Journal: Cells

Article Title: Punicic Acid in Ovarian Cancer: Anticancer Activity and Mechanistic Insights

doi: 10.3390/cells15090792

Figure Lengend Snippet: Differential Effects of Punicic Acid and α-Eleostearic Acid on OC and Normal Cell Lines. ( a ) HEYA8 OC cells and NFT epithelial cell lines (P201 and P211) were treated for 48 h with methanol control (CTL, black bars), 20 µM PunA (medium grey bars), 25 µM PunA (dark grey bars) or 20 µM cisplatin (CP, light grey bars). Data are presented as mean ± SD of six technical replicates, representative of three independent experiments. Statistical significance was determined by one-way ANOVA with Dunnett’s multiple comparisons test versus control. **** p < 0.0001, ns: not significant. ( b ) OC cells and normal cell lines were treated with increasing concentrations of PunA (0, 5, 10, 20, and 40 μM) for 48 h, and cell viability was determined by the MTS assay. Data represent mean ± SD from three independent experiments. Statistical analysis was performed using two-way ANOVA followed by Dunnett’s multiple comparisons test. No significant differences were observed at baseline (0 μM; all p > 0.9999). At 5 μM, HEYA8 cells showed significantly lower viability compared to P201 ( p = 0.0196), P211 ( p = 0.0002), E7 ( p = 0.0005), and 3T3-L1 ( p = 0.0002), with all comparisons reaching p < 0.0001 at higher concentrations. ( c ) Structural comparison of PunA (cis-9, trans-11, cis-13) and α-ESA (cis-9, trans-11, trans-13), both conjugated trienoic FFAs differing only in the π-bond orientation at the omega-5 carbon. ( d ) HEYA8 and P201 cells were treated for 48 h with varying concentrations of PunA and α-ESA. While α-ESA reduced viability in both HEYA8 and P201 cells, PunA showed a dose-dependent reduction in HEYA8 viability with minimal cytotoxicity in P201 cells. Data are presented as mean ± SD of six technical replicates, representative of three independent experiments. Statistical significance was determined by two-way ANOVA with Dunnett’s post hoc test versus control. * p < 0.05, **** p < 0.0001. ( e ) Summary table of PunA IC 50 values between normal cell lines (top four rows) and OC cell lines (bottom three rows).

Article Snippet: The mouse preadipocyte 3T3-L1 cell line (Cat # CL-173) was purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Control, MTS Assay, Comparison

Journal: Cells

Article Title: Punicic Acid in Ovarian Cancer: Anticancer Activity and Mechanistic Insights

doi: 10.3390/cells15090792

Figure Lengend Snippet: Effects of PunA Alone and in Combination with Cisplatin on Cell Viability, Drug Interaction, Resistance, and Lipidic Microenvironment. ( a ) HEYA8 cells were treated with PunA (15 or 30 μM) for 48 h, and cell viability was measured using an MTS assay. Statistical significance was determined by one-way ANOVA with Dunnett’s multiple comparisons test versus control. ( b ) HEYA8 cells were treated with cisplatin (30 μM), PunA (30 μM), or their combination for 48 h, followed by MTS analysis. Statistical significance was determined using one-way ANOVA followed by Šídák’s multiple comparisons test. ( c ) Drug interaction between PunA and cisplatin was evaluated using the Highest Single Agent (HSA) model by comparing predicted and observed inhibition. Data ( a – c ) are presented as mean ± SD of six technical replicates, representative of two independent experiments. * p < 0.05, **** p < 0.0001, ns: not significant. ( d ) Cisplatin-sensitive and -resistant cell lines (41M/41McisR and A2780/A2780cisR) were treated with increasing concentrations of PunA for 48 h (0 μM–60 μM for 41M pair, upper panel; 0 μM–75 μM for A2780 pair, lower panel). Data are presented as mean ± SD of six technical replicates, representative of three independent experiments. Statistical significance was assessed by two-way ANOVA with Dunnett’s post hoc test. For 41M/41McisR, the interaction, PunA factor, and drug-resistance factor were all significant ( p < 0.0001). For A2780/A2780cisR, the interaction was significant ( p = 0.0341), as were the PunA factor ( p < 0.0001) and drug-resistant factor ( p = 0.0019). ( e ) HEYA8 cells were cultured in DF or UD 3T3-L1 conditioned medium and treated with increasing concentrations of PunA (0–100 μM) for 48 h. Cell viability was measured by MTS assay, normalized to untreated controls. Data are presented as mean ± SD of six technical replicates, representative of three independent experiments. Statistical significance was assessed by two-way ANOVA with Dunnett’s post hoc test. The interaction was significant ( p = 0.0003), along with the PunA factor ( p < 0.001) and DF conditioned medium factor ( p = 0.0008).

Article Snippet: The mouse preadipocyte 3T3-L1 cell line (Cat # CL-173) was purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: MTS Assay, Control, Inhibition, Cell Culture

Journal: iScience

Article Title: Induction of adipocyte thermogenic program by Rho-associated coiled-coil containing kinase 2

doi: 10.1016/j.isci.2026.115225

Figure Lengend Snippet: ROCK2-mediated thermogenic gene expression may occur through the JNK/FOXO1/UCP1 axis (A) Western blotting and quantification of FOXO1 in sWAT from male WT mice housed at room or cold temperature for 3 h ( n = 4 mice per group). (B) Ucp1 expression in sWAT from male WT mice housed at 4 °C for 0, 3, 6, or 12 h ( n = 4 mice). (C) Ucp1 expression in sWAT harvested from control mice and then treated with 40 μM isoproterenol (Iso) and/or 4 μM FOXO1 inhibitor (AS1842856) for 6 h ( n = 4 wells per group). (D) Foxo1 expression in sWAT from male control and AdR2KO mice housed at RT or cold for 3 h ( n = 4 mice). (E) Western blotting and quantification of nuclear FOXO1 (nFOXO1), cytoplasmic FOXO1 (cFOXO1), and total FOXO1 (tFOXO1) in sWAT from male control and AdR2KO mice housed at RT or cold for 3 h ( n = 3 mice). (F) Western blotting and quantification of phosphorylated JNK (pJNK) and total JNK (tJNK) in sWAT from male WT mice housed at RT or cold for 3 h ( n = 3 mice). (G) Foxo1 expression in sWAT harvested from control mice and then treated with 40 μM isoproterenol (Iso) and/or 100 μM JNK inhibitor (SP600125) for 6 h ( n = 6 wells per group). (H) Western blotting and quantification of pJNK and tJNK in sWAT from control and AdR2KO mice housed at RT or cold for 3 h ( n = 3 mice). (I) Foxo1 and (J) Ucp1 expression in differentiated primary adipocytes from sWAT of control and AdR2KO mice. After differentiation for 8 days, mature adipocytes were treated with or without 10 μM isoproterenol (Iso), 1 μM AS1842856, or 50 μM SP600125 for 6 h n = 5–6 wells in (I); n = 3–8 wells in (J). (K) The mRNA levels of Mapk8 (encodes JNK1), Foxo1 , Ppargc1a , and Ucp1 in mature mouse adipocyte cells, 3T3-L1. Adipocytes were transfected with siRNAs for 72 h and then treated with 10 μM isoproterenol (Iso) for 6 h ( n = 6 wells per group). (L) The mRNA levels of Foxo1 , Ppargc1a , and Ucp1 in differentiated mouse primary adipocytes from sWAT of control and AdR2KO mice. Adipocytes were transduced with low ( Foxo1L ) and high ( Foxo1H ) MOI of AAV-8 containing mouse Foxo1 plasmid for 7 days and then treated with 20 μM isoproterenol (Iso) for 16 h ( n = 8 wells per group). The immunoblot images are representative of at least three independent experiments. p values were determined by the two-tailed Student’s t test (A and F) or one-way ANOVA with Tukey’s multiple comparisons test (B-E and G-L). All data are presented as the mean ± SEM of at least three independent biological replicates. See also and . AS: AS1842856; SP: SP600125.

Article Snippet: The 3T3-L1 cell line was purchased from the American Type Culture Collection (ATCC) and authenticated by ATCC (catalog number: CL-173).

Techniques: Gene Expression, Western Blot, Expressing, Control, Transfection, Transduction, Plasmid Preparation, Two Tailed Test

Journal: Food Science & Nutrition

Article Title: A Herbal Pair of Taraxacum officinale F.H.Wigg. and Lonicera japonica Thunb. Ameliorates Obesity and Modulates AMPK Signaling

doi: 10.1002/fsn3.71774

Figure Lengend Snippet: Effect of PL‐700 on lipid accumulation in 3T3‐L1 cells. (A) Oil red O staining of fully differentiated 3T3‐L1 adipocytes treated with LIPO‐700 (1, 10, and 100 μg/mL) for 24 h on day 7 of differentiation. The histogram shows quantification of lipid content expressed as a percentage relative to the MDI‐only group. (B) Western blot analysis of p‐AMPK, AMPK, PEPCK, G6Pase, and GAPDH protein expression in 3T3‐L1 adipocytes after 24‐h treatment with LIPO‐700. (C) Western blot analysis of ATGL and HSL protein expression in 3T3‐L1 adipocytes after 24‐h treatment with LIPO‐700. GAPDH was used as a loading control. (D) RT‐PCR analysis of SREBP‐1c, leptin, and LPL mRNA expression in 3T3‐L1 adipocytes following 24‐h LIPO‐700 treatment. GAPDH was used as an internal control. ### p < 0.001, vs. untreated control; * p < 0.05, ** p < 0.01, *** p < 0.001, vs. MDI‐treated group.

Article Snippet: Mouse preadipocyte 3T3‐L1 cells (CVCL_0123; Cat# CL‐173, ATCC, Manassas, VA, USA) were cultured in Dulbecco's Modified Eagle Medium (DMEM; Gibco, Thermo Fisher Scientific, Grand Island, NY, USA) supplemented with 10% calf serum (Gibco, Thermo Fisher Scientific, Grand Island, NY, USA) and 1% penicillin–streptomycin (Gibco, Thermo Fisher Scientific, Grand Island, NY, USA) at 37°C in a humidified atmosphere containing 5% CO 2 (Jackson et al. ; Kaczmarek et al. ).

Techniques: Staining, Western Blot, Expressing, Control, Reverse Transcription Polymerase Chain Reaction