Bone effects of fluoride in animal models in vivo. A review and a recent study.

The effects of fluoride on bone in various animal models are reviewed. In these studies, the doses of fluoride varied from those equivalent to therapeutic doses to toxic doses, and the duration of the treatment was from 15 days to 33 months. No significant modification in serum calcium, phosphorus, and alkaline phosphatase was reported. An increased serum osteocalcin level was noted in ewes. Evidence for hyperparathyroidism was found in some but not all animal models. Studies performed in the rat reported that fluoride had different effects on the periosteal and endosteal bone. An increase in the extent of eroded surfaces was observed in all experimental studies, except one in the mouse. Increases in osteoid parameters and in the number of osteoblasts were noted in mouse, cat, pig, and ewe. Only one study, carried out in dogs, mentioned a decrease in osteoid parameters. Most of the authors reported a mineralization defect due either to a modification in the composition of the bone matrix or to a low calcium intake. The formation period was augmented during fluoride treatment but, at a fluoride dose equivalent to therapeutic doses, this augmentation was mainly due to an increased active formation period. In contrast, at a fivefold greater dose, it was due to an increased inactive formation period. The augmentation of bone volume after fluoride treatment was attributed to an unbalanced coupling between resorption and formation in favor of formation. All these experimental studies support the conclusion that fluoride induces a stimulation of the birthrate of osteoblasts, but at high doses decreases their activity.

In vitro exposure to sodium fluoride does not modify activity or proliferation of human osteoblastic cells in primary cultures.

The anabolic effects of sodium fluoride (NaF) on trabecular bone mass in osteoporosis is now well established. In vivo histologic studies performed in humans and other animals have shown that fluoride induces an increase in osteoblast number at the tissue level. To determine the mechanisms of action of fluoride on osteoblasts, we studied the effects of NaF on short- and long-term cultures of human osteoblastic cells derived from bone explants obtained from 21 donors. In short-term experiments, bone-derived cells were exposed to NaF for 4 days. At doses ranging from 10(-11) to 10(-5) M, NaF did not modify the alkaline phosphatase (AP) activity or osteocalcin secretion. In long-term experiments, half the bone samples from 15 donors were cultured for 4 months in the presence of 10(-5) M NaF and the other half were maintained in NaF-free medium. Observations by light and electron microscopy disclosed no morphologic modification in bone explants after 4 months of exposure to NaF, despite an increase in the bone fluoride content. After the first month of culture, slight but not significant increases were noted in 6 of 10 cases for AP activity, 4 of 10 for osteocalcin secretion, and 5 of 7 for [3H]thymidine incorporation. After 4 months of culture in the presence of NaF, no change in AP activity or cell proliferation was noted. In contrast, the osteocalcin secretion significantly decreased (p < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride increases rat osteoblast function and population after in vivo administration but not after in vitro exposure.

The effects of fluoride on bone tissue are now well documented by in vivo histological studies performed on both human and animal bone biopsies and demonstrating an increase in osteoblast (OB) population. In order to elucidate whether the mechanism of action of fluoride on osteoblasts was direct or indirect, 14 three-week-old Sprague-Dawley rats were selected. Seven animals received 100 ppm fluoride as sodium fluoride (NaF) in drinking water for one month. The other animals, which did not receive fluoride, were considered as controls. At the end of the experiment, femurs and vertebrae were excised and osteoblastic cells were obtained after collagenase digestion separately from each animal. The osteoblastic cells derived from control and NaF-treated rats were exposed in vitro to 10(-5) M NaF. Alkaline phosphatase (AP) activity was measured, and the cellular proliferation was assessed by 3H-thymidine incorporation. Thymidine incorporation and AP activity were significantly higher in osteoblastic cells derived from NaF-treated rats than in cells obtained from control rats (p = 0.05 and p < 0.01, respectively). In contrast, the osteoblast proliferation and activity were not modified after in vitro exposure to NaF in cells derived from control and NaF-treated rats. In conclusion, the function of osteoblasts was not modified after in vitro exposure to fluoride. In contrast, given in vivo to rats for one month, fluoride has a mitogenic effect on osteoblasts and stimulates their activity. These data emphasize the hypothesis that fluoride may act either on osteoprogenitor cells or through an indirect mechanism mediated by a cofactor.

Dose effects on ewe bone remodeling of short-term sodium fluoride administration--a histomorphometric and biochemical study.

The early effects of two doses of sodium fluoride (NaF) on bone remodeling was studied in 14 ewes divided into two groups. Group I received orally 1 mg NaF/kg/day and group II received a five-fold greater dose. No calcium supplement was given. Transiliac bone biopsies and blood samples were taken before treatment (T0) and after 45 (T45) days of treatment. Bone fluoride content significantly increased in group II. In both groups, a significant decrease of serum calcium and phosphorus, and a slight but nonsignificant augmentation in serum parathyroid hormone were noted. Osteoid perimeter and area were significantly increased. The osteoid width significantly increased in both groups, but was twice higher in group II than I. At T45, the osteoblast perimeter increased in both groups. Osteoid perimeter was significantly correlated with serum osteocalcin values (r = 0.74; p less than 0.001) and bone fluoride content (r = 0.64; p less than 0.01). The bone formation rate at tissue level tended to increase in both groups. Concerning the apposition rate, a decrease was noted which was 1.5-fold higher in group II than in I. The increased formation period resulted from a prolonged inactive period in group II. These results point out a stimulatory effect of fluoride on the birth rate of osteoblasts. However, fluoride prolonged the lifespan of osteoblasts that had reduced activity.

In vitro induction of a calcifying matrix by biomaterials constituted of collagen and/or hydroxyapatite: an ultrastructural comparison of three types of biomaterials.

The induction of a calcifying matrix was studied in vitro and compared for three biomaterials (collagen sponge, hydroxyapatite material and a mixture of both (Biostite)) cultured with human osteoblast-like cells. The influence of biomaterials on organic matrix synthesis and the calcification process was analysed at the ultrastructural level (transmission electron microscopy and X-ray microanalysis). Biomaterials were well tolerated by bone cells. Whichever biomaterial was used, osteoblasts proliferated and synthesized a new matrix constituted of fibrillar and non-fibrillar elements. This activity appeared earlier and was more intense with Biostite than with collagen sponge alone. A deposition of a mineral substance in this newly formed matrix was observed with the collagen sponge and Biostite, but never with hydroxyapatite alone. The mineral deposits were identified as hydroxyapatite crystals, similar to those observed and analysed in bone tissue. These in vitro observations clearly demonstrated the property of Biostite to produce a calcified collagenous matrix similar to bone tissue. However, in vivo confirmation is required before extending the use of this biomaterial to periodontology.

[Benefits and risks of fluoride supplements]. / Bénéfices et risques des apports fluorés.

Fluoride is an essential trace element. It is incorporated in calcified tissues. At doses lower than 1.5 mg/day, fluoride has a prophylactic action against dental caries by absorption on the enamel surface. At doses ranging from 15 to 25 mg/day, it is used as a therapeutic agent in adults, in the treatment of post-menopausal osteoporosis with vertebral crush fractures. At high doses, fluoride may induce skeletal fluorosis leading in children to pseudorachitism. In children, fluoride supplement has been recommended for several years, as a prophylactic agent against dental caries. It may be given either as dentifrices and toothwash solutions, easy to use and economic but contraindicated in very young children; or as tablets, sure and effective but requiring a daily intake; or as domestic salts supplemented with 250 mg/kg. Fluoride supplement may also be given through drinking water. Fluoride content is limited to 1.5 mg/l but some mineral waters contain high amounts of fluoride and may have a beneficial effect on bone mass. In order to prevent an excessive fluoride intake in children and in adults, different forms of fluoride supplements must not be associated.

[Cellular culture of osteoblasts and fibroblasts on porous calcium-phosphate bone substitutes]. / Culture cellulaire d'ostéoblastes et de fibroblastes sur substituts osseux poreux en phosphate de calcium.

PURPOSE OF THE STUDY: Calcium phosphate ceramics are synthetic bone substitutes able to fill in bone destruction as a support of the bone growth. This work consisted in an in vitro assessment of osteoblasts and fibroblasts cultures on macroporous calcium-phosphate bone substitutes to analyze the interaction between cells and bone substitute. MATERIALS AND METHODS: The macroporous ceramic was composed of 70% hydroxyapatite and 30% tri-calcium phosphate with known mechanical and physico-chemical properties. Three compounds were processed with different size of macropore and with or without microporosity on their surface. Cells were seeded on discs measuring 10 mm in diameter and 2 mm in thickness. Cellular viability was evaluated by the MTT test for every stage of observation. An histological study to observe the invasion in the depth of discs was performed. Scanning electron microscopy was used to analyze the cellular comportment in contact with the surface of substitutes. RESULTS: An exponential cellular growth was effective on each substitute with the two cellular types. Cells spread on the surface of the compounds covering macropores and colonized the depth of the discs. A size of macropore of 300 microm or more seemed to support this invasion. 15 microm sized interconnections appeared to be effective to allow cell migration between macropores. The cell proliferation was similar on substitutes with or without microporosity. CONCLUSION: Biomaterials currently used as bone substitute are more or less osteoconductive but they have no osteoinductive property. A hybrid association of calcium-phosphate ceramic with osteogenic cells should promote the development of a calcium phosphate compound with osteoinductive capacity.

Influence of magnesium substitution on a collagen-apatite biomaterial on the production of a calcifying matrix by human osteoblasts.

The induction of a calcifying matrix is of great interest in the restoration of bone defects. In a previous in vitro study we demonstrated that a collagen sponge constituted of type I collagen fibrils, chondroitin sulfates, and hydroxyapatite crystals induces an earlier and a more abundant synthesis of a new extracellular calcifying matrix than do other biomaterials such as collagen or hydroxyapatite alone. Bone mineral contains various amounts of magnesium ions, either adsorbed at the surface of apatite crystals or incorporated inside the crystal structure. Magnesium is known to reduce the degradation rate of tricalcium phosphate ceramics and to influence the crystallization of mineral substance. Thus we evaluated two sponges modified with different substituted apatites. The substituted low magnesium-containing apatite sample decreased the osteoinductive properties of the sponge whereas the substituted high magnesium-containing apatite sample had a toxic effect on bone cells and prevented the formation of any extracellular matrix. Such a toxic effect can be explained by the presence of large numbers of magnesium ions released into the culture medium even though at physiological level magnesium is able to promote bone mineralization and to control the growth of hydroxyapatite crystals. Thus collagen sponges containing hydroxyapatite remain one of the most appropriately evaluated biomaterials used for the restoration of periodontal pockets and bone defects.

Fluoride-induced bone changes in lambs during and after exposure to sodium fluoride.

The evolution of bone changes induced by fluoride after the end of exposure was investigated in lambs. Sodium fluoride (NaF) was given orally at a dose of 3.5 mg/kg per day to 14 animals for 120 days. A group of 7 control and 7 treated lambs was slaughtered at the end of NaF administration (T120) and another group 120 days after the end of NaF exposure (T240). At T120, the bone fluoride content (BFC) was very significantly increased in treated animals. The histomorphometric analysis confirmed that fluoride induces an increase in bone formation (the osteoid perimeter and area were 3-fold and 4.5-fold higher respectively in treated than in control animals). The number of osteoblasts was significantly augmented. Serum osteocalcin level was twice as high in treated animals compared with controls. The bone formation rate at the tissue level (BFR) doubled after treatment, but the apposition rate (Aj.AR) was half that in the control group. The mineralization lag time (Mlt) was 120 days in treated animals compared with 42 days in controls. At T240, BFC had decreased by 50% compared with the level at T120, but it was still significantly higher than in controls. The osteoid and osteoblastic parameters were 2 and 1.3 times higher than in control animals. BFR remained significantly increased in treated animals, but Aj.AR and Mlt were similar in control and treated animals. In conclusion, after 4 months of NaF exposure fluoride induced an increase in osteoblast natality and bone formation at the tissue level, associated with a toxic effect at the individual cell level.(ABSTRACT TRUNCATED AT 250 WORDS)