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Effect of pharmacological inhibition of PPARβ in malignant human keratinocytes (SCC13). (A) Cell proliferation quantified using EdU uptake (left) and FACS analysis of the cell cycle progression (right) in control (DMSO) and PPARβ‐inhibited <t>(GSK0660)</t> malignant human keratinocytes (SCC13). (B) Expression of p21 at mRNA (CDKA1A, quantified by qPCR; left) and protein (representative western blot, middle panel; right panel represents the quantification of three independent western blots) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (C) Expression of CHK1, E2F1 and PCNA at the mRNA (qPCR) and protein levels (western blot) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (D) Representative western blots (left) and quantification of three independent western blots (right) of phosphorylated H2AX (γH2AX) (upper panel), of total and phosphorylated CHK1 (middle panels), and of p21 at 0, 0.5, 2, and 6 h after UVB‐exposure in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (E) Percentage of apoptotic cells in response to UVB exposure using FACS analysis of Annexin V on control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. Actin and GAPDH have been used as loading controls. Bars represent mean ± standard deviation from three independent biological replicates (white circles), each with three technical replicates (panels A–C, and E). For western blots (panel D), each white circle corresponds to one independent experiment ( n = 3). (**** p < .0001; ** p < .01; * p < .05, two‐tailed Student's t ‐test for two group comparisons, and two‐way ANOVA with Tukey's multiple‐comparison test for Annexin V FACS).
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Effect of pharmacological inhibition of PPARβ in malignant human keratinocytes (SCC13). (A) Cell proliferation quantified using EdU uptake (left) and FACS analysis of the cell cycle progression (right) in control (DMSO) and PPARβ‐inhibited (GSK0660) malignant human keratinocytes (SCC13). (B) Expression of p21 at mRNA (CDKA1A, quantified by qPCR; left) and protein (representative western blot, middle panel; right panel represents the quantification of three independent western blots) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (C) Expression of CHK1, E2F1 and PCNA at the mRNA (qPCR) and protein levels (western blot) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (D) Representative western blots (left) and quantification of three independent western blots (right) of phosphorylated H2AX (γH2AX) (upper panel), of total and phosphorylated CHK1 (middle panels), and of p21 at 0, 0.5, 2, and 6 h after UVB‐exposure in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (E) Percentage of apoptotic cells in response to UVB exposure using FACS analysis of Annexin V on control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. Actin and GAPDH have been used as loading controls. Bars represent mean ± standard deviation from three independent biological replicates (white circles), each with three technical replicates (panels A–C, and E). For western blots (panel D), each white circle corresponds to one independent experiment ( n = 3). (**** p < .0001; ** p < .01; * p < .05, two‐tailed Student's t ‐test for two group comparisons, and two‐way ANOVA with Tukey's multiple‐comparison test for Annexin V FACS).

Journal: The FASEB Journal

Article Title: Transcriptional and functional regulation of cell cycle and UV response by PPARβ in human skin epidermal cells

doi: 10.1096/fj.202401950R

Figure Lengend Snippet: Effect of pharmacological inhibition of PPARβ in malignant human keratinocytes (SCC13). (A) Cell proliferation quantified using EdU uptake (left) and FACS analysis of the cell cycle progression (right) in control (DMSO) and PPARβ‐inhibited (GSK0660) malignant human keratinocytes (SCC13). (B) Expression of p21 at mRNA (CDKA1A, quantified by qPCR; left) and protein (representative western blot, middle panel; right panel represents the quantification of three independent western blots) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (C) Expression of CHK1, E2F1 and PCNA at the mRNA (qPCR) and protein levels (western blot) in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (D) Representative western blots (left) and quantification of three independent western blots (right) of phosphorylated H2AX (γH2AX) (upper panel), of total and phosphorylated CHK1 (middle panels), and of p21 at 0, 0.5, 2, and 6 h after UVB‐exposure in control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. (E) Percentage of apoptotic cells in response to UVB exposure using FACS analysis of Annexin V on control (DMSO) and PPARβ‐inhibited (GSK0660) SCC13 cells. Actin and GAPDH have been used as loading controls. Bars represent mean ± standard deviation from three independent biological replicates (white circles), each with three technical replicates (panels A–C, and E). For western blots (panel D), each white circle corresponds to one independent experiment ( n = 3). (**** p < .0001; ** p < .01; * p < .05, two‐tailed Student's t ‐test for two group comparisons, and two‐way ANOVA with Tukey's multiple‐comparison test for Annexin V FACS).

Article Snippet: Cells were treated with vehicle DMSO or PPARβ antagonist GSK0660 (MedChemExpress, HY‐12377) at a final concentration of 0.5 μM for 24 h prior to UVB exposure and kept under GSK0660 treatment for the duration of the experiment.

Techniques: Inhibition, Control, Expressing, Western Blot, Standard Deviation, Two Tailed Test, Comparison