fgf receptor fgfr inhibitor su5402  (Tocris)

Journal: Cell death & disease

Article Title: Activation of the JNKs/ATM-p53 axis is indispensable for the cytoprotection of dermal fibroblasts exposed to UVB radiation.

doi: 10.1038/s41419-022-05106-y

Figure Lengend Snippet: Fig. 4 UVB-induced cellular responses are EGFR-Akt mediated in human skin fibroblasts. A Human dermal fibroblasts were pre-incubated with 100 μM suramin for 1 h before exposure to 700 mJ/cm2 of UVB radiation. Then, cells were further incubated at 37 oC for 72 h and cell number was counted after staining with Neutral Red. A representative experiment out of three similar ones is shown. p for statistically significant differences in comparison to the respective samples in the absence of suramin (Student’s t test) is presented in the graph. B Cells were pre-treated with 100 μM suramin for 1 h before exposure to 700 mJ/cm2 of UVB radiation. Protein extracts after 1 and 12 h were subjected to western blot analysis for phospho-Akt (Ser473)/JNKs and phospho-p53, respectively. Representative blots of three independent experiments with similar results are depicted here. The non-phosphorylated forms of the kinases and actin were analyzed to verify equal loading. C A 100 μΜ suramin solution was analyzed spectrophotometrically in a range from 200 to 600 nm in order to assess its absorption spectrum. PBS served as the negative control. Gray shading marks the wavelength range corresponding to the UVB band (280–315 nm). D Cells were pre-incubated for 1 h with the growth factor receptor inhibitors [STI571 for PDGFR (2 μΜ); SU5402 for FGFR and VEGFR (20 μΜ); SB431542 for TGFβR1 (10 μΜ); I-OMe-AG538 for IGFIR (12 μΜ); AG1478 for EGFR (10 μΜ)] and then exposed to 700 mJ/cm2 UVB radiation. Cultures were further incubated at 37 oC for 72 h, cells were detached by trypsinization, stained with Neutral Red and counted in a haemocytometer. p for statistically significant differences in comparison to the respective samples with no inhibitor (Student’s t test) is provided. E Cells were incubated with 10 μΜ of the EGFR inhibitor AG1478 for 1 h, exposed to 700 mJ/cm2 of UVB radiation and further incubated at 37 oC for 1 or 12 h before protein extraction and western blot analysis for phospho-Akt (Ser473)/JNKs or phospho-p53, respectively. Representative blots of three independent experiments are shown here, while total Akt and JNKs, as well as actin were analyzed to validate equal loading. F Cell cultures were treated with 100 ng/ml EGF, exposed to 700 mJ/cm2 of UVB radiation and further incubated at 37 oC for 72 h. Neutral Red-positive cells were counted in a haemocytometer. A representative graph from three independent similar experiments is presented. p for statistically significant differences in comparison to the respective samples in the absence of the growth factor (Student’s t test) is shown.

Article Snippet: The PDGF receptor (PDGFR) kinase inhibitor STI571 was supplied by Novartis AG (Basel, Switzerland), the VEGF receptor (VEGFR) and FGF receptor (FGFR) inhibitor SU5402, the EGFR kinase inhibitor tyrphostin AG1478 and the IGFI receptor (IGFIR) kinase inhibitor tyrphostin I-OMe-AG538 were from CalbiochemMerck KGaA (Darmstadt, Germany), the EGFR inhibitor PD153035 was from MedChemExpress Co., Ltd (Monmouth Junction, NJ, USA) and the TGF-β type I receptor (TGFβR1) kinase inhibitor SB431542 was purchased from Sigma.

Techniques: Incubation, Staining, Comparison, Western Blot, Negative Control, Protein Extraction

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