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 Regulates Adipocyte Lipid Metabolism via ADRA1D. ( A ) Oil red O staining was performed to assess lipid droplet accumulation in 3T3-L1 adipocytes treated with OST. ( B ) Intracellular triglyceride (TG) content was quantified to evaluate lipid accumulation. ( C , D ) RT-qPCR and western blotting were used to measure the mRNA and protein expression levels of FASN, PPARγ, and ADRA1D. (E) Western blot was used to detect the phosphorylation levels of ERK1/2 and PI3K. n = 3. ** P < 0.01, **** P < 0.0001. One-way ANOVA followed by Tukey’s post hoc test was used for multiple group comparisons.

Article Snippet: 3T3-L1 preadipocytes (400,107, Cytion, Eppelheim, Germany) were cultured in Dulbecco’s modified Eagle medium (DMEM; 11,885,084, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (FBS; A5670401, Thermo Fisher Scientific) at 37 °C until reaching 100% confluence.

Techniques: Staining, Quantitative RT-PCR, Western Blot, Expressing, Phospho-proteomics

Journal: Journal of Lipid Research

Article Title: Dopamine D2 receptor upregulates leptin and IL-6 in adipocytes [S]

doi: 10.1194/jlr.M081000

Figure Lengend Snippet: D2R and D3R are expressed in human adipocytes and mouse adipocytes. A, B: The expression of D2R and D3R was determined by Western blot in human subcutaneous adipocytes cells and 3T3-L1 cells. Mouse proximal tubular cells were used as a positive control (PC). PA, preadipocites; A, adipocytes. C:. D2R and D3R mRNA expression was determined by qRT-PCR in human preadipocytes and adipocytes. The data were normalized with GAPDH.

Article Snippet: 3T3-L1 preadipocyte medium (PM-1-L1), 3T3-L1 differentiation medium (DM-2-L1), and 3T3-L1 adipocyte medium (AM-1-L1) were from Zen-Bio, Inc. Differentiated 3T3-L1 adipocyte cells were serum-starved for 2 h and treated for 24 h with 1 μM quinpirole (D 2 R/D 3 R agonist, Sigma-Aldrich), or 1 μM quinpirole plus 1 μM L-741,262 (selective D 2 R antagonist, Sigma-Aldrich), as previously described ( 15 , 16 ).

Techniques: Expressing, Western Blot, Positive Control, Quantitative RT-PCR

Journal: Journal of Lipid Research

Article Title: Dopamine D2 receptor upregulates leptin and IL-6 in adipocytes [S]

doi: 10.1194/jlr.M081000

Figure Lengend Snippet: D2R stimulation increases leptin, IL-6, and TNF α expression in 3T3-L1 cells. A: 3T3-L1 cells were treated with vehicle or quinpirole (1 μM, 24 h), a D2R agonist, in the presence or absence of a D2R antagonist (L-7431,626, 1 μM, 24 h). The expression of adiponectin, visfatin, leptin, and IL-6 was determined by Western blot and normalized with GAPDH expression. Data are expressed as mean ± SEM, one-way ANOVA or Student’s t-test; *P < 0.05; n = 4. B: 3T3-L1 cells were treated with vehicle or quinpirole (1 μM, 15 min, 30 min, or 2 h). The expression of leptin and TNFα was determined by Western blot and normalized with GAPDH expression. Data are expressed as mean ± SEM, one-way ANOVA or Student’s t-test; *P < 0.05; n = 4. C: 3T3-L1 cells were treated with vehicle or quinpirole, a D2R agonist (1μM, 24 h). The mRNA expression of leptin, IL-6, TNFα, NFKB, and MCP-1 were determined by qRT-PCR and normalized with GAPDH. Data are expressed as mean ± SEM, Student’s t-test; *P < 0.05; n = 5. D: Concentration of leptin in the medium was determined by ELISA. Data are expressed as mean ± SEM, Student’s t-test; *P < 0.05; n = 3.

Article Snippet: 3T3-L1 preadipocyte medium (PM-1-L1), 3T3-L1 differentiation medium (DM-2-L1), and 3T3-L1 adipocyte medium (AM-1-L1) were from Zen-Bio, Inc. Differentiated 3T3-L1 adipocyte cells were serum-starved for 2 h and treated for 24 h with 1 μM quinpirole (D 2 R/D 3 R agonist, Sigma-Aldrich), or 1 μM quinpirole plus 1 μM L-741,262 (selective D 2 R antagonist, Sigma-Aldrich), as previously described ( 15 , 16 ).

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

Journal: Journal of Lipid Research

Article Title: Dopamine D2 receptor upregulates leptin and IL-6 in adipocytes [S]

doi: 10.1194/jlr.M081000

Figure Lengend Snippet: Downregulation of D2R decreases leptin, IL-6 in 3T3-L1 cells. 3T3-L1 cells were transfected with D2R siRNA (30 nM, 72 h) or mock (non-silencing) siRNA. A: The expression of D2R, D3R, D4R, leptin, and IL-6 was determined by Western blot and normalized by GAPDH. Data are expressed as mean ± SEM, Student’s t-test; *P < 0.05; n = 5/3. B: The concentration of leptin in the medium was determined by ELISA. Data are expressed as mean ± SEM, Student’s t-test; *P < 0.05; n = 5/3. C: The expression of leptin, IL-6, and TNFα was determined by qRT-PCR and normalized by GAPDH. Student’s t-test; *P < 0.05; n = 5.

Article Snippet: 3T3-L1 preadipocyte medium (PM-1-L1), 3T3-L1 differentiation medium (DM-2-L1), and 3T3-L1 adipocyte medium (AM-1-L1) were from Zen-Bio, Inc. Differentiated 3T3-L1 adipocyte cells were serum-starved for 2 h and treated for 24 h with 1 μM quinpirole (D 2 R/D 3 R agonist, Sigma-Aldrich), or 1 μM quinpirole plus 1 μM L-741,262 (selective D 2 R antagonist, Sigma-Aldrich), as previously described ( 15 , 16 ).

Techniques: Transfection, Expressing, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Journal of Lipid Research

Article Title: Dopamine D2 receptor upregulates leptin and IL-6 in adipocytes [S]

doi: 10.1194/jlr.M081000

Figure Lengend Snippet: PI3K inhibition blunts the stimulatory effect of D2R on leptin expression in 3T3-L1 cells. A: 3T3-L1 cells were treated with vehicle or quinpirole (1 μM; 24 h) in the presence or absence of LY294002 (10 μM; 24 h), an inhibitor of PI3K. The expression of phosphorylated-AKT, AKT, leptin, IL-6, and GAPDH was determined by Western blot; Student’s t-test or ANOVA; *P < 0.05, n = 4. B: The expression of OB-R was determined by Western blot in 3T3-L1 preadipocytes and 3T3-L1 adipocytes treated with quinpirole (1 μM; 24 h). C: 3T3-L1 adipocytes treated with L39A/D40A/F41 (25 nM, 24 h), a leptin antagonist, and quinpirole (1 μM; 24 h). The expression of IL-6 was determined by Western blot and normalized by GAPDH. Student’s t-test or ANOVA; *P < 0.05; n = 4.

Article Snippet: 3T3-L1 preadipocyte medium (PM-1-L1), 3T3-L1 differentiation medium (DM-2-L1), and 3T3-L1 adipocyte medium (AM-1-L1) were from Zen-Bio, Inc. Differentiated 3T3-L1 adipocyte cells were serum-starved for 2 h and treated for 24 h with 1 μM quinpirole (D 2 R/D 3 R agonist, Sigma-Aldrich), or 1 μM quinpirole plus 1 μM L-741,262 (selective D 2 R antagonist, Sigma-Aldrich), as previously described ( 15 , 16 ).

Techniques: Inhibition, Expressing, Western Blot

Journal: Journal of Lipid Research

Article Title: Dopamine D2 receptor upregulates leptin and IL-6 in adipocytes [S]

doi: 10.1194/jlr.M081000

Figure Lengend Snippet: Dopamine decreases leptin expression via adrenergic receptors. 3T3-L1 cells were treated with vehicle or dopamine (1 μM, 24 h), an antagonist of D1R (SCH 23390, 10 µM) and D2R (L741,626, 10 µM), α-adrenergic receptor antagonist (propranolol, 10 µM), and β-adrenergic receptor antagonist (phentolamine, 10 µM) in medium with or without insulin. Protein expression of leptin in the medium was determined by ELISA and normalized by protein concentration, one-way ANOVA; *P < 0.05, control vs. others; #P < 0.05, vs. others; Student’s t-test; †P < 0.05 vs. control, n = 6/11.

Article Snippet: 3T3-L1 preadipocyte medium (PM-1-L1), 3T3-L1 differentiation medium (DM-2-L1), and 3T3-L1 adipocyte medium (AM-1-L1) were from Zen-Bio, Inc. Differentiated 3T3-L1 adipocyte cells were serum-starved for 2 h and treated for 24 h with 1 μM quinpirole (D 2 R/D 3 R agonist, Sigma-Aldrich), or 1 μM quinpirole plus 1 μM L-741,262 (selective D 2 R antagonist, Sigma-Aldrich), as previously described ( 15 , 16 ).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Protein Concentration