2003 Fiscal Year Final Research Report Summary
Effects of Melatonin on regenerative treatment in periodontal disease
| Project/Area Number |
13672194
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Periodontal dentistry
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| Research Institution | Health Sciences University of Hokkaido |
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Principal Investigator |
FUJII Takeo Health Sciences University of Hokkaido, Institute of Medical Science, Associate Professor, 医療科学センター, 助教授 (30173389)
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| Co-Investigator(Kenkyū-buntansha) |
BESSYO Kazuhisa Kyoto University, Graduate School of Medicine, Instructor, 大学院・医学研究科, 助手 (90229138)
SAITO Takashi Health Sciences University of Hokkaido, School of Dentistry, Professor, 歯学部・歯科保存学第二講座, 教授 (40265070)
KOWASHI Yusuke Health Sciences University of Hokkaido, School of Dentistry, Professor, 歯学部・歯科保存学第一講座, 教授 (60014338)
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| Project Period (FY) |
2001 – 2003
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| Keywords | melatonin / fibroblast / IL-8 / phosphophoryn / rhBMP-2 |
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| Research Abstract |
(1) Prostaglandin E2 (PGE2), a potent mediator of inflammation, has been detected in gingival crevicular fluid. The aim of this study was to examine the modulating effect of PGE2 on LPS-induced IL-8 production by human gingival fibroblasts (HGFs) in vitro. HGFs were obtained from medically healthy donors who were clinically free of periodontal disease. Cells were suspended in OMEM supplemented with 5% heat-inactivated FCS, seeded at a cell density of 1x104 cells/well in 96-well microtiter plates and incubated at 37oC in 5% C02 for 3 days. Culture medium was removed and replaced with DMEM containing 1% FCS 24 h prior to the experiments. HGFs were then exposed to various concentrations of LPS from Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans or Escherichia coli. The effect of PGE2 was studied by exposing HGFs cultures to various concentrations of PGE2 for 2 h before adding LPS preparations. Following incubation, culture supernatants were harvested and assayed for IL-8 u
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sing an ELISA kit. The expression of IL-8 mRNA, after stimulation with PGE2, was determined by RT-PCR. IL-8 production by HGFs was shown to be related to the time of incubation and the dose of LPS. A slight increase in IL-8 levels was observed when cells were exposed to PGE2 alone compared to controls. When HGFs were pre-stimulated with PGE2 and then exposed to LPS, IL-8 production was increased two to three times compared to non-treated cells. Herbimycin A completely abolished the secretion of IL-8. These results indicate that PGE2 can modulate cytokine secretion from HGFs and increase IL-8 production from cells exposed to LPS. IL-8 production induced by LPS was dependent on the tyrosine phosphorilation pathway.
(2) Objective: To determine whether phosphophoryn, major non-collagenous protein in dentin, plays a role of a co-factor of rhBMP-2 when rhBMP-2 induces hard tissue formation in vivo. Methods: Atelocollagen absorbed to porous hydroxyapatite was used as the carrier of rhBMP-2 for induction of ectopic bone formation. Five microgram of rhBMP-2 and 1 mg of phosphophoryn were added to apatite/ collagen composite. Then rhBMP-2/apatite/collagen/phosphophoryn was implanted subcutaneously into male Wistar rats (4 week old) at bilateral sites in the back. RhBMP-21 apatite/collagen or rhBMP-2/apatite was also implanted as controls. Rats were sacrificed and the implants were extirpated, at weekly intervals for 3 weeksafter implantation. The implants were analyzed histologically and biochemically. Results: Bone formation was observed at some small parts in all implants 1 week after implantation (week 1 implants). Bone formation by rhBMP-2 combined with apatite/ collagen/ phosphophoryn was more than that by rhBMP-2 combined with apatite/ collagen at week 2. Furthermore, bone was formed at the surface area of the implant, and bone remodeling occurred at week 3. Bone marrow cells took the place of grains of hydroxyapatite at the central part in rhBMP-2/ apatite/ collagen/ phosphophoryn. ALPase activity of rhBMP-21 apatite/ collagen/ phosphophoryn was about two times more than that of rhBMP-2/ apatite/ collagen at week 3. Conclusions: It was indicated that phosphophoryn acts as an accelerate factor of rhBMP-2 for induction of ectopic bone formation, and was suggested that rhBMP-2/ apatite/ collagen/ phosphophoryn composite could be an excellent biomaterial in bone and tooth reconstruction. Less
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Research Products
(5 results)
