Journal: Frontiers in Genetics

Article Title: Exosomes containing long non-coding RNA AGAP2-AS1 promote the differentiation of CD4 + T cells through the miR-424-5p/SGK1 axis in psoriasis

doi: 10.3389/fgene.2025.1521470

Figure Lengend Snippet: ASAP2-AS1 promotes the differentiation of Th1 and Th17 in CD4 + T cells and serves as a sponge for miR-424-5p. (A) Fluorescence microscopy was used to detect the subcellular localization of AGAP2-AS1 in CD4 + T cells. AGAP2-AS1 was labeled with Cy3, cell nuclei were counterstained with DAPI (blue). (B) The expression of AGAP2-AS1 in CD4 + T cells after co-incubation of TNF-α-treated HaCaTs-derived exosomes. (C) CD4 + T cells were transfected with LV-AGAP2-AS1 or LV-NC. The percentage of Th1 and Th17 cells was detected by flow cytometry. (D) The protein levels of IFN-γ and IL-17A are measured by ELISA. (E) Prediction of the binding sequence between AGAP2-AS1 and miR-424-5p and construction of the AGAP2-AS1 MUT sequence. (F) Dual-luciferase reporter gene assay confirmed the binding relationship between AGAP2-AS1 and miR-424-5p in HEK-293T cells. (G) AGAP2-AS1 expression level in CD4 + T cells transfected with miR-424-5p mimics or NC mimics and miR-424-5p inhibitor or NC inhibitor. WT wild type, MUT mutant type. Th, T helper. Experiments were repeated three times. *P < 0.05, **P < 0.01, ***P < 0.001. ns: no significance.

Article Snippet: For intracellular cytokine staining, cells were fixed with fixation buffer (E-CK-A109, Elabscience) and then stained intracellularly with antibodies against human IFN-γ (E-AB-F1196C, Elabscience) and IL-17A (E-AB-F1173D, Elabscience).

Techniques: Fluorescence, Microscopy, Labeling, Expressing, Incubation, Derivative Assay, Transfection, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Binding Assay, Sequencing, Luciferase, Reporter Gene Assay, Mutagenesis

Journal: Frontiers in Genetics

Article Title: Exosomes containing long non-coding RNA AGAP2-AS1 promote the differentiation of CD4 + T cells through the miR-424-5p/SGK1 axis in psoriasis

doi: 10.3389/fgene.2025.1521470

Figure Lengend Snippet: AGAP2-AS1/miR-424-5p regulated CD4 + T cell differentiation via the SGK1 signaling pathway. (A) The relative expression levels of SGK1 in 293T cells that were transfected with siRNA targeting SGK1. (B) SGK1 expression level in CD4 + T cells transfected with LV-AGAP2-AS1 or LV-NC and si-AGAP2-AS1 or si-NC. (C) Western blot confirmed that transfection of AGAP2-AS1 in CD4 + T cells promoted the expression of SGK1, and the miR-424-5p mimics reversed this process. (D) Flow cytometry analysis of the percentages of Th1 and Th17 cells in CD4 + T cells transfected with LV-AGAP2-AS1 and miR-424-5p mimics or NC mimics. (E) The protein levels of IFN-γ and IL-17A are measured by ELISA. (F) Flow cytometry analysis of the percentages of Th1 and Th17 cells in CD4 + T cells transfected with LV-AGAP2-AS1 and SGK1 siRNA or NC siRNA. (G) The protein levels of IFN-γ and IL-17A are measured by ELISA. The experiments were repeated three times. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: For intracellular cytokine staining, cells were fixed with fixation buffer (E-CK-A109, Elabscience) and then stained intracellularly with antibodies against human IFN-γ (E-AB-F1196C, Elabscience) and IL-17A (E-AB-F1173D, Elabscience).

Techniques: Cell Differentiation, Expressing, Transfection, Western Blot, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Veterinary Research

Article Title: Enhancing the toolbox to study IL-17A in cattle and sheep

doi: 10.1186/s13567-017-0426-5

Figure Lengend Snippet: Commercial anti-IL-17A antibodies evaluated by intracellular staining for capacity to bind recombinant bovine and ovine IL-17A

Article Snippet: D.1 , 41809 , R & D Systems IC317P , IgG2b , Recombinant human IL-17A amino acids 20-155 , Human , PE.

Techniques: Staining, Recombinant, Conjugation Assay, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Extensive Histopathological Characterization of Inflamed Bowel in the Dextran Sulfate Sodium Mouse Model with Emphasis on Clinically Relevant Biomarkers and Targets for Drug Development

doi: 10.3390/ijms22042028

Figure Lengend Snippet: List of antibodies used in the study.

Article Snippet: anti-IL-17A , Rat monoclonal, clone TC11-18H10; Novus Biologicals, Littleton, CO, USA , 1:50 , EDTA–Citrate pH 7.8.

Techniques:

Journal: Biomolecules & Therapeutics

Article Title: DW2007 Ameliorates Colitis and Rheumatoid Arthritis in Mice by Correcting Th17/Treg Imbalance and Inhibiting NF-κB Activation

doi: 10.4062/biomolther.2016.018

Figure Lengend Snippet: Effect of DW2007 on Th17 and Treg cell differentiation. (A) Effect on Th17 and Treg cell differentiation. Splenocytes were stimulated with anti-CD3 (1 μg/mL) and anti-CD28 (1 μg/mL) with or without IL-6 (20 ng/mL) plus TGF-β (1 ng/mL) in the presence of DW2007 (AC) for 5 days. Staining of cell surface CD4 antigen and intracellular IL-17 was performed and analyzed using flow cytometry. (B) Effect on RORγt, Foxp3, IL-17, and IL-10 expression by qRT-PCR. (C) Effect on IL-17 and IL-10 expression by ELISA. All data indicate mean ± SD (n=6). # p <0.05 vs. normal control. * p <0.05 vs. cells stimulated with anti-CD3/anti-CD28 and IL-6/TGFβ.

Article Snippet: The reaction mixture were filtered and T cells were isolated using a Pan T cell Isolation Kit II, fixed, stained with anti-Foxp3 or anti-IL-17A antibodies (Biogems, CA, USA) and analyzed by flow cytometry.

Techniques: Cell Differentiation, Staining, Flow Cytometry, Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Control

Journal: Biomolecules & Therapeutics

Article Title: DW2007 Ameliorates Colitis and Rheumatoid Arthritis in Mice by Correcting Th17/Treg Imbalance and Inhibiting NF-κB Activation

doi: 10.4062/biomolther.2016.018

Figure Lengend Snippet: Effect of DW2007 on the differentiation to Th17 and Treg cells in wild-type mice with DSS-induced colitis. (A) Effect on Th17 (CD4 + IL17 + ) cell differentiation. (B) Effect on Treg (CD4 + Foxp3 + ) cell differentiation. DSS, except in the control group, was orally administered to mice treated with saline, DW2007 (AC), or sulfasalazine (SS). Test agents (AC10, 10 mg/kg DW2007; AC20, 20 mg/kg DW2007; or SS50, 50 mg/kg SS) or saline were orally administered for 3 days after DSS treatment. The mice were killed 18 h after the final administration of test agents. The CD4 + T cells were isolated from the lamina propria of the colon using Pan T cell isolation kit. So, we performed single staining for flow cytometry (Th17 cells: IL-17, Treg cells: Foxp3). Th17 and Treg cells were then analyzed by flow cytometry. All values are mean ± SD (n=6). # p <0.05 vs. control group. * p <0.05 vs. DSS group.

Article Snippet: The reaction mixture were filtered and T cells were isolated using a Pan T cell Isolation Kit II, fixed, stained with anti-Foxp3 or anti-IL-17A antibodies (Biogems, CA, USA) and analyzed by flow cytometry.

Techniques: Cell Differentiation, Control, Saline, Isolation, Cell Isolation, Staining, Flow Cytometry

Journal: Biomolecules & Therapeutics

Article Title: DW2007 Ameliorates Colitis and Rheumatoid Arthritis in Mice by Correcting Th17/Treg Imbalance and Inhibiting NF-κB Activation

doi: 10.4062/biomolther.2016.018

Figure Lengend Snippet: Effects of DW2007 on the expression of iNOS and COX-2, activation of NF-κB (A), and expression of inflammatory cytokines (B) and in TLR4 −/− mice with DSS-induced colitis. DSS, except in the normal control group (NOR), was orally administered to mice treated with saline (DSS), DW2007 (AC), or sulfasalazine (SS). Test agents (AC10, 10 mg/kg DW2007; AC20, 20 mg/kg AC; or SS50, 50 mg/kg SS) or saline were orally administered for 3 days after DSS treatment. iNOS, COX-2, and NF-κB were determined by immunoblotting. TNF-α, IL-1β, IL-6, IL-10, and IL-17 were determined by ELISA. All values are mean ± SD (n=6). # p <0.05 vs. control group. * p <0.05 vs. DSS group.

Article Snippet: The reaction mixture were filtered and T cells were isolated using a Pan T cell Isolation Kit II, fixed, stained with anti-Foxp3 or anti-IL-17A antibodies (Biogems, CA, USA) and analyzed by flow cytometry.

Techniques: Expressing, Activation Assay, Control, Saline, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Biomolecules & Therapeutics

Article Title: DW2007 Ameliorates Colitis and Rheumatoid Arthritis in Mice by Correcting Th17/Treg Imbalance and Inhibiting NF-κB Activation

doi: 10.4062/biomolther.2016.018

Figure Lengend Snippet: Effect of DW2007 on the expression of T cell differentiation (A) and its cytokines (B) in CIA mice. Arthritis was induced by the first intradermal injection of bovine type II collagen and the second intradermal injection of collagen with complete Freund’s adjuvant at the base of the tail of DBA/1 J mice. Test agents (AC, 20 mg/kg DW2007; IB, 50 mg/kg ibuprofen; or CIA, vehicle alone) were orally administrated daily for 20 days from day 1 after the second injection. T cells were isolated from the spleen. Th17 and Treg cells were then analyzed by a flow cytometer. Cytokines were determined in the paw joint tissues by real-time PCR (for IL-10, IL-17a, IFN-γ, Foxp3, and β-actin). All data indicate mean ± SD (n=7). # p <0.05 vs. control group. * p <0.05 vs. CIA group.

Article Snippet: The reaction mixture were filtered and T cells were isolated using a Pan T cell Isolation Kit II, fixed, stained with anti-Foxp3 or anti-IL-17A antibodies (Biogems, CA, USA) and analyzed by flow cytometry.

Techniques: Expressing, Cell Differentiation, Injection, Adjuvant, Isolation, Flow Cytometry, Real-time Polymerase Chain Reaction, Control

Journal: Biomolecules & Therapeutics

Article Title: DW2007 Ameliorates Colitis and Rheumatoid Arthritis in Mice by Correcting Th17/Treg Imbalance and Inhibiting NF-κB Activation

doi: 10.4062/biomolther.2016.018

Figure Lengend Snippet: Effect of DW2007 on the expression of iNOS and COX-2, activation of NF-κB (A), expression of inflammatory cytokines (B), and production of PGE2 and NO (C) in CIA mice. Arthritis was induced by the first intradermal injection of bovine type II collagen and the second intradermal injection of collagen with complete Freund’s adjuvant at the base of the tail of DBA/1 J mice. Test agents (AC, 20 mg/kg DW2007; IB, 50 mg/kg ibuprofen; or CIA, vehicle alone) were orally administrated daily for 20 days from day 1 after the second injection. Proteins were determined in the paw joint tissues by immunoblotting. Cytokines were determined in the paw joint tissues by ELISA (IL-1β, IL-10, IL-17a, TNF-α, and PGE2). NO was measured using Griess reagent. All data indicate mean ± SD (n=7). # p <0.05 vs. control group. * p <0.05 vs. CIA group.

Article Snippet: The reaction mixture were filtered and T cells were isolated using a Pan T cell Isolation Kit II, fixed, stained with anti-Foxp3 or anti-IL-17A antibodies (Biogems, CA, USA) and analyzed by flow cytometry.

Techniques: Expressing, Activation Assay, Injection, Adjuvant, Western Blot, Enzyme-linked Immunosorbent Assay, Control