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BBB Seminar: Daniela Elena Costea

Carcinoma-associated fibroblasts promote a cancer stem cell-like phenotype through TGFβ1 secretion

Daniela Elena Costea
Department of Biomedicine, University of Bergen

We previously showed the ability of cancer associated fibroblasts (CAFs) to support invasion of oral squamous cell carcinoma (OSCC) cells. We have now investigated the ability of CAFs to induce clonogenic and tumorigenic abilities using several cell lines derived from oral dysplasia and OSCC, as well as primary cells from OSCC patients. Stem cell properties and molecular changes resulting from in vitro growth or following xenotransplantation in NOD/SCID mice of cells either alone or with CAFs were assessed. Colony- and sphere-forming abilities were significantly enhanced (p<0.05) when pre-malignant and primary OSCC cells were grown with CAFs. FACS and qPCR showed that stem cell-related markers (CD44, Oct4A, Bmi-1) were up-regulated several fold in pre-malignant and primary OSCC cells grown with CAFs. CAFs induced the low-clonogenic, low-tumorigenic, and low-metastatic (CD44low) fraction of primary OSCC cells into highly clonogenic, tumorigenic and metastatic cells (CD44high). CAFs secreted significantly higher amounts of TGFβ1 than normal fibroblasts, and TGFβ1 and CAF-conditioned medium (CAF-CM) increased colony- and sphere-formation in OSCC cell lines, and the expression of stem cell-related markers CD44 and βcatenin. In addition, both TGFβ1 and CAF-CM increased the subpopulation of CD44high in OSCC cell lines, while the small molecule TGFβ1 inhibitor SB431542 decreased it. Of interest, TGFβ1 increased the clonogenic and tumorigenic abilities and expression of stem cell markers preferentially in the low clonogenic OSCC cells. CD44low cells expressed higher levels of TGFβ1 receptors and responded to TGFβ1 treatment by a higher level of phosphorylation of Smad proteins than CD44high cells, indicating that this subpopulation is more prone, and indeed responded better to TGFβ1 treatment. Of interest, TGFβ1 had a negative effect on clonogenicity and expression of stem-cell related markers in pre-malignant cells, indicating once more the dual role of TGFβ1 in carcinogenesis. In conclusion, this study provides functional and molecular evidence that CAFs enhance self-renewal and a tumorigenic ability of premalignant and malignant oral cells, and that TGFβ1 is an essential mediator of this effect in malignant but not pre-malignant cells, by preferentially activating and de-differentiating the non-stem cancer cells.

Chair: Donald Gullberg <donald.gullberg@biomed.uib.no>, Department of Biomedicine