Bone Mineral Apposition Rate Research Articles

Is resveratrol an estrogen agonist in growing rats?

Trans-3,4,5-trihydroxystilbene (resveratrol), a polyphenolic compound found in juice and wine from dark-skinned grape cultivars, was recently shown to bind to estrogen receptors in vitro, where it activated transcription of estrogen-responsive reporter genes. The purpose of this 6-day study in weanling rats was to determine the dose response (1, 4, 10, 40, and 100 microg/day) effects of orally administered resveratrol on estrogen target tissues. The solvent (10% ethanol) had no significant effect on any measurement or derived value. 17Beta-estradiol treatment (100 microg/day) decreased the growth rate, final body weight, serum cholesterol, and radial bone growth (periosteal bone formation and mineral apposition rates) at the tibia-fibula synostosis. In the uterus, 17beta-estradiol treatment increased wet weight, epithelial cell height, and steady state messenger RNA levels for insulin-like growth factor I. In contrast, resveratrol treatment had no significant effect on body weight, serum cholesterol, radial bone growth, epithelial cell height, or messenger RNA levels for insulin-like growth factor I. Resveratrol treatment resulted in slight increases in uterine wet weight, but significance was achieved at the 10-microg dose only. A second experiment was performed to determine whether a high dose of resveratrol (1000 microg/day) antagonizes the ability of estrogen to lower serum cholesterol. As was shown for the lower doses, resveratrol had no effect on body weight, uterine wet weight, uterine epithelial cell height, cortical bone histomorphometry, or serum cholesterol. 17Beta-estradiol significantly lowered serum cholesterol, and this response was antagonized by cotreatment with resveratrol. These in vivo results suggest, in contrast to prior in vitro studies, that resveratrol has little or no estrogen agonism on reproductive and nonreproductive estrogen target tissues and may be an estrogen antagonist.

Is Resveratrol an Estrogen Agonist in Growing Rats?

Trans-3,4,5-trihydroxystilbene (resveratrol), a polyphenolic compound found in juice and wine from dark-skinned grape cultivars, was recently shown to bind to estrogen receptors in vitro, where it activated transcription of estrogen-responsive reporter genes. The purpose of this 6-day study in weanling rats was to determine the dose response (1, 4, 10, 40, and 100 μg/day) effects of orally administered resveratrol on estrogen target tissues. The solvent (10% ethanol) had no significant effect on any measurement or derived value. 17β-Estradiol treatment (100 μg/day) decreased the growth rate, final body weight, serum cholesterol, and radial bone growth (periosteal bone formation and mineral apposition rates) at the tibia-fibula synostosis. In the uterus, 17β-estradiol treatment increased wet weight, epithelial cell height, and steady state messenger RNA levels for insulin-like growth factor I. In contrast, resveratrol treatment had no significant effect on body weight, serum cholesterol, radial bone growth, epithelial cell height, or messenger RNA levels for insulin-like growth factor I. Resveratrol treatment resulted in slight increases in uterine wet weight, but significance was achieved at the 10-μg dose only. A second experiment was performed to determine whether a high dose of resveratrol (1000μ g/day) antagonizes the ability of estrogen to lower serum cholesterol. As was shown for the lower doses, resveratrol had no effect on body weight, uterine wet weight, uterine epithelial cell height, cortical bone histomorphometry, or serum cholesterol. 17β-Estradiol significantly lowered serum cholesterol, and this response was antagonized by cotreatment with resveratrol. These in vivo results suggest, in contrast to prior in vitro studies, that resveratrol has little or no estrogen agonism on reproductive and nonreproductive estrogen target tissues and may be an estrogen antagonist.

Short-Term Treatment of Recombinant Murine Interleukin-4 Rapidly Inhibits Bone Formation in Normal and Ovariectomized Mice

Estrogen deficiency contributes to an increase in bone resorption and bone formation characterized by a high rate of bone turnover. Interleukin-4 (IL-4) is a rapid and potent inhibitor of bone resorption. We examined the short term in vivo effects of recombinant murine IL-4 (rmIL-4) on bone remodeling in normal and ovariectomized mice. Eight-week-old mice were randomized into the following five groups: (1) sham-operated mice (sham); (2) sham-operated mice infused with rmIL-4; (3) ovariectomized mice (ovx); (4) ovx infused with rmIL-4; and (5) ovx replaced by 10 or 20 μg of 17β-estradiol (E 2) for 14 or 28 days after ovariectomy, respectively. rmIL-4 at a dose of 5 μg/day was infused into ovx and sham for 3 days prior to sacrifice. Analyses were performed 14 and 28 days after operation. An increase in serum alkaline phosphatase and urinary deoxypyridinoline levels induced by ovariectomy was inhibited by the 3-day infusion of rmIL-4. In ovx, serum and urinary IL-6 levels were also increased significantly 14 days after ovariectomy, which were restored by E 2 but not by rmIL-4. Histomorphometrical analysis of trabecular bone revealed that the 3-day infusion of rmIL-4 inhibited the high rate of bone turnover induced by ovariectomy, such as an increase in the osteoclastic surface (Oc.S/BS), number of osteoclasts per mm bone surface (N.Oc/BS), mineralized surface per mm bone surface (MS/BS), and bone mineral apposition rate (MAR). A significant decrease in the bone volume (BV/TV) observed in ovx was not modulated by a 3-day infusion of rmIL-4 prior to sacrifice. In sham, rmIL-4 also caused a significant decrease in the Oc.S/BS, N.Oc/BS, MS/BS, and MAR, but the BV/TV was not modulated by rmIL-4. We conclude that short term infusion of rmIL-4 in vivo rapidly inhibits not only bone resorption but also its formation in both sham-operated and ovariectomized growing mice, resulting in a low rate of bone turnover without modulating bone volume.

Distribution of mineralization indices of modeling and remodeling over eight months in middiaphyseal cross sections of femurs from adult swine

Background This study assessed the distribution of active mineral formation sites within the middiaphyseal femoral cross section of swine and determined the extent to which various subsections represented bone formation activity in the entire cross section. Methods Twenty adult female swine (sows) were injected with two double (10-day intervals) labels 8 months apart. Labels involved fluorochrome markers of active mineral formation sites. Intact femoral middiaphyseal cross sections were embedded in polymethylmethacrylate, cut, and ground to 80 μm for analysis. Each specimen was subdivided into 16 anatomical and eight geometric subsections. Labeled mineralizing surface and mineral apposition rate were determined in the periosteal and endocortical envelopes. The number of labeled osteons per unit area of bone and osteonal mineral apposition rate were determined in the intracortical envelope. Results Periosteal mineralizing surface followed a bimodal distribution with highest surface activation on the anterior and posterior segments. Periosteal mineral apposition rate followed a modal distribution with highest apposition rates in the posterior portion of the cross section. The distribution of forming osteons was modal with highest frequencies of labeled osteons in the posterior segment. No significant regional differences were detected for osteonal mineral apposition rate, endocortical mineralizing surface, or endocortical mineral apposition rate. The location of either a single or a combination of two to four subsections that best predicted mineralizing surface and mineral apposition rate in the entire cross section differed with each trait and envelope. Fifty percent of the entire area as alternate anatomical subsections was required to predict >90% of variation in all traits evaluated. Overall, the predictability of mineralizing surface and mineral apposition rate was similar for geometrically defined subsections. Conclusion At least 50% of the cross-sectional area from alternate anatomical subsections must be measured to predict >90% of variation in periosteal, intracortical, and endocortical mineralizing surface and mineral apposition rate in the sow middiaphyseal femur. Anat. Rec. 250:136–145, 1998. © 1998 Wiley-Liss, Inc.

Distribution of mineralization indices of modeling and remodeling over eight months in middiaphyseal cross sections of femurs from adult swine.

This study assessed the distribution of active mineral formation sites within the middiaphyseal femoral cross section of swine and determined the extent to which various subsections represented bone formation activity in the entire cross section. Twenty adult female swine (sows) were injected with two double (10-day intervals) labels 8 months apart. Labels involved fluorochrome markers of active mineral formation sites. Intact femoral middiaphyseal cross sections were embedded in polymethylmethacrylate, cut, and ground to 80 microm for analysis. Each specimen was subdivided into 16 anatomical and eight geometric subsections. Labeled mineralizing surface and mineral apposition rate were determined in the periosteal and endocortical envelopes. The number of labeled osteons per unit area of bone and osteonal mineral apposition rate were determined in the intracortical envelope. Periosteal mineralizing surface followed a bimodal distribution with highest surface activation on the anterior and posterior segments. Periosteal mineral apposition rate followed a modal distribution with highest apposition rates in the posterior portion of the cross section. The distribution of forming osteons was modal with highest frequencies of labeled osteons in the posterior segment. No significant regional differences were detected for osteonal mineral apposition rate, endocortical mineralizing surface, or endocortical mineral apposition rate. The location of either a single or a combination of two to four subsections that best predicted mineralizing surface and mineral apposition rate in the entire cross section differed with each trait and envelope. Fifty percent of the entire area as alternate anatomical subsections was required to predict > 90% of variation in all traits evaluated. Overall, the predictability of mineralizing surface and mineral apposition rate was similar for geometrically defined subsections. At least 50% of the cross-sectional area from alternate anatomical subsections must be measured to predict > 90% of variation in periosteal, intracortical, and endocortical mineralizing surface and mineral apposition rate in the sow middiaphyseal femur.

Effects of 17β-estradiol administration on cortical and cancellous bone of ovariectomized rats with and without hypophysectomy

Pituitary hormones are essential for bone growth and bone turnover. Hypophysectomy (HX) diminishes mitogenesis and abolishes the high bone turnover rate induced by ovariectomy (OV). It is not known whether the suppressive effect of estrogen on bone resorption is diminished or abolished by HX. The present study investigates the effects of 17β-estradiol (E 2) (20 μg/wk) on cortical and cancellous bone mass and bone turnover as measured by histomorphometry in HX + OV (HO) rats. Sprague-Dawley rats at 2 months of age were OV or HO and the experiment was performed over a 6 week period. Hypophysectomy + OV (HO) resulted in a cessation of periosteal bone formation, and longitudinal bone growth and a decrease in cancellous bone volume. The tibial dry weight and tibial density were significantly lower in the HO than in the intact or OV groups. Administration of E 2 to HO rats partially prevented cancellous bone loss, whereas the same dosage of E 2 fully prevented cancellous bone loss in rats with OV alone. Nevertheless, cancellous bone volume was higher in the HO + E 2 than in the HO-alone groups. Estradiol administration in HO rats did not suppress cancellous bone formation rate or the eroded surface as much as it did in the OV rats. The suppressive effect of E 2 on periosteal bone formation rate and mineral apposition rate was also diminished in HO rats. However, factorial ANOVA showed that the effects of E 2 on increasing cancellous bone volume and decreasing periosteal bone formation rate and mineral apposition rate were still significant in the HO rats. Tibial dry weight and tibial density did not differ between HO and HO + E 2 groups. In conclusion, we have demonstrated that the estrogen-induced effects of preventing cancellous bone loss, of suppressing bone formation, and resorption as seen in OV rats was diminished but not abolished in HO rats.

Changes in quantitative bone histomorphometry in aging healthy men.

Changes in bone mineral metabolism with aging in healthy men and the roles of various factors in the pathogenesis of age-related changes in quantitative bone histomorphometry in men are poorly defined. To clarify these changes and factors, serum and urinary biochemical parameters and iliac crest bone biopsies were evaluated in 43 healthy men, aged 20-80 yr. The static histomorphometric parameters, cancellous bone volume and osteoblast-osteoid interface, decreased by 40.0% and 19.2%, respectively, between 20-80 yr of age. The dynamic histomorphometric parameters, double and single labeled osteoid, also decreased by 18.6% and 18.0%, respectively, over this period. None of the other static or dynamic histomorphometric parameters changed with age in this population sample of healthy men. Univariate analysis of static bone histomorphometric parameters and biochemical parameters revealed significant correlations between osteoid surface and intact PTH (r = 0.37; P = 0.015); osteoclast surface and serum total testosterone (r = 0.36; P = 0.016), estradiol (r = 0.40; P = 0.009), and FSH (r = 0.49; P = 0.001); osteoblast-osteoid interface and serum phosphate (r = 0.31; P = 0.046); and cortical thickness and serum total calcium (r = 0.38; P = 0.013). Univariate analysis of dynamic bone histomorphometric parameters and biochemical parameters revealed correlations between mineral apposition rate and serum total testosterone (r = 0.32; P = 0.037); total volume-referent bone formation rate and serum osteocalcin (r = 0.43; P = 0.004), total testosterone (r = 0.47; P = 0.001), estradiol (r = 0.35; P = 0.023), and dehydroepinadrosterone sulfate (r = 0.31; P = 0.045); and mean wall thickness and serum total calcium (r = 0.36; P = 0.019) and creatinine clearance (r = 0.42; P = 010). Mineralization lag time and serum phosphate (r = -0.39; P = 0.012) and urinary total pyridinoline (r = 0.36; P = 0.023), and mean wall thickness and urinary total pyridinoline (r = -0.38; P = 0.013), were inversely correlated. Multiple regression analysis using all-subset analysis comparing cancellous bone volume to serum and urinary biochemical parameters in these men indicated that the log free androgen index and body weight best predicted the age-related decline in iliac crest cancellous bone volume (r2 = 0.19; P = 0.015). Multiple regression analysis by the same method, comparing bone density at different skeletal sites to bone histomorphometric parameters, indicated that lumbar spine bone mineral density (BMD) was best predicted by cancellous bone volume and mineral apposition rate (r2 = 0.31; P = 0.001), femoral neck BMD by cancellous bone volume and osteoid surface (r2 = 0.19; P = 0.020), femoral greater trochanter BMD by cortical thickness and single labeled osteoid surface (r2 = 0.13; P = 0.060), and total body BMD by cancellous bone volume and surface-based bone formation rate (r2 = 0.31; P = 0.001). In summary, cancellous bone volume, osteoblast-osteoid interface, and double and single labeled osteoid decreased with age in this sample of healthy men. The lack of detectable change in bone density at some skeletal sites in these men may be due to the small sample size or other confounding factors. Multivariate analysis suggests that different combinations of histomorphometric parameters predict bone density at different skeletal sites, and that cancellous bone volume predicts bone density at the lumbar spine, femoral neck, and total body, but not at the femoral greater trochanter. We conclude that alterations in several biochemical parameters are important in the pathogenesis of age-related bone loss in healthy men.

Lack of an effect of sodium zeolite A on rat tibia histomorphometry

Cell culture studies suggest that the aluminum silicate polymer sodium Zeolite A (SZA) increases bone cell proliferation and extracellular matrix production. This study in rats investigated the short-term (2 weeks) and long-term (18 weeks) in vivo effects of SZA on growth rate (weight gain) and tibia histomorphometry. In separate short-term experiments, female (experiment 1) or male (experiment 2) Sprague-Dawley rats grown and maintained on normal calcium diets were gavaged daily during a 2 week treatment period with 30 mg/kg, 100 mg/kg, or 500 mg of SZA/kg of rat body weight. In the long-term study (experiment 3) ovariectomized (OVX) rats were fed a low calcium diet containing 0, 1.80, and 9.00 g of SZA/kg of diet (0, 125, and 617 mg/kg of body weight, respectively). Short- and long-term treatment of growing rats with SZA resulted in a dose-dependent increase in bone aluminum. In the first experiment, growing intact female rats showed no significant SZA dose-dependent response in growth rate (weight gain) or histomorphometry of cortical bone in the tibial diaphysis or cancellous bone in the secondary spongiosa of the tibial metaphysis. In the second experiment, growing male rats, with right hind limbs immobilized by unilateral sciatic neurotomy, showed no SZA dose-dependent response in growth rate. The longitudinal growth of cancellous bone in the tibia of the denervated limb and the intact contralateral limb were not influenced by sciatic neurectomy and/or by treatment with SZA. Histomorphometry demonstrated that cortical bone mass and formation was reduced in the sciatic-sectioned limb when compared with the contralateral intact limb of vehicle-treated rats, as evidenced by significant reductions in static measurements of cortical bone area (-8.5%) and cross-sectional area (-4.8%) and in calculations of the periosteal formation rate (-33.8%) and mineral apposition rate (-31.6%), and the endocortical formation rate (-35.5%) and mineral apposition rate (-37.9%). The cancellous bone mass of denervated limbs of vehicle-treated rats was also deficient, as evidenced by decreased cancellous bone area (-39.1%) and perimeter (-31.9%). The bone mineral apposition rate was decreased (-26.7%) indicating reduced osteoblast activity. Treatment with SZA did not influence these indices in the tibiae of either sciatic-sectioned limbs or contralateral intact limbs. In the long-term experiment, OVX resulted in a dramatic 88% decrease in cancellous bone volume which was prevented by treatment with 17 beta-estradiol and not influenced by treatment with Zeolite A. The increases in osteoblast and osteoclast number following OVX were not influenced by SZA. The results indicate that SZA treatment has no anabolic effect on cortical and cancellous bone formation and mass in normal growing female rats and that this compound does not protect against osteopenia due to reduced load bearing in the growing male rat or gonadal hormone deficiency in adult female rats.

Topographical variation within the articular cartilage and subchondral bone of the normal ovine knee joint: a histological approach

Topographical variation in the articular cartilage and subchondral bone of the normal ovine knee was examined using histological techniques. The articular cartilage was examined grossly, then histological sections were cut and the cartilage thickness and chondrocyte density were measured. Bone mineral density, thickness of the subchondral bone plate (SBP) and volume and surface histomorphometrical parameters and mineral apposition rate were calculated for the subchondral bone. It was found that the articular cartilage on the tibial plateaux was thicker, less cellular, and overlay a thicker SBP than that on the femoral condyles. Similarly, the cartilage in the medial joint compartments was thicker, less cellular and overlying a thicker less dense SBP than that in the lateral joint compartments. There was no variation in bone histomorphometric parameters or mineral apposition rate between regions. Biomechanical testing has shown that loading is not uniform throughout the normal human knee joint. The present results suggest that loading within the ovine knee is also nonuniform, with the central regions of the tibial plateaux bearing greater loads than the femoral condyles, and the medial joint compartment being loaded more than the lateral one. The articular cartilage and subchondral bone have adapted in order to best withstand these variations in loading. These histological findings, plus the topographical variations in cartilage biochemistry reported by Read et al. (Topographical variation in composition, PG-biosynthesis and swelling pressure of cartilages of loaded tibio-femoral joints (Abstract). Proceedings of the Combined Meeting of the Orthopaedic Research Societies of USA, Japan and Canada.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of exogenous growth hormone on bone mineralization and remodeling and on plasma calcitriol in intact pigs

Growing pigs were given subcutaneous (SC) injections of 40 μg porcine GH/kg body weight (BW) or its vehicle twice daily for 2 months. Animals were pair-fed with a diet containing 1.1% Ca, 0.6% P, and 1000 IU vitamin D 3/kg. At slaughter, several bone characteristics including histomorphometric data (using double tetracycline labeling) were measured on tibia and metacarpals. GH accelerated growth, with greater ( p < 0.01) tibial and metacarpal weights, greater tibial length ( p < 0.01) and diameters (outside and inside, p < 0.01), and greater tibial ash weight ( p < 0.02) in GH-treated pigs than in controls. The similar values of apparent bone density (weight/volume) and ash/bone volume or ash/dry matter in the two groups suggest adequate coupling between bone growth and mineralization in GH-treated pigs. Histomorphometric data for the distal metacarpal metaphysis indicated greater trabecular bone volume ( p < 0.01), osteoblastic surface ( p < 0.01), and mineral apposition rate ( p < 0.05) in GH-treated pigs. The osteoclast surface, lacuna depth, and osteoid-related parameters in GH-treated and control pigs were similar. The plasma PTH of the two groups of pigs were similar throughout the experiment. These data and the elevated plasma alkaline phosphatase activity ( p < 0.05) in GH-treated pigs suggest that GH specifically affects bone formation. GH had no effect on the plasma 25-OH vitamin D 3 but 1,25(OH) 2 vitamin D 3 (calcitriol) was higher ( p < 0.01) in treated pigs throughout the experiment. This suggests that calcitriol may help adapt bone mineralization to accelerated bone formation during growth hormone treatment. Plasma phosphate ( p < 0.05) and insulin-like growth factor (IGF-I, p < 0.01) were elevated, whereas plasma calcium did not change in the GH-treated pigs. Because the plasma PTH of the GH-treated pigs was unchanged, the elevated plasma P may result from directly or indirectly, via IGF-I stimulation of phosphate tubular reabsorption by GH. Thus exogenous GH increases bone mass via bone formation in growing pigs. GH is also involved in vitamin D and phosphorous metabolism.

The effect of 17 β-estradiol treatment on the mass and the turnover of bone in ovariectomized rats taking a mild dose of thyroxin

We performed the dosing experiment to establish whether estrogen administration has any beneficial effects on the mass and the turnover of bone in ovariectomized rats taking a mild dose of thyroxin. Thirty-five Wistar rats, 28 weeks of age, received ovariectomies (OVX) or sham operations and were divided into five groups. Group I was the sham group, Groups 2–5 were ovariectomized. Group 2 was the OVX-control, Group 3 treated with thyroxin 30 μg/kg/day (T4), Group 4, 17 β-estradiol 0.3 mg/kg/week (E2), and Group 5, the combination of T4 and E2. The duration of the experiment was 12 weeks. At the end of the experiment, serum chemistries were measured. Bone minerals in the femur were determined with single photon absorptiometry and bone turnover was assessed histomorphometrically. Alkaline-phosphatase increased in Group 3 (OVX-T4), but it reduced in Groups 4 (OVX-E2) and 5 (OVX-T4 + E2). Bone minerals decreased in Groups 2 (OVX) and 3. In Group 4, it was preserved at the same level as in Group 1. Group 5 showed a significant increase of bone mass compared with Group 1. Eroded surface and osteoid surface increased in Groups 2 and 3 and they were reduced in Groups 4 and 5. Bone volume and mineral apposition rate were at a maximum in Group 5. This study demonstrated that 17 β-estradiol was capable of preventing the bone mass decrease by regulating the turnover in ovariectomized rats taking a mild dose of thyroxin. Osteoblast function appeared to be stimulated in combination with 17 β-estradiol and thyroxin.

Estrogen does not increase bone formation in growing rats.

The purpose of these experiments was to test the hypothesis that estrogen has an anabolic effect on cancellous bone formation in long bones of growing rats. In the present studies, estrogen decreased measurements related to cancellous bone formation, including osteoblast number, fluorochrome-labeled bone perimeter, and incorporation of [3H]proline into osteoid. Steady state mRNA levels for bone matrix proteins and mineral apposition rate were unchanged or reduced. The [3H]thymidine labeling index for osteoblasts was very low and was reduced by estrogen treatment. Finally, estrogen decreased the rate of removal of tetracycline previously incorporated into cancellous bone as well as the growth-dependent decrease in cancellous bone area, indicating that hormone treatment reduced net bone resorption. In summary, no evidence was secured to support the hypothesis that estrogen has an anabolic action on bone formation; the results obtained in these studies consistently demonstrated a pronounced inhibitory action of the hormone on bone turnover.

Estrogen does not increase bone formation in growing rats

The purpose of these experiments was to test the hypothesis that estrogen has an anabolic effect on cancellous bone formation in long bones of growing rats. In the present studies, estrogen decreased measurements related to cancellous bone formation, including osteoblast number, fluorochrome-labeled bone perimeter, and incorporation of [3H]proline into osteoid. Steady state mRNA levels for bone matrix proteins and mineral apposition rate were unchanged or reduced. The [3H]thymidine labeling index for osteoblasts was very low and was reduced by estrogen treatment. Finally, estrogen decreased the rate of removal of tetracycline previously incorporated into cancellous bone as well as the growth-dependent decrease in cancellous bone area, indicating that hormone treatment reduced net bone resorption. In summary, no evidence was secured to support the hypothesis that estrogen has an anabolic action on bone formation; the results obtained in these studies consistently demonstrated a pronounced inhibitory action of the hormone on bone turnover.

Fabrication of Vascularized Bone Grafts Using Ceramic Chambers

The present study was designed to create a vascularized bone graft combining the osteogenetic potential of bone marrow cells, and vascular bundle implantation in a hydroxyapatite chamber, using New Zealand white rabbits. A hydroxyapatite chamber was molded into a cylindrical shape, and hydroxyapatite fillers were soaked in an autogenous bone-marrow-cell suspension at a concentration of 1.6 x 10(8)/ml. In one group, the implant was packed with allogenic demineralized bone matrix powder (DBM) and implanted in the mid-thigh subcutaneously, with the epigastric vessels running through the chamber. In a second group, the chamber with bone marrow cells and DBM was implanted subcutaneously, without vascular bundle implantation. The control group consisted of a chamber without DBM, soaked in venous blood and implanted in the opposite thigh, with the epigastric vessels running through the chamber. Cross-sectional bone area and mineral apposition rate were measured, in addition to newly-formed vessels. After 3 weeks, chambers implanted with bone marrow cells and DBM demonstrated consistent bone formation in the pores of the chamber walls and within the chambers. No evidence of bone formation was noted in the chambers soaked with venous blood. The results indicated that vascular-bundle implantation promoted earlier bone formation with neovascularization in the chambers with bone-marrow cells and DBM. Microangiograms revealed vascular connections between the vascular bundle and soft tissue surrounding the chamber through newly-formed vessels in the chamber wall. These findings support the concept of creating a preformed vascularized bone graft, to reconstruct segmental bone defects.

Preliminary findings on the effects of sex hormones on bone metabolism in anorexia nervosa

We performed tetracycline double-labeled bone biopsies and serial bone densitometry on four patients with anorexia nervosa to determine the effects of various conditions of sex hormone replacement and body weight status. We found negligible bone mineral apposition rate in two patients with anorexia nervosa who were taking estrogen and progestin pills while weighing only 79% of normal. However, bone mineral apposition rate was normal in two patients with anorexia nervosa who took no sex hormones and remained amenorrheic while weighing 87% of normal. Histomorphometric analysis revealed a decrease in intracortical trabecular area by 57% and an increase in active osteoblastic surface area by 300%, comparing the subjects who were taking hormone replacement to those who were not. There was no direct evidence of increased resorption in any subject. Six months after biopsy, the two subjects taking sex hormones decreased lumbar bone mineral density by 1.9%, while the two taking no sex hormones remained amenorrheic, but increased bone mineral density by 1.3%. Therefore, we propose that sex hormone therapy in the presence of persistently low body weight might be of limited benefit for bone mineral apposition and density.

Response of human cancellous bone to identically structured commercially pure titanium and cobalt chromium alloy porous-coated cylinders

In order to better understand the human cancellous bone ingrowth response into porous coated biomaterials, identically structured commercially pure (CP) titanium and cobalt chromium alloy cylindrical implants were placed into the distal femoral condyles of five human patients. Bone indices (BIs), a measure of bone ingrowth and mineral apposition rates (MARs), were determined for cancellous bone around each implant. The mean BI for CP titanium implants measured 48·4 ± 21·5% while that for cobalt chrome alloy implants measured 26·7 ± 19·7% ( p < 0·05). Mean MARS surrounding CP titanium at 0·92 ± 0–32μ/day were similar to those surrounding cobalt chromium at 0·94 ± 0·39 μm/day ( p > 0·05), however, cancellous bone advanced towards CP titanium and away from cobalt chromium. Based on bone index and the direction of cancellous bone remodeling, the implant model used in this study suggests that there are differences in the response of human cancellous bone to CP titanium and cobalt chromium alloy in porous coated implants.

Comparative effects of clodronate and calcitonin on bone in metastatic breast cancer: A histomorphometric study

We studied the effects of long-term treatment with clodronate, calcitonin or placebo on bone in 36 normocalcaemic women with osteolytic metastases due to breast cancer. Clodronate (1.6 g daily given to 12 patients) induced a significant decrease in osteoclast surface and osteoclast number, and a significant fall in serum calcium and urinary excretion of calcium and hydroxyproline, an effect not noted after treatment with calcitonin (100 U in 12 patients) or in 12 placebo-treated patients. Treatment with clodronate did not abnormally suppress bone turnover nor impair mineralisation, as measured by bone formation and mineral apposition rates.

Tissue-specific expression of bone proteins in femora of growing rats.

Total cellular RNA was extracted from bone cells of three different femoral compartments of 2-mo-old rats. The intact femora were first incubated with collagenase to obtain periosteal cells. The bisected periosteum-free diaphyses and metaphyses were then incubated with collagenase to obtain enriched populations of endosteal and cancellous bone cells, respectively. The total cellular RNA from these three tissues was separated by size using agarose gel electrophoresis, transferred to nylon filters, hybridized to 32P-labeled cDNA probes for glyceraldehyde-3-phosphate dehydrogenase (GAP), pre-pro-alpha (I) type I collagen (collagen), osteocalcin (BGP), and alkaline phosphatase (AP), and the cDNA/mRNA hybrids were visualized by radioautography. Bone matrix deposition was measured in each tissue compartment by tetracycline-based dynamic bone histomorphometry. The bone formation and apposition rates were greatest in the periosteum and least in metaphysis. Mean mRNA levels for collagen and BGP were positively correlated with mean bone formation and mineral apposition rates. Interestingly, mean AP mRNA levels were not correlated with indexes of bone formation. These results demonstrate that the steady-state mRNA levels for bone matrix proteins in femora show pronounced site specificity and correlate with the rates of bone matrix deposition.

Effects of long-term daily administration of prostaglandin-E2 on maintaining elevated proximal tibial metaphyseal cancellous bone mass in male rats

The effects of long-term prostaglandin E2 (PGE2) on cancellous bone in proximal tibial metaphysis were studied in 7-month-old male Sprague-Dawley rats given daily subcutaneous injections of 0, 1, 3, and 6 mg PGE2/kg/day and sacrificed after 60, 120, and 180 days. Histomorphometric analyses were performed on double fluorescent-labeled undecalcified bone specimens. After 60 days of treatment, PGE2 produced diffusely labeled trabecular bone area, increased trabecular bone area, eroded and labeled trabecular perimeter, mineral apposition rate, and bone formation rate at all dose levels when compared with age-matched controls. In rats given PGE2 for longer time periods (120 and 180 days), trabecular bone area, diffusely labeled trabecular bone area, labeled perimeter, mineral apposition, and bone formation rates were sustained at the elevated levels achieved earlier at 60-day treatment. The eroded perimeter continued to increase until 120 days, then plateau. The observation that continuous systemic PGE2 administration to adult male rats elevated metaphyseal cancellous bone mass to 3.5-fold of the control level within 60 days and maintained it for another 120 days indicates that the powerful skeletal anabolic effects of PGE2 can be sustained with continuous administration.

口蓋深形成術施行後の治癒経過に関する実験的研究

To elucidate postoperative healing at the lateral site of palatal vault osteotomy by the modified method of Yoshizawa and Takahashi, operation was conducted experimentally on 25 adult dogs. One or two months later, the site was examined by soft X-ray radiography and contact micro-radiography, and ground sections of undecalcified bone specimens were examined after Cole's hematoxylin-eosin stain and Villanueva bone stain. These sections were then subjected to bone histomorphometry.The direction of bone growth on cut surfaces varied depending on the postoperative arrangement of segments of hard palate bones separated by osteotomy. New bone formation proved to begin at a point where the space between the interior and exterior cutends were narrowest. In hard palate segments, bone resorption was noted on the nasal surface, and bone formation, on the oral surface.The shape of a cut end of the original bone was classified into 4 types, and newly formed bone, into 5 types. With the exterior cut end of the original bone, most commonly, the upper part of the cut end projected (upper projecting type), and new bone formed over the projecting surface (upper forming type). With the interior cut end of the original bone, the type such that the lower part of the cut end projected (lower projecting type) was usually found, and new bone mostly formed over the projecting surface (lower forming type), except in the case of the no forming type.Bone histomorphometry was carried out as 3 parameters, new bone volume, corrected new bone volume and mineral appositional rate. New bone volume on an exterior cut end was greater than that on an interior cut end. The volume 2 months after osteotomy was greater than that at 1 month. Corrected new bone volume on the exterior cut end was almost the same as that on the interior cut end. The mineral appositional rate 1month after osteotomy was slightly faster than that at 2 months. The rate on the exterior cut end was faster than on the interior cut end.