Effects of minocycline and doxycycline on cell survival and gene expression in human gingival and periodontal ligament cells.
OBJECTIVE
Periodontitis is an infectious disease in the gingival crevice caused by periodontopathic bacteria, including Porphyromonas gingivalis, Prevotella intermedia, Actinobacillus actinomycetemcomitans, and Tennerella forsythensis, and antibacterial agents are directly administered to the site of infection to treat it. To maximize the therapeutic effects while reducing the adverse effects, the antibacterial agents should be administered at concentrations greater than their MIC(90) doses required to inhibit the growth of 90% of periodontopathic bacteria and the administration should not damage the periodontal tissue. One approach for estimating cellular damage in the periodontal tissue caused by the administration is to assay cytological damages following exposures of cultured human cells derived from periodontal tissues to antibacterial agents. In the present study, we investigated the cytotoxic effect of minocycline (MINO) and doxycycline (DOX) by using a human gingival fibroblast cell line, a human gingival epithelial cell line, and a human periodontal ligament fibroblast cell line. We also used these cell lines to study the effect of MINO or DOX on the mRNA and protein expressions of genes associated with the differentiation of fibroblasts and the proliferation, differentiation, or cellular adhesion important to the epithelial regeneration of the periodontal attachment.


METHODS
The cytotoxic effect of MINO or DOX was measured as a decrease in cell survivals. The effects of these antibiotics on the mRNA and protein expressions in the cell lines were studied by reverse transcription-polymerase chain reaction (RT-PCR) and western blot analyses, respectively.


RESULTS
The maximum concentration of MINO or DOX that has little effect on the cell survivals and the mRNA and protein expressions of genes for alkaline phosphatase, type I procollagen, keratinocyte growth factor receptor, keratin 18 or 8/18, integrin beta1, integrin beta4, and laminin 5gamma2 was 10 or 30 microm, respectively, which are greater than their MIC(90) doses against periodontopathic bacteria described above.


CONCLUSIONS
These findings suggest that little, if any, cellular damage would be expected with topical administration of MINO or DOX to the periodontal pocket at a dose equivalent to the MIC(90). It is important to note, however, that the extrapolation of these findings to in vivo conditions has yet to be undertaken.
Suzuki
Ayako
A
Department of Pharmacology, The Nippon Dental University, School of Dentistry at Tokyo, Japan.
Yagisawa
Junko
J
Kumakura
Shin-ichi
S
Tsutsui
Takeki
T
D000287:Administration, Topical; D000900:Anti-Bacterial Agents / Q000008:administration & dosage / Q000633:toxicity*; D015153:Blotting, Western; D002461:Cell Line, Transformed; D002470:Cell Survival / Q000187:drug effects; D004318:Doxycycline / Q000008:administration & dosage / Q000633:toxicity*; D004847:Epithelial Cells / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D005881:Gingiva / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D008911:Minocycline / Q000008:administration & dosage / Q000633:toxicity*; D010513:Periodontal Ligament / Q000166:cytology / Q000187:drug effects*; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000900:Anti-Bacterial Agents; D012333:RNA, Messenger; D008911:Minocycline; D004318:Doxycycline