Culture, Staining, Confocal Microscopy, Labeling

Journal: Virology

Article Title: Identification of a Novel Signaling Complex Containing Host Chemokine Receptor CXCR4, Interleukin-10 Receptor, and Human Cytomegalovirus US27

doi: 10.1016/j.virol.2020.06.006

Figure Lengend Snippet: HEK293 cells stably expressing US27 (top panel) or US28 (bottom panel) were grown on glass bottom chamber slides and co-stained with anti-FLAG mouse antibody, anti-CXCR4 goat polyclonal antibody, and anti-IL-10Rα rabbit polyclonal antibody, followed by Alexa Fluor 514 goat-anti-mouse secondary antibody, Alexa Fluor 647 donkey-anti-goat secondary antibody, and Alexa Fluor 594 goat-anti-rabbit secondary antibody, respectively. US27 or US28 (as indicated) is represented in green, CXCR4 in magenta, IL-10R in red, and areas of colocalization are represented in yellow in the merged images. Representative images are shown as maximum intensity projections from Z-stacks. Nuclei appear blue due to DAPI staining. Scale bar = 10 μm.

Article Snippet: After blocking, anti-CXCR4 goat polyclonal (1:100, Santa Cruz) and anti-IL-10Rα rabbit polyclonal antibody (1:100, Santa Cruz) were diluted in incubation buffer (1% BSA, 0.3% Triton X-100, and 0.01% sodium azide in PBS) and added onto slides with overnight incubation at 4°C.

Techniques: Stable Transfection, Expressing, Staining

Journal: Virology

Article Title: Identification of a Novel Signaling Complex Containing Host Chemokine Receptor CXCR4, Interleukin-10 Receptor, and Human Cytomegalovirus US27

doi: 10.1016/j.virol.2020.06.006

Figure Lengend Snippet: A) HEK293 cells were grown in glass chamber slides and stained with primary antibodies against CXCR4 and IL-10R, followed by oligonucleotide-conjugated secondary antibodies for PLA. Maximum intensity projections from Z-stack images are shown. Nuclei appear blue due to DAPI staining. Scale bar = 10 μm. B) Quantification of discrete spots for CXCR4:IL-10R interactions, average of 10 cells. Error bars, SEM. *, p < 0.001 by paired Student’s t-test. C) 293-US27 cells were grown in glass chamber slides and stained with primary antibodies against CXCR4, IL-10R, OSMR, and/or US27 followed by oligonucleotide-conjugated secondary antibodies for PLA as in A.

Article Snippet: After blocking, anti-CXCR4 goat polyclonal (1:100, Santa Cruz) and anti-IL-10Rα rabbit polyclonal antibody (1:100, Santa Cruz) were diluted in incubation buffer (1% BSA, 0.3% Triton X-100, and 0.01% sodium azide in PBS) and added onto slides with overnight incubation at 4°C.

Techniques: Staining

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 1 Protein expression of IL-10, IL-10Rα, and STAT3 by IHC in cancerous colonic tissues (n = 120 patients; 20 × objective; Scale bar = 15 μm) and the IHC arbitrary scores for each protein are shown as boxplots according to tumour locations and cancer stage (a = P < 0.01 compared to early-stage right-sided normal colonic tissues; b = P < 0.01 compared to early-stage right-sided cancerous colonic tissues; c = P < 0.01 compared to early-stage left-sided normal colonic tissues; d = P < 0.01 compared to early-stage left-sided cancerous colonic tissues; e = P < 0.01 compared to late-stage right-sided normal colonic tissues; f = P < 0.01 compared to late-stage right-sided cancerous colonic tissues; g = P < 0.01 compared with late-stage left-sided normal colonic tissues)

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Expressing

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 2 Protein detection of total and phosphorylated STAT3 by Western blotting alongside their relative expression (mean ± SD) in non-treated control cells (CT) and after treatment with varying concentrations (0.1, 1, and 40 ng/ml) of IL-10 protein and anti-IL-10Rα monoclonal antibody, and a single concentration of STAT3 inhibitor V (2 µM) for 24 h in the (a) HT29 and (b) SW620 human colon cancer cell lines (mean ± SD; a = P < 0.01 compared to the CT group; b = P < 0.01 compared to 0.1 ng/mL of IL-10 protein; c = P < 0.01 compared to 1 ng/mL of IL-10 protein; d = P < 0.01 compared to 40 ng/mL of IL-10 protein; e = P < 0.01 compared to 0.1 ng/mL of anti-IL-10Rα antibody; f = P < 0.01 compared to 1 ng/mL of anti-IL-10Rα antibody; and g = P < 0.01 compared to 40 ng/mL of anti-IL-10Rα antibody). The uncropped Western blot membranes for total and phosphorylated STAT3 are provided in Suppl. Figure 4

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Western Blot, Expressing, Control, Concentration Assay

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 3 Distribution of cells (mean ± SD) in the different phases of cell cycle in non-treated control cells (CT) and after treatment with varying concentrations (0.1, 1, and 40 ng/ml) of IL-10 protein and anti-IL-10Rα monoclonal antibody, or a single concentration of STAT3 inhibitor (2 µM) for 24 h in the (a) HT29 and (b) SW620 human colon cancer cell lines (mean ± SD; a = P < 0.01 compared to the CT group; b = P < 0.01 compared to 0.1 ng/mL of IL-10 protein; c = P < 0.01 compared to 1 ng/mL of IL-10 protein; d = P < 0.01 compared to 40 ng/mL of IL-10 protein; e = P < 0.01 compared to 0.1 ng/mL of anti-IL-10Rα antibody; f = P < 0.01 compared to 1 ng/mL of anti-IL-10Rα antibody; and g = P < 0.01 compared to 40 ng/mL of anti-IL-10Rα antibody)

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Control, Concentration Assay

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 4 a Heatmap showing relative mRNA expression (mean ± SD) of CCND1, CCND3, p21, and p27 genes alongside (b) their relative protein expression (mean ± SD) by Western blotting, in non-treated control cells (CT) and after treatment with varying concentrations (0.1, 1, and 40 ng/ml) of IL-10 protein and anti-IL-10Rα monoclonal antibody, or a single concentration of STAT3 inhibitor V (2 µM) for 24 h in the HT29 and SW620 human colon cancer cell lines (mean ± SD; a = P < 0.01 compared to the CT group; b = P < 0.01 compared to 0.1 ng/mL of IL-10 protein; c = P < 0.01 compared to 1 ng/mL of IL-10 protein; d = P < 0.01 compared to 40 ng/mL of IL-10 protein; e = P < 0.01 compared to 0.1 ng/mL of anti-IL-10Rα antibody; f = P < 0.01 compared to 1 ng/mL of anti-IL-10Rα antibody; and g = P < 0.01 compared to 40 ng/mL of anti-IL-10Rα antibody). The uncropped Western blot membranes for CCND1, CCND3, p21, and p27 proteins are provided in Suppl. Figure 4

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Expressing, Western Blot, Control, Concentration Assay

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 5 Percentage (mean ± SD) of living, early and late apoptotic, and dead cells in non-treated control cells (CT) and after treatment with varying concentrations (0.1, 1, and 40 ng/ml) of IL-10 protein and anti-IL-10Rα monoclonal antibody, or a single concentration of STAT3 inhibitor V (2 µM) for 24 h in the (a) HT29 and (b) SW620 human colon cancer cell lines (mean ± SD; a = P < 0.01 compared to the CT group; b = P < 0.01 compared to 0.1 ng/mL of IL-10 protein; c = P < 0.01 compared to 1 ng/mL of IL-10 protein; d = P < 0.01 compared to 40 ng/mL of IL-10 protein; e = P < 0.01 compared to 0.1 ng/mL of anti-IL-10Rα antibody; f = P < 0.01 compared to 1 ng/mL of anti-IL-10Rα antibody; and g = P < 0.01 compared to 40 ng/mL of anti-IL-10Rα antibody)

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Control, Concentration Assay

Journal: Discover oncology

Article Title: Increased protein expression of interleukin-10 and its signalling molecules in colon cancer progression: potential prognostic and therapeutic targets.

doi: 10.1007/s12672-025-02452-z

Figure Lengend Snippet: Fig. 6 a Heatmap showing relative mRNA expression (mean ± SD) of PCNA, survivin, and caspase-3 genes alongside (b) their relative protein expression (mean ± SD) by Western blotting, in non-treated control cells (CT) and after treatment with varying concentrations (0.1, 1, and 40 ng/ml) of IL-10 protein and anti-IL-10Rα monoclonal antibody, or a single concentration of STAT3 inhibitor V (2 µM) for 24 h in the HT29 and SW620 human colon cancer cell lines (mean ± SD; a = P < 0.01 compared to the CT group; b = P < 0.01 compared to 0.1 ng/mL of IL-10 protein; c = P < 0.01 compared to 1 ng/mL of IL-10 protein; d = P < 0.01 compared to 40 ng/mL of IL-10 protein; e = P < 0.01 compared to 0.1 ng/mL of anti-IL-10Rα antibody; f = P < 0.01 compared to 1 ng/mL of anti-IL-10Rα antibody; g = P < 0.01 compared to 40 ng/mL of anti-IL-10Rα antibody). The uncropped Western blot membranes for PCNA, survivin, and caspase-3 proteins are provided in Suppl. Figure 4

Article Snippet: The mouse monoclonal IgG antibody (#MAB274) that blocks IL-10Rα was purchased from R&D Systems Inc (Minneapolis, MN, USA).

Techniques: Expressing, Western Blot, Control, Concentration Assay

Journal: Nature Communications

Article Title: Adipocyte CLDN5 promotes thermogenesis and energy expenditure through regulation of IL10 expression

doi: 10.1038/s41467-025-61371-3

Figure Lengend Snippet: a qRT-PCR analysis of Il10 in iWAT from Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice, n = 8. b Western blot analysis with densitometric quantification of IL10 in BAT and iWAT from Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice ( n = 3). c IL10 ELISA assays in culture medium collected from adipocytes isolated from BAT and iWAT of Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice ( n = 5). d , e qRT-PCR analysis of IL10 target genes in differentiated brown ( d ) and beige ( e ) adipocytes treated with dbcAMP while co-cultured with CD36 + primary adipocytes isolated from BAT and iWAT of Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice ( n = 4). f IL10 ELISA assays in culture medium collected from AAV-transfected adipocytes isolated from wild-type mice ( n = 4). g – j qRT-PCR analysis of IL10 target genes ( g and h , n = 3) and OCR ( i and j ) in differentiated brown and beige adipocytes treated with dbcAMP along with either an IL10 neutralizing antibody or control IgG while co-cultured with CD36 + primary adipocytes isolated from BAT and iWAT of Cldn5 flox/flox ; Fabp4 -Cre mice. i , IgG ( n = 3), anti-IL10 ( n = 4). j , IgG ( n = 4), anti-IL10 ( n = 5). k , l qRT-PCR analysis of IL10 target genes in differentiated brown ( k ) and beige ( l ) adipocytes treated with dbcAMP along with either an IL10Rα blocking antibody or control IgG while co-cultured with CD36 + primary adipocytes isolated from BAT and iWAT of Cldn5 flox/flox ; Fabp4 -Cre mice ( n = 3). Data are expressed as the mean ± SEM. Statistical analyses were performed using two-sided Mann–Whitney test (BAT in c ) or two-sided unpaired t test ( a , b , iWAT in c , d – l ). Ads, Adipocytes. Source data are provided as a Source Data file.

Article Snippet: In order to determine whether CLDN5-deficient adipocytes communicate with thermogenic cells through paracrine substances, CD36 + primary adipocytes isolated from Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice were co-cultured with the differentiated brown and beige adipocytes for 48 h. To assess whether CLDN5-depleted cells influence thermogenic cells through IL10, IL10 neutralizing antibody (Abcam, AB9969; 1 μg/mL) or IL10Rα blocking antibody (Novus, NBP3-07330; 1 μg/mL), along with their corresponding negative control antibodies IgG (1 μg/mL) were introduced into the CD36 + adipocyte culture medium.

Techniques: Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Isolation, Cell Culture, Transfection, Control, Blocking Assay, MANN-WHITNEY

Journal: Nature Communications

Article Title: Adipocyte CLDN5 promotes thermogenesis and energy expenditure through regulation of IL10 expression

doi: 10.1038/s41467-025-61371-3

Figure Lengend Snippet: a , b Schematic of experimental design in Cldn5 flox/flox ; Fabp4 -Cre mice, with in situ BAT ( a ) or iWAT ( b ) injections of Il10ra siRNA (si- Il10ra ) or control siRNA (si- Ctrl ), maintained at 4 °C for 7 days. c – g Analysis in BAT after siRNA injections: qRT-PCR analysis of Il10ra ( c , n = 5), Western blot analysis with densitometric quantification of PGC1α, IL10Rα, and UCP1 ( d , n = 3), qRT-PCR analysis of IL10 target genes ( e , n = 5), HE staining with lipid droplet size quantification ( f , n = 5), and rectal temperature ( g , n = 7 for si- Ctrl , n = 9 for si- Il10ra ). h - l Analysis in iWAT after siRNA injections: qRT-PCR analysis of Il10ra ( h , n = 5), Western blot with densitometric quantification analyses of PGC1α, IL10Rα, and UCP1 ( i , n = 3), qRT-PCR analyses of IL10 target genes ( j , n = 5), HE staining with lipid droplet size quantification ( k , n = 5), and rectal temperature ( l , n = 5 for si- Ctrl , n = 7 for si- Il10ra ). m, n Schematic of experimental design in Cldn5 flox/flox ; Fabp4 -Cre mice, with in situ BAT ( m ) or iWAT ( n ) injections of AAV-sh- Ctrl or AAV-sh- Il10 , housed for 3 weeks before 7 days at 4 °C. o - s Analysis in BAT AAV-injected mice: qRT-PCR analysis of Il10 in BAT, liver, and muscle ( o , n = 6), Western blot analysis with densitometric quantification of IL10, PGC1α, and UCP1 ( p , n = 3), qRT-PCR analysis of IL10 target genes ( q , n = 6), HE staining with lipid droplet size quantification ( r , n = 5), rectal temperature ( s , n = 10). t –x Analysis in iWAT AAV-injected mice: qRT-PCR analysis of Il10 in iWAT, liver, and muscle ( t , n = 6), Western blot analysis with densitometric quantification of IL10, PGC1α, and UCP1 ( u , n = 3), qRT-PCR analysis of IL10 target genes in iWAT ( v , n = 6 for AAV-sh- Ctrl , n = 7 for AAV-sh- Il10 ), HE staining with lipid droplet size quantification ( w , n = 5), and rectal temperature ( x , n = 10). Data are expressed as the mean ± SEM. Statistical analyses were performed using two-sided unpaired t test. Source data are provided as a Source Data file. Panels a and m were created in BioRender . Panels b and n were created in BioRender .

Article Snippet: In order to determine whether CLDN5-deficient adipocytes communicate with thermogenic cells through paracrine substances, CD36 + primary adipocytes isolated from Cldn5 flox/flox and Cldn5 flox/flox ; Fabp4 -Cre mice were co-cultured with the differentiated brown and beige adipocytes for 48 h. To assess whether CLDN5-depleted cells influence thermogenic cells through IL10, IL10 neutralizing antibody (Abcam, AB9969; 1 μg/mL) or IL10Rα blocking antibody (Novus, NBP3-07330; 1 μg/mL), along with their corresponding negative control antibodies IgG (1 μg/mL) were introduced into the CD36 + adipocyte culture medium.

Techniques: In Situ, Control, Quantitative RT-PCR, Western Blot, Staining, Injection