Inhibition of growth of human gingival fibroblasts by chimeric DNA-RNA hammerhead ribozyme targeting transforming growth factor-beta 1.
BACKGROUND
Transforming growth factor (TGF)-beta1 is involved in the pathogenesis of both drug-induced gingival overgrowth and hereditary gingival fibromatosis. Ribozymes enzymatically cleave target mRNAs and are expected to be utilized as the basis of novel nucleic acid-based therapies. We designed a chimeric DNA-RNA ribozyme targeting TGF-beta1 mRNA and examined its effect on growth of gingival fibroblasts in culture.


METHODS
Chimeric DNA-RNA hammerhead ribozyme with sequence complementary to the loop structure of human TGF-beta1 mRNA was used. We evaluated transfer of the chimeric ribozyme by hemagglutinating virus of Japan (HVJ)-envelope into cultured human gingival fibroblasts in vitro and rat gingival tissues in vivo. We then examined effects of the chimeric ribozyme to TGF-beta1 on proliferation and DNA synthesis in human gingival fibroblasts. We also examined effects of the chimeric ribozyme to TGF-beta1 on expression of TGF-beta1, type IV collagens, and fibronectin mRNAs and expression of TGF-beta1 protein in human gingival fibroblasts.


RESULTS
Chimeric ribozyme was sufficiently distributed into human fibroblasts in vitro and rat gingivae in vivo. Chimeric ribozyme to TGF-beta1 significantly inhibited expression of TGF-beta1, type IV collagen, and fibronectin mRNAs and TGF-beta1 protein in human gingival fibroblasts. Mismatch ribozyme had no effect on expression of these molecules. Chimeric ribozyme to TGF-beta1 also significantly inhibited proliferation and DNA synthesis in gingival fibroblasts.


CONCLUSIONS
Chimeric DNA-RNA ribozyme targeting TGF-beta1 may be a useful gene therapy agent for treatment of gingival hyperplasia.
Yusa
Junko
J
Department of Periodontology, The Nippon Dental University, School of Dentistry, Tokyo, Japan.
Fukuda
Noboru
N
Sato
Soh
S
Matsmoto
Koichi
K
Mugishima
Hideo
H
Kamoi
Kyuichi
K
D000293:Adolescent; D000818:Animals; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D016326:Extracellular Matrix Proteins / Q000037:antagonists & inhibitors; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D005881:Gingiva / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D019214:Gingival Overgrowth / Q000517:prevention & control*; D006801:Humans; D008297:Male; D016337:RNA, Catalytic / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D053773:Transforming Growth Factor beta1
D016326:Extracellular Matrix Proteins; D016337:RNA, Catalytic; C508960:TGFB1 protein, human; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C427204:hammerhead ribozyme