The effect of the IGF-I treated gingival and periodontal ligament fibroblast on osteoblasts / 대한치과교정학회지

Insulin-like growth factor I (IGF-I) has the local tissue regulating actions. In bone, IGF-I increases the replication of osteoblastic lineage, probably preosteoblasts, and enhances osteoblastic collagen synthesis and matrix composition rates. The purpose of this study was to investigate the local regulatory effect of IGF-I on periodontium totally, both in an autocrine and paracrine manner. To examine the effect of IGF-I directly on osteoblast (OB) of test rats, and indirectly on OB via periodontal ligament fibroblast (PDLF), and the effect of gingival fibroblast (GF) on OB via cellular paracrine manner for the understanding of humoral action of adjacent tissue, GF and PDLF were obtained from male Sprague-Dawley rats of six to eight weeks of age. OB was obtained from frontal and parietal calvarial bone of Sprague-Dawley 21day-old-fetus. After each cell was incubated 24 hours, for collecting conditioned medium, different concentrations of IGF-I (1,10,100 ng/ml,1ml/well) was adding in the GF, PDLF cells, and the supernatant from these cultures was put into the primary OB culture with 1x104cell/ml/well. The experimental group was divided into six groups - control OB, IGF-I treated OB, OB culture with conditioned medium from PDLF, OB culture with conditioned medium from IGF-I treated PDLF, OB culture with conditioned medium from GF, OB culture with conditioned medium from IGF-I treated GF. After final IGF-I treatment, OB was incubated for 24 hours, and alkaline phosphatase activity assay, BMP expression, cell proliferation measurement using MTT assay, total protein measurement, collagen synthesis assay using western blot, and examination of bone nodule synthesis were done. Alkaline phosphatase expressions were increased in the group of PDLF-IGF-I supernatant treatment. Direct IGF-I treatment with concentrations of 100ng/ml showed increased viable cell number measured by MTT assay. And IGF-I treatment did not increase total protein amount. The entire experimental group showed BMP2, 4 expression in western blot, and there was no significant difference between control and experimental groups. These results suggested that supernatant from PDLF effects on increasing cellular activities of OB regardless of IGF-I, and at high concentration, IGF-I increases OB cell proliferation.

Effect of various cytokines on the production of prostaglandin E2 leukotriene B4 and collagenase in human periodontal ligament fibroblasts in vitro / 대한치과교정학회지

This experiment was designed to study possible roles of interleukin-1beta, interleukin-6 and tumor necrosis factor-alpha in bone remodeling by measuring their effects on PGE2, LTB4 and collagenase production when they were administered to human periodontal ligament fibroblasts. Human periodontal ligament fibroblasts were collected from first premolars extracted for orthodontic treatment. They were incubated in the environment of 37degrees C, 5% CO2, and 100% humidity. They were treated with 0.25% trypsin-EDTA solution and centrifuged. PDL cells in the fifth to seventh passage were used for the experiment. Cells were seeded onto the culture dishes and when they were successfully attached, human recombinant interieukin-lbeta, interleukin-6, and tumor necrosis factor-alpha were administered, alone or in combination. They were incubated for 4, 8 and 24 hours and the levels of PGE2, LTB4 and collagenase released into the culture media were assessed by enzymeimmunoassay and collagenase activity assay. The conclusions are as follows: 1. IL-1beta and TNF-a were very active in stimulating the production of PGE2 and collagenase by human periodontal ligament fibroblasts, while IL -6 increased LTB4 production. 2. IL-1beta significantly increased PGE2 , but LTB4 production was not increased. IL-1beta is thought to act mainly via the cyclooxygenase pathway of arachidonic acid metabolism. 3. IL-6 tended to inhibit IL- 1beta in the production of PGE2 and collagense whereas IL-6 and TNF-alpha showed additive effect in the level of PGE2. The above cytokines increased the release of at least one of PGE2, L TB4 and collagenase. It suggests that cytokines are involved in bone remodeling process by stimulating PDL fibroblasts to produce various bone-resorptive agents. The roles of cytokines in bone remodeling as a whole would need further study.