Fig. 3
Internalized PTEN alters signaling, retains polarity, and inhibits the EMT and invasion of 344SQ cells. a Immunoblot analysis of indicated proteins in 344SQ cell lysates after incubation with CM for the indicated times. b Immunoblot analysis of PTEN in 344SQ cells after incubation with ApoSQ-exposed CM at various dilutions (from 1/5 to 1) with control CM for 12 h. The qPCR analysis of PTEN mRNA is shown in (c). The immunoblot analysis of indicated proteins in 344SQ cells after the addition of TGF-β1 (10 ng/ml) is shown in (d) and that after the addition of CM is shown in (e). f–l RAW cells were transfected with PTEN siRNA (#1 siPTEN) before ApoSQ cell stimulation for 24 h. f Immunoblot analysis of PTEN in RAW cells. g–l CM was added to 344SQ cells with or without TGF-β1 (10 ng/ml) for the indicated times. g, h, j Immunoblot analysis of indicated proteins in 344SQ cells. i Confocal microscopic images of 3D acini at 12 h after treatment with TGF-β1 (10 ng/ml). Normal acini of 344SQ cells were grown in 3D Matrigel containing CM and stained with anti-β-catenin (green) and DAPI. Scale bars: 20 μm. k qPCR analysis of Snai1 and Zeb1 mRNAs in 344SQ cells. l The numbers of invaded cells were analyzed to assess their invasive ability using Matrigel-coated Transwells. NS not significant; ***P < 0.001. Data are from three independent experiments (mean ± s.e.m. in (c, k, l)) or one experiment representative of three independent experiments with similar results, as shown in (a, b, d–j)
