Showing papers in "Calcified Tissue International in 1983"
TL;DR: Taking into account all factors, 1-hydroxypentylidene-1,1-bisphosphonate and AHPrBP seem to be the most active compounds to inhibit bone resorption.
Abstract: A variety of bisphosphonates with aliphatic side chains of increasing length, as well as 3-amino-1-hydroxypropylidene-1,1-bisphosphonate (AHPrBP, formerly APD), dichloromethylene-bisphosphonate (Cl2MBP, formerly Cl2MDP), and dibromomethylene bisphosphonate (Br2MBP, formerly Br2MDP), were compared in vitro and in vivo to find (a) a possible relationship between structure and activity in order to give some indication about their mechanism(s) of action on bone and (b) the most efficient and safe compound having an effect on bone resorption. Some relationship was found between inhibition of calcium phosphate precipitation in vitro and of mineralization in vivo. No correlation existed, however, between any parameter measured and bone resorption. The number of calvaria cells in culture was decreased by compounds with a chain length greater than 5-C, by AHPrBP, Cl2MBP, and Br2MBP. Lactate production by these cells in vitro was increased by the long chain bisphosphonates and AHPrBP, and was decreased by Cl2MBP. No good correlation existed between the inhibition of bone resorption measured in vitro on calvaria and that seen in vivo on rat tibiae metaphyses. The latter was inhibited the most efficiently by the bisphosphonates longer than 5-C and by AHPrBP; these were 10 times more effective than Cl2MBP. Taking into account all factors, 1-hydroxypentylidene-1,1-bisphosphonate and AHPrBP seem to be the most active compounds to inhibit bone resorption.
297 citations
TL;DR: It is concluded that corticosteroid therapy results in a reduction of the MWT of trabecular bone packets and, consequently, of TBV in these patients, where the OAR was normal, the reduction in MWT was apparently caused by a shortening of the lifespan of the active osteoblastic population at the basic multicellular unit (BMU) level.
Abstract: We have compared the mean wall thickness (MWT) and active formation periods (sigma f(A] of trabecular bone packets in iliac crest biopsies from 20 patients (7 male, 13 female) with corticosteroid-induced osteoporosis (CS-OP) and 20 age- and sex-matched controls. The trabecular bone volume (TBV) of the CS-OP patients (9.6% +/- 2.2% [SD]) was significantly reduced compared to controls (19.3% +/- 5.1%). The MWT of CS-OP patients (32.7 +/- 4.3 micron) was also significantly lower than the control value (48.0 +/- 6.2 micron). There was a positive correlation between MWT and TBV in both groups. The mineralization rate (M) of the CS-OP patients (0.54 +/- 0.25 micron/day) was within the normal range, and since there was no increase in osteoid seam thickness, so therefore was the osteoblastic appositional rate (OAR). The active formation period of trabecular bone packets (sigma f(A) = MWT/M) was significantly lower in the CS-OP patients (55.9 +/- 14.4 days) than in the control group (68.1 +/- 9.4 days). MWT and sigma f(A) both decreased with age in the control group, whereas in the CS-OP group they were independent of age. We conclude that corticosteroid therapy results in a reduction of the MWT of trabecular bone packets and, consequently, of TBV. In these patients, where the OAR was normal, the reduction in MWT was apparently caused by a shortening of the life-span of the active osteoblastic population at the basic multicellular unit (BMU) level.
218 citations
TL;DR: The major advances of the past several years in bone measurement have direct applicat ions for manned space missions, for paraplegics, for patients confined to bed by disease and for the large population of relatively hypodynamic elderly individuals.
Abstract: The major advances of the past several years in bone measurement have direct applicat ions for manned space missions, for paraplegics, for patients confined to bed by disease and for the large population of relatively hypodynamic elderly individuals. A meeting held in San Francisco (June 16, 1982) focussed on newer measurement methods and on major results obtained by both histological and noninvasive approaches. The specific aim was to provide the National Aeronautics and Space Administration (NASA) with information to help implement appropriate programs of research. G. Donald Whedon, National Institute of Health, summarized the history of research on bone loss in immobilization and space flight. Negative calcium balance of 150-200 mg/day continued for up to 20 -30 weeks in young bed rest subjects; in the Skylab astronauts, the pattern and degree of calcium loss was similar to that in bed rest but with much inter-individual variation. Calcium losses in patients with spinal cord injuries appeared somewhat higher than in bed rest subjects, and higher losses were found in subjects with complete versus incomplete spinal cord lesions. Calciuria usually declined to the normal range by 30 weeks, but in some patients, a modest elevation was evident even after one year (N. Eric Naftchi, New York University). Losses in paraplegic patients were evidenced h i s t o m o r p h o m e t r i c a l l y by a 33% reduc t i on of trabecular bone volume in iliac crest biopsy over 25 weeks (P. Minaire and C. Alexandre, Hospital Regional, St. Etienne, France). There was both an in-
190 citations
TL;DR: For a given animal age and degree of mineralization, crystallinity increases with animal age, indicating that changes in bone mineral occur even after mineralization is complete or nearly complete.
Abstract: The crystallinity of bone mineral at different stages of maturation has been measured by quantitative X-ray diffraction methods. Crystallinity measurements were made on tibial middiaphyses from 17-day embryonic chicks, newlyformed periosteal bone from embryonic chicks, and density-fractionated bone from post-hatch chickens from 5 weeks to 2 years of age. For a given animal age and degree of mineralization, crystallinity increases with animal age, indicating that changes in bone mineral occur even after mineralization is complete or nearly complete.
164 citations
TL;DR: It is shown that BGP is chemotactic for cultured human breast cancer cells, human and mouse monocytes, and for cultured rat osteosarcoma cells which have the characteristics of osteoblasts, and a synthetic peptide, which is identical to the carboxyterminal peptide cleaved from BGP when digested by trypsin, is also chemOTactic for the same cells.
Abstract: We have found that the gamma-carboxyglutamic acid (GLA)-containing protein from bone (BGP, osteocalcin) has chemotactic activity in vitro for a number of cells which are found adjacent to endosteal bone surfaces in vivo. Using the Boyden chamber technique for measuring cell chemotaxis in vitro, we have shown that BGP is chemotactic for cultured human breast cancer cells, human and mouse monocytes, and for cultured rat osteosarcoma cells which have the characteristics of osteoblasts. The migration of these cells in response to BGP is undirectional and not due to spontaneous or random migration. A synthetic peptide (Phe-Tyr-Gly-Pro-Val), which is identical to the carboxy-terminal peptide cleaved from BGP when digested by trypsin, is also chemotactic for the same cells. BGP retains its chemotactic activity after conversion of the gamma-carboxyglutamic acid residues to glutamic acid, indicating that this biological effect requires neither gamma-carboxyglutamate nor the ability of BGP to bind calcium. Since BGP is released from bone during states of increased bone turnover, it is possible that this chemotactic effect of the protein may be a mechanism for recruitment of these cells to sites of active bone remodeling.
160 citations
TL;DR: The density values obtained for the proximal humerus differ from those in the literature which represent the femur, however, they are more readily compared with the results of clinical densitometry and may have greater clinical applications.
Abstract: Study of the humeral cortex of 89 acute cadavers showed that an important factor contributing to the physiologic bone loss of aging is increasing bone porosity. Mean cortical porosity increases in both sexes with age, from 4.6% in men and 4% in women at 40 years of age to 10% and more at age 80. In the population studied, no significant difference of porosity was observed between men and women. Apparent mineral density is linked to porosity, and decreases markedly with age in women. Changes in men are lesser in magnitude and show a larger difference of density values. Correction of the apparent mineral density, by a factor reflecting the proportion of vascular and resorption spaces in the cortical bone, produces a true mineral density which does not vary significantly with age in either sex. The density values obtained for the proximal humerus differ from those in the literature which represent the femur. However, they are more readily compared with the results of clinical densitometry and may have greater clinical applications.
123 citations
TL;DR: Results are interpreted as indicating that osteoprogenitor cells and osteoblasts are target cells for the direct action of 1,25-D3 on fetal bone.
Abstract: Thaw-mount autoradiographic studies after injection of3H-1,25-D3 were conducted on 18-and 20-day-old rat fetuses. In maxillary bones, ribs, and tibia, nuclear concentration of radioactivity was found in osteoprogenitor cells and osteoblasts. Osteocytes and chondrocytes in epiphyseal plates were either unlabeled or weakly labeled. In competition experiments, nuclear concentration of radioactivity was blocked by the injection of a high dose of nonradioactive 1,25-D3 prior to the administration of the labeled hormone, but not by a similar dose of nonradioactive 25-D3. The results are interpreted as indicating that osteoprogenitor cells and osteoblasts are target cells for the direct action of 1,25-D3 on fetal bone.
107 citations
TL;DR: The thermodynamic analysis of the solution composition data suggests that ACP converts into a nonstoichiometric apatite when the OCP-like intermediary is formed, and a stoichiometric Apatite is formed when no O CP- like intermediary is involved.
Abstract: The hydrolysis of previously prepared amorphous calcium phosphate (ACP) was studied in a solution “saturated” with ACP; this eliminated the initial consumption of acid due to ACP dissolution. The procedure established that conversion of a high-concentration ACP slurry to an apatite involves two processes: the first process consumes acid and indicates the formation of a more acidic calcium phosphate intermediary with the solubility of octacalcium phosphate (OCP); the second process consumes base and indicates the conversion of the intermediary to apatite and, possibly, direct conversion of ACP to apatite. The thermodynamic analysis of the solution composition data suggests that ACP converts into a nonstoichiometric apatite when the OCP-like intermediary is formed, and a stoichiometric apatite is formed when no OCP-like intermediary is involved.
105 citations
TL;DR: The linear attenuation coefficient (μ in cm1) of trabecular bone was modeled for different conditions of bone and marrow composition in order to assess their influence on computed tomography (CT) quantitation.
Abstract: The linear attenuation coefficient (mu in cm-1) of trabecular bone was modeled for different conditions of bone and marrow composition in order to assess their influence on computed tomography (CT) quantitation. A large relative change (10% of TBV at 15% TBV) of bone concentration resulted in small changes of mu: 2.3% at 60 keV, 3.4% at 44 keV, 5.2% at 29 keV. Relative changes of trabecular bone volume (TBV) on the order of 3% could be detected in vivo by CT were it not for errors of relocation and for compositional influences on accuracy. The mu (and density) depended critically not only on amounts of bone substance and marrow but on their compositions. Normal variation in the composition of bone substance produced an uncertainty in mu equivalent to 0.5 to 1% TBV. Increases of yellow marrow produced a decrease of mu which could be mistaken for a decrease of bone concentration. The biological variation (90% confidence limit) of marrow composition gives an uncertainty at 15% TBV of about 2.4% TBV at 60 keV, 1.7% at 44 keV, and 1.3% at 29 keV. These correspond to relative uncertainties of 16, 11, and 9% respectively. These factors help explain the large accuracy errors (30%) observed in all studies of trabecular bone where single-energy CT was used. Marrow composition also can affect precision of bone measurement. Systematic shifts of red and yellow marrow could mask biological changes such as those occurring with aging or treatment.
99 citations
TL;DR: Though hydroxypyridinium residues became the major aldehyde-mediated crosslinks of adult dentin collagen, significant levels of reducible crosslinks remain throughout the tooth's adult life.
Abstract: The hydroxypyridinium crosslinks of collagen are believed to derive from reducible, divalent crosslinks. To study this concept further, both types of crosslink were quantified as a function of age in dentin, a tissue thought to have minimal collagen turnover. Human (5, 15, 28 and 56 years) and bovine (fetal and adult) root dentin was analyzed by a procedure that measures both hydroxypyridinium and reducible crosslinks on the amino acid analyzer. In human dentin, hydroxypyridinium crosslinks increased with age and became the predominant crosslinks as the two reducible residues, dehydrodihydroxylysinonorleucine and dehydrohydroxylysinonorleucine, diminished. Similarly in adult bovine dentin, hydroxypyridinium residues were sixfold more concentrated than in fetal bovine dentin. Borohydride treatment of tissue did not influence the measured content of hydroxypyridinium residues. The analyses also ruled out natural reduction as a stabilizing reaction for the divalent, reducible crosslinks. Though hydroxypyridinium residues became the major aldehyde-mediated crosslinks of adult dentin collagen, significant levels of reducible crosslinks remain throughout the tooth's adult life.
93 citations
TL;DR: In this group of animals the mineral apposition rate was reduced at the metaphyseal endosteum but increased at the periosteUM, indicating different control mechanisms at the two sites, and aluminum administration caused no skeletal changes in nonuremic animals.
Abstract: The effects of intraperitoneal aluminum chloride (1.5 mg aluminum/kg/day for 9 weeks) were studied in normal and uremic rats. Parameters measured included tissue aluminum, serum vitamin D metabolites, and quantitative bone histology.
TL;DR: It is reported here that prostaglandin I2 (PGI2) and dibutyryl cyclic AMP induce an identical change in osteoclastic behavior, which suggests that PGI2 and CT cause cytoplasmic quiescence by increasing the intracellular level of cyclIC AMP, a view compatible with the known ability of CT to increase cyclicAMP in bone.
Abstract: We separated osteoclasts from bone and observed the effect of several known and potential mediators of the control of bone resorption on their cytoplasmic motility. We already found that calcitonin (CT), a hormone that inhibits bone resorption, regularly causes complete inhibition of cytoplasmic motility, specific for osteoclasts, through a trypsin-sensitive membrane receptor [1]. We report here that prostaglandin I2 (PGI2) and dibutyryl cyclic AMP induce an identical change in osteoclastic behavior. We found that theophylline, which inhibits intracellular cyclic AMP degradation, and which itself had no effect on osteoclastic motility, potentiated the cytoplasmic inhibition caused by CT, PGI2, and cyclic AMP. This suggests that PGI2 and CT cause cytoplasmic quiescence by increasing the intracellular level of cyclic AMP, a view compatible with the known ability of CT to increase cyclic AMP in bone [2]. Parathyroid hormone (PTH), PGE2, and 1,25 dihydroxycholecalciferol (1,25 (OH)2D3), hormones known to stimulate osteoclasts, did not stimulate the activity of either active or quiescent isolated osteoclasts. The undoubted ability of these hormones to stimulate osteoclastic activity in vivo may therefore be mediated through a primary hormonal interaction with another cell type.
TL;DR: The results indicated that although the responses of the two bone systems to the calcemic hormones were qualitatively similar, media and culture conditions could markedly affect the sensitivity.
Abstract: The relative responses of fetal rat limb bones and neonatal mouse calvaria to parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) were examined in organ culture. Limb bones were cultured in dishes in BGJ+1 mg/ml bovine serum albumin (bSA) or DMEM+15% heat-inactivated horse serum (hS). Calvaria were either cultured in dishes with one of the above two media or in roller tubes in DMEM+hS. The order of sensitivity to PTH was: calvaria in roller tubes>limb bones in dishes in DMEM+hS>limb bones in dishes in BGJ+bSA ≥ calvaria in dishes in DMEM+hS>calvaria in dishes in BGJ+bSA. The most sensitive system (i.e., calvaria in roller tubes) showed significant resorption in response to 10−10M PTH at 48 h. The least sensitive system did not respond to 3×10−8M PTH after the same length of time. The greater response in DMEM+hS compared with BGJ+bSA appeared to be due to the protein component of the medium. The order of sensitivity to 1,25(OH)2D3 was: calvaria in roller tubes=long bones in culture dishes in BGJ+bSA>long bones in culture dishes in DMEM+hS>calvaria in dishes in DMEM+hS>calvaria in dishes in BGJ+bSA. The most sensitive systems showed resorption in response to 10−11M 1,25(OH)2D3 by 72 h. The least sensitive system failed to respond to 10−9M 1,25(OH)2D3 after the same length of time. The nature of the protein constituent did not seem to influence the response of the limb bones to 1,25(OH)2D3. The results indicated that although the responses of the two bone systems to the calcemic hormones were qualitatively similar, media and culture conditions could markedly affect the sensitivity.
TL;DR: Apatites containing Na, Mg, and CO3 in amounts occurring in tooth enamel mineral were synthesized by precipitation from aqueous solutions by using X-ray diffraction techniques, which showed only apatitic reflections, which were somewhat broader than those of enamel.
Abstract: Apatites containing Na, Mg, and CO3 in amounts occurring in tooth enamel mineral were synthesized by precipitation from aqueous solutions. The X-ray diffraction patterns showed only apatitic reflections, which were somewhat broader than those of enamel. X-ray Guinier photographs resulted in diffuse reflections from which the lattice parameters could not be determined. The infrared (IR) absorption spectrum of the synthetic samples was practically identical with that of tooth enamel. However, after heating the samples for 1/2 h at a temperature between 300 and 600°C, no absorption peak was found at 2340 cm−1 as in the IR spectrum of tooth enamel after heating.
TL;DR: The hypothesis that an abnormal osteocytic function, which may be part of the VDRR phenotype, is involved along with overall impaired bone mineralization in the etiology of the periosteocytic lesion is supported.
Abstract: Hypomineralized periosteocytic lesions (HPL) are peculiar bone features associated with osteomalacia in vitamin D-resistant rickets (VDRR). To determine whether HPL result from defective bone mineralization, tetracycline double-labeled cortical bone specimens from VDRR children were analyzed before (n=13) and after treatment with phosphate supplements combined with vitamin D2 (Pi+D, n=20) or 1,25-dihydroxyvitamin D3 (Pi+1,25) (n=22). On microradiographs of undecalcified sections, the percentage of osteocytes with HPL was determined separately in the old interstitial bone, the well calcified resting osteons, and in the less mineralized growing osteons. Using histomorphometric methods, dynamic parameters of cortical bone mineralization were assessed on the same samples. In untreated patients, HPL were more prominent in the less calcified bone areas, in accordance with the reported decrease with bone aging. In contrast to therapy with Pi+D, Pi+1,25 reduced HPL frequency in all cortical areas in correlation with improvement of dynamic parameters of bone mineralization. In addition, HPL frequency progressively decreased with the duration of treatment, further demonstrating that the lesion resulted in part from the defective cortical bone mineralization. However, HPL frequency was unrelated to the severity of osteomalacia in untreated children and the lesion persisted in more than 20% of young osteocytes despite complete correction of bone mineralization parameters. The data support the hypothesis that an abnormal osteocytic function, which may be part of the VDRR phenotype, is involved along with overall impaired bone mineralization in the etiology of the periosteocytic lesion.
TL;DR: The suppressive effects of EGF on MC3T3-El cells in vitro suggest that this hormone may be involved in bone remodelling in vivo as well, and not mediated by prostaglandins.
Abstract: The effect of epidermal growth factor (EGF) on clone MC3T3-El cells that have osteoblastic activity was examined by phase-contrast microscopy and electron microscopy; hydroxyproline content, collagen synthesis, collagen pattern, and alkaline phosphatase (ALP) activity were also determined. We found that EGF (0.4 ng/ml) transformed the cells from their normal polygonal shape to a spindle-like morphology by 8 h. This hormone also caused dose-related suppression of hydroxyproline content and ALP activity which was detectable 2 days and 1 day, respectively, after EGF addition. Indomethacin did not affect hydroxyproline content and ALP activity, suggesting that the effect of EGF on the cells may not be mediated by prostaglandins. Epidermal growth factor at concentrations of 2 to 50 ng/ml significantly decreased collagen synthesis in the cells, whereas protein synthesis was stimulated. Electron microscopy demonstrated that collagen fiber formation was also reduced by EGF; an immature type of fibril was observed compared with the typical cross-striated one in the controls. Moreover, the hormone treatment also resulted in the appearance of type III collagen in addition to the type I already present in the cells. These suppressive effects of EGF on MC3T3-El cellsin vitro suggest that this hormone may be involved in bone remodellingin vivo as well.
TL;DR: A (sub)population of cells obtained from newborn rat calvaria by (sequential) collagenase digestion is grown to confluence in serum-containing medium and are osteoblast-like with respect to high alkaline phosphatase activity and marked responsiveness to parathormone.
Abstract: A (sub)population of cells obtained from newborn rat calvaria by (sequential) collagenase digestion is grown to confluence in serum-containing medium. These cells are osteoblast-like with respect to high alkaline phosphatase activity and marked responsiveness (cAMP) to parathormone. Insulin-like growth factors (IGFs) enhance net incorporation of the labeled precursors thymidine, uridine, and glucose into the respective macromolecules DNA, RNA, and glycogen. Human IGF I is five times as potent as IGF II in evoking these anabolic responses in cultured rat calvaria cells. In contrast to insulin, the factors are effective in concentrations in which they are present in serum.
TL;DR: It is proposed that in bone there are two types of receptors, one governing demineralization via regulation of the calcium influx and one governing adenylate cyclase activity, and the receptors are different from each other with respect to their affinities toward the agonists and the antagonist.
Abstract: The effects of bPTH-(1-84), bPTH-(1-34), [Nle-8, Nle-18, Tyr-34] bPTH-(1-34), bPTH-(1-34) amide (NTA 1-34, desamino bPTH-(1-34), bPTH-(2-34), bPTH-(3-34), and [Nle-8, Nle-18, Tyr-34] bPTH-(3-34) amide (NTA 3-34) were tested in cultured bone cells, isolated from the osteoblast layers of fetal chicken calvaria (cyclic AMP) and in fetal rat calvaria (cyclic AMP, Ca release, and lactate production). Only bPTH-(1-84), bPTH-(1-34), and NTA 1-34 increased cyclic AMP production in a doserelated manner, both in calvaria and in bone cells, whereas all fragments (except NTA 3-34) stimulated bone resorption, the order of decreasing potency being bPTH-(1-84), NTA 1-34, bPTH-(1-34), desamino bPTH-(1-34), bPTH-(2-34), bPTH-(3-34). As in human cells, the antagonist NTA 3-34 inhibited specifically and in a dose-dependent way the cyclic AMP response of maximal concentrations of both bPTH-(1-84) and bPTH-(1-34) in rat calvaria and in chicken bone cells, when measured after short (15 min) and longer (1 1/2–16 h) incubation periods. In addition, measured after 4 h of incubation, NTA 3-34 completely inhibited bPTH-(1-84)-stimulated Ca release using maximal and submaximal concentrations. However, after 6–24 h of incubation, NTA 3-34 had no effect on bPTH-(1-84)-stimulated Ca and lactate release, even at an antagonist/agonist ratio up to 12.5 M, perhaps due to its lower affinity for the PTH receptor. From these findings we propose that (a) in bone there are two types of receptors, one governing demineralization via regulation of the calcium influx and one governing adenylate cyclase activity, and (b) the receptors are different from each other with respect to their affinities toward the agonists and the antagonist.
TL;DR: The present results point to an interaction of EGF with osteoblasts, which has been shown to stimulate bone resorption and increase incorporation of3H-labeled thymidine into acid-precipitable macromolecules.
Abstract: Studies were carried out to identify and characterize the receptors for epidermal growth factor (EGF) in osteoblast-rich newborn rat calvarial cells and in 4 clonal lines derived from a transplantable rat osteogenic sarcoma with a well-characterized osteoblast-like phenotype. The cells were grown in monolayer culture in replicate wells; 40,000-50,000 cpm 125I-labeled mouse EGF with a specific activity of 100-120 microCi/micrograms was added to each well. Binding studies were carried out at 37 degrees C. Binding of 125I-labeled EGF was specific, saturable, reversible, and pH dependent. Maximum binding occurred 2 h after addition of the tracer. Thereafter, cell-bound radioactivity decreased to reach a plateau of 15-20% of maximum binding at 24 h. This observation is consistent with internalization and processing of the receptor-hormone complex as has been shown with other EGF target cells. Scatchard analyses revealed a single class of high-affinity binding sites in the normal and malignant osteoblast-like cells. Dissociation constants (KD) in the clonal lines ranged from 2.3 X 10(-10)M to 4.7 X 10(-10)M with receptor number per cell ranging from 25,000 to 33,000. The calvarial cells had a KD of 2.0 X 10(-10)M with 14,000 receptors per cell. In both the normal and malignant cell strains, EGF was found to increase incorporation of 3H-labeled thymidine into acid-precipitable macromolecules. EGF has been shown to stimulate bone resorption; however, studies in organ cultures have not identified the target cell for EGF. The present results point to an interaction of EGF with osteoblasts.
TL;DR: The mechanical properties and structural changes in the monkey tibia with disuse osteoporosis and during subsequent recovery and even after 15 months of recovery, bone mineral content did not necessarily return to normal levels.
Abstract: The mechanical properties and structural changes in the monkey tibia with disuse osteoporosis and during subsequent recovery are investigated. Bone mending stiffness is evaluated in relation to microscopic changes in cortical bone and Norland bone mineral analysis. Restraint in the semireclined position is found to produce regional losses of bone most obviously in the anterior-proximal tibiae. After six months of restraint, the greatest losses of bone mineral in the proximal tibiae range from 23 percent to 31 percent; the largest changes in bone stiffness range from 36 percent to 40 percent. Approximately eight and one-half months of recovery are required to restore the normal bending properties. Even after 15 months of recovery, however, the bone mineral content does not necessarily return to normal levels. Histologically, resorption cavities in cortical bone are seen within one month of restraint; by two and one-half months of restraint there are large resorption cavities subperiosteally, endosteally, and intracortically. After 15 months of recovery, the cortex consists mainly of first-generation haversian systems. After 40 months, the cortex appears normal, with numerous secondary and tertiary generations of haversian systems.
TL;DR: The results from this study indicate that vitamin D does not appear to play a major role in fetal skeletal development, however, after birth, vitamin D becomes progressively more important with age for normal bone development, mineralization, and endochondral growth.
Abstract: The role of vitamin D in early skeletal development was studied by measuring serum calcium and phosphorus, osseous tissue quantity and mineralization, and endochondral bone elongation in rat fetuses and pups from vitamin D-replete and vitamin D-deficient mothers. At the 20th day of pregnancy there was a slight, yet significant, increase in the amount of osteoid on trabecular bone surfaces in fetuses from vitamin D-deficient mothers. The fetal bones otherwise appeared normal in spite of severe skeletal changes in the vitamin D-deficient mothers. After parturition, the importance of vitamin D in skeletal development becomes progressively more obvious. Serum calcium levels were slightly, yet significantly, lower in vitamin D-deficient than in vitamin D-replete pups and these levels continued to fall in the vitamin D-deficient pups through lactation and after weaning. At 3 days postpartum, there was a small, yet significant, increase in the amount of osteoid on bone surfaces of the vitamin D-deficient pups. The relative amounts of osteoid in the vitamin D-deficient pups continued to increase through lactation and after weaning when compared with vitamin D-replete pups. By the 14th day of lactation and at later periods, there were significant reductions in metaphyseal mineralized tissues in the vitamin D-deficient pups when compared with the vitamin D-replete pups. At weaning and after weaning, there were substantial increases in growth plate thickness and decreases in longitudinal bone growth in the vitamin D-deficient pups when compared with the vitamin D-replete pups. The results from this study indicate that vitamin D does not appear to play a major role in fetal skeletal development. However, after birth, vitamin D becomes progressively more important with age for normal bone development, mineralization, and endochondral growth.
TL;DR: It is demonstrated that TBV is lost from iliac crest throughout adult life, and the large spread in TBV indicates further that factors operating during adolescence or even earlier may have an important impact on skeletal mass.
Abstract: We measured the trabecular bone volume (TBV) of 62 iliac crest biopsies taken from women admitted to lymphoma protocols at Stanford University between 1970-1981 All subjects were active, cycling premenopausal women, with bone marrows that were negative for tumor Disease status was stage III or less in 90% of the subjects Trabecular bone volume was negatively correlated with age, and the annual predicted loss of bone was 014-018% TBV, or 07% of the original bone volume In addition, there was a substantial range of normal TBV at any given age, evident even during adolescence This study demonstrates that TBV is lost from iliac crest throughout adult life The large spread in TBV indicates further that factors operating during adolescence or even earlier may have an important impact on skeletal mass
TL;DR: The results point to the possible importance of prostacyclin either in bone resorption or in some other local function, e.g., regulation of bone blood flow, in both normal and malignant osteoblasts.
Abstract: The metabolism of arachidonic acid to its cyclo-oxygenase products was studied in monolayer cultures of osteoblast-rich rat calvarial cells and of clonal cell lines from a rat osteogenic sarcoma, enriched in the osteoblast phenotype. Prostanoids were measured by radioimmunoassay after extraction of media and fractionation by high pressure liquid chromatography. In both normal and malignant osteoblasts the major cyclo-oxygenase product was 6-oxo-prostaglandin F1α, the hydration product of prostacyclin, with lesser amounts of prostaglandin E2 and prostaglandin F2α. No significant thromboxane B2 was detected. Prostaglandins are thought to have a local role in the regulation of bone resorption. These results point to the possible importance of prostacyclin either in bone resorption or in some other local function, e.g., regulation of bone blood flow.
TL;DR: Highly phosphorylated noncollagenous proteins (NCP) with molecular weights of ∼70–100,000 daltons have been purified from rat and bovine dentin, finding racemization presents no problem for shorter lived mammals, but could be partly responsible for purification difficulties with human dentin.
Abstract: Highly phosphorylated noncollagenous proteins (NCP) with molecular weights of ∼70–100,000 daltons have been purified from rat and bovine dentin. Efforts to isolate phosphoprotein from human teeth have not yielded consistent results, and failures have been attributed to proteolysis due to preparative techniques. Diagenetic reactions affecting metabolically stable proteinsin vivo also can interfere in protein purification. Racemization is one of the reactions known to take place in human dentin. EDTA extraction of dentin from an age-graded series of human teeth has yielded an EDTA-soluble NCP fraction having an aspartic acid racemization rate 3 X that in unfractionated dentin and 8 X the rate in EDTA-insoluble protein. D-Aspartic acid is accumulating in EDTA-S protein at a rate of 0.22% yr−1. For humans, more than 13% of the aspartyl residues in NCP will be the D-enantiomer by 60 years of age. While racemization presents no problem for shorter lived mammals, such as rats, it could be partly responsible for purification difficulties with human dentin.
TL;DR: It is suggested that chemotactic protein(s) have a significant role in the recruitment of osteoprogenitor cells to a site of bone repair.
Abstract: When demineralized bone matrix powder is implanted subcutaneously in the rat, the early responses involve the appearance and proliferation of mesenchymal cells at the site of implantation, followed by cartilage and bone formation. The ability of cells to migrate to the implant suggests that chemotaxis may be a critical event in this process. Therefore, using the modified Boyden chamber assay, we tested extracts of demineralized bone matrix for chemotactic activity. We have identified and partially purified, on molecular sieve chromatography, a heat labile and trypsin-sensitive protein (Mr=60,000–70,000) that is a potent chemoattractant for mouse calvaria, osteoblast-like cells (MMB-1), but not for monocytes (putative osteoclast precursors). These findings suggest that chemotactic protein(s) have a significant role in the recruitment of osteoprogenitor cells to a site of bone repair.
TL;DR: The stimulatory role of progesterone in combination with estradiol in bone formation and mineralization was demonstrated and ornithine decarboxylase activity was significantly suppressed by the combination of Estradiol and progester one.
Abstract: The influence of estradiol and progesterone, alone or in combination, on the discrete phases of matrix-induced endochondral bone formation was investigated. Administration of estradiol and progesterone in combination increased mesenchymal cell proliferation, as indicated by [3H] thymidine incorporaton into acid precipitable material. However, ornithine decarboxylase (ODC) activity was significantly suppressed by the combination of estradiol and progesterone. Also, this treatment did not influence the35SO4 incorporation into proteoglycans on day 7. Mineralization of newly induced bone was quantitated by alkaline phosphatase,45Ca incorporation into bone mineral and calcium content, and was found to be significantly increased by progesterone alone and in combination with estradiol in both matrix-induced plaques and tibial metaphysis. These results demonstrated the stimulatory role of progesterone in combination with estradiol in bone formation and mineralization.
TL;DR: The data indicate that 1,25(OH)2D3 can regulate bone turnover as well as mineral homeostasis in the young mouse, and produces a dose-dependent stimulation of bone mineralization rate in response to increased osteoclastic bone resorption.
Abstract: The effects of continuous administration of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] on mineral and bone metabolism have been examined in the normal mouse. Four doses (0.05-0.25 micrograms/kg/day) of 1,25(OH)2D3 were infused continuously for 4 weeks in 21-day-old intact animals. Mineral and skeletal changes were evaluated by analytical methods and by histomorphometric analysis of endosteal bone formation and resorption parameters. All doses of 1,25(OH)2D3 increased the fractional osteoclastic surface and the osteoclast number in conjunction with increased hydroxyproline excretion. 1,25(OH)2D3 induced a dose-dependent elevation of the calcification rate, reduction of the mean osteoid seam thickness, and shortening of the mineralization lag time. In addition, there was a dose-related increase in the extent of tetracycline double-labeled osteoid surface and a concurrent rise in the fractional osteoblastic surface associated with elevated serum alkaline phosphatase levels. Increased bone formation appeared to have been balanced by increased bone resorption since the trabecular bone volume remained unchanged. Except at the highest dose given, serum calcium and phosphate concentrations remained normal in spite of increased bone mobilization and presumably enhanced intestinal absorption of minerals. Urinary cAMP and TmP/GFR remained normal, suggesting that parathormone secretion was not altered. The results show that continuous 1,25(OH)2D3 infusion in the young mouse produces a dose-dependent stimulation of bone mineralization rate in response to increased osteoclastic bone resorption. The data indicate that 1,25(OH)2D3 can regulate bone turnover as well as mineral homeostasis in the young mouse.
TL;DR: The osteoblast is established as a target cell for EGF action on bone with a dose-dependent increase in cell proliferation by calvarial cells.
Abstract: Normal and malignant osteoblast-like cells in culture have been shown to possess specific, high affinity receptors for epidermal growth factor (EGF). In this study, the mitogenic response to EGF was examined in a clonal line of a rat osteogenic sarcoma (UMR 106) and in osteoblast-rich newborn rat calvarial cells. Twenty-four hour treatment of UMR 106 cells with EGF in doses ranging from 10−12
m to 2 × 10−8
m stimulated the incorporation of [3H]thymidine and DNA synthesis in a dose-dependent manner. This short-term stimulatory effect was sustained in long-term culture with a dose-dependent increase in cell proliferation by calvarial cells. A lag period of 8 h occurred before significant stimulation of [3H]thymidine incorporation was observed. Commitment to increased incorporation of [3H]thymidine required a minimum of 6 h continuous incubation with EGF. These results establish the osteoblast as a target cell for EGF action on bone.
TL;DR: In this paper, a matrix vesicle-enriched fraction was isolated from different zones of epiphyseal cartilage by nonenzymatic methods involving tissue homogenization, differential centrifugation, and isosmotic percoll gradient fractionation.
Abstract: Matrix vesicle-enriched fractions were isolated from different zones of epiphyseal cartilage by nonenzymatic methods involving tissue homogenization, differential centrifugation, and isosmotic Percoll gradient fractionation. Uptakes of both32Pi and45Ca were studied concomitantly over periods from 20 min to 24 h. Percoll density gradients separated epiphyseal microsomes into two alkaline phosphatase-rich fractions: a low-density noncalcifiable fraction (P-I), and a higher-density fraction (P-II) which readily mineralized. The P-II fraction was found only in calcifying regions of the growth plate. Based on chemical and physical properties and enzyme activities, both fractions were similar except that P-II contained significantly higher levels of mineral ions than did P-I, and had lower levels of alkaline phosphatase. The mineral appeared to be primarily in a noncrystalline form. Metabolism of32Pi and45Ca by P-II followed a complex kinetic pattern in which accumulation of large amounts of both ions was preceded by an initial limited burst of uptake and a lag-phase of variable duration. During mineral ion loading, the density of the P-II fraction progressively increased as evidenced by co-migration of45Ca,32Pi, and alkaline phosphatase to increasingly higher densities. During the period of early mineral deposition (1–5 h), Ca/P uptake ratios were very low (1.0–1.2) and X-ray diffraction patterns showed a predominantly amorphous pattern. This suggests that the mineral accumulated in matrix vesicles is initially some form of noncrystalline calcium monohydrogenphosphate. L-tetramisole, a potent inhibitor of alkaline phosphatase, inhibited accumulation of both45Ca and32Piin the absence of organic P substrates,32Pi being preferentially inhibited over45Ca. This finding, coupled with recent studies on the behavior of alkaline phosphatase at physiological pH, suggests that the protein is not acting as a phosphohydrolase, but rather as a Pi-binding or transport agent in vesicle-mediated calcification.
TL;DR: The association between increased numbers of mast cells and postmenopausal osteoporosis is confirmed and the number of marrow mast cells in undecalcified section of iliac crest bone is compared to values from 10 normal women and 12 normal men.
Abstract: Because mast cell proliferation has been reported in disorders of abnormal bone remodeling, we quantitated the number of mast cells in undecalcified section of iliac crest bone from 16 untreated women with postmenopausal osteoporosis and contrasted the findings to values from 10 normal women and 12 normal men. The mean number of marrow mast cells was greater in normal women than men (1.17 vs 0.40 cells per mm2, P less than 0.05). Compared to the normal women, osteoporotic women had a greater number of mast cells in the marrow (3.38 vs 1.17 cells per mm2, P less than 0.01). However, there was no statistically significant correlation between the number of marrow mast cells and either the patient's age or the severity of the bone lesions, as assessed by histomorphometry. Our findings confirm the association between increased numbers of mast cells and postmenopausal osteoporosis.