il 4 (R&D Systems)

R&D Systems il 4

a Serum IL-31 levels were determined in dcSSc patients ( n = 55), lcSSc patients ( n = 19), and healthy controls ( n = 14) by a specific <t>ELISA</t> kit. The horizontal lines represent the mean values. The broken line represents the cut-off value (mean + 2 SD of the healthy controls). ** p < 0.01 and *** p < 0.001 vs. healthy controls. Significance was evaluated by two-tailed Mann–Whitney U test. Exact p values = 0.00002 (SSc vs. Ctrl), 0.000008 (dcSSc vs. Ctrl), 0.008 (lcSSc vs. Ctrl). Ctrl; healthy controls. b Correlations between serum IL-31 levels and the following parameters were analyzed in SSc patients ( n = 74): MRSS, %VC, %DLco, serum IL-4 levels, serum IL-6 levels, and serum IL-13 levels. Correlations were assessed by Spearman’s rank correlation test. Exact p values = 0.000000005 (MRSS); 0.005 (%VC); 0.033 (%DLco); 0.012 (IL-4); 0.00003 (IL-6); 0.026 (IL-13). Source data are provided as a Source Data file.

Il 4, supplied by R&D Systems, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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hil 4 (R&D Systems)

R&D Systems hil 4

Hil 4, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/hil 4/product/R&D Systems
Average 94 stars, based on 1 article reviews

hil 4 - by Bioz Stars, 2026-03

94/100 stars

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a Serum IL-31 levels were determined in dcSSc patients ( n = 55), lcSSc patients ( n = 19), and healthy controls ( n = 14) by a specific ELISA kit. The horizontal lines represent the mean values. The broken line represents the cut-off value (mean + 2 SD of the healthy controls). ** p < 0.01 and *** p < 0.001 vs. healthy controls. Significance was evaluated by two-tailed Mann–Whitney U test. Exact p values = 0.00002 (SSc vs. Ctrl), 0.000008 (dcSSc vs. Ctrl), 0.008 (lcSSc vs. Ctrl). Ctrl; healthy controls. b Correlations between serum IL-31 levels and the following parameters were analyzed in SSc patients ( n = 74): MRSS, %VC, %DLco, serum IL-4 levels, serum IL-6 levels, and serum IL-13 levels. Correlations were assessed by Spearman’s rank correlation test. Exact p values = 0.000000005 (MRSS); 0.005 (%VC); 0.033 (%DLco); 0.012 (IL-4); 0.00003 (IL-6); 0.026 (IL-13). Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a Serum IL-31 levels were determined in dcSSc patients ( n = 55), lcSSc patients ( n = 19), and healthy controls ( n = 14) by a specific ELISA kit. The horizontal lines represent the mean values. The broken line represents the cut-off value (mean + 2 SD of the healthy controls). ** p < 0.01 and *** p < 0.001 vs. healthy controls. Significance was evaluated by two-tailed Mann–Whitney U test. Exact p values = 0.00002 (SSc vs. Ctrl), 0.000008 (dcSSc vs. Ctrl), 0.008 (lcSSc vs. Ctrl). Ctrl; healthy controls. b Correlations between serum IL-31 levels and the following parameters were analyzed in SSc patients ( n = 74): MRSS, %VC, %DLco, serum IL-4 levels, serum IL-6 levels, and serum IL-13 levels. Correlations were assessed by Spearman’s rank correlation test. Exact p values = 0.000000005 (MRSS); 0.005 (%VC); 0.033 (%DLco); 0.012 (IL-4); 0.00003 (IL-6); 0.026 (IL-13). Source data are provided as a Source Data file.

Article Snippet: Human samples were analyzed using the specific ELISA kits for IL-4 (D4050, R&D Systems), IL-6 (D6050, R&D Systems), IL-13 (D1300B, R&D Systems), TGF-β1 (DB100B, R&D Systems), CTGF (DY9190-05, R&D Systems), MMP-9 (DMP900, R&D Systems), IL-33 (D3300B, R&D Systems), CCL2 (DCP00, R&D Systems), IL-31 (ab119546, abcam), MMP-1 (ab215083, abcam), IL-31RA (MBS923904, MyBioSource, San Diego, CA, USA), and type I collagen (EC1-E205, Adjusted Cell Experiment Laboratory, Kanagawa, Japan).

Techniques: Enzyme-linked Immunosorbent Assay, Two Tailed Test, MANN-WHITNEY

a Relative mRNA expression levels of Il31 and Il31ra were analyzed by real-time PCR in the skin of SSc patients ( n = 10) and healthy controls ( n = 6). Exact p values = 0.0002 ( Il31 ); 0.0002 ( Il31ra ). Relative fold differences = 2.37 ( Il31 ); 3.89 ( Il31ra ). b Representative double-immunofluorescence images for IL-31 (top, red), IL-31RA (bottom, red), FSP-1 (green), and nuclei (DAPI, blue) in the skin of SSc patients and healthy controls (scale bar=20 µm). The results shown are representative of three independent experiments with similar results. c , d . Relative mRNA levels ( c ) and protein levels ( d ) of IL-31 and IL-31RA in DFs from SSc patients and healthy controls ( n = 7, respectively) were assessed by real-time PCR and ELISA, respectively. Exact p values = 0.007 ( Il31 ); 0.0006 ( Il31ra ); 0.004 (IL-31); 0.0006 (IL-31RA). Relative fold differences = 2.05 ( Il31 ); 3.13 ( Il31ra ). e. Relative mRNA expression levels of Il31 and Il31ra were examined by real-time PCR in SSc DFs stimulated with IL-4 or IL-13 ( n = 5, respectively). Exact p values (IL-4 0.1 ng/ml vs. media, IL-4 1 ng/ml vs. media, IL-4 10 ng/ml vs. media, IL-13 0.1 ng/ml vs. media, IL-13 1 ng/ml vs. media, IL-13 10 ng/ml vs. media) = 0.841, 0.008, 0.008, 0.151, 0.841, 0.548 ( Il31 ); 0.310, 0.008, 0.008, 0.999, 0.008, 0.008 ( Il31ra ). Relative fold differences (IL-4 0.1 ng/ml, IL-4 1 ng/ml, IL-4 10 ng/ml, IL-13 0.1 ng/ml, IL-13 1 ng/ml, IL-13 10 ng/ml) = 1.02, 2.92, 6.78, 1.13, 0.98, 1.08 ( Il31 ); 1.16, 5.36, 13.91, 1.01, 3.15, 4.76 ( Il31ra ). Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. healthy controls ( a , c , and d ) or media ( e ). Significance was determined by two-tailed Mann-Whitney U test. Ctrl, healthy controls. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a Relative mRNA expression levels of Il31 and Il31ra were analyzed by real-time PCR in the skin of SSc patients ( n = 10) and healthy controls ( n = 6). Exact p values = 0.0002 ( Il31 ); 0.0002 ( Il31ra ). Relative fold differences = 2.37 ( Il31 ); 3.89 ( Il31ra ). b Representative double-immunofluorescence images for IL-31 (top, red), IL-31RA (bottom, red), FSP-1 (green), and nuclei (DAPI, blue) in the skin of SSc patients and healthy controls (scale bar=20 µm). The results shown are representative of three independent experiments with similar results. c , d . Relative mRNA levels ( c ) and protein levels ( d ) of IL-31 and IL-31RA in DFs from SSc patients and healthy controls ( n = 7, respectively) were assessed by real-time PCR and ELISA, respectively. Exact p values = 0.007 ( Il31 ); 0.0006 ( Il31ra ); 0.004 (IL-31); 0.0006 (IL-31RA). Relative fold differences = 2.05 ( Il31 ); 3.13 ( Il31ra ). e. Relative mRNA expression levels of Il31 and Il31ra were examined by real-time PCR in SSc DFs stimulated with IL-4 or IL-13 ( n = 5, respectively). Exact p values (IL-4 0.1 ng/ml vs. media, IL-4 1 ng/ml vs. media, IL-4 10 ng/ml vs. media, IL-13 0.1 ng/ml vs. media, IL-13 1 ng/ml vs. media, IL-13 10 ng/ml vs. media) = 0.841, 0.008, 0.008, 0.151, 0.841, 0.548 ( Il31 ); 0.310, 0.008, 0.008, 0.999, 0.008, 0.008 ( Il31ra ). Relative fold differences (IL-4 0.1 ng/ml, IL-4 1 ng/ml, IL-4 10 ng/ml, IL-13 0.1 ng/ml, IL-13 1 ng/ml, IL-13 10 ng/ml) = 1.02, 2.92, 6.78, 1.13, 0.98, 1.08 ( Il31 ); 1.16, 5.36, 13.91, 1.01, 3.15, 4.76 ( Il31ra ). Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. healthy controls ( a , c , and d ) or media ( e ). Significance was determined by two-tailed Mann-Whitney U test. Ctrl, healthy controls. Source data are provided as a Source Data file.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Two Tailed Test, MANN-WHITNEY

a , b DFs from dcSSc patients and healthy controls were cultured for 24 h with rhIL-31 (0, 10, 25, 50, or 100 ng/ml) to examine Col1a1 and Col1a2 mRNA expression ( a ) and type I collagen production ( b ) by real-time PCR and ELISA, respectively. Exact p values (Ctrl + IL-31 10 ng/ml vs. Ctrl, Ctrl + IL-31 25 ng/ml vs. Ctrl, Ctrl + IL-31 50 ng/ml vs. Ctrl, Ctrl + IL-31 100 ng/ml vs. Ctrl, SSc + IL-31 10 ng/ml vs. SSc, SSc + IL-31 25 ng/ml vs. SSc, SSc + IL-31 50 ng/ml vs. SSc, SSc + IL-31 100 ng/ml vs. SSc) = 0.690, 0.095, 0.056, 0.008, 0.032, 0.016, 0.008, 0.008 ( Col1a1 ); 0.421, 0.421, 0.222, 0.016, 0.008, 0.008, 0.008, 0.008 ( Col1a2 ); 0.508, 0.127, 0.841, 0.008, 0.008, 0.008, 0.008, 0.008 (Type I collagen). Relative fold differences (Ctrl + IL-31 10 ng/ml, Ctrl + IL-31 25 ng/ml, Ctrl + IL-31 50 ng/ml, Ctrl + IL-31 100 ng/ml, SSc + IL-31 10 ng/ml, SSc + IL-31 25 ng/ml, SSc + IL-31 50 ng/ml, SSc + IL-31 100 ng/ml) = 1.10, 1.16, 1.19, 1.53, 1.47, 2.10, 2.24, 2.71, 2.86 ( Col1a1 ); 1.11, 1.10, 1.31, 1.54, 1.83, 2.35, 2.54, 2.75, 3.25 ( Col1a2 ). c Protein levels of TGF-β1, CTGF, MMP-1, MMP-3, and MMP-9 were assessed by ELISA in the supernatants of dcSSc DFs treated with or without rhIL-31 (50 ng/ml). Exact p values = 0.008 (TGF-β1), 0.587 (CTGF), 0.008 (MMP-1), 0.016 (MMP-3), 0.024 (MMP-9). d Representative western blot analysis of α-SMA in dcSSc DFs treated with or without rhIL-31 (50 ng/ml; left). Quantification of protein expression normalized to β-actin (right). Exact p value = 0.008. e Relative mRNA expression level of Acta2 was evaluated by real-time PCR in dcSSc DFs treated with or without rhIL-31 (50 ng/ml). Exact p value = 0.008. Relative fold difference = 2.85. f BrdU incorporation was quantified by ELISA in dcSSc DFs treated with or without rhIL-31 (50 ng/ml). The absorbance at 450 nm was measured. Exact p value = 0.008. g DFs from dcSSc patients were treated with or without rhIL-31 (50 ng/ml) and analyzed for apoptosis by flow cytometry. Representative dot plots were shown (left). Annexin-V + , 7-AAD − cells were considered early apoptotic cells, and Annexin-V + , 7-AAD + cells were considered late apoptotic cells, respectively (right). Exact p values (IL-31 vs. media) = 0.008 (early apoptosis); 0.008 (late apoptosis). h DFs from dcSSc patients and healthy controls were pretreated with mitomycin C, scratched to make a wound, and incubated for 24 h with rhIL-31 (50 ng/ml) or media alone. Representative microscopic images were shown (left, scale bar=500 µm). Red lines show the borders of the wounds. Wound closure was expressed as the percentage of wound reduction from the original wound (right). Exact p values (Ctrl + media vs. Ctrl + IL-31, Ctrl + media vs. SSc + media, Ctrl + media vs. SSc + IL-31, Ctrl + IL-31 vs. SSc + media, Ctrl + IL-31 vs. SSc + IL-31, SSc + media vs. SSc + IL-31) = 0.005, 0.0000001, 0.00000000006, 0.00008, 0.000000004, 0.00002. i , j DFs obtained from dcSSc patients were cultured with rhIL-31 (50 ng/ml) or media alone for 24 h, and expression levels of IL-6, IL-33, and CCL2 were measured by real-time PCR ( i ) and ELISA ( j ). Exact p values = 0.008 ( Il6 ); 0.032 ( Il33 ); 0.008 ( Ccl2 ); 0.008 (IL-6); 0.032 (IL-33); 0.008 (CCL2). Relative fold differences = 1.99 ( Il6 ); 1.37 ( Il33 ); 2.39 ( Ccl2 ). n = 5. Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. unstimulated Ctrl or SSc fibroblasts ( a , b ) or media ( c–g , i , and j ). Significance was determined by two-tailed Mann-Whitney U test ( a–g , i , and j ) and one-way analysis of variance followed by Tukey’s post hoc comparison test (h). Ctrl, healthy controls; OD, optical density. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a , b DFs from dcSSc patients and healthy controls were cultured for 24 h with rhIL-31 (0, 10, 25, 50, or 100 ng/ml) to examine Col1a1 and Col1a2 mRNA expression ( a ) and type I collagen production ( b ) by real-time PCR and ELISA, respectively. Exact p values (Ctrl + IL-31 10 ng/ml vs. Ctrl, Ctrl + IL-31 25 ng/ml vs. Ctrl, Ctrl + IL-31 50 ng/ml vs. Ctrl, Ctrl + IL-31 100 ng/ml vs. Ctrl, SSc + IL-31 10 ng/ml vs. SSc, SSc + IL-31 25 ng/ml vs. SSc, SSc + IL-31 50 ng/ml vs. SSc, SSc + IL-31 100 ng/ml vs. SSc) = 0.690, 0.095, 0.056, 0.008, 0.032, 0.016, 0.008, 0.008 ( Col1a1 ); 0.421, 0.421, 0.222, 0.016, 0.008, 0.008, 0.008, 0.008 ( Col1a2 ); 0.508, 0.127, 0.841, 0.008, 0.008, 0.008, 0.008, 0.008 (Type I collagen). Relative fold differences (Ctrl + IL-31 10 ng/ml, Ctrl + IL-31 25 ng/ml, Ctrl + IL-31 50 ng/ml, Ctrl + IL-31 100 ng/ml, SSc + IL-31 10 ng/ml, SSc + IL-31 25 ng/ml, SSc + IL-31 50 ng/ml, SSc + IL-31 100 ng/ml) = 1.10, 1.16, 1.19, 1.53, 1.47, 2.10, 2.24, 2.71, 2.86 ( Col1a1 ); 1.11, 1.10, 1.31, 1.54, 1.83, 2.35, 2.54, 2.75, 3.25 ( Col1a2 ). c Protein levels of TGF-β1, CTGF, MMP-1, MMP-3, and MMP-9 were assessed by ELISA in the supernatants of dcSSc DFs treated with or without rhIL-31 (50 ng/ml). Exact p values = 0.008 (TGF-β1), 0.587 (CTGF), 0.008 (MMP-1), 0.016 (MMP-3), 0.024 (MMP-9). d Representative western blot analysis of α-SMA in dcSSc DFs treated with or without rhIL-31 (50 ng/ml; left). Quantification of protein expression normalized to β-actin (right). Exact p value = 0.008. e Relative mRNA expression level of Acta2 was evaluated by real-time PCR in dcSSc DFs treated with or without rhIL-31 (50 ng/ml). Exact p value = 0.008. Relative fold difference = 2.85. f BrdU incorporation was quantified by ELISA in dcSSc DFs treated with or without rhIL-31 (50 ng/ml). The absorbance at 450 nm was measured. Exact p value = 0.008. g DFs from dcSSc patients were treated with or without rhIL-31 (50 ng/ml) and analyzed for apoptosis by flow cytometry. Representative dot plots were shown (left). Annexin-V + , 7-AAD − cells were considered early apoptotic cells, and Annexin-V + , 7-AAD + cells were considered late apoptotic cells, respectively (right). Exact p values (IL-31 vs. media) = 0.008 (early apoptosis); 0.008 (late apoptosis). h DFs from dcSSc patients and healthy controls were pretreated with mitomycin C, scratched to make a wound, and incubated for 24 h with rhIL-31 (50 ng/ml) or media alone. Representative microscopic images were shown (left, scale bar=500 µm). Red lines show the borders of the wounds. Wound closure was expressed as the percentage of wound reduction from the original wound (right). Exact p values (Ctrl + media vs. Ctrl + IL-31, Ctrl + media vs. SSc + media, Ctrl + media vs. SSc + IL-31, Ctrl + IL-31 vs. SSc + media, Ctrl + IL-31 vs. SSc + IL-31, SSc + media vs. SSc + IL-31) = 0.005, 0.0000001, 0.00000000006, 0.00008, 0.000000004, 0.00002. i , j DFs obtained from dcSSc patients were cultured with rhIL-31 (50 ng/ml) or media alone for 24 h, and expression levels of IL-6, IL-33, and CCL2 were measured by real-time PCR ( i ) and ELISA ( j ). Exact p values = 0.008 ( Il6 ); 0.032 ( Il33 ); 0.008 ( Ccl2 ); 0.008 (IL-6); 0.032 (IL-33); 0.008 (CCL2). Relative fold differences = 1.99 ( Il6 ); 1.37 ( Il33 ); 2.39 ( Ccl2 ). n = 5. Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. unstimulated Ctrl or SSc fibroblasts ( a , b ) or media ( c–g , i , and j ). Significance was determined by two-tailed Mann-Whitney U test ( a–g , i , and j ) and one-way analysis of variance followed by Tukey’s post hoc comparison test (h). Ctrl, healthy controls; OD, optical density. Source data are provided as a Source Data file.

Techniques: Cell Culture, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Western Blot, BrdU Incorporation Assay, Flow Cytometry, Incubation, Two Tailed Test, MANN-WHITNEY, Comparison

a Representative western blot analysis of STAT1, STAT3, STAT5 and their phosphorylated forms in dcSSc DFs treated with rhIL-31 (50 ng/ml; left). Quantification of protein expression normalized to β-actin (right). Exact p values (10 min vs. 0 min, 20 min vs. 0 min) = 0.008, 0.008 (pSTAT1/STAT1); 0.008, 0.008 (pSTAT3/STAT3); 0.016, 0.040 (pSTAT5/STAT5). Relative fold differences (10 min, 20 min) = 3.63, 3.34 (pSTAT1/STAT1); 2.11, 7.23 (pSTAT3/STAT3); 1.36, 1.35 (pSTAT5/STAT5). b Representative ChIP assay on dcSSc DFs assessing the binding of phosphorylated STAT3 to the Col1a2 enhancer (left). Immunoprecipitated chromatin was analyzed by real-time PCR using specific primers for the HS4 region of the Col1a2 enhancer (right). Exact p value = 0.029. n = 4 biologically independent experiments. c Protein levels of type I collagen, IL-6, IL-33, and CCL2 were evaluated by ELISA in the supernatants of dcSSc DFs that were transfected with IL-31RA siRNA and treated with rhIL-31 (50 ng/ml). Exact p values (media vs. IL-31 + Ctrl siRNA, media vs. IL-31 + IL-31RA siRNA, IL-31 + Ctrl siRNA vs. IL-31 + IL-31RA siRNA) = 0.00000009, 0.111, 0.0000008 (Type I collagen); 0.0000009, 0.202, 0.000007 (IL-6); 0.002, 0.766, 0.005 (IL-33); 0.000003, 0.222, 0.00002 (CCL2). d Protein levels of Type I collagen were measured by ELISA in the supernatants of dcSSc DFs that were transfected with OSMR siRNA or treated with inhibitors for STAT1 (STAT1-I: Fludarabine; 50 µM), STAT3 (STAT-I: Stattic; 5 µM), or STAT5 (STAT5-I: CAS 285986-31-4; 50 µM), followed by the treatment with rhIL-31 (50 ng/ml). Exact p values = 0.238 (IL-31 + OSMR siRNA vs. IL-31), 0.175 (IL-31 + STAT1-I vs. IL-31), 0.008 (IL-31 + STAT3-I vs. IL-31), 0.548 (IL-31 + STAT5-I vs. IL-31). n = 5 biologically independent experiments, unless otherwise noted. Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using two-tailed Mann-Whitney U test ( a , b , and d) and one-way analysis of variance followed by Tukey’s post hoc comparison test ( c ). pSTAT1, phosphorylated STAT1; pSTAT3, phosphorylated STAT3; pSTAT5, phosphorylated STAT5. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a Representative western blot analysis of STAT1, STAT3, STAT5 and their phosphorylated forms in dcSSc DFs treated with rhIL-31 (50 ng/ml; left). Quantification of protein expression normalized to β-actin (right). Exact p values (10 min vs. 0 min, 20 min vs. 0 min) = 0.008, 0.008 (pSTAT1/STAT1); 0.008, 0.008 (pSTAT3/STAT3); 0.016, 0.040 (pSTAT5/STAT5). Relative fold differences (10 min, 20 min) = 3.63, 3.34 (pSTAT1/STAT1); 2.11, 7.23 (pSTAT3/STAT3); 1.36, 1.35 (pSTAT5/STAT5). b Representative ChIP assay on dcSSc DFs assessing the binding of phosphorylated STAT3 to the Col1a2 enhancer (left). Immunoprecipitated chromatin was analyzed by real-time PCR using specific primers for the HS4 region of the Col1a2 enhancer (right). Exact p value = 0.029. n = 4 biologically independent experiments. c Protein levels of type I collagen, IL-6, IL-33, and CCL2 were evaluated by ELISA in the supernatants of dcSSc DFs that were transfected with IL-31RA siRNA and treated with rhIL-31 (50 ng/ml). Exact p values (media vs. IL-31 + Ctrl siRNA, media vs. IL-31 + IL-31RA siRNA, IL-31 + Ctrl siRNA vs. IL-31 + IL-31RA siRNA) = 0.00000009, 0.111, 0.0000008 (Type I collagen); 0.0000009, 0.202, 0.000007 (IL-6); 0.002, 0.766, 0.005 (IL-33); 0.000003, 0.222, 0.00002 (CCL2). d Protein levels of Type I collagen were measured by ELISA in the supernatants of dcSSc DFs that were transfected with OSMR siRNA or treated with inhibitors for STAT1 (STAT1-I: Fludarabine; 50 µM), STAT3 (STAT-I: Stattic; 5 µM), or STAT5 (STAT5-I: CAS 285986-31-4; 50 µM), followed by the treatment with rhIL-31 (50 ng/ml). Exact p values = 0.238 (IL-31 + OSMR siRNA vs. IL-31), 0.175 (IL-31 + STAT1-I vs. IL-31), 0.008 (IL-31 + STAT3-I vs. IL-31), 0.548 (IL-31 + STAT5-I vs. IL-31). n = 5 biologically independent experiments, unless otherwise noted. Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using two-tailed Mann-Whitney U test ( a , b , and d) and one-way analysis of variance followed by Tukey’s post hoc comparison test ( c ). pSTAT1, phosphorylated STAT1; pSTAT3, phosphorylated STAT3; pSTAT5, phosphorylated STAT5. Source data are provided as a Source Data file.

Techniques: Western Blot, Expressing, Binding Assay, Immunoprecipitation, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Transfection, Two Tailed Test, MANN-WHITNEY, Comparison

a BLM-SSc and PBS-treated control (PBS-Ctrl) mice were administered with either rmIL-31 or saline as a sham daily from day 1 to 14, and were analyzed on day 15. b Representative histological sections of the skin and lungs stained with hematoxylin and eosin (left) and Masson trichrome (right) were shown (horizontal scale bars=100 μm in skin; 20 µm in lung). Vertical bars with arrows represent dermal thickness. Dermal thickness and lung fibrosis score were assessed histologically. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.003, 0.00001, 0.0000000005, 0.049, 0.00000006, 0.000004 (dermal thickness); 0.009, 0.0000003, 0.00000000004, 0.0001, 0.000000001, 0.000003 (lung fibrosis score). c Hydroxyproline contents of skin and lung samples. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.036, 0.00007, 0.0000001, 0.027, 0.00001, 0.004 (skin); 0.015, 0.00003, 0.0000001, 0.025, 0.00002, 0.011 (lung). d Relative mRNA expression levels of Col1a1 and Col1a2 in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.004, 0.000000002, 0.00000000004, 0.0000004, 0.000000003, 0.005 ( Col1a1 , skin); 0.033, 0.000000002, 0.000000000002, 0.00000006, 0.00000000002, 0.000005 ( Col1a2 , skin); 0.046, 0.00004, 0.00000001, 0.011, 0.0000006, 0.0003 ( Col1a1 , lung); 0.006, 0.000002, 0.000000007, 0.002, 0.000001, 0.004 ( Col1a2 , lung). Relative fold differences (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.86, 3.78, 4.61 ( Col1a1 , skin); 1.86, 4.87, 7.03 ( Col1a2 , skin); 1.47, 2.05, 2.91 ( Col1a1 , lung); 1.54, 2.14, 2.69 ( Col1a2 , lung). e Protein levels of type I collagen in the skin and lungs were assessed by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.048, 0.00004, 0.00000001, 0.011, 0.0000006, 0.0003 (skin); 0.029, 0.000005, 0.000000004, 0.002, 0.0000002, 0.0004 (lung). f Relative mRNA expression levels of Il31 and Il31ra in the skin and lungs were evaluated by real-time PCR. Exact p values = 0.008 ( Il31 , skin); 0.008 ( Il31ra , skin); 0.008 ( Il31 , lung); 0.008 ( Il31ra , lung). Relative fold differences = 5.49 ( Il31 , skin); 3.48 ( Il31ra , skin); 2.51 ( Il31 , lung); 1.98 ( Il31ra , lung). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test ( b – e ) and two-tailed Mann-Whitney U test (f). The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a BLM-SSc and PBS-treated control (PBS-Ctrl) mice were administered with either rmIL-31 or saline as a sham daily from day 1 to 14, and were analyzed on day 15. b Representative histological sections of the skin and lungs stained with hematoxylin and eosin (left) and Masson trichrome (right) were shown (horizontal scale bars=100 μm in skin; 20 µm in lung). Vertical bars with arrows represent dermal thickness. Dermal thickness and lung fibrosis score were assessed histologically. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.003, 0.00001, 0.0000000005, 0.049, 0.00000006, 0.000004 (dermal thickness); 0.009, 0.0000003, 0.00000000004, 0.0001, 0.000000001, 0.000003 (lung fibrosis score). c Hydroxyproline contents of skin and lung samples. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.036, 0.00007, 0.0000001, 0.027, 0.00001, 0.004 (skin); 0.015, 0.00003, 0.0000001, 0.025, 0.00002, 0.011 (lung). d Relative mRNA expression levels of Col1a1 and Col1a2 in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.004, 0.000000002, 0.00000000004, 0.0000004, 0.000000003, 0.005 ( Col1a1 , skin); 0.033, 0.000000002, 0.000000000002, 0.00000006, 0.00000000002, 0.000005 ( Col1a2 , skin); 0.046, 0.00004, 0.00000001, 0.011, 0.0000006, 0.0003 ( Col1a1 , lung); 0.006, 0.000002, 0.000000007, 0.002, 0.000001, 0.004 ( Col1a2 , lung). Relative fold differences (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.86, 3.78, 4.61 ( Col1a1 , skin); 1.86, 4.87, 7.03 ( Col1a2 , skin); 1.47, 2.05, 2.91 ( Col1a1 , lung); 1.54, 2.14, 2.69 ( Col1a2 , lung). e Protein levels of type I collagen in the skin and lungs were assessed by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.048, 0.00004, 0.00000001, 0.011, 0.0000006, 0.0003 (skin); 0.029, 0.000005, 0.000000004, 0.002, 0.0000002, 0.0004 (lung). f Relative mRNA expression levels of Il31 and Il31ra in the skin and lungs were evaluated by real-time PCR. Exact p values = 0.008 ( Il31 , skin); 0.008 ( Il31ra , skin); 0.008 ( Il31 , lung); 0.008 ( Il31ra , lung). Relative fold differences = 5.49 ( Il31 , skin); 3.48 ( Il31ra , skin); 2.51 ( Il31 , lung); 1.98 ( Il31ra , lung). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test ( b – e ) and two-tailed Mann-Whitney U test (f). The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Techniques: Control, Saline, Staining, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Comparison, Two Tailed Test, MANN-WHITNEY

a Relative mRNA expression levels of Il4 , Il6 , Il10 , Il17a , Tnf , Tgfb1 , and Ifng in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.866, 0.039, 0.00006, 0.162, 0.0003, 0.021 ( Il4 , skin); 0.403, 0.00000000006, 0.0000000000002, 0.0000000003, 0.0000000000004, 0.000001 ( Il6 , skin); 0.034, 0.004, 0.000003, 0.699, 0.0007, 0.007 ( Il10 , skin); 0.926, 0.000007, 0.0000005, 0.00002, 0.000001, 0.344 ( Il17a , skin); 0.977, 0.012, 0.063, 0.005, 0.029, 0.832 ( Tnf , skin); 0.572, 0.0001, 0.00000001, 0.001, 0.00000008, 0.0001 ( Tgfb1 , skin); 0.428, 0.00007, 0.002, 0.001, 0.040, 0.361 ( Ifng , skin); 0.00008, 0.003, 0.0000002, 0.286, 0.008, 0.0002 ( Il4 , lung); 0.023, 0.00001, 0.0000001, 0.007, 0.00002, 0.038 ( Il6 , lung); 0.002, 0.227, 0.00003, 0.085, 0.231, 0.002 ( Il10 , lung); 0.192, 0.000008, 0.000001, 0.0004, 0.00005, 0.686 ( Il17a , lung); 0.641, 0.013, 0.041, 0.127, 0.321, 0.937 ( Tnf , lung); 0.967, 0.0002, 0.0000005, 0.0005, 0.000001, 0.014 ( Tgfb1 , lung); 0.973, 0.00003, 0.0001, 0.00007, 0.0003, 0.868 ( Ifng , lung). Relative fold differences (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.55, 3.12, 5.45 ( Il4 , skin); 2.14, 13.09, 19.11 ( Il6 , skin); 2.59, 3.15, 5.16 ( Il10 , skin); 1.22, 3.63, 4.24 ( Il17a , skin); 0.89, 1.97, 1.74 ( Tnf , skin); 1.34, 2.57, 4.12 ( Tgfb1 , skin); 1.68, 3.71, 2.98 ( Ifng , skin); 3.59, 2.81, 5.20 ( Il4 , lung); 5.56, 10.92, 15.12 ( Il6 , lung); 2.29, 1.56, 2.85 ( Il10 , lung); 2.35, 5.74, 6.45 ( Il17a , lung); 1.27, 1.81, 1.68 ( Tnf , lung); 1.16, 2.98, 4.22 ( Tgfb1 , lung); 1.22, 4.34, 3.95 ( Ifng , lung). b Protein levels of TGF-β1 were evaluated by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.924, 0.00006, 0.0000003, 0.0002, 0.0000007, 0.022 (skin); 0.805, 0.00003, 0.0000004, 0.0002, 0.000002, 0.076 (lung). c , d Relative mRNA levels and protein levels of Ctgf were evaluated by real-time PCR ( c ) and ELISA ( d ), respectively. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.912, 0.000000003, 0.0000000005, 0.000000007, 0.000000001, 0.391 ( Ctgf , skin); 0.967, 0.00000007, 0.000000009, 0.0000001, 0.00000002, 0.393 ( Ctgf , lung); 0.972, 0.00006, 0.00002, 0.00003, 0.00001, 0.941 (CTGF, skin); 0.933, 0.005, 0.006, 0.017, 0.019, 0.999 (CTGF, lung). Relative fold difference (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.14, 3.79, 4.13 (skin); 1.12, 3.86, 4.30 (lung). e Relative mRNA expression levels of Mmp3 , Mmp9 , Mmp13 , Timp1 , Timp2 , and Timp3 were assessed in the lungs of BLM-SSc mice by real-time PCR. Exact p values = 0.008 ( Mmp3 ); 0.016 ( Mmp9 ); 0.008 ( Mmp13 ); 0.008 ( Timp1 ); 0.008 ( Timp2 ); 0.008 ( Timp3 ). Relative fold differences = 0.79 ( Mmp3 ); 0.72 ( Mmp9 ); 0.68 ( Mmp13 ); 2.06 ( Timp1 ); 1.42 ( Timp2 ); 1.26 ( Timp3 ). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test (a-d) and two-tailed Mann–Whitney U test ( e ). The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a Relative mRNA expression levels of Il4 , Il6 , Il10 , Il17a , Tnf , Tgfb1 , and Ifng in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.866, 0.039, 0.00006, 0.162, 0.0003, 0.021 ( Il4 , skin); 0.403, 0.00000000006, 0.0000000000002, 0.0000000003, 0.0000000000004, 0.000001 ( Il6 , skin); 0.034, 0.004, 0.000003, 0.699, 0.0007, 0.007 ( Il10 , skin); 0.926, 0.000007, 0.0000005, 0.00002, 0.000001, 0.344 ( Il17a , skin); 0.977, 0.012, 0.063, 0.005, 0.029, 0.832 ( Tnf , skin); 0.572, 0.0001, 0.00000001, 0.001, 0.00000008, 0.0001 ( Tgfb1 , skin); 0.428, 0.00007, 0.002, 0.001, 0.040, 0.361 ( Ifng , skin); 0.00008, 0.003, 0.0000002, 0.286, 0.008, 0.0002 ( Il4 , lung); 0.023, 0.00001, 0.0000001, 0.007, 0.00002, 0.038 ( Il6 , lung); 0.002, 0.227, 0.00003, 0.085, 0.231, 0.002 ( Il10 , lung); 0.192, 0.000008, 0.000001, 0.0004, 0.00005, 0.686 ( Il17a , lung); 0.641, 0.013, 0.041, 0.127, 0.321, 0.937 ( Tnf , lung); 0.967, 0.0002, 0.0000005, 0.0005, 0.000001, 0.014 ( Tgfb1 , lung); 0.973, 0.00003, 0.0001, 0.00007, 0.0003, 0.868 ( Ifng , lung). Relative fold differences (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.55, 3.12, 5.45 ( Il4 , skin); 2.14, 13.09, 19.11 ( Il6 , skin); 2.59, 3.15, 5.16 ( Il10 , skin); 1.22, 3.63, 4.24 ( Il17a , skin); 0.89, 1.97, 1.74 ( Tnf , skin); 1.34, 2.57, 4.12 ( Tgfb1 , skin); 1.68, 3.71, 2.98 ( Ifng , skin); 3.59, 2.81, 5.20 ( Il4 , lung); 5.56, 10.92, 15.12 ( Il6 , lung); 2.29, 1.56, 2.85 ( Il10 , lung); 2.35, 5.74, 6.45 ( Il17a , lung); 1.27, 1.81, 1.68 ( Tnf , lung); 1.16, 2.98, 4.22 ( Tgfb1 , lung); 1.22, 4.34, 3.95 ( Ifng , lung). b Protein levels of TGF-β1 were evaluated by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.924, 0.00006, 0.0000003, 0.0002, 0.0000007, 0.022 (skin); 0.805, 0.00003, 0.0000004, 0.0002, 0.000002, 0.076 (lung). c , d Relative mRNA levels and protein levels of Ctgf were evaluated by real-time PCR ( c ) and ELISA ( d ), respectively. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.912, 0.000000003, 0.0000000005, 0.000000007, 0.000000001, 0.391 ( Ctgf , skin); 0.967, 0.00000007, 0.000000009, 0.0000001, 0.00000002, 0.393 ( Ctgf , lung); 0.972, 0.00006, 0.00002, 0.00003, 0.00001, 0.941 (CTGF, skin); 0.933, 0.005, 0.006, 0.017, 0.019, 0.999 (CTGF, lung). Relative fold difference (PBS-Ctrl + IL-31, BLM-SSc + sham, BLM-SSc + IL-31) = 1.14, 3.79, 4.13 (skin); 1.12, 3.86, 4.30 (lung). e Relative mRNA expression levels of Mmp3 , Mmp9 , Mmp13 , Timp1 , Timp2 , and Timp3 were assessed in the lungs of BLM-SSc mice by real-time PCR. Exact p values = 0.008 ( Mmp3 ); 0.016 ( Mmp9 ); 0.008 ( Mmp13 ); 0.008 ( Timp1 ); 0.008 ( Timp2 ); 0.008 ( Timp3 ). Relative fold differences = 0.79 ( Mmp3 ); 0.72 ( Mmp9 ); 0.68 ( Mmp13 ); 2.06 ( Timp1 ); 1.42 ( Timp2 ); 1.26 ( Timp3 ). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test (a-d) and two-tailed Mann–Whitney U test ( e ). The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Comparison, Two Tailed Test, MANN-WHITNEY, Control

a – c The frequencies of Th1, Th2, Th17, and Treg cells in BLM-SSc and PBS-treated control (PBS-Ctrl) mice treated with either rmIL-31 or sham were analyzed by flow cytometry in spleen ( a ), lung ( b ), and lung-draining lymph nodes ( c ). Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.960, 0.417, 0.996, 0.145, 0.841, 0.625 (Th1, spleen); 0.017, 0.0002, 0.00000000007, 0.342, 0.00000001, 0.0000003 (Th2, spleen); 0.795, 0.005, 0.000006, 0.059, 0.00006, 0.041 (Th17, spleen); 0.001, 0.0000001, 0.008, 0.002, 0.926, 0.0003 (Treg, spleen); 0.521, 0.005, 0.00002, 0.079, 0.0002, 0.043 (Th2/Th1 ratio, spleen); 0.206, 0.00005, 0.0007, 0.003, 0.044, 0.521 (Th17/Treg ratio, spleen); 0.007, 0.011, 0.000004, 0.996, 0.006, 0.004 (Th2/Treg ratio, spleen); 0.966, 0.932, 0.996, 0.999, 0.904, 0.849 (Th1, lung); 0.018, 0.0003, 0.00000002, 0.233, 0.000003, 0.00009 (Th2, lung); 0.633, 0.005, 0.00002, 0.059, 0.0002, 0.039 (Th17, lung); 0.005, 0.0000002, 0.00002, 0.0002, 0.050, 0.048 (Treg, lung); 0.031, 0.0007, 0.0000002, 0.264, 0.00002, 0.0007 (Th2/Th1 ratio, lung); 0.363, 0.0002, 0.000003, 0.005, 0.00005, 0.125 (Th17/Treg ratio, lung); 0.034, 0.0002, 0.00000005, 0.082, 0.000004, 0.0005 (Th2/Treg ratio, lung); 0.939, 0.488, 0.961, 0.221, 0.719, 0.772 (Th1, lung-draining lymph nodes); 0.038, 0.000009, 0.00000000006, 0.002, 0.000000001, 0.0000003 (Th2, lung-draining lymph nodes); 0.839, 0.010, 0.00001, 0.051, 0.00005, 0.013 (Th17, lung-draining lymph nodes); 0.057, 0.000001, 0.001, 0.0002, 0.230, 0.009 (Treg, lung-draining lymph nodes); 0.225, 0.00008, 0.000000005, 0.004, 0.00000006, 0.00003 (Th2/Th1 ratio, lung-draining lymph nodes); 0.398, 0.00003, 0.000001, 0.0005, 0.00001, 0.244 (Th17/Treg ratio, lung-draining lymph nodes); 0.011, 0.00000007, 0.000000000007, 0.00002, 0.0000000002, 0.0000005 (Th2/Treg ratio, lung-draining lymph nodes). d Serum levels of IL-4 and IL-6 were assessed by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.129, 0.001, 0.000002, 0.103, 0.0001, 0.019 (IL-4); 0.085, 0.032, 0.00007, 0.954, 0.012, 0.033 (IL-6). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a – c The frequencies of Th1, Th2, Th17, and Treg cells in BLM-SSc and PBS-treated control (PBS-Ctrl) mice treated with either rmIL-31 or sham were analyzed by flow cytometry in spleen ( a ), lung ( b ), and lung-draining lymph nodes ( c ). Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.960, 0.417, 0.996, 0.145, 0.841, 0.625 (Th1, spleen); 0.017, 0.0002, 0.00000000007, 0.342, 0.00000001, 0.0000003 (Th2, spleen); 0.795, 0.005, 0.000006, 0.059, 0.00006, 0.041 (Th17, spleen); 0.001, 0.0000001, 0.008, 0.002, 0.926, 0.0003 (Treg, spleen); 0.521, 0.005, 0.00002, 0.079, 0.0002, 0.043 (Th2/Th1 ratio, spleen); 0.206, 0.00005, 0.0007, 0.003, 0.044, 0.521 (Th17/Treg ratio, spleen); 0.007, 0.011, 0.000004, 0.996, 0.006, 0.004 (Th2/Treg ratio, spleen); 0.966, 0.932, 0.996, 0.999, 0.904, 0.849 (Th1, lung); 0.018, 0.0003, 0.00000002, 0.233, 0.000003, 0.00009 (Th2, lung); 0.633, 0.005, 0.00002, 0.059, 0.0002, 0.039 (Th17, lung); 0.005, 0.0000002, 0.00002, 0.0002, 0.050, 0.048 (Treg, lung); 0.031, 0.0007, 0.0000002, 0.264, 0.00002, 0.0007 (Th2/Th1 ratio, lung); 0.363, 0.0002, 0.000003, 0.005, 0.00005, 0.125 (Th17/Treg ratio, lung); 0.034, 0.0002, 0.00000005, 0.082, 0.000004, 0.0005 (Th2/Treg ratio, lung); 0.939, 0.488, 0.961, 0.221, 0.719, 0.772 (Th1, lung-draining lymph nodes); 0.038, 0.000009, 0.00000000006, 0.002, 0.000000001, 0.0000003 (Th2, lung-draining lymph nodes); 0.839, 0.010, 0.00001, 0.051, 0.00005, 0.013 (Th17, lung-draining lymph nodes); 0.057, 0.000001, 0.001, 0.0002, 0.230, 0.009 (Treg, lung-draining lymph nodes); 0.225, 0.00008, 0.000000005, 0.004, 0.00000006, 0.00003 (Th2/Th1 ratio, lung-draining lymph nodes); 0.398, 0.00003, 0.000001, 0.0005, 0.00001, 0.244 (Th17/Treg ratio, lung-draining lymph nodes); 0.011, 0.00000007, 0.000000000007, 0.00002, 0.0000000002, 0.0000005 (Th2/Treg ratio, lung-draining lymph nodes). d Serum levels of IL-4 and IL-6 were assessed by ELISA. Exact p values (PBS-Ctrl + sham vs. PBS-Ctrl + IL-31, PBS-Ctrl + sham vs. BLM-SSc + sham, PBS-Ctrl + sham vs. BLM-SSc + IL-31, PBS-Ctrl + IL-31 vs. BLM-SSc + sham, PBS-Ctrl + IL-31 vs. BLM-SSc + IL-31, BLM-SSc + sham vs. BLM-SSc + IL-31) = 0.129, 0.001, 0.000002, 0.103, 0.0001, 0.019 (IL-4); 0.085, 0.032, 0.00007, 0.954, 0.012, 0.033 (IL-6). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. Significance was determined using one-way analysis of variance followed by Tukey’s post hoc comparison test. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control. Source data are provided as a Source Data file.

Techniques: Control, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Comparison

a BLM-SSc and PBS-treated control (PBS-Ctrl) mice were administrated with anti-mouse IL-31RA antibody (αIL-31RA) or isotype control IgG (iso) on days 1, 8, and 15, and sacrificed on day 22 for evaluation. b Representative histological sections of the skin and lungs stained with hematoxylin and eosin (left) and Masson trichrome (right) were shown (horizontal scale bars=100 μm in skin; 20 µm in lung). Vertical bars with arrows represent dermal thickness. Dermal thickness and lung fibrosis score were assessed histologically. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00004, 0.028, 0.004 (dermal thickness); 0.0000002, 0.00002, 0.00004 (lung fibrosis score). c Hydroxyproline contents in skin and lung samples. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00005, 0.052, 0.003 (skin); 0.00004, 0.011, 0.012 (lung). d Relative mRNA expression levels of Col1a1 and Col1a2 in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.0000000003, 0.00009, 0.00000002 ( Co11a1 , skin); 0.00000002, 0.005, 0.0000006 ( Co11a2 , skin); 0.0000004, 0.014, 0.00002 ( Col1a1 , lung); 0.0000002, 0.001, 0.00005 ( Col1a2 , lung). Relative fold differences (BLM-SSc + iso, BLM-SSc + αIL-31RA) = 4.21, 2.00 ( Col1a1 , skin); 5.64, 2.27 ( Col1a2 , skin); 2.62, 1.50 ( Col1a1 , lung); 3.08, 1.86 ( Col1a2 , lung). e Protein expression of type I collagen in the skin and lungs was evaluated by ELISA. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.0000002, 0.021, 0.000007 (skin); 0.0000005, 0.026, 0.00002 (lung). f The mRNA expression of Il4 , Il6 , Il10 , Il17a , Tnf , Tgfb1 , and Ifng in the skin and lungs of these mice was evaluated by real-time PCR. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00001, 0.002, 0.015 ( Il4 , skin); 0.0000005, 0.0003, 0.0007 ( Il6 , skin); 0.0006, 0.083, 0.037 ( Il10 , skin); 0.0003, 0.001, 0.625 ( Il17a , skin); 0.00004, 0.00005, 0.996 ( Tnf , skin); 0.00000001, 0.00006, 0.00001 ( Tgfb1 , skin); 0.000003, 0.000005, 0.848 ( Ifng , skin); 0.0000001, 0.0001, 0.0001 ( Il4 , lung); 0.000003, 0.003, 0.001 ( Il6 , lung); 0.0005, 0.251, 0.010 ( Il10 , lung); 0.000000007, 0.000000009, 0.927 ( Il17a , lung); 0.003, 0.002, 0.997 ( Tnf , lung); 0.0000009, 0.001, 0.0005 ( Tgfb1 , lung); 0.00004, 0.00002, 0.943 ( Ifng , lung). Relative fold differences (BLM-SSc + iso, BLM-SSc + αIL-31RA) = 4.19, 2.83 ( Il4 , skin); 18.09, 10.01 ( Il6 , skin); 5.75, 3.15 ( Il10 , skin); 5.05, 4.38 ( Il17a , skin); 2.57, 2.55 ( Tnf , skin); 3.52, 2.14 ( Tgfb1 , skin); 4.51, 4.30 ( Ifng , skin); 5.00, 3.01 ( Il4 , lung); 17.72, 8.92 ( Il6 , lung); 2.56, 1.50 ( Il10 , lung); 7.20, 7.05 ( Il17a , lung); 2.17, 2.19 ( Tnf , lung); 4.14, 2.48 ( Tgfb1 , lung); 5.34, 5.54 ( Ifng , lung). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. One-way analysis of variance followed by Tukey’s post hoc comparison test was used. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control; αIL-31RA, anti-mouse IL-31RA antibody; iso, isotype control IgG. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a BLM-SSc and PBS-treated control (PBS-Ctrl) mice were administrated with anti-mouse IL-31RA antibody (αIL-31RA) or isotype control IgG (iso) on days 1, 8, and 15, and sacrificed on day 22 for evaluation. b Representative histological sections of the skin and lungs stained with hematoxylin and eosin (left) and Masson trichrome (right) were shown (horizontal scale bars=100 μm in skin; 20 µm in lung). Vertical bars with arrows represent dermal thickness. Dermal thickness and lung fibrosis score were assessed histologically. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00004, 0.028, 0.004 (dermal thickness); 0.0000002, 0.00002, 0.00004 (lung fibrosis score). c Hydroxyproline contents in skin and lung samples. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00005, 0.052, 0.003 (skin); 0.00004, 0.011, 0.012 (lung). d Relative mRNA expression levels of Col1a1 and Col1a2 in the skin and lungs were evaluated by real-time PCR. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.0000000003, 0.00009, 0.00000002 ( Co11a1 , skin); 0.00000002, 0.005, 0.0000006 ( Co11a2 , skin); 0.0000004, 0.014, 0.00002 ( Col1a1 , lung); 0.0000002, 0.001, 0.00005 ( Col1a2 , lung). Relative fold differences (BLM-SSc + iso, BLM-SSc + αIL-31RA) = 4.21, 2.00 ( Col1a1 , skin); 5.64, 2.27 ( Col1a2 , skin); 2.62, 1.50 ( Col1a1 , lung); 3.08, 1.86 ( Col1a2 , lung). e Protein expression of type I collagen in the skin and lungs was evaluated by ELISA. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.0000002, 0.021, 0.000007 (skin); 0.0000005, 0.026, 0.00002 (lung). f The mRNA expression of Il4 , Il6 , Il10 , Il17a , Tnf , Tgfb1 , and Ifng in the skin and lungs of these mice was evaluated by real-time PCR. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00001, 0.002, 0.015 ( Il4 , skin); 0.0000005, 0.0003, 0.0007 ( Il6 , skin); 0.0006, 0.083, 0.037 ( Il10 , skin); 0.0003, 0.001, 0.625 ( Il17a , skin); 0.00004, 0.00005, 0.996 ( Tnf , skin); 0.00000001, 0.00006, 0.00001 ( Tgfb1 , skin); 0.000003, 0.000005, 0.848 ( Ifng , skin); 0.0000001, 0.0001, 0.0001 ( Il4 , lung); 0.000003, 0.003, 0.001 ( Il6 , lung); 0.0005, 0.251, 0.010 ( Il10 , lung); 0.000000007, 0.000000009, 0.927 ( Il17a , lung); 0.003, 0.002, 0.997 ( Tnf , lung); 0.0000009, 0.001, 0.0005 ( Tgfb1 , lung); 0.00004, 0.00002, 0.943 ( Ifng , lung). Relative fold differences (BLM-SSc + iso, BLM-SSc + αIL-31RA) = 4.19, 2.83 ( Il4 , skin); 18.09, 10.01 ( Il6 , skin); 5.75, 3.15 ( Il10 , skin); 5.05, 4.38 ( Il17a , skin); 2.57, 2.55 ( Tnf , skin); 3.52, 2.14 ( Tgfb1 , skin); 4.51, 4.30 ( Ifng , skin); 5.00, 3.01 ( Il4 , lung); 17.72, 8.92 ( Il6 , lung); 2.56, 1.50 ( Il10 , lung); 7.20, 7.05 ( Il17a , lung); 2.17, 2.19 ( Tnf , lung); 4.14, 2.48 ( Tgfb1 , lung); 5.34, 5.54 ( Ifng , lung). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. One-way analysis of variance followed by Tukey’s post hoc comparison test was used. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control; αIL-31RA, anti-mouse IL-31RA antibody; iso, isotype control IgG. Source data are provided as a Source Data file.

Techniques: Control, Staining, Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Comparison

a – c The frequencies of Th1, Th2, Th17, and Treg cells in BLM-SSc and PBS-treated control (PBS-Ctrl) mice administrated with anti-mouse IL-31RA antibody (αIL-31RA) or isotype control IgG (iso) were analyzed by flow cytometry in spleen ( a ), lung ( b ), and lung-draining lymph nodes ( c ). Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.649, 0.769, 0.978 (Th1, spleen); 0.0006, 0.269, 0.010 (Th2, spleen); 0.010, 0.085, 0.463 (Th17, spleen); 0.021, 0.026, 0.994 (Treg, spleen); 0.006, 0.674, 0.027 (Th2/Th1 ratio, spleen); 0.005, 0.020, 0.712 (Th17/Treg ratio, spleen); 0.0002, 0.042, 0.019 (Th2/Treg ratio, spleen); 0.547, 0.830, 0.878 (Th1, lung); 0.000004, 0.032, 0.0002 (Th2, lung); 0.002, 0.005, 0.768 (Th17, lung); 0.042, 0.040, 0.999 (Treg, lung); 0.00001, 0.052, 0.0006 (Th2/Th1 ratio, lung); 0.006, 0.010, 0.957 (Th17/Treg ratio, lung); 0.0000003, 0.003, 0.00004 (Th2/Treg ratio, lung); 0.785, 0.830, 0.996 (Th1, lung-draining lymph nodes); 0.000005, 0.004, 0.002 (Th2, lung-draining lymph nodes); 0.015, 0.012, 0.988 (Th17, lung-draining lymph nodes); 0.008, 0.007, 0.999 (Treg, lung-draining lymph nodes); 0.00000005, 0.0001, 0.00005 (Th2/Th1 ratio, lung-draining lymph nodes); 0.0008, 0.0006, 0.983 (Th17/Treg ratio, lung-draining lymph nodes); 0.000002, 0.001, 0.002 (Th2/Treg ratio, lung-draining lymph nodes). d Serum levels of IL-4 and IL-6 were assessed by ELISA. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00001, 0.0007, 0.031 (IL-4); 0.0000008, 0.0005, 0.0009 (IL-6). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. One-way analysis of variance followed by Tukey’s post hoc comparison test was used. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control; αIL-31RA, anti-mouse IL-31RA antibody; iso, isotype control IgG. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Interleukin-31 promotes fibrosis and T helper 2 polarization in systemic sclerosis

doi: 10.1038/s41467-021-26099-w

Figure Lengend Snippet: a – c The frequencies of Th1, Th2, Th17, and Treg cells in BLM-SSc and PBS-treated control (PBS-Ctrl) mice administrated with anti-mouse IL-31RA antibody (αIL-31RA) or isotype control IgG (iso) were analyzed by flow cytometry in spleen ( a ), lung ( b ), and lung-draining lymph nodes ( c ). Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.649, 0.769, 0.978 (Th1, spleen); 0.0006, 0.269, 0.010 (Th2, spleen); 0.010, 0.085, 0.463 (Th17, spleen); 0.021, 0.026, 0.994 (Treg, spleen); 0.006, 0.674, 0.027 (Th2/Th1 ratio, spleen); 0.005, 0.020, 0.712 (Th17/Treg ratio, spleen); 0.0002, 0.042, 0.019 (Th2/Treg ratio, spleen); 0.547, 0.830, 0.878 (Th1, lung); 0.000004, 0.032, 0.0002 (Th2, lung); 0.002, 0.005, 0.768 (Th17, lung); 0.042, 0.040, 0.999 (Treg, lung); 0.00001, 0.052, 0.0006 (Th2/Th1 ratio, lung); 0.006, 0.010, 0.957 (Th17/Treg ratio, lung); 0.0000003, 0.003, 0.00004 (Th2/Treg ratio, lung); 0.785, 0.830, 0.996 (Th1, lung-draining lymph nodes); 0.000005, 0.004, 0.002 (Th2, lung-draining lymph nodes); 0.015, 0.012, 0.988 (Th17, lung-draining lymph nodes); 0.008, 0.007, 0.999 (Treg, lung-draining lymph nodes); 0.00000005, 0.0001, 0.00005 (Th2/Th1 ratio, lung-draining lymph nodes); 0.0008, 0.0006, 0.983 (Th17/Treg ratio, lung-draining lymph nodes); 0.000002, 0.001, 0.002 (Th2/Treg ratio, lung-draining lymph nodes). d Serum levels of IL-4 and IL-6 were assessed by ELISA. Exact p values (PBS-Ctrl + iso vs. BLM-SSc + iso, PBS-Ctrl + iso vs. BLM-SSc + αIL-31RA, BLM-SSc + iso vs. BLM-SSc + αIL-31RA) = 0.00001, 0.0007, 0.031 (IL-4); 0.0000008, 0.0005, 0.0009 (IL-6). n = 5. Data are shown as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001. One-way analysis of variance followed by Tukey’s post hoc comparison test was used. The results shown are representative of three independent experiments with similar results. PBS-Ctrl, PBS-treated control; αIL-31RA, anti-mouse IL-31RA antibody; iso, isotype control IgG. Source data are provided as a Source Data file.

Techniques: Control, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Comparison

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