Effect of transforming growth factor-beta1 on expression of the connective tissue growth factor (CCN2/CTGF) gene in normal human gingival fibroblasts and periodontal ligament cells.
OBJECTIVE
Connective tissue growth factor (CCN2/CTGF) plays an important role in wound healing and regulation of the extracellular matrix in periodontal tissue. However, the functional relationship between altered transforming growth factor-beta1 levels and CCN2/CTGF has not been extensively investigated in human gingival fibroblasts and periodontal ligament cells. This study investigated the effects of transforming growth factor-beta1 on the expression of the CCN2/CTGF gene in human gingival fibroblasts and periodontal ligament cells in vitro.


METHODS
Cells were isolated from normal periodontal tissues and cultured in Dulbecco's modified Eagle's minimal essential medium/F12 containing 10% fetal bovine serum. Subconfluent cells were maintained under serum deprivation for 24 h then treated with Dulbecco's modified Eagle's minimal essential medium/F12 containing 0.5% fetal bovine serum (control) and 0.1, 1, 5 or 10 ng/mL of transforming growth factor-beta1 for 24, 48 or 72 h. The effects of transforming growth factor-beta1 on CCN2/CTGF mRNA expression were measured by reverse transcription-polymerase chain reaction. CCN2/CTGF protein was quantitatively analyzed using enzyme-liked immunosorbent assay. Subcellular distribution of CCN2/CTGF protein in both human gingival fibroblasts and periodontal ligament cells was observed using immunofluorescence microscopy.


RESULTS
In both human gingival fibroblasts and periodontal ligament cells, the expression of CCN2/CTGF mRNA and CCN2/CTGF protein was significantly increased, in a dose- and time-dependent manner, in the presence of transforming growth factor-beta1. Moreover, immunofluorescence analysis indicated that immunoreactivity to CCN2/CTGF showed a granular pattern of protein localization.


CONCLUSIONS
The expression of CCN2/CTGF mRNA and protein was induced by transforming growth factor-beta1 in human gingival fibroblasts and periodontal ligament cells. These results suggest that CCN2/CTGF plays an important role in wound healing and in the regeneration of periodontal tissue.
Takeuchi
H
H
Department of Periodontology, School of Life Dentistry at Tokyo, Nippon Dental University, 1-9-20 Fujimi, Chiyoda-ku, Tokyo, Japan.
Kubota
S
S
Murakashi
E
E
Fukada
T
T
Hashimoto
S
S
Takigawa
M
M
Numabe
Y
Y
D000328:Adult; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor / Q000096:biosynthesis / Q000235:genetics*; D005347:Fibroblasts / Q000187:drug effects; D015870:Gene Expression; D005881:Gingiva / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D010513:Periodontal Ligament / Q000166:cytology / Q000187:drug effects*; D012038:Regeneration / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*; D014945:Wound Healing / Q000502:physiology; D055815:Young Adult
D053773:Transforming Growth Factor beta1; D055513:Connective Tissue Growth Factor