Smad7 blocks transforming growth factor-β1-induced gingival fibroblast-myofibroblast transition via inhibitory regulation of Smad2 and connective tissue growth factor.
BACKGROUND
Transforming growth factor-β1 (TGF-β1), its downstream signaling mediators (Smad proteins), and specific targets, including connective tissue growth factor (CTGF), play important roles in tissue remodeling and fibrosis via myofibroblast activation. We investigated the effect of overexpression of Smad7, a TGF-β1 signaling inhibitor, on transition of gingival fibroblast to myofibroblast. Moreover, we analyzed the participation of CTGF on TGF-β1-mediated myofibroblast transformation.


METHODS
To study the inhibitory effect of Smad7 on TGF-β1/CTGF-mediating gingival fibroblast transition into myofibroblasts, we stably overexpressed Smad7 in normal gingival fibroblasts and in myofibroblasts from hereditary gingival fibromatosis (HGF). Myofibroblasts were characterized by the expression of the specific marker isoform α of the smooth muscle actin (α-SMA) by Western blot, flow cytometry, and immunofluorescence. Enzyme-linked immunosorbent assay for type I collagen was performed to measure myofibroblast activity. CTGF's role on myofibroblast transformation was examined by enzyme-linked immunosorbent assay and small interference RNA.


RESULTS
TGF-β1 induced the expression of α-SMA and CTGF, and small interference RNA-mediating CTGF silencing prevented fibroblast-myofibroblast switch induced by TGF-β1. In Smad7-overexpressing fibroblasts, ablation of TGF-β1-induced Smad2 phosphorylation marked decreased α-SMA, CTGF, and type I collagen expression. Similarly, HGF transfectants overexpressing Smad7 demonstrated low levels of α-SMA and phospho-Smad2 and significant reduction on CTGF and type I collagen production.


CONCLUSIONS
CTGF is critical for TGF-β1-induced gingival fibroblast-myofibroblast transition, and Smad7 overexpression is effective in the blockage of myofibroblast transformation and activation, suggesting that treatments targeting myofibroblasts by Smad7 overexpression may be clinically effective in gingival fibrotic diseases, such as HGF.
Department of Oral Diagnosis, School of Dentistry, State University of Campinas, Piracicaba, São Paulo, Brazil.
Lays M
LM
Sobral
Patrick Franz
PF
Montan
Karina Gottardello
KG
Zecchin
Hercílio
H
Martelli-Junior
Pablo Agustin
PA
Vargas
Edgard
E
Graner
Ricardo D
RD
Coletta
D054338:Cell Transdifferentiation / Q000502:physiology*; D055513:Connective Tissue Growth Factor / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D005881:Gingiva / Q000166:cytology; D006801:Humans; D058628:Myofibroblasts / Q000166:cytology* / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D051899:Smad2 Protein / Q000378:metabolism; D051906:Smad7 Protein / Q000502:physiology*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
D051899:Smad2 Protein; D051906:Smad7 Protein; D053773:Transforming Growth Factor beta1; D055513:Connective Tissue Growth Factor