Smad3 has a critical role in TGF-beta-mediated growth inhibition and apoptosis in colonic epithelial cells.

Published

Journal Article

BACKGROUND: Smad proteins play a key role in TGF-beta signaling that regulates cell proliferation, differentiation, and apoptosis. Mice deficient in Smad3 develop colonic adenocarcinoma. MATERIALS AND METHODS: We developed a Smad3-deficient colonocyte cell line that was used to study TGF-beta-mediated growth inhibition and induction of apoptosis was compared to young adult mouse colonocyte (YAMC) control cells. Growth inhibition was assessed by cell count and ((3)H)-thymidine incorporation assay. Transcriptional response to TGF-beta was measured by transfecting the reporters p3TP-Lux and p(CAGA)(9)-MLP-luc. TGF-beta-induced apoptosis was assessed using ELISA and Hoechst staining. Mediators of cell-cycle arrest and apoptosis were assayed by Western blot. RESULTS: Smad3-/- cells were resistant to TGF-beta-mediated growth inhibition compared to control cells. Ninety-eight percent of cell count growth inhibition observed in YAMC cells, while 34% inhibition was observed in Smad3-/- cells after TGF-beta treatment. ((3)H)-thymidine incorporation was inhibited by 61% in YAMC cells, while Smad3-/- cells showed 25% inhibition after TGF-beta treatment. Smad3-/- cells were deficient in luciferase reporter induction by TGF-beta. TGF-beta induced apoptosis 8-fold in YAMC cells, but had no effect on apoptosis in Smad3-/- cells. p21(Cip11) and PAI-1 are induced in YAMC cells by TGF-beta, but unchanged in Smad3-/- cells. TGF-beta decreases cyclin D1 levels in YAMC cells but does not affect levels in Smad3-/- cells. CONCLUSIONS: Our findings suggest that the loss of Smad3 contributes to resistance of TGF-beta growth inhibition and apoptosis in colonic epithelium. This may represent a mechanism by which cells are able to escape antiproliferative controls and embark on a pathway toward neoplasia.

Full Text

Duke Authors

Cited Authors

  • Mithani, SK; Balch, GC; Shiou, S-R; Whitehead, RH; Datta, PK; Beauchamp, RD

Published Date

  • April 2004

Published In

Volume / Issue

  • 117 / 2

Start / End Page

  • 296 - 305

PubMed ID

  • 15047135

Pubmed Central ID

  • 15047135

Electronic International Standard Serial Number (EISSN)

  • 1095-8673

International Standard Serial Number (ISSN)

  • 0022-4804

Digital Object Identifier (DOI)

  • 10.1016/s0022-4804(03)00335-4

Language

  • eng