Fig. 3. MrgprF overexpression inhibited tumor cell migration.
a, b MrgprF overexpression inhibited A375 cell migration by wound healing (a) and trans-well assays (b) in indicated A375 cells. Quantification data were also indicated for each assay. c Indicated cell extracts were probed with indicated antibodies to examine the protein expressions of E-cadherin, N-cadherin, and Vimentin. d Representative images showing the morphological change in A375 after MrgprF overexpression under bright field. Scale bar = 50 μm. e Immunofluorescence staining of Phalloidine in indicated cells. White head arrows pointing to the pseudopodia like structure. Scale bar = 25 μm. f MrgprF knockdown in A375 promoted cell migration ability as determined by trans-well assay. Quantification data is also shown. Scale bar = 50 μm. g Establishment of mouse MrgprF (mMrgprF) overexpression in mouse B16 cells, verified by Real-time RT-PCR (top) and immunoblot (bottom). Green arrow: exogenous mMrgprF-Flag; black arrow: endogenous mMrgprF. h, i Lung metastasis model using B16 cells in C57BL/6 mice. Representative images for the lung tissues (h) and the quantification result for the metastatic tumor numbers (i) are shown. j, k Representative IHC staining images for MrgprF, E-cadherin, N-cadherin and Ki67. k Quantification data for (j). l Immunofluorescence staining of phalloidine in HaCaT cells after MrgprF knockdown. White head arrows pointing to the pseudopodia like structure. Scale bar = 25 μm. m–o HaCaT cells stably expressing control or MRGPRF targeting shRNAs were subcutaneously injected (2 × 107 cells/mouse) into nude mice, and representative xenograft tumor images (m), tumor-free mice percentage (n) and H&E staining images of the xenograft tumors (o) are shown. Scale bar = 50 μm. Quantified results for all the immunoblots are indicated below, which are normalized to the β-actin signal, compared to reciprocal control. Bars are the mean value ± SD. *P < 0.05, **P < 0.01, ***P < 0.001