Periosteal Bone Surfaces Research Articles

Bone and hypopituitarism: not only a mass issue

Bone and hypopituitarism: not only a mass issue

Comparative effects of teriparatide and strontium ranelate in the periosteum of iliac crest biopsies in postmenopausal women with osteoporosis

The periosteum contains osteogenic cells that regulate the outer shape of bone and contribute to determine its cortical thickness, size and position. We assessed the effects of subcutaneous injections of teriparatide (TPTD, 20 μg/day) or oral strontium ranelate (SrR, 2 g/day) in postmenopausal women with osteoporosis on new bone formation activity at the periosteal and endosteal bone surfaces using dynamic histomorphometric measurements. Evaluable tetracycline-labeled transiliac crest bone biopsies were analyzed from 27 patients in the TPTD group, and 22 in the SrR group after six months of treatment. Measurements were conducted on the thicker and thinner cortices separately, and comparisons between the thicker, thinner and combined cortices were carried out. At the combined periosteal cortex, the mineralization surface as a percent of bone surface (MS/BS%) was greater for TPTD (mean ± SE: 8.08 ± 1.22%) than SrR (3.22 ± 1.05%) ( p < 0.005). The difference in mineral apposition rate (MAR) between TPTD (0.35 ± 0.06 μm/day) and SrR (0.14 ± 0.06 μm/day) was also significant ( p < 0.05), while that of bone formation rate per bone surface (BFR/BS) between TPTD (0.014 ± 0.004 mm 3/mm 2/year) and SrR (0.004 ± 0.003 mm 3/mm 2/year) was not ( p = 0.057). Statistically significant differences between the two treatments were also observed for MS/BS%, BFR/BS, MAR and the double-labeled perimeter in the periosteum of the thicker, but not thinner, iliac crest cortices. The comparison between the thicker and thinner cortices of both periosteal and endosteal surfaces showed statistically significant differences for MAR and the double-labeled perimeter for TPTD treated women. There were no statistically significant differences in any bone formation dynamic measurements between the two cortices in the SrR group. In conclusion, most of the bone formation and mineralization variables were significantly higher for TPTD- than SrR-treated women at both the periosteal and endosteal combined cortices. The response to TPTD for dynamic bone formation measurements in the periosteal surface was greater for the thicker than thinner cortex, but this difference was not significant in SrR treated patients. This may reflect a greater ability of TPTD to enhance responsiveness of bone to the mechanical loading environment. These effects on bone formation may underlie the improvement in bone quality in patients with osteoporosis treated with TPTD.

ASB Clinical Biomechanics Award Paper 2010: Virtual pre-operative reconstruction planning for comminuted articular fractures

Background Highly comminuted intra-articular fractures are complex and difficult injuries to treat. Once emergent care is rendered, the definitive treatment objective is to restore the original anatomy while minimizing surgically induced trauma. Operations that use limited or percutaneous approaches help preserve tissue vitality, but reduced visibility makes reconstruction more difficult. A pre-operative plan of how comminuted fragments would best be re-positioned to restore anatomy helps in executing a successful reduction. Methods In this study, the methods for virtually reconstructing a tibial plafond fracture were developed and applied to clinical cases. Building upon previous benchtop work, novel image analysis techniques and puzzle solving algorithms were developed for clinical application. Specialty image analysis tools were used to segment the fracture fragment geometries from CT data. The original anatomy was then restored by matching fragment native (periosteal and subchondral) bone surfaces to an intact template, generated from the uninjured contralateral limb. Findings Virtual reconstructions obtained for ten tibial plafond fracture cases had average alignment errors of 0.39 (0.5 standard deviation) mm. In addition to precise reduction planning, 3D puzzle solutions can help identify articular deformities and bone loss. Interpretation The results from this study indicate that 3D puzzle solving provides a powerful new tool for planning the surgical reconstruction of comminuted articular fractures.

Strain driven transport for bone modeling at the periosteal surface

Bone modeling and remodeling has been the subject of extensive experimental studies. There have been several mathematical models proposed to explain the observed behavior, as well. A different approach is taken here in which the bone is treated from a macroscopic view point. In this investigation, a one-dimensional analytical model is used to shed light on the factors which play the greatest role in modeling or growth of cortical bone at the periosteal surface. It is presumed that bone growth is promoted when increased amounts of bone nutrients, such as nitric oxide synthase (NOS) or messenger molecules, such as prostaglandin E 2 (PGE 2), seep out to the periosteal surface of cortical bone and are absorbed by osteoblasts. The transport of the bone nutrients is assumed to be a strain controlled process. Equations for the flux of these nutrients are written for a one-dimensional model of a long bone. The obtained partial differential equation is linearized and solved analytically. Based upon the seepage of nutrients out of the bone, the effect of loading frequency, number of cycles and strain level is examined for several experiments that were found in the literature. It is seen that bone nutrient seepage is greatest on the tensile side of the bone; this location coincides with the greatest amount of bone modeling.

Identification of a possible engraved Venus from Předmostí, Czech Republic

One Gravettian feminine representation – the schematic Venus of Předmostí – is so different from all the others that it has always been regarded as unique. The engraving, which was closely examined for the purpose of comparison in this study, represents a woman composed of geometric shapes, including a triangular head with interior lattice-work, concentric ellipses for the breasts, belly and pelvis, a double ladder for the remaining arm, hatch marks, and a thigh made up of parallel vertical lines. This article presents the discovery and discusses the authenticity of a second, feminine anthropomorph engraved on a bone shaft fragment, which is labelled as coming from the same site and is highly similar to the one described at the end of the nineteenth century. The modern history of Předmostí reveals that the site was used as a quarry for the extraction of loess and limestone, and that tons of mammoth bones were extracted to produce spodium. Remarkable objects that were found both during early archaeological excavations and the site’s commercial exploitation went towards private collections. The history of the piece under study traces it back through a succession of owners to the collection of René de Poilloüe de Saint-Périer (b.1877–d.1950) and Raymonde-Suzanne de Saint-Périer (b.1890–d.1978). The engraving is on the periosteal surface of a limb bone from a very large mammal, probably a proboscidian. Microscopic analysis of the bone surface and engravings identify the chronology of the grooves and their relations to the stages of the bone’s alteration and fracturing, starting with: 1) heavy weathering of the bone surface that produced longitudinal cracks, which probably led to its breakage, 2) engraving of the feminine representation, 3) intense mechanical and chemical attacks that smoothed all of the bone’s surfaces and wore down the engravings, 4) covering with a consolidation agent, and, most recently, 5) some abrasion, which resulted in the creation of a few straight lines. Since no evidence is found to suggest that the highly worn appearance of the engravings composing the feminine representation resulted from modifications that were meant to artificially age the periosteal surface of the bone, we conclude that the engraving can be plausibly attributed to the Gravettian and that further analyses are warranted.

A Novel Closed-Loop Electromechanical Stimulator to Enhance Osseointegration with Immediate Loading of Dental Implant Restorations

The degree of osseomechanical integration of dental implants is acutely sensitive to their mechanical environment. Bone, both as a tissue and structure, adapts its mass and architecture in response to loading conditions. Therefore, application of predefined controlled loads may be considered as a treatment option to promote early maturation of bone/implant interface prior to or in conjunction with crown/prosthesis attachment. Although many studies have established that the magnitude, rate of the applied strain, and frequency have significant effects on the osteogenic response, the actual specific relationships between strain parameters and frequency have not yet been fully defined. The purpose of this study was to develop a stimulator to apply defined mechanical stimuli to individual dental implants in vivo immediately after implantation, exploring the hypothesis that immediate controlled loading could enhance implant integration. An electromechanical device was developed, based on load values obtained using a two-dimensional finite element analysis of the bone/implant interface generating 1000 to 4000 pe and operated at 30 and 3 Hz respectively. The device was then tested in a cadaveric pig mandible, and periosteal bone surface strains were recorded for potential future comparison with a three-dimensional finite element model to determine loading regimens to optimize interface strains and iterate the device for clinical use.

Histomorphometric changes in the wing bones of the fruit bat,Pteropus poliocephalus, (Megachiroptera: Pteropidae) in relation to increased bone strain and the failure of a good (?) hypothesis

Fluorochrome labelling of bone formation was used to examine the effect of exercise (flight) on the wing skeleton of fruit bats, Pteropus poliocephalus, over a 194 day period. The bats in this study had been born and raised in captivity and it was hypothesised that the large increases in bone strain that accompanied active flight would result in bone formation at the periosteal bone surface, leading to increased mechanical stiffness and strength. This hypothesis was not supported by the results. Bone formation rates, percentage mineralising surface and mineral apposition rates at the mid-shaft periosteal surface of the radius, metacarpal III and metacarpal V were small. The proximal phalanx of digit V did not display any bone formation at this surface. Bone appositional activity was not significantly different between baseline, control (non-flight) and treatment (flight) groups at any time-point of the experiment. Apposition, although limited, occurred primarily at the endocortical surface in all bones of all animals. No correlation was found between activity and bone formation. Active intracortical remodelling (a total of four secondary osteons) was only seen in three individuals.There was evidence of earlier remodelling activity in most bones, although there was no evidence of any secondary remodelling in the proximal phalanx.

Lipopolysaccharide induces increased bone resorption and homing of osteoclast progenitors to periosteal bone surface

Lipopolysaccharide induces increased bone resorption and homing of osteoclast progenitors to periosteal bone surface

The cell biology of parathyroid hormone in osteoblasts

Continuous exposure to parathyroid hormone (PTH) is associated with catabolic effects, whereas intermittent exposure to low doses of PTH is associated with anabolic effects. By controlling osteoblast function, PTH increases bone formation on cancellous, endocortical, and periosteal bone surfaces. In general, PTH does not affect the replication of uncommitted osteoblast progenitors but suppresses proliferation of committed osteoprogenitors. Intermittent PTH promotes osteoblast differentiation, in part, by its ability to promote exit from the cell cycle, to activate Wnt signaling in osteoblasts, and to inhibit the Wnt antagonist sclerostin in osteocytes. Insulin-like growth factor-1 is also required for the actions of PTH to increase osteoblast numbers. Intermittent PTH prolongs osteoblast survival in rodents by mechanisms that involve activation and proteolytic degradation of Runx2. PTH's ability to orchestrate a dynamic range of signaling cascades that determine osteoblast fate may explain both its catabolic and beneficial actions on the skeleton.

Ultrasound mimics the effect of mechanical loading on bone formation in vivo on rat ulnae

While the effect of ultrasound as an extreme example of low-magnitude high-frequency stimulation has been explored in the response of bone to injury, little is known about its effect on normal bone. This experiment was designed to test the hypothesis that ultrasound exerts a similar influence on bone as mechanical stimulation at a physiological level. Three groups of female Wistar rats were anaesthetised (6 per group). In one group, the left ulna was loaded cyclically in vivo 40 times, repeated on a further 5 occasions on alternate days. In a second group, transcutaneous low-intensity pulsed ultrasound stimulation was applied to the left ulnae for the same duration as the period of loading. In a third group, loading and ultrasound stimulation were applied concurrently. The right ulna served as non-loaded control in each animal. At the end of the experiment after 14 days, both ulnae were removed. Induced bone formation was assessed by measuring the proportion of medial periosteal bone surface with double label (dLS/BS, %) and by calculation of mineral apposition rate (MAR) from the inter-label distance. All three treatments induced a significant periosteal response, increasing dLS/BS values from <10% in control limbs to >80% in treated limbs. Increases in MAR of experimental ulnae versus contralateral control ulnae were 2.9 (±0.9), 8.6 (±2.4) and 8.7 μm (±3.2) for the ultrasound only, ultrasound and load, and load only groups, respectively. The effects of loading plus ultrasound were not significantly different from ultrasound alone. These data suggest that ultrasound is able to induce changes in bone that share at least some features with mechanical loading.

Strontium ranelate does not stimulate bone formation in ovariectomized rats

Strontium ranelate (SrR) is suggested to function as a dual-acting agent in the treatment of postmenopausal osteoporosis with anti-resorptive and anabolic skeletal benefits. We evaluated the effects of SrR on the skeleton in ovariectomized (OVX) rats and evaluated the influence of dietary calcium. Three-month old virgin female rats underwent ovariectomy (OVX, n = 50) or SHAM surgery (SHAM, n = 10). Four weeks post-surgery, rats were treated daily by oral gavage with distilled water (10 ml/kg/day) or SrR (25 or 150 mg/kg/day) for 90 days. Separate groups of animals for each dose of SrR were fed a low (0.1%) or normal (1.19%) calcium (Ca) diet. Static and dynamic histomorphometry, DXA, mu-CT, mechanical testing, and serum and skeletal concentrations of strontium were assessed. SrR at doses of 25 and 150 mg/kg/day did not increase bone formation on trabecular or periosteal bone surfaces, and failed to inhibit bone resorption of trabecular bone regardless of Ca intake. There were no improvements in bone mass, volume or strength with either dose of SrR given normal Ca. These findings demonstrate that SrR at dosages of 25 and 150 mg/kg/day did not stimulate an anabolic bone response, and failed to improve the bone biomechanical properties of OVX rats.

Assessing compliance, acceptance, and tolerability of teriparatide in patients with osteoporosis who fractured while on antiresorptive treatment or were intolerant to previous antiresorptive treatment: An 18-month, multicenter, open-label, prospective study

Background: Teriparatide (parathyroid hormone [1-34] [ribosomal DNA origin]) stimulates new bone formation on trabecular and cortical (periosteal and/or endosteal) bone surfaces by preferential stimulation of osteoblastic activity over osteoclastic activity. It has been found to significantly reduce vertebral fractures by 65%, and nonvertebral fragility fractures by 53% in treatment-naive postmenopausal women who have previously suffered a vertebral fracture. Objective: This study examined the compliance, acceptance, and adherance of SC teriparatide 20 pg QD. Methods: In this 18-month, multicenter, openlabel, prospective study, women with postmenopausal osteoporosis, and men >30 years of age with either idiopathic or hypogonadal osteoporosis (with low bone mass [T-score of -1 or worse] and ≥1 fragility fracture), who had experienced a treatment-related adverse event (AE) or an inadequate response while receiving antiresorptive treatment, and who were willing to receive open-label teriparatide for ≥18 months were eligible. Compliance was defined as self-reported use of ≥80% of study medication. Acceptance of the injection pen was determined by scores obtained from questionnaires and rating scales measuring patients' perception. Patients self-reported on injection discomfort, ease of use, and the overall injection administration. Acceptance was assessed at baseline, and 3, 6, and 18 months. AEs were recorded at each clinical visit from the patients' self-reports. At the 3-month visit, a serum calcium level was drawn ≥16 hours after the previous teriparatide dose. Results: In this study, 116 patients—102 women with postmenopausal osteoporosis and 14 men (12 with idiopathic osteoporosis and 2 with hypogonadal osteoporosis)—were assessed for inclusion in the study. The mean (SD) age was 68.8 (11.1) years (range, 40-89 years) and the mean (SD) weight was 60.5 (11.7) kg (range, 37-87 kg). Seventy-three percent of the patients in this study had baseline spinal T-scores ≤-2.5, and 72% had fractured during treatment with an osteoporosis medication. Reported compliance was 89% at 6 months and 82% at 18 months. At baseline, 42% of patients were concerned about injection discomfort, and 43% were somewhat concerned with daily injections, while 7% were quite concerned. At 6 months, most patients reported much less concern (49%) or no concern (42%). Patient perceptions associated with learning how to use the pen injection, attaching the needle, holding the pen, and injecting the dose, improved during the first 6 months of the study. The most commonly reported AEs were dizziness, 12 (10.3%); nausea, 12 (10.3%); back pain, 9 (7.8%); and muscle cramps, 9 (7.8%). No AEs were believed to be associated with the use of the pen injection or teriparatide. Five patients had mildly elevated serum calcium concentrations (maximum value 2.8 mmol/L) at 3 months. However, all were normal on repeat testing ~4 weeks thereafter. Conclusions: This study found that teriparatide pen injection was well accepted in these patients, and acceptance rates improved during the first 6 months of treatment and, thereafter, improved slightly for ~18 months. Reported compliance remained high throughout the study (82%-89%). Teriparatide pen injection was a viable treatment in these osteopenic or osteoporotic patients with fragility fractures.

The skeletal responsiveness to mechanical loading is enhanced in mice with a null mutation in estrogen receptor-β

Mechanical loading caused by physical activity can stimulate bone formation and strengthen the skeleton. Estrogen receptors (ERs) play some role in the signaling cascade that is initiated in bone cells after a mechanical load is applied. We hypothesized that one of the ERs, ER-beta, influences the responsiveness of bone to mechanical loads. To test our hypothesis, 16-wk-old male and female mice with null mutations in ER-beta (ER-beta(-/-)) had their right forelimbs subjected to short daily loading bouts. The loading technique used has been shown to increase bone formation in the ulna. Each loading bout consisted of 60 compressive loads within 30 s applied daily for 3 consecutive days. Bone formation was measured by first giving standard fluorochrome bone labels 1 and 6 days after loading and using quantitative histomorphometry to assess bone sections from the midshaft of the ulna. The left nonloaded ulna served as an internal control for the effects of loading. Mechanical loading increased bone formation rate at the periosteal bone surface of the mid-ulna in both ER-beta(-/-) and wild-type (WT) mice. The ulnar responsiveness to loading was similar in male ER-beta(-/-) vs. WT mice, but for female mice bone formation was stimulated more effectively in ER-beta(-/-) mice (P < 0.001). We conclude that estrogen signaling through ER-beta suppresses the mechanical loading response on the periosteal surface of long bones.

Effects Of a One-Month Treatment With PTH(1–34) on Bone Formation on Cancellous, Endocortical, and Periosteal Surfaces of the Human Ilium

Using bone histomorphometry, we found that a 1-month treatment with PTH(1-34) [hPTH(1-34)] stimulated new bone formation on cancellous, endocortical, and periosteal bone surfaces. Enhanced bone formation was associated with an increase in osteoblast apoptosis. The precise mechanisms by which hPTH(1-34) increases bone mass and improves bone structure are unclear. Using bone histomorphometry, we studied the early effects of treating postmenopausal women with osteoporosis with hPTH(1-34). Tetracycline-labeled iliac crest bone biopsies were obtained from 27 postmenopausal women with osteoporosis who were treated for 1 month with hPTH(1-34), 50 microg daily subcutaneously. The results were compared with tetracycline-labeled biopsies from a representative control group of 13 postmenopausal women with osteoporosis. The bone formation rate on the cancellous and endocortical surfaces was higher in hPTH(1-34)-treated women than in control women by factors of 4.5 and 5.0, respectively. We also showed a 4-fold increase in bone formation rate on the periosteal surface, suggesting that hPTH(1-34) has the potential to increase bone diameter in humans. On the cancellous and endocortical surfaces, the increased bone formation rate was primarily caused by stimulation of formation in ongoing remodeling units, with a modest amount of increased formation on previously quiescent surfaces. hPTH(1-34)-stimulated bone formation was associated with an increase in osteoblast apoptosis, which may reflect enhanced turnover of the osteoblast population and may contribute to the anabolic action of hPTH(1-34). These findings provide new insight into the cellular basis by which hPTH(1-34) improves cancellous and cortical bone architecture and geometry in patients with osteoporosis.

Low-dose estrogen treatment suppresses periosteal bone formation in response to mechanical loading

Estrogen and exercise influence cortical bone formation. Both affect bone during growth, but with complex interactions. We hypothesized that estrogen reduces the osteogenic response caused by exercise at the periosteal surface of bone, while it enhances bone formation on the endocortical surface. To test our hypothesis, 16 young (8 weeks old) male Sprague-Dawley rats were randomized into two groups: (1) low-dose 17-α ethynylestradiol treatment+bone loading (EE2) or (2) vehicle-treated+bone loading (vehicle). We applied controlled loading to the right ulna at a peak force of 17 N, 2 min/day, 3 days/week for 5 weeks to simulate exercise. The left nonloaded ulna served as an internal control for loading. Mechanical loading increased cortical area (7.7%) and bone mineral content (8%) in the vehicle-treated group (P<0.05) but only slightly increased cortical area in the EE2 group (P=0.08). Histomorphometry showed 1 week of mechanical loading increased periosteal bone formation rate by 29% in the vehicle group and this response was reduced (P<0.05) to only 15% in the EE2 group. At the endocortical surface, there were no differences in the loading response between the vehicle and EE2-treated groups. We conclude low-dose EE2 suppresses the mechanical loading response on the periosteal surface of long bones, but had no effect on the loading response at the endocortical bone surface in growing male rats.

In situ measurement of solute transport in the bone lacunar-canalicular system.

Solute transport through the bone lacunar-canalicular system is believed to be essential for osteocyte survival and function but has proved difficult to measure. We report an approach that permits direct measurement of real-time solute movement in intact bones. By using fluorescence recovery after photobleaching, the movement of a vitally injected fluorescent dye (sodium fluorescein) among individual osteocytic lacunae was visualized in situ beneath the periosteal surface of mouse cortical bone at depths up to 50 microm with laser scanning confocal microscopy. Transport was analyzed by using a two-compartment mathematical model of solute diffusion that accounted for the characteristic anatomical features of the lacunar-canalicular system. The diffusion coefficient of fluorescein (376 Da) was determined to be 3.3 +/- 0.6 x 10(-6) cm2/sec, which is 62% of its diffusion coefficient in water and is similar to diffusion coefficients measured for comparably sized molecules in cartilage. The diffusion of fluorescein in bone is also consistent with the presence of an osteocyte pericellular matrix whose structure resembles that proposed for the endothelial glycocalyx [Squire, J. M., Chew, M., Nneji, G., Neal, C., Barry, J. & Michel, C. (2001) J. Struct. Biol. 136, 239-255]. To our knowledge, this is the first instance where the dynamics of molecular movement has been measured directly in the bone lacunar-canalicular system. This in situ imaging approach should also facilitate the analysis of convection-based transport mechanisms in bones of living animals.

Beta-Arrestin2 regulates the differential response of cortical and trabecular bone to intermittent PTH in female mice.

Cytoplasmic arrestins regulate PTH signaling in vitro. We show that female beta-arrestin2(-/-) mice have decreased bone mass and altered bone architecture. The effects of intermittent PTH administration on bone microarchitecture differed in beta-arrestin2(-/-) and wildtype mice. These data indicate that arrestin-mediated regulation of intracellular signaling contributes to the differential effects of PTH at endosteal and periosteal bone surfaces. The effects of PTH differ at endosteal and periosteal surfaces, suggesting that PTH activity in these compartments may depend on some yet unidentified mechanism(s) of regulation. The action of PTH in bone is mediated primarily by intracellular cAMP, and the cytoplasmic molecule beta-arrestin2 plays a central role in this signaling regulation. Thus, we hypothesized that arrestins would modulate the effects of PTH on bone in vivo. We used pDXA, muCT, histomorphometry, and serum markers of bone turnover to assess the skeletal response to intermittent PTH (0, 20, 40, or 80 mug/kg/day) in adult female mice null for beta-arrestin2 (beta-arr2(-/-)) and wildtype (WT) littermates (7-11/group). beta-arr2(-/-) mice had significantly lower total body BMD, trabecular bone volume fraction (BV/TV), and femoral cross-sectional area compared with WT. In WT females, PTH increased total body BMD, trabecular bone parameters, and cortical thickness, with a trend toward decreased midfemoral medullary area. In beta-arr2(-/-) mice, PTH not only improved total body BMD, trabecular bone architecture, and cortical thickness, but also dose-dependently increased femoral cross-sectional area and medullary area. Histomorphometry showed that PTH-stimulated periosteal bone formation was 2-fold higher in beta-arr2(-/-) compared with WT. Osteocalcin levels were significantly lower in beta-arr2(-/-) mice, but increased dose-dependently with PTH in both beta-arr2(-/-) and WT. In contrast, whereas the resorption marker TRACP5B increased dose-dependently in WT, 20-80 mug/kg/day of PTH was equipotent with regard to stimulation of TRACP5B in beta-arr2(-/-). In summary, beta-arrestin2 plays an important role in bone mass acquisition and remodeling. In estrogen-replete female mice, the ability of intermittent PTH to stimulate periosteal bone apposition and endosteal resorption is inhibited by arrestins. We therefore infer that arrestin-mediated regulation of intracellular signaling contributes to the differential effects of PTH on cancellous and cortical bone.

Nail and bone surface as biomarkers for acute fluoride exposure in rats.

When acute exposure to fluoride is thought to be the cause of death, confirmation often depends on the analysis of some body fluid or tissue. The aim of this study was to evaluate the use of nails and the periosteal surface of bone as indicators of acute exposure to fluoride. Six groups of rats were given a single oral dose of fluoride (50 mg/kg body weight), while the control group was given deionized water. The rats were killed at 2, 4, 8, 16, 24, and 48 h after fluoride administration. Plasma and nails (the proximal halves) were collected and analyzed for fluoride with an ion-specific electrode after hexamethyldisiloxane-facilitated diffusion. A circular area of the femur (4.52 mm(2)) was etched with 0.5M HCl for 15 s, and, after the addition of a buffer, the solution was analyzed with an ion-specific electrode. Peak plasma concentration occurred at 2 h, followed by progressively declining concentrations. Peak nail fluoride concentrations occurred at 8 h. The mean nail concentrations at 8, 16, and 24 h were significantly higher than that of the control group. Bone surface concentrations were significantly higher than that of the control group at 4 h and thereafter. Thus, the proximal portion of nails and bone surface are suitable biomarkers for acute fluoride exposure in rats.

Local biochemical markers of bone turnover: relationship to subsequent density of healing alveolar bone defects.

This pilot study was designed to test whether biochemical markers of bone turnover in washes of periosteal or trabecular alveolar bone surfaces could be correlated with increases in bone density of an adjacent healing implant socket. Ten subjects had a canula inserted into the alveolar crest and sterile phosphate-buffered saline was washed over the periosteal and trabecular surfaces and collected. Surgical flaps were reflected, 5 mm diameter bone cores were removed from the bone wash site, and standardized radiographs were taken. The sites were allowed to heal for 12 weeks, and radiographs were repeated. Bone washes of the healing sites were also collected after 2 and 12 weeks. Washes were analysed for bone turnover markers osteocalcin (OC; radioimmunoassay) and C-terminal telopeptide of Type 1 collagen (ICTP; enzyme-linked immunosorbent assay (ELISA)), and blood component albumin (ALB; ELISA). Changes in bone density during healing were determined by radiographic absorptiometry. OC/ALB and ICTP/ALB ratios were higher for trabecular than periosteal washes at baseline (p<or=0.01). Trabecular OC/ALB and ICTP/ALB were inversely correlated with increasing bone density of the healing bone core socket (r=-0.72, p=0.03; Pearson's correlation coefficient). Biochemical markers of bone turnover in bone washes of specific alveolar bone sites may prove helpful in predicting how the bone density will increase around healing dental implants.

PHOSPHO1—A novel phosphatase specifically expressed at sites of mineralisation in bone and cartilage

Mineralisation of bone and cartilage is essential for skeletal development and function. We have previously reported a novel gene (PHOSPHO1); a member of the large haloacid dehalogenase superfamily of hydrolases which has an active site indicative of a phosphatase. Its high expression in skeletal tissues has led us to speculate that PHOSPHO1 may be involved in the mineralisation process. Therefore, in this study, we have determined that PHOSPHO1 is localized to sites of mineralisation in both cartilage and bone. Recombinant derived PHOSPHO1 protein was produced and affinity purified PHOSPHO1 antiserum was generated and used to immunostain a range of skeletal and soft avian tissues. In addition, PHOSPHO1 gene expression was determined in SaOS-2 and MG-63 osteoblast-like cells by RT-PCR. In diaphyseal cortical bone, immunohistochemistry localized PHOSPHO1 protein to the osteoid layer of the periosteum, forming surfaces of growing osteons, and newly formed osteocytes, whereas the endosteum and closed osteons were negative. In growth plate cartilage, immunoreactivity was limited to the early hypertrophic chondrocytes and the ossification groove of Ranvier. Cartilage remnants and trabecular bone within the primary spongiosa exhibited strong immunoreactivity on their mineralising surfaces. In 17-day-old embryonic calvaria, the osteoid present on the intramembranous and periosteal bone surfaces stained positively for PHOSPHO1. All soft tissues examined were negative. PHOSPHO1 gene expression was detected in mineralising SaOS-2 but not in the non-mineralising MG-63 osteoblast-like cells and gene expression levels were unchanged by dexamethasone, estradiol, 1,25-dihydroxyvitamin D3 or PTHrP treatment. Western analysis of chick growth plate cell lysate yielded bands (30.4 and 28.6 kD) corresponding to transcripts initiated at each of two possible initiation codons indicating the presence of alternative transcripts for PHOSPHO1 in growth cartilage. These results confirm that the PHOSPHO1 protein and gene expression profile is consistent with a role for PHOSPHO1 in bone and cartilage matrix mineralisation.