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. 2016 Dec 2;48(12):e275. doi: 10.1038/emm.2016.107

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Copyright © 2016 The Author(s)

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BAMBI functions as a tumor suppressor in vitro and in vivo. (a) BAMBI-mediated morphological changes in HepG2 cells stably overexpressing the BAMBI protein (upper panels). Immunoblot analysis of β-catenin and TGF-β1 in the stable transfectants overexpressing the c-Myc-tagged BAMBI protein (lower panels). (b) The BAMBI-expressing HepG2 cells showed minimal colony growth in soft agar compared with that of the vector control (upper panels). The colonies shown are 15 days old. Quantification was performed in triplicate (lower panel). The values represent the mean±s.d. from three independent experiments. **P<0.01. (c) Growth of the tumor masses in BAMBI-expressing HepG2 cells and vector control cells injected into the left shoulders (red arrows) of nude mice. Vector control cell lines injected into the right shoulder (blue arrows), grew quickly and formed tumor masses, whereas the BAMBI-expressing cells did not (upper panels). The tumor volume was measured as a function of time (lower panel). Each value represents the mean±s.d. (d) BAMBI expression in SH-J1 cells infected with either Ad-LacZ or Ad-BAMBI (left). The Ad-BAMBI-infected SH-J1 cells showed minimal colony growth in soft agar compared with the parent or Ad-LacZ-infected cells (right upper panels). The colonies shown are 15 days old. Quantification was performed in triplicate (right lower panel). The values represent the mean±s.d. of three independent experiments. **P<0.01. (e) Bioluminescent images of the subcutaneous tumor masses (n=5 mice per group, representative anterior–posterior images, 1 min exposure time). SH-J1 cells infected with Ad-LacZ subcutaneously injected into the right shoulder (blue arrows) grew quickly and formed tumor masses, but the Ad-BAMBI-infected cells injected into the left shoulder (red arrows) formed rare, small tumor masses (upper panels). Quantitative measurement of photon flux (lower panel). The values represent the mean±s.d. (f) Silencing of TGF-β1/BAMBI/β-catenin protein expression by lentiviral delivery of shRNA (upper panels) in HepG2 cells and their clonogenic ability in the soft agar (middle panels) and quantification (lower panel). The values represent the mean±s.d. of three independent experiments. **P<0.01. (g) A schematic model of the inhibition of BAMBI signaling by ctHBx. HBx profoundly stimulates Wnt/β-catenin signaling, and overexpression of BAMBI suppresses both TGF-β1 and β-catenin signaling and subsequently inhibits tumorigenicity. , inhibition; , stimulation.

Inline graphic — BAMBI functions as a tumor suppressor in vitro and in vivo. (a) BAMBI-mediated morphological changes in HepG2 cells stably overexpressing the BAMBI protein (upper panels). Immunoblot analysis of β-catenin and TGF-β1 in the stable transfectants overexpressing the c-Myc-tagged BAMBI protein (lower panels). (b) The BAMBI-expressing HepG2 cells showed minimal colony growth in soft agar compared with that of the vector control (upper panels). The colonies shown are 15 days old. Quantification was performed in triplicate (lower panel). The values represent the mean±s.d. from three independent experiments. **P<0.01. (c) Growth of the tumor masses in BAMBI-expressing HepG2 cells and vector control cells injected into the left shoulders (red arrows) of nude mice. Vector control cell lines injected into the right shoulder (blue arrows), grew quickly and formed tumor masses, whereas the BAMBI-expressing cells did not (upper panels). The tumor volume was measured as a function of time (lower panel). Each value represents the mean±s.d. (d) BAMBI expression in SH-J1 cells infected with either Ad-LacZ or Ad-BAMBI (left). The Ad-BAMBI-infected SH-J1 cells showed minimal colony growth in soft agar compared with the parent or Ad-LacZ-infected cells (right upper panels). The colonies shown are 15 days old. Quantification was performed in triplicate (right lower panel). The values represent the mean±s.d. of three independent experiments. **P<0.01. (e) Bioluminescent images of the subcutaneous tumor masses (n=5 mice per group, representative anterior–posterior images, 1 min exposure time). SH-J1 cells infected with Ad-LacZ subcutaneously injected into the right shoulder (blue arrows) grew quickly and formed tumor masses, but the Ad-BAMBI-infected cells injected into the left shoulder (red arrows) formed rare, small tumor masses (upper panels). Quantitative measurement of photon flux (lower panel). The values represent the mean±s.d. (f) Silencing of TGF-β1/BAMBI/β-catenin protein expression by lentiviral delivery of shRNA (upper panels) in HepG2 cells and their clonogenic ability in the soft agar (middle panels) and quantification (lower panel). The values represent the mean±s.d. of three independent experiments. **P<0.01. (g) A schematic model of the inhibition of BAMBI signaling by ctHBx. HBx profoundly stimulates Wnt/β-catenin signaling, and overexpression of BAMBI suppresses both TGF-β1 and β-catenin signaling and subsequently inhibits tumorigenicity. , inhibition; , stimulation.