Increased intracortical bone remodeling during lactation in beagle dogs

There are substantial changes in skeletal and mineral metabolism during pregnancy and lactation. The purpose of this study was to determine the changes in intracortical bone remodeling and turnover during lactation in beagle dogs. A femur and rib were obtained from dogs near the end of lactation or soon after weaning and compared with nonlactating controls. Rib cortical bone had much higher bone turnover rates than did femoral diaphyseal cortical bone. The number of single-labeled osteons and the number of resorption spaces were significantly greater during lactation in both the rib and the femur. Additionally, the mineral apposition rate, basic multicellular unit activation frequency, and bone turnover rates were greater in the femoral cortical bone from the lactating dogs than from the controls. These data demonstrate that during lactation, intracortical bone remodeling increases, and this may provide a mechanism for the skeleton to be responsive to the calcium requirements of the mother. In addition, these data may help explain the transient decreases in cortical bone mineral density that are reported to occur during human lactation.     

Mineralization in rat metaphyseal bone exhibits a circadian stage dependency

Density gradient fractionation analysis of rat metaphyseal bone was used to delineate the biorhythmic changes in bone matrix mineralization. Seventy-two 4-week-old rats were entrained to 12-hr light, 12-hr dark cycles (light, 0800-2000 hr; darkness, 2000-0800 hr) for 4 weeks. All animals were fed ad lib. on Purina laboratory rat chow and tap water. Groups of 10-12 rats were killed by cervical dislocation at 4-hr intervals during a 24-hr period, and the tibias were then biopsied and frozen in liquid N2. Metaphyseal bone was fractionated via bromoformtoluene density gradients into specific gravity fractions ranging from 1.7 to 2.8. Density gradient fractions were analyzed for concentrations of calcium and inorganic phosphorus. Chronograms indicated that the accumulation of both calcium and inorganic phosphorus into the newly forming/least-dense mineral moieties of bone (1.3-1.7 sp grav) showed a single peak in the biorhythm of the rat. A statistically significant circadian rhythm of mineralization was detected for calcium (P less than 0.001) and inorganic phosphorus (P less than 0.039), with peaks during the environmental dark span. These results suggest that the physiological phasing of bone mineralization in the light-dark synchronized rat, is similar to that previously noted for cartilage mineralization and is antiphasal to the midday peak in bone collagen synthesis.     

Bone material properties and mineral matrix contributions to fracture risk or age in women and men

The strength of bone is related to its mass and geometry, but also to the physical properties of the tissue itself. Bone tissue is composed primarily of collagen and mineral, each of which changes with age, and each of which can be affected by pharmaceutical treatments designed to prevent or reverse the loss of bone. With age, there is a decrease in collagen content, which is associated with an increased mean tissue mineralization, but there is no difference in cross-link levels compared to younger adult bone. In osteoporosis, however, there is a decrease in the reducible collagen cross-links without an alteration in collagen concentration; this would tend to increase bone fragility. In older people, the mean tissue age (MTA) increases, causing the tissue to become more highly mineralized. The increased bone turnover following menopause may reduce global MTA, and would reduce overall tissue mineralization. Bone strength and toughness are positively correlated to bone mineral content, but when bone tissue becomes too highly mineralized, it tends to become brittle. This reduces its toughness, and makes it more prone to fracture from repeated loads and accumulated microcracking. Most approved pharmaceutical treatments for osteoporosis suppress bone turnover, increasing MTA and mineralization of the tissue. This might have either or both of two effects. It could increase bone volume from refilling of the remodeling space, reducing the risk for fracture. Alternatively, the increased MTA could increase the propensity to develop microcracks, and reduce the toughness of bone, making it more likely to fracture. There may also be changes in the morphology of the mineral crystals that could affect the homogeneity of the tissue and impact mechanical properties. These changes might have large positive or negative effects on fracture incidence, and could contribute to the paradox that both large and small increases in density have about the same effect on fracture risk. Bone mineral density measured by DXA does not discriminate between density differences caused by volume changes, and those caused by changes in mineralization. As such, it does not entirely reflect material property changes in aging or osteoporotic bone that contribute to bone's risk for fracture.     

Demonstration of the capacity of nacre to induce bone formation by human osteoblasts maintained in vitro.

Nacre implanted in vivo in bone is osteogenic suggesting that it may possess factor(s) which stimulate bone formation. The present study was undertaken to test the hypothesis that nacre can induce mineralization by human osteoblasts in vitro. Nacre chips were placed on a layer of first passage human osteoblasts. None of the chemical inducers generally required to obtain bone formation in vitro was added to the cultures. Osteoblasts proliferated and were clearly attracted by nacre chips to which they attached. Induction of mineralization appeared preferentially in bundles of osteoblasts surrounding the nacre chips. Three-dimensional nodules were formed by a dense osteoid matrix with cuboidal osteoblasts at the periphery and osteocytic-like cells in the center. These nodules contained foci with features of mineralized structures and bone-like structures, both radiodense to X-ray. Active osteoblasts (e.m.) with abundant rough endoplasmic reticulum, extrusion of collagen fibrils and budding of vesicles were observed. Matrix vesicles induced mineral deposition. Extracellular collagen fibrils appeared cross-banded and electrodense indicating mineralization. These results demonstrate that a complete sequence of bone formation is reproduced when human osteoblasts are cultured in the presence of nacre. This model provides a new approach to study the steps of osteoblastic differentiation and the mechanisms of induction of mineralization.     

Curcumin analogue UBS109 prevents bone loss in breast cancer bone metastasis mouse model: involvement in osteoblastogenesis and osteoclastogenesis

Bone metastasis of breast cancer typically leads to osteolysis, which causes severe pathological bone fractures and hypercalcemia. Bone homeostasis is skillfully regulated through osteoblasts and osteoclasts. Bone loss with bone metastasis of breast cancer may be due to both activation of osteoclastic bone resorption and suppression of osteoblastic bone formation. This study was undertaken to determine whether the novel curcumin analogue UBS109 has preventive effects on bone loss induced by breast cancer cell bone metastasis. Nude mice were inoculated with breast cancer MDA-MB-231 bone metastatic cells (10⁶ cells/mouse) into the head of the right and left tibia. One week after inoculation, the mice were treated with control (vehicle), oral administration (p.o.) of UBS109 (50 or 150 mg/kg body weight), or intraperitoneal administration (i.p.) of UBS109 (10 or 20 mg/kg body weight) once daily for 5 days per week for 7 weeks. After UBS109 administration for 7 weeks, hind limbs were assessed using an X-ray diagnosis system and hematoxylin and eosion staining to determine osteolytic destruction. Bone marrow cells obtained from the femurs and tibias were cultured to estimate osteoblastic mineralization and osteoclastogenesis ex vivo and in vitro. Remarkable bone loss was demonstrated in the tibias of mice inoculated with breast cancer MDA-MB-231 bone metastatic cells. This bone loss was prevented by p.o. administration of UBS109 (50 and 150 mg/kg body weight) and i.p. treatment of UBS109 (10 and 20 mg/kg) in vivo. Culture of bone marrow cells obtained from the bone tissues of mice with breast cancer cell bone metastasis showed suppressed osteoblastic mineralization and stimulated osteoclastogenesis ex vivo. These changes were not seen after culture of the bone marrow cells obtained from mice treated with UBS109. Moreover, UBS109 was found to stimulate osteoblastic mineralization and suppress lipopolysaccharide (LPS)-induced osteoclastogenesis in bone marrow cells obtained from normal nude mice in vitro. These findings suggest that the novel curcumin analogue UBS109 prevents breast cancer cell bone metastasis-induced bone loss by stimulating osteoblastic mineralization and suppressing osteoclastogenesis.     

Correlation between loss of alkaline phosphatase activity and accumulation of calcium during matrix vesicle-mediated mineralization

Activity of the bone/liver/kidney isozyme of alkaline phosphatase (AP) is known to be critical for mineralization in developing bone, although its role is unclear. The work now reported explores changes in the activity of this Zn2+-containing enzyme that occur during Ca2+ accumulation by matrix vesicles (MV). A marked loss (up to 65-70%) in AP activity was found to accompany Ca2+ accumulation by MV. These two events were highly correlated, both temporally and quantitatively. Investigation into possible causes revealed that the decline in AP activity during Ca2+ uptake was not due to action of proteases but rather resulted from interaction with the developing mineral phase, loss of metal ions (Zn2+ and Mg2+) from the active site of the enzyme, and concomitant irreversible denaturation of the enzyme. Protease inhibitors did not protect AP from loss of activity during mineralization; in contrast, protease treatments, which progressively destroyed the ability of MV to accumulate Ca2+ actually reduced loss of AP activity. These findings clearly demonstrate that AP is present at the site of MV mineralization and that its catalytic activity is profoundly reduced by the mineralization process.     

Biomechanical consequences of developmental changes in trabecular architecture and mineralization of the pig mandibular condyle

The purpose of the present study was to examine the changes in apparent mechanical properties of trabecular bone in the mandibular condyle during fetal development and to investigate the contributions of altering architecture, and degree and distribution of mineralization to this change. Three-dimensional, high-resolution micro-computed tomography (microCT) reconstructions were utilized to assess the altering architecture and mineralization during development. From the reconstructions, inhomogeneous finite element models were constructed, in which the tissue moduli were scaled to the local degree of mineralization of bone (DMB). In addition, homogeneous models were devised to study the separate influence of architectural and DMB changes on apparent mechanical properties. It was found that the bone structure became stiffer with age. Both the mechanical and structural anisotropies pointed to a rod-like structure that was predominantly oriented from anteroinferior to posterosuperior. Resistance against shear, also increasing with age, was highest in the sagittal plane. The reorganization of trabecular elements, which occurred without a change in bone volume fraction, contributed to the increase in apparent stiffness. The increase in DMB, however, contributed more dominantly. Incorporating the observed inhomogeneous distribution of mineralization decreased the apparent stiffness, but increased the mechanical anisotropy. This denotes that there might be a directional dependency of the DMB of trabecular elements, i.e. differently orientated trabecular elements might have different DMBs. In conclusion, the changes in DMB and its distribution are important to consider when studying mechanical properties during development and should be considered in other situations where differences in DMB are expected.     

Effects of four-week feed restriction on toxicological parameters in beagle dogs

This study was conducted to examine any changes caused by feed restriction in dogs to contribute to safety evaluation in toxicity studies. Two male 7-month-old beagle dogs/group were fed 300 (control), 150 (50% of control), or 70 g/animal of diet daily (23% of control) for 4 weeks. Effects of feed restriction, except for clinical signs, were noted depending on the feed dosage in almost all examinations. The principal outcomes were: decreased body weight and water consumption, ECG changes (decreased heart rate and prolonged QTc), and hematopoietic and lymphopoietic suppression (decreased reticulocyte ratio or white blood cell count in hematology, decreased nucleated cell count in bone marrow, decreased erythroid parameters in myelography, and hypocellularity of bone marrow and thymic atrophy in histopathology). In addition, some changes were noted in urinalysis (decreased urine volume and sodium and potassium excretion), blood chemistry (decreased ALP and inorganic phosphorus and increased creatinine), organ weights, and gastric histopathology. These results provide important reference data for distinguishing the primary effects of test compounds from secondary effects of decreased food consumption in toxicity studies in beagle dogs.     

An in vitro assay of bone development using fetal long bones of mice: morphological studies

The purpose of this study was to examine the morphological changes in an in vitro system in which the two elements of bone modelling, formation and resorption, could be studied simultaneously. Pregnant mice were killed on days 15, 16 and 17 of gestation, the fetuses were removed and the radii and ulnae dissected free of soft tissue. The bones were cultured for 6 days in media (BGJ) supplemented with 20% fetal calf serum and 150 micrograms/ml vitamin C. Growth and mineralization were estimated by measuring the total length of the bone, and diaphysis, and by light and transmission electron microscopy (TEM). The results of this study indicate that there is a continuous measurable increase in the total length of fetal mouse long bones over the 6 days of culture. These bones show a continuous growth of periosteal bone, with mesenchymal tissue penetrating into the diaphyseal shaft, and development of bone marrow like tissue. TEM examination showed differentiation of mesenchymal cells to osteoblasts, formation of new bone matrix and bone mineralization similar to that found in developmentally matched controls. In the cartilagenous epiphyses, however, many hydroxyapatite crystals were not associated with matrix vesicles. In addition, some of the chondrocytes of the hypertrophic zone appeared to be dedifferentiating into mesenchymal cells with osteoblast-like features. In spite of the lack of osteoclasts in the 15- and 16-day explants, osteoclasts appeared in the diaphysis after 2 and 4 days in culture. Our results suggest that this system can serve as a good model for the study of bone formation and resorption as they occur, simultaneously, during bone modelling.(ABSTRACT TRUNCATED AT 250 WORDS)     

Transmission FT-IR microspectroscopy of mineral phases in calcified tissues

Fourier-transform infrared microspectroscopy (FT-IRM) was used to study bone mineralization processes in an in vivo model and in enamel in osteogenesis imperfecta. Finally, the ability of FT-IRM to map new bone formed in implanted macroporous calcium phosphate biomaterial from sections was reported for the first time. FTIRM allowed the correlation of the microstructure of bone formation in the in vivo model with modifications in carbonate and phosphate environments of the mineral phases during maturation. FT-IRM analysis on enamel sections revealed changes in the mineral environment of carbonate and phosphate ions and probably in the size of enamel crystals. These modifications contributed to the fragility of enamel in osteogenesis imperfecta. The infrared functional group imaging of a part of implanted biomaterial and the bone ingrowth provided the visualization of chemical modifications occurring in biomaterial implants at 20 μm spatial resolution. The use of FT-IRM, in conjunction with appropriate sampling methods and data analysis should provide further insight into the molecular structure of mineral phases of calcified tissues and help to elucidate mineralization processes, skeletal disorders and properties of the biomaterials used as bone substitute.     

Connective tissue mineralization in Abcc6-/- mice, a model for pseudoxanthoma elasticum

Pseudoxanthoma elasticum (PXE) is a heritable multisystem disorder characterized by ectopic mineralization. However, the structure of the mineral deposits, their interactions with the connective tissue matrix, and the details of the progressive maturation of the mineral crystals are currently unknown. In this study, we examined the mineralization processes in Abcc6(-/-) mice, a model system for PXE, by energy dispersive X-ray and Fourier transform infrared imaging spectroscopy (FT-IRIS). The results indicated that the principal components of the mineral deposits were calcium and phosphate which co-localized within the histologically demonstrable lesions determined by topographic mapping. The Ca/P ratio increased in samples with progressive mineralization reaching the value comparable to that in endochondral bone. A progressive increase in mineralization was also reflected by increased mineral-to-matrix ratio determined by FT-IRIS. Determination of the mineral phases by FT-IRIS suggested progressive maturation of the mineral deposits from amorphous calcium phosphate to hydroxyapatite. These results provide critical information of the mechanisms of mineralization in PXE, with potential pharmacologic implications.     

Osteomimicry of Mammary Adenocarcinoma Cells In Vitro; Increased Expression of Bone Matrix Proteins and Proliferation within a 3D Collagen Environment

Bone is the most common site of metastasis for breast cancer, however the reasons for this remain unclear. We hypothesise that under certain conditions mammary cells possess osteomimetic capabilities that may allow them to adapt to, and flourish within, the bone microenvironment. Mammary cells are known to calcify within breast tissue and we have recently reported a novel in vitro model of mammary mineralization using murine mammary adenocarcinoma 4T1 cells. In this study, the osteomimetic properties of the mammary adenocarcinoma cell line and the conditions required to induce mineralization were characterized extensively. It was found that exogenous organic phosphate and inorganic phosphate induce mineralization in a dose dependent manner in 4T1 cells. Ascorbic acid and dexamethasone alone have no effect. 4T1 cells also show enhanced mineralization in response to bone morphogenetic protein 2 in the presence of phosphate supplemented media. The expression of several bone matrix proteins were monitored throughout the process of mineralization and increased expression of collagen type 1 and bone sialoprotein were detected, as determined by real-time RT-PCR. In addition, we have shown for the first time that 3D collagen glycosaminoglycan scaffolds, bioengineered to represent the bone microenvironment, are capable of supporting the growth and mineralization of 4T1 adenocarcinoma cells. These 3D scaffolds represent a novel model system for the study of mammary mineralization and bone metastasis. This work demonstrates that mammary cells are capable of osteomimicry, which may ultimately contribute to their ability to preferentially metastasize to, survive within and colonize the bone microenvironment.     

Effects of Decreased Occlusal Loading during Growth on the Mandibular Bone Characteristics

Background

Bone mass and mineralization are largely influenced by loading. The purpose of this study was to evaluate the reaction of the entire mandibular bone in response to decreased load during growth. It is hypothesized that decreased muscular loading will lead to bone changes as seen during disuse, i.e. loss of bone mass.

Methods and Findings

Ten 21-day-old Wistar strain male rats were divided into two groups (each n=5) and fed on either a hard- or soft-diet for 11 weeks. Micro-computed tomography was used for the investigation of bone mineralization, bone volume, bone volume fraction (BV/TV) and morphological analysis. Mandibular mineralization patterns were very consistent, showing a lower degree of mineralization in the ramus than in the corpus. In the soft-diet group, mineralization below the molars was significantly increased (p<0.05) compared to the hard diet group. Also, bone volume and BV/TV of the condyle and the masseter attachment were decreased in the soft-diet group (p<0.05). Morphological analysis showed inhibited growth of the ramus in the soft-diet group (p<0.05).

Conclusion

Decreased loading by a soft diet causes significant changes in the mandible. However, these changes are very region-specific, probably depending on the alterations in the local loading regime. The results suggest that muscle activity during growth is very important for bone quality and morphology.     

Changes in Structural-Mechanical Properties and Degradability of Collagen during Aging-associated Modifications

During aging, changes occur in the collagen network that contribute to various pathological phenotypes in the skeletal, vascular, and pulmonary systems. The aim of this study was to investigate the consequences of age-related modifications on the mechanical stability and in vitro proteolytic degradation of type I collagen. Analyzing mouse tail and bovine bone collagen, we found that collagen at both fibril and fiber levels varies in rigidity and Young''s modulus due to different physiological changes, which correlate with changes in cathepsin K (CatK)-mediated degradation. A decreased susceptibility to CatK-mediated hydrolysis of fibrillar collagen was observed following mineralization and advanced glycation end product-associated modification. However, aging of bone increased CatK-mediated osteoclastic resorption by ∼27%, and negligible resorption was observed when osteoclasts were cultured on mineral-deficient bone. We observed significant differences in the excavations generated by osteoclasts and C-terminal telopeptide release during bone resorption under distinct conditions. Our data indicate that modification of collagen compromises its biomechanical integrity and affects CatK-mediated degradation both in bone and tissue, thus contributing to our understanding of extracellular matrix aging.     

Occurrence of osteoblast necroses during ossification of long bone cortices in mouse fetuses

Previous investigations concerned with in vitro osteogenesis and mineralization have revealed some indication of a participation of cell necroses in the course of calcification. These observations were confirmed by in vivo investigations on desmoid ossification in fetal mouse calvariae, where abundant necrotic osteoblasts were found at the mineralization border and in the osteoid. In the present study, ossification of long bone cortices from fetal mice was investigated by use of electron microscopy. Specimens obtained from the collection of the Institute of Anatomy, Free University of Berlin (mouse fetuses, forearm; rat fetuses, forearm) were reinvestigated for control purposes. In all cases, mineralization of osteoid was accompanied by cell necroses. Cell degeneration was characterized by swelling of the endoplasmic reticulum and loss of the plasma membrane resulting in freely distributed vesicular structures. Cell debris was incorporated within the mineral. Initially, cell necroses in the perichondrium occurred in the region surrounding the hypertrophic cartilage and the matrix of which showed spots of endochondral mineralization. Necrotic osteoblasts occurred simultaneously with mineralization of the osteoid. During further ossification of the long bone cortices, the number of necrotic cells increased markedly. In addition to necrotic cells, healthy osteoblasts, osteocytes and perichondral tissue were present, indicating that an artifact can be excluded. The importance of cell necroses in the process of mineralization is as yet unclear. Possibly, the cells act as calcium and/or phosphate stores, which are liberated by cell death to increase the amount of mineral constituents at sites of mineralization.     

Near-infrared-labeled tetracycline derivative is an effective marker of bone deposition in mice

Bone-specific compounds have been used effectively for the detection of bone mineralization, growth, and morphological changes. These agents typically contain iminodiacetic acid groups that can form complexes with apatite and fluoresce in the visible spectrum. We exploited a subset of these chemical chelators to produce a near-infrared (NIR) optical bone marker for preclinical animal imaging. By conjugating target compounds to IRDye 800CW, we extended the effective fluorescence signal detection to the NIR region without affecting the compound’s ability to function as a marker of the mineralization process. Calcein and a tetracycline derivative (BoneTag agent [BT]) bound specifically to differentiated mineralized osteoblast cultures, with the latter exhibiting 6-fold higher signal intensities. Subsequent in vivo testing demonstrated effective skeletal labeling with IRDye 800CW BT. We were able to identify a changing mineralization front in bone sections from (i) normal growing mice injected with IRDye 800CW BT 6 weeks prior to the administration of IRDye 680 BT and (ii) an osteoporosis mouse model comparing cortical bone in sham-treated and ovariectomized mice. These results provide evidence that the NIR-labeled BT is effective as a general marker of skeletal features and an indicator of the bone mineralization and remodeling processes.     

Age-related changes in microarchitecture and mineralization of cancellous bone in the porcine mandibular condyle

The mandibular condyle is considered a good model for developing cancellous bone because of its rapid growth and high rate of remodeling. The aim of the present study was to analyze the simultaneous changes in microarchitecture and mineralization of cancellous bone during development in a three-dimensional fashion. Eight mandibular condyles of pigs aged 8 weeks prepartum to 108 weeks postpartum were scanned using microCT with an isotropic spatial resolution of 10 microm. The number of trabeculae decreased during development, whereas both the trabecular thickness and the distance between the trabeculae increased. The bone surface to volume ratio decreased during development, possibly limiting the amount of (re)modeling. Both the mean degree of mineralization and intratrabecular differences in mineralization between the surfaces and cores of trabecular elements increased during development. The trabecular surfaces were more highly mineralized in the older condyles compared to the younger ones. Together with the observed decrease in the relative size of trabecular surface, this finding suggests a decrease in (re)modeling activity during development. In accordance with the general growth and development of the pig, it was concluded that most developmental changes in cancellous bone occur until the age of 40 weeks postpartum.     

低强度脉冲超声波辐照与泡沫TiC/Ti对犬节段性骨缺修复的促进作用

目的探讨低强度脉冲超声波辐照对节段性骨缺损修复效果的影响。方法将直径12 mm长20mm泡沫TiC/Ti植入6只Beagle犬的左侧胫骨节段性骨缺损区。随机分为超声组和对照组,超声组采用低强度脉冲超声波辐照（频率1.5 MHz、强度30 mW/cm2、脉冲宽度200μs、脉冲周期1 kHz、20 min/次、1次/d）,对照组为不开功率源的假辐照,术后4、8周后分别行X线检查及骨密度测定,观察及分析材料周围骨愈合情况。结果 6只beagle犬均进入结果分析。术后4周超声组骨早期成熟度优于对照组,表现在材料周围骨痂影密度增高,骨痂影由两端向中央生长;对照组仅见骨痂区密度低,还可见部分骨痂缺如。术后8周超声组新生骨痂面积优于对照组,骨干结构相对稳定;对照组骨缺损区未闭合,在骨干两侧看到少量骨痂,愈合较差。骨密度测定结果显示,4周时超声组高于对照组,两组间存在统计学差异;8周时超声组略高于对照组,但两组间没有统计学差异。结论通过联合应用低强度脉冲超声波辐照与人工骨材料修复可提高新骨形成速度及骨组织密度,缩短节段性骨缺损的骨愈合时间。     

Vitamin K deficiency inhibits mineralization and enhances deformity in vertebrae of haddock (Melanogrammus aeglefinus L.)

Vitamin K has been known to regulate bone formation through osteocalcin synthesis by osteoblasts, which is important for mineralization and bone structure. The mechanism underlying the relationship of vitamin K with the changes of microanatomy is not fully understood, and our goal is to test whether bone deformities develop in association with vitamin K deficiency. Fish were fed a semi-purified diet containing either devoid (0.00 mg/kg diet) or adequate (40.0 mg/kg diet supplemented but 20.8 mg/kg analyzed) levels of vitamin K (menadione sodium bisulphite) for 20 weeks. At the end of 8 and 20 weeks, fish were subjected to gross examination and X-ray, and mineral content of the vertebrae was measured. The vertebrae were also subjected to histological, histomorphometric and enzyme histochemical examinations to determine the bone formation and resorption. Vitamin K deficiency primarily decreased bone mineralization and subsequently a decrease in bone mass thus resulted in an increased susceptibility to bone deformity. The occurrence of bone deformities coincided with an increased amount of osteoid tissue and decreased bone mineral content. Number of osteoblasts and osteoclasts were not affected by dietary vitamin K. In conclusion, vitamin K deficiency can impair bone mineralization and enhances bone deformities.