Influence of TGF-β1 on tumor transition in oral cancer cell and BMSC co-cultures.
OBJECTIVE
TGF-β1 signaling modulates epithelial mesenchymal transitions (EMT) of head and neck squamous cell carcinoma (HNSCC). Bone marrow mesenchymal stromal cells (BMSC) are able to exert a regulating influence on the expression of markers of EMT in HNSCC cells. It was thus the aim of this study to test the hypothesis that TGF-β1 modulates the interactions of tumor transition between BMSCs and HNSCC, affecting the expression of E-cadherin, Vimentin, Snail, Twist, MMP14 and beta-catenin. Furthermore, we analyzed alterations in the AKT-signaling of tumor and stroma cells.


METHODS
BMSCs were isolated from iliac bone marrow aspirates and co-cultured in trans-well permeable membrane wells with tumor cells of the established HNSCC cell line PCI-13. Following the induction with TGF-β1 under serum free conditions the expression of Vimentin and E-Cadherin was assessed via immunofluorescence. A quantitative RT-PCR analysis of tumor transition markers E-cadherin, Vimentin, Snail, Twist, MMP14 and beta-catenin was performed. Changes in AKT-Signaling were identified via protein analysis.


RESULTS
In non-induced co-cultures, BMSC were able to suppress Vimentin in PCI-13 as a marker of tumor transition. In TGF-β1 induced co-cultures PCI-13 significantly increased the expression of Vimentin, Twist, Snail, MMP14, GSK3a, PRAS40, 4E-BP1, and AMPKa compared to monolayer controls. TGF-β1 co-cultured BMSC demonstrated a significant increase of Snail, PRAS40, mTOR, GSK3a/b, Bad, PDK1 and 4E-BP1.


CONCLUSIONS
TGF-β1 was able to attenuate the modulating influence of BMSC in co-culture and drive the co-culture towards a progressive tumor transition, affecting the expression of markers of EMT, AKT-Signaling and proliferative checkpoints.
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany. Electronic address: florian.boehrnsen@med.uni-goettingen.de.
F
F
Böhrnsen
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany; Department of Preventive Dentistry, Periodontology and Cardiology, University Medical Center Göttingen, Germany.
F
F
Godek
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany.
J
J
Kiesel
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany.
F J
FJ
Kramer
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany.
P
P
Brockmeyer
Clinic of Oral and Maxillofacial Surgery, University Medical Center Göttingen, Germany.
H
H
Schliephake
D000293:Adolescent; D000328:Adult; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology; D002471:Cell Transformation, Neoplastic / Q000187:drug effects*; D002648:Child; D018920:Coculture Techniques; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D006258:Head and Neck Neoplasms / Q000378:metabolism / Q000473:pathology; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008875:Middle Aged; D009062:Mouth Neoplasms / Q000378:metabolism / Q000473:pathology*; D060888:Real-Time Polymerase Chain Reaction; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*; D055815:Young Adult
D053773:Transforming Growth Factor beta1
BMSC; Co-culture; EMT; HNSCC; TGF-beta; Transition