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. 2022 May 14;23(10):5499. doi: 10.3390/ijms23105499

Figure 4.

Figure 4

LEF1 down-regulation in early HGPS iPSCs-keratinocytes differentiation. (A) Western blot analysis of total and non-phospho (active) β-catenin expression during normal and HGPS iPSCs-keratinocytes induction. Experiments were repeated at least three times, and representative data are shown as indicated. (B) Western blot analysis of nuclear and cytosolic non-phospho (active) β-catenin level in early-differentiating (week 1) normal and HGPS iPSCs. Regulator of Chromosome Condensation 1 (RCC1) and S6 were used as a nuclear marker and the overall loading control, respectively. Experiments were repeated at least three times, and representative data are shown as indicated. (C) Quantitative RT-PCR analysis of the relative expression of WNT transcription factors LEF1, TCF7, TCF7L1, and TCF7L2 before (day 0) and after induction with RA and BMP-4 (day 5) in normal and HGPS iPSCs differentiation. Data were normalized to endogenous ACTB mRNA and to the average of Normal day 0. Data are presented as mean ± SD (n = 3). * p < 0.05, ** p < 0.01, ns, not significant, two-way ANOVA followed by Tukey’s multiple comparisons test. (D) Quantitative RT-PCR analysis of the relative expression of LEF1 during normal and HGPS iPSCs-keratinocytes induction. Data were normalized to endogenous ACTB mRNA and to the average of Normal week 1. Data are presented as mean ± SD (n = 3). ** p < 0.01, *** p < 0.001, ns, not significant, two-way ANOVA followed by Sidak’s multiple comparisons test. (E) Western blot analysis of LEF1 expression during normal and HGPS iPSCs-keratinocytes induction. Experiments were repeated at least three times, and representative data are shown as indicated.