Journal: Scientific Reports

Article Title: Osthol ameliorates obesity-associated lipid metabolic disorders by inhibiting ADRA1D-dependent Th17 cell differentiation

doi: 10.1038/s41598-025-20719-x

Figure Lengend Snippet: OST Attenuates Systemic Inflammation and Downregulates Th17-Related Factor Expression. ( A ) KEGG pathway enrichment analysis of downstream targets of OST. This figure is based on data from KEGG (Kyoto Encyclopedia of Genes and Genomes) at https://www.kegg.jp/ ; © Kanehisa Laboratories. ( B ) Flow cytometry was used to evaluate the proportion of CD4⁺IL-17A⁺ cells in the spleen. ( C ) Immunohistochemical staining of liver tissues was performed to assess IL-17A expression. ( D ) Immunofluorescence staining was conducted to detect CD4⁺RORγt⁺ double-positive cells in liver sections. ( E ) Serum levels of IL-6, IL-1β, TNF-α, and IL-17A were measured using ELISA. n = 6. **** P < 0.0001. One-way ANOVA followed by Tukey’s post hoc test was used for multiple group comparisons.

Article Snippet: Cells were then fixed and permeabilized, followed by intracellular staining with a phycoerythrin (PE)-conjugated anti-IL-17A antibody (F21IL1702, Lianke Biotech, Hangzhou, China).

Techniques: Expressing, Flow Cytometry, Immunohistochemical staining, Staining, Immunofluorescence, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Osthol ameliorates obesity-associated lipid metabolic disorders by inhibiting ADRA1D-dependent Th17 cell differentiation

doi: 10.1038/s41598-025-20719-x

Figure Lengend Snippet: OST Inhibits Th17 Cell Differentiation in Vitro. ( A ) Flow cytometry was used to assess the proportion of CD4⁺IL-17A⁺ cells in a Th17-polarized cell model treated with different doses of OST. ( B ) RT-qPCR was performed to evaluate mRNA expression levels of RORγt and IL-17A. ( C ) Western blot was performed to evaluate protein expression levels of RORγt, IL-17A, IL-17RA, TRAF6, and Act1. ( D ) ELISA was used to quantify the secretion of IL-6, IL-1β, TNF-α, IL-10, and TGF-β in the culture supernatants. n = 3. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. One-way ANOVA followed by Tukey’s post hoc test was used for multiple group comparisons.

Article Snippet: Cells were then fixed and permeabilized, followed by intracellular staining with a phycoerythrin (PE)-conjugated anti-IL-17A antibody (F21IL1702, Lianke Biotech, Hangzhou, China).

Techniques: Cell Differentiation, In Vitro, Flow Cytometry, Quantitative RT-PCR, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Osthol ameliorates obesity-associated lipid metabolic disorders by inhibiting ADRA1D-dependent Th17 cell differentiation

doi: 10.1038/s41598-025-20719-x

Figure Lengend Snippet: OST Suppresses Th17 Differentiation via an ADRA1D-Dependent Mechanism. ( A ) Volcano plot of differentially expressed genes (DEGs) in the GSE110729 obesity dataset. ( B ) Venn diagram showing the intersection between OST’s potential targets and DEGs from the GSE110729 dataset. ( C ) RT-qPCR was performed to assess the expression of ADRA1D, IL-17A, and RORγt in Th17-polarized CD4⁺ T cells with or without ADRA1D overexpression. ( D ) Western blot was conducted to evaluate the protein expression of ADRA1D, IL-17A, RORγt, IL-17RA, TRAF6, and Act1. ( E ) ELISA was used to measure the levels of IL-6, IL-1β, TNF-α, IL-10, and TGF-β in the culture supernatants. n = 3. *** P < 0.001, **** P < 0.0001. One-way ANOVA followed by Tukey’s post hoc test was used for multiple group comparisons.

Article Snippet: Cells were then fixed and permeabilized, followed by intracellular staining with a phycoerythrin (PE)-conjugated anti-IL-17A antibody (F21IL1702, Lianke Biotech, Hangzhou, China).

Techniques: Quantitative RT-PCR, Expressing, Over Expression, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Osthol ameliorates obesity-associated lipid metabolic disorders by inhibiting ADRA1D-dependent Th17 cell differentiation

doi: 10.1038/s41598-025-20719-x

Figure Lengend Snippet: ADRA1D Overexpression Attenuates the Metabolic and Immunomodulatory Effects of OST in Vivo. ( A ) Body weight changes were monitored in HFD-fed mice treated with OST. (B) Weights of adipose tissue and liver were measured to assess the effect on organ hypertrophy. ( C ) Serum levels of TG, TC, FFA, ALT, and AST were evaluated by biochemical assays. ( D ) Flow cytometry was performed to assess the proportion of CD4⁺IL-17A⁺ cells in the spleen. ( E ) Immunofluorescence staining of liver tissue was used to detect CD4⁺RORγt⁺ double-positive cell infiltration. ( F ) Immunohistochemistry was performed to evaluate hepatic IL-17A protein expression. ( G ) Western blotting was conducted to assess ADRA1D protein expression in liver tissue. ( H ) Western blot was used to detect the phosphorylation levels of ERK1/2 and PI3K in liver tissue. ( I ) H&E staining of adipose tissue was used to assess adipocyte morphology. (J) Oil red O staining of liver sections was performed to evaluate hepatic lipid deposition. n = 6. ** P < 0.01, **** P < 0.0001. For three or more groups, one-way or two-way analysis of variance (ANOVA) was applied, followed by Tukey’s post hoc test. P < 0.05 was considered statistically significant.

Article Snippet: Cells were then fixed and permeabilized, followed by intracellular staining with a phycoerythrin (PE)-conjugated anti-IL-17A antibody (F21IL1702, Lianke Biotech, Hangzhou, China).

Techniques: Over Expression, In Vivo, Flow Cytometry, Immunofluorescence, Staining, Immunohistochemistry, Expressing, Western Blot, Phospho-proteomics