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PubMed1982

Lung cancer and smoking in a group of iron ore miners.

Several studies have shown that miners, in both uranium and nonuranium mines, have an increased lung cancer mortality, probably caused by exposure to radon and its daughters. The excess mortality has been observed primarily among smoking miners but some recent studies have also indicated a considerably increased risk among nonsmoking miners. This study, among a group of iron ore miners, was undertaken to further elucidate the somewhat unclear and presumably complex relationship of mining, smoking, and lung cancer. The results show a 16-fold increase in lung cancer mortality among miners versus nonminers. Even nonsmoking miners seem to be at a rather high risk of developing lung cancer, but there was a tendency for the most heavy smoking miners to die earlier and to have a slightly shorter induction-latency period for development of lung cancer than was found among the nonsmoking miners.

Adult↗
PubMed1983

An epidemiological study of salt miners in diesel and nondiesel mines.

A cross-sectional study of 5 NaCl mines and 259 miners addressed the following questions: 1) Is there an association of increased respiratory symptoms, radiographic findings, and reduced pulmonary function with exposure to nitrogen dioxide (NO2) and/or respirable particulate (RP) among these miners? 2) Is there increased morbidity of these miners compared to other working populations? Personal samples of NO2 and respirable particulate for jobs in each mine were used to estimate cumulative exposure. NO2 is used as a surrogate measure of diesel exposure. Cough was associated with age and smoking, dyspnea with age; neither symptom was associated with exposure (years worked, estimated cumulative NO2 or RP exposure). Phlegm was associated with age, smoking, and exposure. Reduced pulmonary function (FVC, FEV1, peak, flow, FEF50, FEF75) showed no association with exposure. There was one case of small rounded and one case of small irregular opacities; pneumoconiosis was not analyzed further. Compared to underground coal miners, above ground coal miners, potash miners, and nonmining workers, the study population after adjustment for age and smoking generally showed no increased prevalence of cough, phlegm, dyspnea, or obstruction (FEV1/FVC less than 0.7). Obstruction in younger salt miners and phlegm in older salt miners was elevated compared to nonmining workers. Mean predicted pulmonary function was reduced 2-4% for FEV1 and FVC, 7-13% for FEF50, and 18-22% for FEF75 below all comparison populations.

Adult↗
PubMed2005

Architecture and mineralization of developing trabecular bone in the pig mandibular condyle.

Architecture and mineralization are important determinants of trabecular bone quality. To date, no quantitative information is available on changes in trabecular bone architecture and mineralization of newly formed bone during development. Three-dimensional architecture and mineralization of the trabecular bone in the mandibular condyle from six pigs of different developmental ages were investigated with micro-CT. Anteriorly in the condyle, a more advanced state of remodeling was observed than posteriorly, where more active growth takes place. Posteriorly, the bone volume fraction increased with age (r=0.87; P<0.05) by an increase of trabecular thickness (r=0.88; P<0.05), while the number of trabeculae declined (r=-0.86; P<0.05). Anteriorly, despite an increase in trabecular thickness (r=0.97; P<0.001), there was no change in bone volume fraction due to a simultaneous decline in trabecular number (r=-0.84; P<0.05) and increase in trabecular separation (r=0.95; P<0.01). Posteriorly, rods were remodeled into plates as expressed by the structure model index (r=-0.97; P<0.001), whereas anteriorly, a plate-like structure was already present in early stages. The trabecular structure had a clear orientation throughout the developmental process. The global degree of mineralization increased both anteriorly (r=0.86; P<0.05) and posteriorly (r=0.89; P<0.05). We suggest that the degree of mineralization does not depend on the bone volume, but on the thickness of the trabeculae as the mineralized centers of trabeculae were getting larger and more highly mineralized with age compared to their appositional layers. This indicates that besides apposition of new bone material on the surface of trabeculae, the mineralized tissue in their centers still changes and matures.

Animals↗
PubMed2004

Quantitative microcomputed tomography analysis of mineralization within three-dimensional scaffolds in vitro.

Synthetic and naturally derived scaffold biomaterials in combination with osteogenic cells or bioactive factors have the potential to serve as bone graft substitutes. Porous poly(l-lactide-co-dl-lactide) (PLDL) scaffolds with mechanical properties comparable to trabecular bone and an oriented, interconnected porosity designed to enhance internal mass transport were recently developed. In this study, PLDL scaffolds were seeded with rat calvarial or rat stromal cells and cultured up to 8 weeks in media containing osteogenic supplements. Cell-seeded human demineralized trabecular bone matrix (DTBM) scaffolds were included for comparison. All constructs were imaged weekly from 4 to 8 weeks using microcomputed tomography (micro-CT) to nondestructively quantify the amount and distribution of mineralized matrix formation. The total mineralized matrix volume increased with time in culture for all construct groups. DTBM constructs contained significantly more mineralized matrix than PLDL constructs. However, an analysis of the acellular DTBM scaffolds exposed to osteogenic media revealed partial remineralization of the demineralized matrix whereas no mineralization was detected in acellular PLDL scaffolds. Differences in mineral distribution were also evident with cell-mediated mineralization found throughout the PLDL constructs but localized to the periphery of the DTBM constructs for both cell types. Expression of bone marker genes indicating osteoblast differentiation was demonstrated in all groups at 8 weeks using a quantitative reverse transcription polymerase chain reaction. Osteocalcin expression was significantly higher for calvarial cell constructs compared to stromal cell constructs, regardless of the type of scaffold. This study demonstrated that micro-CT imaging may be used to nondestructively and quantitatively monitor mineralization within three-dimensional scaffolds in vitro. PLDL scaffolds with an oriented microarchitecture were shown to support cell attachment, differentiation, and cell-mediated mineralization comparable to natural DTBM scaffolds.

Animals↗
PubMed2005

Enhanced intrinsic biomechanical properties of osteoblastic mineralized tissue on roughened titanium surface.

The biological mechanisms underlying bone-titanium integration and biomechanical properties of the integrated bone are poorly understood. This study assesses intrinsic biomechanical properties of mineralized tissue cultured on titanium having different surface topographies. The osteoblastic phenotypes associated with mineral deposition and collagen synthesis underlying the biomechanical modulation are also reported. Rat bone marrow-derived osteoblastic cells were cultured either on the machined titanium disc or acid-etched titanium disc. Nano-indentation study of day 28 culture revealed that the mineralized tissue on the acid-etched surface shows 3-3.5 times greater hardness than that on the machined surface (p < 0.01). Elastic modulus of the mineralized tissue was also 2.5-3 times greater on the acid-etched surface than on the machined surface (p < 0.01). After 28 days of culture, mineralized nodule area was significantly lower on the acid-etched surface than on the machined surface (p = 0.0105), while total calcium deposition did not differ between the two surfaces, indicating denser mineral deposition on the acid-etched surface. Osteopontin and osteocalcin gene expressions assayed by the reverse transcriptase-polymerase chain reaction were upregulated in the acid-etched titanium culture. Collagen synthesis measured by Sirius red stain-based colorimetry was 1.5-10 times higher on the acid-etched surface than on the machined surface in the initial culture period of day 1 to day 14 (p < 0.0001). The amount of collagen synthesis corresponded with the enhanced gene expression of prolyl 4-hydroxylase, a key enzyme for post-translational modification of collagen chains. Scanning electron microscopic images revealed that tissue cultured on the acid-etched titanium exhibited plate-like, compact surface morphology, while the tissue on the machined titanium appeared porous and was covered by fibrous and punctate structures. We conclude that culturing osteoblasts on rougher titanium surfaces enhances hardness and elastic modulus of the mineralized tissue, associated with condensed mineralization, accelerated collagen synthesis, and upregulated expression of selected bone-related genes.

Animals↗
PubMed1991

Structural studies of the mineral phase of calcifying cartilage.

The calcified cartilage of the epiphyseal growth plate of young calves has been studied by x-ray diffraction. Fourier transform infrared spectroscopy, magic angle 31P nuclear magnetic resonance spectroscopy, and chemical composition. The powdered tissue was separated by density centrifugation as a function of mineral content and thus qualitatively of the age of the calcium-phosphorus mineral phase. The individual density centrifugation fractions were examined separately. X-ray diffraction of the samples, especially of the lowest density fractions, revealed very poorly crystalline apatite. Fourier transform infrared spectroscopy and 31P nuclear magnetic resonance spectroscopy revealed the presence of significant amounts of nonapatitic phosphate ions. The concentration of such nonapatitic phosphates increases during the early stages of mineralization but then decreases as the mineral content steadily rises until full mineralization is achieved. The total concentration of carbonate ions was found to be much lower in calcified cartilage than in bone from the same organ (scapula). The carbonate ions are located in both A sites (OH-) and B sites (PO4(3-)), with a distribution similar to that found in bone mineral. However, discrepancies between infrared resolution factors of phosphate and carbonate bands are consistent with a heterogeneous distribution of carbonate ions in poorly organized domains of the solid phase of calcium phosphate. These initial studies permit one to characterize the calcium phosphate mineral phase as a very poorly crystalline, immature calcium phosphate apatite, rich in labile nonapatitic phosphate ions, with a low concentration of carbonate ions compared with bone mineral of the same animal, indeed from the bone of the same organ (scapula).

Animals↗
PubMed1992

beta-Glycerophosphate-induced mineralization of osteoid does not alter expression of extracellular matrix components in fetal rat calvarial cell cultures.

When fetal rat calvarial cells are cultured in medium containing vitamin C, osteoid nodules develop after approximately 15 days of culture. Upon addition of an organic phosphate (beta-glycerophosphate, beta GP), these nodules mineralize. We have now used this system to explore the suggestion made by others that a negative feedback may exist between matrix mineralization on the one hand and the synthesis of alkaline phosphatase and bone matrix collagen on the other by analyzing the synthesis of these proteins and the levels of their mRNAs in mineralizing and nonmineralizing cultures. Our results indicate that in the osteoid nodule-bone nodule system, matrix mineralization did not affect the mRNA levels for osteopontin, type I collagen, bone sialoprotein, or osteocalcin. Synthesis of total protein and collagen and the osteocalcin content of culture media were also not different in the mineralizing and nonmineralizing cultures. However, alkaline phosphatase mRNA was increased in early mineralizing cultures and alkaline phosphatase activity in the cell layer was also increased in mineralizing cultures. Thus, the hypothesis that a direct negative feedback exists between mineralization and matrix protein synthesis is not supported by our experiments.

Alkaline Phosphatase↗
PubMed1993

Effect of age and osteoarthritis on bone mineral in rhesus monkey vertebrae.

We have used vertebrae of free-ranging rhesus macaques to study the effect of age and osteoarthritis on bone mineralization and bone density and to relate these findings to weight, sex, parity, and mineral chemistry. Bone mineralization was determined using the density fractionation technique and bone density using dual-photon absorptiometry. Arthritis was determined osteologically. We found a relationship between mineralization, age, and osteophytes, such that mineralization rose with age in nonarthritics and decreased with age in arthritics. This could also be seen when the females were examined separately. In males, only an increase in mineralization with age could be seen. In females mineralization decreases with parity. Also in females, DPA density decreases with age and increases with parity. No relationships with DPA density could be seen using males and females together or males alone. In conclusion, we have shown that normal skeletal aging in rhesus monkeys is accompanied by an increase in mineralization similar to that in other species, but this is not true in the presence of osteoarthritis. In the females parity has an important effect because it seems to build up bone mass even though the bone present may be undermineralized.

Absorptiometry, Photon↗
PubMed1993

Effect of basic fibroblast growth factor on the growth and differentiation of adult stromal bone marrow cells: enhanced development of mineralized bone-like tissue in culture.

Rat stromal bone marrow cells (SBMC) were shown to produce mineralized bone-like tissue in culture in the presence of dexamethasone, ascorbic acid, and beta-glycerophosphate. The addition of 3 ng/ml of basic fibroblast growth factor (bFGF) resulted in a significant increase in formation of mineralized tissue. The present study was aimed at assessing the effect of bFGF on the proliferation and differentiation of SBMC and on the sequential development of mineralized bone-like tissue in culture. Transmission electron microscopy of bFGF-treated cultures demonstrated the development of a multilayered structure resembling mineralized bone tissue consisting of cell layers embedded within a heavy extracellular matrix. The matrix was rich in bundles of collagen fibers associated with extensive mineral deposits consisting of hydroxyapatite as determined by infrared spectrophotometry. The addition of 3 ng/ml of bFGF resulted in significant enhancement of [3H]thymidine and [3H]proline incorporation and protein accumulation by 12-, 2.5-, and 2.5-fold, respectively. bFGF treatment increased cAMP responsiveness, alkaline phosphatase activity, osteocalcin level, 45Ca2+ deposition, and mineralized-like tissue formation and induced the earlier expression of these markers in the treated culture. A biphasic sequence of events was observed during the development of mineralized bone-like tissue in bFGF-treated and control cultures. The first phase is characterized by cell proliferation and matrix accumulation and is reflected by a progressive increase in [3H]thymidine and [3H]proline incorporation until day 11. The second phase, which follows, is characterized by a sharp decline in cell proliferation and matrix accumulation and a concomitant expression of osteoblast differentiation as reflected by the progressive increase in alkaline phosphatase activity, mineral deposition, and osteocalcin expression. Treatment of cultures with bFGF accentuated this biphasic sequence of events. These results indicate that bFGF has the capacity to stimulate both the growth and the biochemical functions of SBMC obtained from a young adult animal.

Alkaline Phosphatase↗
PubMed1993

Differential effects of fluoride during initiation and progression of mineralization of osteoid nodules formed in vitro.

Osteoid nodules form in cultures of fetal rat calvarial (RC) cells grown in medium containing 10% FBS and 50 micrograms/ml of ascorbic acid. When 10 mM beta-glycerophosphate (beta-GP) is added, osteoid nodules mineralize in two phases: an initiation phase, which is dependent upon alkaline phosphatase activity for conversion of beta-GP to P(i), and a progression phase that proceeds independently of alkaline phosphatase activity and does not require exogenous phosphate. We have now used this system to investigate the effects of fluoride (F-) on mineralization. In cultures in which osteoid was formed and mineralization initiated in the presence of F-, a dose-dependent inhibition of the initiation of mineralization occurred over a concentration range of 25-500 microM F- (p < 0.001 in all cases). The initiation of mineralization was not inhibited if F- was removed from the cultures at the time when mineralization was initiated with beta-GP. In osteoid nodules grown in the absence of F-, addition of F- resulted in a dose-dependent inhibition of the initiation of mineralization, with significant decreases in 45Ca uptake occurring at F- concentrations of 3 microM (p < 0.01) and higher. However, if F- was added to cultures after mineralization was initiated in the absence of F-, a stimulation of 45Ca uptake was observed at F- concentrations of 250 microM and above (p < 0.001). F- (1-1000 microM) did not affect the conversion of beta-GP to P(i) or alkaline phosphatase activity in the cultures.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed2006

Evidence that both 1alpha,25-dihydroxyvitamin D3 and 24-hydroxylated D3 enhance human osteoblast differentiation and mineralization.

Vitamin D plays a major role in the regulation of mineral homeostasis and affects bone metabolism. So far, detailed knowledge on the vitamin D endocrine system in human bone cells is limited. Here we investigated the direct effects of 1alpha,25-(OH)2D3 on osteoblast differentiation and mineralization. Also, we studied the impact of 24-hydroxylation, generally considered as the first step in the degradation pathway of vitamin D, as well as the role of the nuclear and presumed membrane vitamin D receptor (VDR). For this we used a human osteoblast cell line (SV-HFO) that has the potency to differentiate during culture forming a mineralized extracellular matrix in a 3-week period. Transcriptional analyses demonstrated that both 1alpha,25-(OH)2D3 and the 24-hydroxylated metabolites 24R,25-(OH)2D3 and 1alpha,24R,25-(OH)3D3 induced gene transcription. All metabolites dose-dependently increased alkaline phosphatase (ALP) activity and osteocalcin (OC) production (protein and RNA), and directly enhanced mineralization. 1Alpha,24R,25-(OH)3D3 stimulated ALP activity and OC production most potently, while for mineralization it was equipotent to 1alpha,25-(OH)2D3. The nuclear VDR antagonist ZK159222 almost completely blocked the effects of all metabolites. Interestingly, 1beta,25-(OH)2D3, an inhibitor of membrane effects of 1alpha,25-(OH)2D3 in the intestine, induced gene transcription and increased ALP activity, OC expression and mineralization. In conclusion, not only 1alpha,25-(OH)2D3, but also the presumed 24-hydroxylated "degradation" products stimulate differentiation of human osteoblasts. 1Alpha,25-(OH)2D3 as well as the 24-hydroxylated metabolites directly enhance mineralization, with the nuclear VDR playing a central role. The intestinal antagonist 1beta,25-(OH)2D3 acts in bone as an agonist and directly stimulates mineralization in a nuclear VDR-dependent way.

24,25-Dihydroxyvitamin D 3↗
PubMed1995

Postembedding colloidal-gold immunocytochemistry of noncollagenous extracellular matrix proteins in mineralized tissues.

Immunocytochemistry is a powerful tool for investigating protein secretion, extracellular matrix assembly, and cell-matrix and matrix-matrix/mineral relationships. When applied to the tissues of bones (bone and calcified cartilage) and teeth (dentin, cementum, and enamel), where calcium phosphate-containing extracellular matrices are the predominant structural component related to their weight-bearing and masticatory roles, respectively, data from immunocytochemical studies have been prominent in advancing our understanding of mineralized tissue modeling and remodeling. The present review on the application of postembedding, colloidal-gold immunocytochemistry to mineralized tissues focuses on the advantages of this approach and relates them to conceptual, theoretical, and experimental data currently available discussing matrix-mineral interactions and extracellular matrix formation and turnover in these tissues. More specifically, data are summarized regarding the distribution and role of noncollagenous proteins in different mineralized tissues, particularly in the context of how they interface with mineral, and how this relationship might be affected by the various tissue-processing steps and immunocytochemical strategies commonly implemented to examine the distribution and function of tissue proteins. Furthermore, a technical discussion is presented that outlines several different possibilities for epitope exposure in mineralized tissues during preparation of thin sections for transmission electron microscopy. Cell biological concepts of protein secretion by cells of the mineralized tissues, and subsequent extracellular matrix assembly and organization, are illustrated by examples of high-resolution, colloidal-gold immunolabeling for osteopontin, bone sialoprotein, and osteocalcin in the collagen-based mineralized tissues and for enamel protein (amelogenin) in enamel.

Animals↗
PubMed2006

Evolutionary genetics of vertebrate tissue mineralization: the origin and evolution of the secretory calcium-binding phosphoprotein family.

Three principal mineralized tissues are present in teeth; a highly mineralized surface layer (enamel or enameloid), body dentin, and basal bone. Similar tissues have been identified in the dermal skeleton of Paleozoic jawless vertebrates, suggesting their ancient origin. These dental tissues form on protein matrix and their mineralization is controlled by distinctive proteins. We have shown that many secretory calcium-binding phosphoproteins (SCPPs) are involved in tetrapod tissue mineralization. These SCPPs all originated from the common ancestral gene SPARCL1 (secreted protein, acidic, cysteine-rich like 1) that initially arose from SPARC. The SCPP family also includes a bird eggshell matrix protein, mammalian milk casein, and salivary proteins. The eggshell SCPP plays crucial roles in rigid eggshell production, milk SCPPs in efficient lactation and in the evolution of complex dentition, and salivary SCPPs in maintaining tooth integrity. A comparative analysis of the mammalian, avian, and amphibian genomes revealed a tandem duplication history of the SCPP genes in tetrapods. Although these tetrapod SCPP genes are fewer in teleost genomes, independent parallel duplication has created distinct SCPP genes in this lineage. These teleost SCPPs are also used for enameloid and dentin mineralization, implying essential roles of SCPPs for dental tissue mineralization in osteichthyans. However, the SCPPs used for tetrapod enamel and teleost enameloid, as well as tetrapod dentin and teleost dentin, are all different. Thus, the evolution of vertebrate mineralized tissues seems to be explained by phenogenetic drift: while mineralized tissues are retained during vertebrate evolution, the underlying genetic basis has extensively drifted.

Animals↗
PubMed1998

Microbial mineralization of VC and DCE under different terminal electron accepting conditions.

Production of 14CO2 from [1,2-14C] dichloroethene (DCE) or [1,2-14C] vinyl chloride (VC) was quantified in aquifer and stream-bed sediment microcosms to evaluate the potential for microbial mineralization as a pathway for DCE and VC biodegradation under aerobic, Fe(III)-reducing, SO4-reducing, and methanogenic conditions. Mineralization of [1,2-14C] DCE and [1,2-14C] VC to 14CO2 decreased under increasingly reducing conditions, but significant mineralization was observed for both sediments even under anaerobic conditions. VC mineralization decreased in the order of aerobic > Fe(III)-reducing > SO4-reducing > methanogenic conditions. For both sediments, VC mineralization was greater than DCE mineralization under all electron-accepting conditions examined. For both sediments, DCE mineralization was at least two times greater under aerobic conditions than under anaerobic conditions. Although significant microbial mineralization of DCE was observed under anaerobic conditions, recovery of 14CO2 did not differ substantially between anaerobic treatments.

Journal Article↗
PubMed1992

Kinetics of beta-glycerophosphate-induced endochondral mineralization in vitro. Calcium accumulation, alkaline phosphatase activity, and effects of levamisole.

Isolated mesenchymal limb bud cells from day-12 mouse embryos grown at high density in organoid culture at the medium/air interphase differentiate into chondrocytes and form cartilage nodules. Upon addition of beta-glycerophosphate (beta-GP), cartilage undergoes endochondral mineralization. This beta-GP-induced mineralization was investigated by measuring the calcium content in the cultures and the activity of alkaline phosphatase (AP) in the cell mass and the medium. Calcium incorporation depended on the amount of beta-GP added. After continuous treatment, mineralization began on day 8 of the culture period and increased linearly until day 15. In long-term cultures, periodical treatment for 6 days caused an increase in mineralization the older the cultures were, but the slope of increase was proportionately less steep. Treatment at the latest period on days 19-24 resulted in a markedly reduced mineralization. After short-term treatment (48 hours), mineralization increased also the older the cultures were and proceeded during further cultivation in beta-GP-free medium. This kinetic behavior indicates a dependency of mineralization on cartilage maturation in this in vitro system. AP activity increased enormously and nearly logarithmically in the cell mass in beta-GP-free medium, whereas beta-GP treatment inhibited this drastic increase. In the medium, considerable activities of AP were also measurable from day 10 onward. It increased in beta-GP-free medium up to day 14, but was diminished after mineralization had been induced. Levamisole inhibited AP activity dose dependently when added directly to the enzyme-containing medium (100% inhibition at 10(-3) M).(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed1995

Mineral concentration gradients in rat femoral diaphyses measured by X-ray microtomography.

The bone mineral concentrations of five rat femora were measured as a function of distance from the distal metaphysis by quantitative X-ray microtomography (XMT) at a resolution of approximately 23 x 23 x 15 microns3. Assuming the mineral phase of bone to be hydroxyapatite, Ca10 (PO4)6(OH)2, the mean cortical mineral concentration (CM) per transverse section was found to range from 1.33 to 1.47 g cm-3. Detectable variations in the bone mineral concentration between sections of femora from different animals could not be attributed to the age when the particular animal was sacrificed. An increase in CM with distance, L, from the distal growth plate was observed and a saturating exponential equation, CM = a - be-alpha L, was used to describe the changes. Each section of bone tissue was considered as a population of elementary volumes of bone (EVB) and L was related to the age of the EVB (TEVB). A simple model for the mineralization process of an EVB was then proposed. Each newly formed EVB accumulated mineral rapidly to give an initial mineral concentration of approximately 1.3 g cm-3 (parameter a-b). Their mineral concentrations then increased asymptotically to approximately 1.5 g cm-3 (parameter a) with a time constant of approximately 330 days. This slow maturation process is attributed to Ostwald ripening of the bone crystals with further crystal growth using ions from the extracellular fluid.

Analysis of Variance↗
PubMed1996

Vitamin K2 promotes 1alpha,25(OH)2 vitamin D3-induced mineralization in human periosteal osteoblasts.

The effect of vitamin K on mineralization by human periosteal osteoblasts was investigated in the absence and presence of 1alpha, 25 dihydroxyvitamin D3 (1,25(OH)2D3). Vitamin K1 and K2, but not vitamin K3, at 2.5 microM enhanced in vitro mineralization when cells were cultured with vitamin K for 20 days after reaching confluence in vitro. Vitamin K2 (2-methyl-3-all-trans-tetraphenyl-1, 4-naphthoquinone : menatetrenone) was the most potent of these vitamin K analogs; it slightly inhibited alkaline phosphatase (ALP) activity. Human osteoblasts were mineralized and showed the enhanced ALP activity on treatment with 10(-9) M of 1,25(OH)2D3 for 20 or 25 days after confluence. Vitamin K2 promoted the 1,25(OH)2D3-induced mineralization, but slightly inhibited the 1,25(OH)2D3-induced ALP activity. Moreover, vitamin K2 enhanced the 1,25(OH)2D3-induced osteocalcin accumulation in the cells and the extracellular matrix (cell layer), but inhibited the osteocalcin content in the medium produced by the 1,25(OH)2D3 treatment. However, vitamin K2 alone did not induce osteocalcin production in the human osteoblasts. On Northern blot analysis, osteocalcin mRNA expression on 1, 25(OH)2D3-treated cells was enhanced by vitamin K2 treatment, but vitamin K2 alone did not induce osteocalcin mRNA expression. Warfarin blocked both the 1,25(OH)2D3-induced osteocalcin production and the accumulation in the cell layer, and also blocked the 1, 25(OH)2D3 plus vitamin K2-induced osteocalcin production and the accumulation in the cell layer. The 1,25(OH)2D3-induced mineralization promoted by vitamin K2 was probably due to the enhanced accumulation of osteocalcin induced by vitamin K2 in the cell layer. However, we concluded that the mineralization induced by vitamin K2 alone was due to the accumulation of osteocalcin in bovine serum on the cell layer, since osteocalcin extracted from the cell layer was not identified by specific antiserum against human osteocalcin, which does not cross-react with bovine osteocalcin. These results suggest that the mechanism underlying the mineralization induced by vitamin K2 in the presence of 1,25(OH)2D3 was different from that of vitamin K2 alone, and that osteocalcin plays an important role in mineralization by osteoblasts in vitro.

Adult↗
PubMed1985

Mineral and matrix alterations in the bones of incisors-absent (ia/ia) osteopetrotic rats.

The bones of incisors-absent (ia/ia) osteopetrotic rats differ from those of their normal littermates (ia/+) in histologic and radiographic appearance and in mechanical properties. This study examined how the mineral and matrices of osteopetrotic metaphyses and calvaria differed from normal controls. Bones of 11-day-old and 52-day-old osteopetrotic animals had higher ash (mineral) contents that age-matched controls; osteopetrotic metaphyses had elevated hexosamine contents, indicative of the persistence of cartilage. Calcium acidic phospholipid phosphate complexes, involved in initiation of hydroxyapatite formation in vivo and in vitro, were significantly reduced in content in all osteopetrotic bones. These results suggest that in the osteopetrotic rat, where osteoclast activity is defective, new mineral formation is reduced and replaced by accretion of mineral on existing crystals. Fractionation of osteopetrotic bone particles by density centrifugation demonstrated that the osteopetrotic bone was much more mineralized than that of age-matched controls. X-ray diffraction analysis of the mineralized fractions indicated a lack of growth of mineral crystals during maturation of the osteopetrotic animals. The absence of remodeling was apparent from the elevated Ca:P ratios of the highly mineralized osteopetrotic bone fractions, and from the high hexosamine content of these dense fractions. These observations may explain some of the unusual mechanical properties of osteopetrotic bone.

Animals↗