title: Isolation and characterization of myofibroblast cell lines from oral squamous cell carcinoma.
abstract: Oral squamous cell carcinoma (OSCC) invasion is followed by several stromal events such as inflammatory and immune cell infiltration, neo-vascularization, fibroblast activation and occasionally myofibroblast emergence. Our previous studies demonstrated that myofibroblasts in the stroma of OSCC are associated with a more aggressive behavior, leading to shorter patient overall survival. Therefore, we evaluated whether OSCC-associated myofibroblasts have different characteristics compared to OSCC-associated fibroblasts. OSCC myofibroblast cell lines were isolated, cultured and characterized on the basis of the expression of specific isoform α of smooth muscle actin (α-SMA) and of the excessive production of type I collagen. To assess the proliferative potential of the cell lines, growth curves were constructed, whereas the production and activity of matrix metalloproteinases (MMP) were analyzed by ELISA and enzymography, respectively. Myofibroblast clones were positive for α-SMA and vimentin, and negative for pan-cytokeratin and CD34. In long time cultures, western blotting, flow cytometry and ELISA analysis revealed constant α-SMA expression and elevated production of type I collagen. There were no differences on proliferative potential between fibroblast and myofibroblast clones, but myofibroblast cells secreted significantly higher levels of MMP-1, -2, -9 and -13. Furthermore, MMP-2 gelatinolytic activity was significantly higher in myofibroblast clones. The results of this study suggest that myofibroblasts may contribute to OSCC invasion through elevation of MMP synthesis.
authors.affiliation: Department of Oral Diagnosis, School of Dentistry, State University of Campinas, Piracicaba, São Paulo, Brazil.
authors.forename: Lays Martin
authors.initials: LM
authors.lastname: Sobral
authors.forename: Karina Gottardello
authors.initials: KG
authors.lastname: Zecchin
authors.forename: Sibele
authors.initials: S
authors.lastname: Nascimento de Aquino
authors.forename: Marcio Ajudarte
authors.initials: MA
authors.lastname: Lopes
authors.forename: Edgard
authors.initials: E
authors.lastname: Graner
authors.forename: Ricardo D
authors.initials: RD
authors.lastname: Coletta
mesh_terms: D000199:Actins / Q000378:metabolism; D017209:Apoptosis; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology*; D049109:Cell Proliferation; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007124:Immunoenzyme Techniques; D020782:Matrix Metalloproteinases / Q000378:metabolism; D009062:Mouth Neoplasms / Q000378:metabolism / Q000473:pathology*; D009130:Muscle, Smooth / Q000378:metabolism / Q000473:pathology; D058628:Myofibroblasts / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017154:Stromal Cells / Q000378:metabolism / Q000473:pathology*; D014407:Tumor Cells, Cultured
chemical_list: D000199:Actins; D012333:RNA, Messenger; D020782:Matrix Metalloproteinases