Invasive breast cancer induces laminin-332 upregulation and integrin β4 neoexpression in myofibroblasts to confer an anoikis-resistant phenotype during tissue remodeling

Breast Cancer Res. 2012 Jun 6;14(3):R88. doi: 10.1186/bcr3203.

Abstract

Introduction: Although development of anoikis-resistant myofibroblasts during tissue remodeling is known to be associated with tumor invasion, the mechanism by which myofibroblasts become resistant to anoikis is unknown. We previously demonstrated laminin-332 upregulation in the fibrosis around invasive ductal carcinoma (IDC). Because laminin-332 promotes cell survival through binding to integrins, we hypothesized that invasive breast cancer cells confer an anoikis-resistant phenotype on myofibroblasts by upregulating laminin-332 expression during tissue remodeling. Here, we demonstrate that invasive breast cancer cells induce laminin-332 upregulation and integrin β4 neoexpression in myofibroblasts to confer an anoikis-resistant phenotype.

Methods: Three types of fibroblasts were isolated from the tumor burden, the fibrosis, and normal tissue of patients with early stage IDC (less than 10 mm diameter), designated cancer-associated fibroblasts (CAFs), interface fibroblasts (InFs), and normal breast fibroblasts (NBFs), respectively. To investigate direct and indirect crosstalk with tumor cells, fibroblasts were co-cultured with invasive MDA-MB-231 or noninvasive MCF7 cells or in conditioned medium. Anoikis resistance of fibroblasts was measured by cell viability and caspase-3 activity after incubation on poly-HEMA coated plates for 72 hours. Involvement of laminin-332/integrin α3β1 or α6β4 signaling in anoikis resistance was confirmed by treatment with purified laminin-332 or blocking antibodies against laminin-332, integrin β1, or integrin β4.

Results: MDA-MB-231 cells induced laminin-332 upregulation and integrin β4 neoexpression in fibroblasts, leading to anoikis resistance. InFs showed a higher endogenous level of laminin-332 than did CAFs and NBFs. After stimulation with MDA-MB-231-conditioned medium, laminin-332 expression of InFs was dramatically increased and maintained under anoikis conditions. Laminin-332 upregulation was also observed in CAFs and NBFs, but at a lower level than in InFs. Laminin-332 induced Akt (Ser473) phosphorylation by binding to integrin α3β1. Integrin β4 neoexpression induced laminin-332-independent Rac1 activation and promoted anoikis resistance in fibroblasts approximately twofold more effectively than did laminin-332, regardless of the type of fibroblast. In addition, integrin β4 expression suppressed fibroblast aggregation in conditions of anoikis.

Conclusion: Invasive breast cancer cells confer an anoikis-resistant phenotype on myofibroblasts during tissue remodeling by inducing laminin-332 upregulation and integrin β4 neoexpression. Interface fibroblasts appear to be the primary myofibroblasts that interact with invasive tumor cells during tissue remodeling.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anoikis*
  • Antibodies, Blocking
  • Breast Neoplasms / metabolism*
  • Breast Neoplasms / pathology*
  • Carcinoma, Ductal, Breast / metabolism*
  • Carcinoma, Ductal, Breast / pathology
  • Caspase 3 / metabolism
  • Cell Adhesion Molecules / immunology
  • Cell Adhesion Molecules / metabolism*
  • Cell Communication
  • Cell Line, Tumor
  • Cell Survival
  • Female
  • Fibrosis
  • Humans
  • Integrin alpha3beta1 / metabolism
  • Integrin beta1 / immunology
  • Integrin beta4 / immunology
  • Integrin beta4 / metabolism*
  • MCF-7 Cells
  • Myofibroblasts / metabolism*
  • Phosphorylation
  • Proto-Oncogene Proteins c-akt / metabolism
  • Up-Regulation
  • rac1 GTP-Binding Protein / metabolism

Substances

  • Antibodies, Blocking
  • Cell Adhesion Molecules
  • Integrin alpha3beta1
  • Integrin beta1
  • Integrin beta4
  • kalinin
  • Proto-Oncogene Proteins c-akt
  • Caspase 3
  • rac1 GTP-Binding Protein