Journal: International Journal of Molecular Sciences

Article Title: Derivation of Porcine Extra-Embryonic Endoderm Cell Lines Reveals Distinct Signaling Pathway and Multipotency States

doi: 10.3390/ijms222312918

Figure Lengend Snippet: Signaling dependence analysis of pXEN cells. ( A ) The morphology and JC1 staining of pXEN cells cultured in LCDM, LCDM+10μm SD1008, LCDM+10μm SU5402 and LCDM+10μm SB431542. ( B ) Quantitative RT-PCR analysis of FGF, TGFβ and LIF signaling related genes in pXEN cells, porcine nESCs and porcine embryo fibroblasts (PEF). Relative expression reflected as a fold difference in pXEN cells and porcine nESCs compared to PEF, PEF = 1. Data are depicted as mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 versus Control ( t -test). ( C ) The morphology of pXEN cells cultured in LCDM and LCDM supplemented with 1, 5 and 10μM concentration of PD0325901. ( D ) Western blotting analysis of the phosphorylation status of ERK, STAT3 andSMAD2/3, and the expression of ERK, STAT3 andSMAD2/3 in pXEN cells and porcine nESCs. ( E ) The quantification of proteins after Western blotting. Scale bar, 100μm.

Article Snippet: The cells were cultured in medium supplemented with or without the JAK inhibitor SD1008 (Tocris), the FGF receptor (FGFR) inhibitor SU5402 (Tocris), Mek inhibitor PD0325901 (Sigma) and the TGFβ receptor inhibitor SB431542 (Tocris) for LIF, FGF and TGFβ signal pathway identification, respectively.

Techniques: Staining, Cell Culture, Quantitative RT-PCR, Expressing, Control, Concentration Assay, Western Blot, Phospho-proteomics

Journal: PLoS ONE

Article Title: Differential effects of amnion and chorion membrane extracts on osteoblast-like cells due to the different growth factor composition of the extracts

doi: 10.1371/journal.pone.0182716

Figure Lengend Snippet: (A) MG-63 cells were cultured in OIM supplemented with DMSO (0.1% v/v) and were pretreated with various concentrations of SU5402 (0.1–1 μM) or SB505124 (0.5–1 μM) for 1 h, followed by treatment with 400 μg/mL CME. After 9 days, mineralization was determined by performing Alizarin red S staining. Alizarin red S stain was extracted using 10% cetylpyridinium chloride, and absorbance was measured at 570 nm. (B) For the calcium assay, the cells were pretreated with DMSO (0.1% v/v) and various concentrations of SU5402 (0.1–1 μM) or SB505124 (0.5–1 μM) alone or their combination for 1 h, followed by treatment with 200 μμg/mL CME. Calcium assay was performed after 9 days. (C) Expression of genes encoding ALP and IBSP was determined on day 4 by performing quantitative real-time RT-PCR under the experimental condition described in (A): SU5402: FGF specific inhibitor, SB505124: TGFβ-1 specific inhibitor, SU+SB: SU5402+SB505124. Data are presented as the mean ± SD of multiple repeated experiments; * p < 0.05, ** p < 0.01, and # p < 0.001 versus only CME, indicate statistical significance.

Article Snippet: Recombinant human bFGF, TGFβ-1, EGF, and BMP2 were purchased from Peprotech (Rocky Hill, NJ, USA), and inhibitors of FGF (SU5402), TGFβ-1 (SB505124), and EGF (PD153035) were purchased from Santa Cruz Biotechnology (Dallas, Texas, USA).

Techniques: Cell Culture, Staining, Calcium Assay, Expressing, Quantitative RT-PCR