Journal: Oncology Letters

Article Title: Anti-IL-20 monoclonal antibody suppresses hepatocellular carcinoma progression

doi: 10.3892/ol.2018.9402

Figure Lengend Snippet: Expression of IL-20 in HCC and adjacent non-cancerous tissues. (A) IL-20 mRNA levels were examined by reverse transcription-quantitative polymerase chain reaction. **P<0.01, as indicated. (B) Protein levels of IL-20 were confirmed via immunohistochemistry. Magnification, ×100 and ×200 as indicated. (C) Association between IL-20 expression and overall survival time. IL, interleukin; HCC, hepatocellular carcinoma.

Article Snippet: Mouse recombinant IL-20 (mIL-20) (300 ng/ml; R&D Systems), anti-IL-20 mAb (3 mg/ml) or mIL-20 plus anti-IL-20 mAb (dilution, 1:10) was added to the culture system.

Techniques: Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Immunohistochemistry

Journal: Oncology Letters

Article Title: Anti-IL-20 monoclonal antibody suppresses hepatocellular carcinoma progression

doi: 10.3892/ol.2018.9402

Figure Lengend Snippet: Growth, migration and invasion of Hep1-6 cells. (A) A Cell Counting Kit-8 assay was used to evaluate Hep1-6 proliferation. (B) A plate colony formation assay was used to evaluate Hep1-6 clonogenic ability. Transwell assays were performed to assess the (C) migration and (D) invasion of Hep1-6 cells (magnification, ×100). The four groups were treated with mIL-20, anti-IL-20 mAb, mIL-20 plus mAb or PBS (untreated control). *P<0.05 and **P<0.01 vs. the untreated group. IL, interleukin; mAb, monoclonal antibody; mIL-20, mouse recombinant IL-20; OD, optical density.

Article Snippet: Mouse recombinant IL-20 (mIL-20) (300 ng/ml; R&D Systems), anti-IL-20 mAb (3 mg/ml) or mIL-20 plus anti-IL-20 mAb (dilution, 1:10) was added to the culture system.

Techniques: Migration, Cell Counting, Colony Assay, Recombinant

Journal: Oncology Letters

Article Title: Anti-IL-20 monoclonal antibody suppresses hepatocellular carcinoma progression

doi: 10.3892/ol.2018.9402

Figure Lengend Snippet: Effect of anti-IL-20 monoclonal antibody on tumorigenesis in nude mice. The tumor (A) size, (B) weight (**P<0.01 vs. PBS) and (C) volume were measured. (D) Western blot analysis of MMP-9, TGF-β, p-STAT3 and p-JNK levels in the implanted tumors. Analysis of (E) MMP-9 and TGF-β and (F) p-STAT3 and p-JNK levels shown in panel D. **P<0.01, as indicated. mIgG, membrane immunoglobulin G; IL, interleukin; IL-20 mAb, anti-IL-20 monoclonal antibody; p-, phosphorylated; MMP, matrix metalloproteinase; TGF, tumor growth factor; STAT, signal transducer and activator of transcription; JNK, Jun N-terminal kinase; IgG, immunoglobulin G.

Article Snippet: Mouse recombinant IL-20 (mIL-20) (300 ng/ml; R&D Systems), anti-IL-20 mAb (3 mg/ml) or mIL-20 plus anti-IL-20 mAb (dilution, 1:10) was added to the culture system.

Techniques: Western Blot, Membrane

Journal: Arthritis & rheumatology (Hoboken, N.J.)

Article Title: Decreased Interleukin-20 Expression in Scleroderma Skin Contributes to Cutaneous Fibrosis

doi: 10.1002/art.38380

Figure Lengend Snippet: Effect of interleukin-20 (IL-20) on collagen expression. A, Normal fibroblasts were treated with IL-20 (100 ng/ml) for 12 hours. Levels of collagen mRNA were determined by real-time polymerase chain reaction (PCR) (n = 7 samples). ⅞* = P < 0.05 versus untreated fibroblasts (set to 1.0). B, Normal fibroblasts were treated with IL-20 (100 ng/ml) for 24 hours. Cell lysates were subjected to immunoblotting. The protein levels of type I collagen quantitated by scanning densitometry and corrected for the levels of β-actin in the same samples are shown relative to those in untreated fibroblasts (set to 1.0) (n = 3 samples). * = P < 0.05 versus untreated fibroblasts. C, Fibroblasts were incubated in the presence or absence of IL-20 for 12 hours before the addition of 2.5 μg/ml actinomycin D for 6 or 12 hours. IL-20 mRNA expression was analyzed by real-time PCR. D, Fibroblasts were incubated in the presence or absence of IL-20 for 24 hours before the addition of cycloheximide (10 μg/ml). Cells were harvested at the indicated time points after cycloheximide was administered, and immunoblotting was performed. The protein levels of type I collagen quantitated by scanning densitometry and corrected for the levels of β-actin in the same samples are shown relative to those in untreated fibroblasts (set to 1.0) (n = 3 samples). E, The indicated α2(I) collagen promoter deletion constructs were transfected into normal fibroblasts in the absence or presence of IL-20 (100 ng/ml) for 24 hours (n = 3 samples). The bar graph represents fold stimulation of chloramphenicol acetyltransferase activities stimulated with IL-20 relative to those not stimulated with IL-20 (set to 1.0). * = P < 0.05 versus cells not stimulated with IL-20. Values are the mean ± SEM. c/EBPβ = CCAAT/enhancer binding protein β; AP-1 = activator protein 1; CBF = CCAAT-binding transcription factor.

Article Snippet: Recombinant human IL-19, IL-20, and IL-24 and mouse IL-20 were obtained from R&D Systems.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Incubation, Construct, Transfection, Binding Assay

Journal: Arthritis & rheumatology (Hoboken, N.J.)

Article Title: Decreased Interleukin-20 Expression in Scleroderma Skin Contributes to Cutaneous Fibrosis

doi: 10.1002/art.38380

Figure Lengend Snippet: Effect of interleukin-20 (IL-20) on Fli-1 expression. A, Normal fibroblasts were incubated in the presence or absence of IL-20 for 12 hours. Messenger RNA levels were determined by real-time polymerase chain reaction (PCR) (n = 7 samples). * = P < 0.05 versus untreated fibroblasts (set to 1.0). B, Normal fibroblasts were treated with IL-20 for 24 hours. Immunoblotting was performed using antibody to Ets-1 or Fli-1. Protein levels of Ets-1 and Fli-1 were quantitated as described in Figure 1B, and the ratio of Ets-1:Fli-1 protein expression is shown. * = P < 0.05 versus untreated fibroblasts (set to 1.0) (n = 3 samples). C, Normal fibroblasts were treated with IL-20 (100 ng/ml) for 24 hours. Levels of phosphorylated and acetylated Fli-1 were determined as described in Patients and Methods. D, Normal fibroblasts were incubated in the presence or absence of IL-20 for 12 hours. Cellular DNA was sheared, and chromatin (input DNA) was immunoprecipitated with anti–Fli-1 antibody or IgG isotype control antibody. The presence of α2(I) collagen promoter fragments in the precipitates was detected using PCR followed by agarose gel electrophoresis. E, Levels of Smad3, endoglin, and caveolin 1 mRNA were determined as described in A. * = P < 0.05 versus untreated fibroblasts (set to 1.0) (n = 7 samples). F, Normal fibroblasts were treated with IL-20 (100 ng/ml) for 12 hours in the presence or absence of Fli-1 small interfering RNA (siRNA). The expression of type I collagen was determined by immunoblotting. Values are the mean ± SEM.

Article Snippet: Recombinant human IL-19, IL-20, and IL-24 and mouse IL-20 were obtained from R&D Systems.

Techniques: Expressing, Incubation, Real-time Polymerase Chain Reaction, Western Blot, Immunoprecipitation, Control, Agarose Gel Electrophoresis, Small Interfering RNA

Journal: Arthritis & rheumatology (Hoboken, N.J.)

Article Title: Decreased Interleukin-20 Expression in Scleroderma Skin Contributes to Cutaneous Fibrosis

doi: 10.1002/art.38380

Figure Lengend Snippet: Interleukin-20 (IL-20) levels in sera and skin tissues from patients with rheumatic diseases. A, Shown are serum IL-20 levels as determined by enzyme-linked immunosorbent assay. Serum samples were obtained from 33 patients with systemic sclerosis (SSc), 9 patients with scleroderma spectrum disorders (SSD), 10 patients with systemic lupus erythematosus (SLE), 12 patients with dermatomyositis (DM), and 15 normal subjects. Symbols represent individual samples; bars show the mean. * = P < 0.05. B, Total RNA was extracted from skin tissues obtained from 12 SSc patients and 6 normal subjects. IL-20 mRNA expression was analyzed by real-time polymerase chain reaction (PCR). Symbols represent individual samples; bars show the mean. * = P < 0.05. C, Paraffin-embedded sections of normal and SSc-involved skin were subjected to immunohistochemical analysis for IL-20. Original magnification × 200 (n = 3 samples). D, Paraffin-embedded sections of normal and SSc-involved skin were subjected to immunohistochemical analysis for IL-20 receptor (IL-20R). Original magnification × 400 (n = 3 samples). E, Total RNA was extracted, and IL-20R mRNA expression was analyzed by real-time PCR as described in B. Symbols represent individual samples; bars show the mean. F, Cell lysates were obtained from cultured normal and SSc dermal fibroblasts and subjected to immunoblotting with antibody to IL-20R. Results are representative of 5 normal and 5 SSc fibroblasts. G, Expression of IL-20R in cultured normal dermal fibroblasts and SSc fibroblasts was visualized by immunofluorescence microscopy. Original magnification × 1,000. dcSSc = diffuse cutaneous SSc; lcSSc = limited cutaneous SSc.

Article Snippet: Recombinant human IL-19, IL-20, and IL-24 and mouse IL-20 were obtained from R&D Systems.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Real-time Polymerase Chain Reaction, Immunohistochemical staining, Cell Culture, Western Blot, Immunofluorescence, Microscopy

Journal: Arthritis & rheumatology (Hoboken, N.J.)

Article Title: Decreased Interleukin-20 Expression in Scleroderma Skin Contributes to Cutaneous Fibrosis

doi: 10.1002/art.38380

Figure Lengend Snippet: Effect of IL-20 on collagen expression in transforming growth factor β (TGFβ)–stimulated normal fibroblasts or SSc fibroblasts. A, Normal fibroblasts were treated with IL-20 (100 ng/ml) and/or TGFβ (2 ng/ml) for 24 hours. The expression of type I collagen was evaluated by immunoblotting. B, Normal fibroblasts were treated with IL-20 in the presence of TGFβ (2 ng/ml) for 24 hours. Immunoblotting was performed using antibody to Ets-1 or Fli-1. C, Normal fibroblasts were treated with IL-20 (100 ng/ml) for 12 hours in the presence of TGFβ (2 ng/ml). Real-time PCR was performed to evaluate levels of Smad3, endoglin, and caveolin 1 mRNA (n = 7 samples). * = P < 0.05 versus untreated fibroblasts (set to 1.0). D, SSc fibroblasts were treated with IL-20 for 24 hours. Immunoblotting was performed using antibody to type I collagen. E, SSc fibroblasts were treated with IL-20 for 24 hours. Immunoblotting was performed using antibody to Ets-1 or Fli-1. F, SSc fibroblasts were treated with IL-20 (100 ng/ml) for 12 hours. Real-time PCR was performed to evaluate levels of Smad3, endoglin, and caveolin 1 mRNA (n = 7 samples). * = P < 0.05 versus untreated fibroblasts (set to 1.0). Values are the mean ± SEM. See Figure 3 for other definitions.

Article Snippet: Recombinant human IL-19, IL-20, and IL-24 and mouse IL-20 were obtained from R&D Systems.

Techniques: Expressing, Western Blot, Real-time Polymerase Chain Reaction

Journal: Arthritis & rheumatology (Hoboken, N.J.)

Article Title: Decreased Interleukin-20 Expression in Scleroderma Skin Contributes to Cutaneous Fibrosis

doi: 10.1002/art.38380

Figure Lengend Snippet: Effect of interleukin-20 (IL-20) on bleomycin (BLM)–induced skin fibrosis in vivo. A, Paraffin-embedded sections of skin were subjected to immunohistochemical analysis for IL-20. Top, Phosphate buffered saline (PBS)–treated wild-type mouse. Bottom, Bleomycin-treated mouse. Results shown are representative of 5 samples. Original magnification × 200. B, PBS- or bleomycin-treated mouse skin was injected with control PBS (left) or IL-20 (right) and stained with hematoxylin and eosin. Double-headed arrows indicate thickness of the dermis. Results shown are representative of 5 samples. Bars = 0.1 mm. C, Paraffin-embedded sections of bleomycin-treated mouse skin injected with control PBS or IL-20 were subjected to immunohistochemical analysis for Fli-1 or Smad3. Results shown are representative of 5 samples. Original magnification × 400. D, Dermal thickness was measured in PBS- or bleomycin-treated mouse skin injected with control PBS or IL-20 (n = 5 samples per group). Symbols represent individual samples; bars show the mean. * = P < 0.05. E, The amount of collagen content in skin sections was quantified using an assay kit (n = 5 samples per group). Symbols represent individual samples; bars show the mean. * = P < 0.05.

Article Snippet: Recombinant human IL-19, IL-20, and IL-24 and mouse IL-20 were obtained from R&D Systems.

Techniques: In Vivo, Immunohistochemical staining, Saline, Injection, Control, Staining

Journal: Experimental eye research

Article Title: IL-20 Promotes Epithelial Healing of the Injured Mouse Cornea

doi: 10.1016/j.exer.2016.11.006

Figure Lengend Snippet: IL-20 and IL-20RA are detected in the mouse cornea. (A) Binding of TRITC-anti-IL-20RA or TRITC-IgG to normal mouse corneal epithelium. (B) Binding of FITC-anti-IL-20 or FITC-IgG to normal mouse corneal epithelium, and (C) to stromal keratocytes. (D) ELISA determinations of IL-20 from extracted corneas without wounding and at various times after central epithelial abrasion (n=10 corneas pooled at each timepoint; ** p<0.01). (E) Human epithelial cell line (HTCEpi cells) stained with FITC-anti-IL-20 or FITC-IgG and (F) with TRITC-anti-IL-20RA or TRITC-IgG.

Article Snippet: Neutralization of endogenous IL-20 IL-20 function was blocked by topical application of a neutralizing goat-anti-mouse IL-20 antibody (R&D Systems, Minneapolis, MN).

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Staining