Journal: EMBO Molecular Medicine
Article Title: Impaired migration and lung invasion of human melanoma by a novel small molecule targeting the transmembrane domain of death receptor p75 NTR
doi: 10.1038/s44321-025-00297-1
Figure Lengend Snippet: ( A ) Representative Western blot analysis (IB) of fascin in p75 NTR immunoprecipitates (IP) and whole cell lysates (WCL) of A875 melanoma cells after overnight treatment with NGF or Np75-4A22 as indicated. The histogram on the right shows quantification expressed as average of three independent experiments, each performed in triplicate ± SD. # P = 0.0091 vs. control (first bar); ** P = 0.0088 vs. NGF alone (second bar) (one-way ANOVA followed by Tukey’s multiple comparisons). ( B ) Representative Western blot analysis of fascin in Triton-insoluble extracts of A875 melanoma cells after overnight treatment with NGF or Np75-4A22 as indicated. The histogram on the right shows quantification expressed as average of three independent experiments, each performed in triplicate ± SD. # P = 0.0412 vs. control (first bar); ** P = 0.0082 vs. NGF alone (second bar) (one-way ANOVA followed by Tukey’s multiple comparisons). ( C ) Photomicrographs of cultured A875 control (NT) and knock-down (shp75) cells treated with NGF or Np75-4A22 as indicated and stained with fluorophor-conjugated phalloidin (green), to reveal actin filaments, and DAPI (blue). White arrows denote some of the filopodia in NGF-treated cells. Scale bar, 5 μM. ( D , E ) Quantification of filopodia per cell in A875 NT ( D ) and A875 shp75 ( E ) cells treated with 100 ng/ml NGF or 1 or 5 μM Np75-4A22 as indicated. Average of three independent experiments, each performed in triplicate, are shown ± SEM. ### P = 0.0004 vs. vehicle (first bar); *** P = 0.0001 vs. NGF alone (second bar). .
Article Snippet: For immunoprecipitation, A875 cell extracts were incubated for 16 h at 4 °C on a rotating wheel with 0.8 μg of anti-p75 NTR antibody (ANT-007, Alomone) and then incubated with Sepharose protein-G beads (GE Healthcare).
Techniques: Western Blot, Control, Cell Culture, Knockdown, Staining