MicroRNA‐31a‐5p from aging BMSCs links bone formation and resorption in the aged bone marrow microenvironment

The alteration of age‐related molecules in the bone marrow microenvironment is one of the driving forces in osteoporosis. These molecules inhibit bone formation and promote bone resorption by regulating osteoblastic and osteoclastic activity, contributing to age‐related bone loss. Here, we observed that the level of microRNA‐31a‐5p (miR‐31a‐5p) was significantly increased in bone marrow stromal cells (BMSCs) from aged rats, and these BMSCs demonstrated increased adipogenesis and aging phenotypes as well as decreased osteogenesis and stemness. We used the gain‐of‐function and knockdown approach to delineate the roles of miR‐31a‐5p in osteogenic differentiation by assessing the decrease of special AT‐rich sequence‐binding protein 2 (SATB2) levels and the aging of BMSCs by regulating the decline of E2F2 and recruiting senescence‐associated heterochromatin foci (SAHF). Notably, expression of miR‐31a‐5p, which promotes osteoclastogenesis and bone resorption, was markedly higher in BMSCs‐derived exosomes from aged rats compared to those from young rats, and suppression of exosomal miR‐31a‐5p inhibited the differentiation and function of osteoclasts, as shown by elevated RhoA activity. Moreover, using antagomiR‐31a‐5p, we observed that, in the bone marrow microenvironment, inhibition of miR‐31a‐5p prevented bone loss and decreased the osteoclastic activity of aged rats. Collectively, our results reveal that miR‐31a‐5p acts as a key modulator in the age‐related bone marrow microenvironment by influencing osteoblastic and osteoclastic differentiation and that it may be a potential therapeutic target for age‐related osteoporosis.     

Osteoclast‐specific Dicer gene deficiency suppresses osteoclastic bone resorption

Osteoclasts are unique cells that resorb bone, and are involved in not only bone remodeling but also pathological bone loss such as osteoporosis and rheumatoid arthritis. The regulation of osteoclasts is based on a number of molecules but full details of these molecules have not yet been understood. MicroRNAs are produced by Dicer cleavage an emerging regulatory system for cell and tissue function. Here, we examine the effects of Dicer deficiency in osteoclasts on osteoclastic activity and bone mass in vivo. We specifically knocked out Dicer in osteoclasts by crossing Dicer flox mice with cathepsin K‐Cre knock‐in mice. Dicer deficiency in osteoclasts decreased the number of osteoclasts (N.Oc/BS) and osteoclast surface (Oc.S/BS) in vivo. Intrinsically, Dicer deficiency in osteoclasts suppressed the levels of TRAP positive multinucleated cell development in culture and also reduced NFATc1 and TRAP gene expression. MicroRNA analysis indicated that expression of miR‐155 was suppressed by RANKL treatment in Dicer deficient cells. Dicer deficiency in osteoclasts suppressed osteoblastic activity in vivo including mineral apposition rate (MAR) and bone formation rate (BFR) and also suppressed expression of genes encoding type I collagen, osteocalcin, Runx2, and Efnb2 in vivo. Dicer deficiency in osteoclasts increased the levels of bone mass indicating that the Dicer deficiency‐induced osteoclastic suppression was dominant over Dicer deficiency‐induced osteoblastic suppression. On the other hand, conditional Dicer deletion in osteoblasts by using 2.3 kb type I collagen‐Cre did not affect bone mass. These results indicate that Dicer in osteoclasts controls activity of bone resorption in vivo. J. Cell. Biochem. 109: 866–875, 2010. © 2009 Wiley‐Liss, Inc.     

The roles of bone‐derived exosomes and exosomal microRNAs in regulating bone remodelling

Pathological destructive bone diseases are primarily caused by the failure of a lifelong self‐renewal process of the skeletal system called bone remodelling. The mechanisms underlying this process include enhanced osteoclast activity and decreased generation of the osteoblast lineage. Intercellular interaction and crosstalk among these cell types are crucial for the maintenance of bone remodelling, either through the secretion of growth factors or direct cell–cell physical engagement. Recent studies have revealed that exosomes derived from bone cells, including osteoclasts, osteoblasts and their precursors, play pivotal roles on bone remodelling by transferring biologically active molecules to target cells, especially in the processes of osteoclast and osteoblast differentiation. Here, we review the contents of bone‐derived exosomes and their functions in the regulatory processes of differentiation and communication of osteoclasts and osteoblasts. In addition, we highlight the characteristics of microRNAs of bone‐derived exosomes involved in the regulation of bone remodelling, as well as the potential clinical applications of bone‐derived exosomes in bone remodelling disorders.     

Up‐regulated CST5 inhibits bone resorption and activation of osteoclasts in rat models of osteoporosis via suppression of the NF‐κB pathway

Here, we aim at exploring the effect of CST5 on bone resorption and activation of osteoclasts in osteoporosis (OP) rats through the NF‐κB pathway. Microarray analysis was used to screen the OP‐related differentially expressed genes. Osteoporosis was induced in rats by intragastric retinoic acid administration. The serum levels of tartrate‐resistant acid phosphatase (TRAP), bone alkaline phosphatase (BALP) and osteocalcin (OC) and the expression of CD61 on the surface of osteoclasts were examined. The number of osteoclasts and the number and area of resorption pits were detected. Besides, the pathological changes and bone mineral density in bone tissues of rats were assessed. Also, the relationship between CST5 and the NF‐κB pathway was identified through determining the expression of CST5, RANKL, RANK, OPG, p65 and IKB. Poorly expressed CST5 was indicated to affect the OP. CST5 elevation and inhibition of the NF‐κB pathway decreased serum levels of TRAP, BALP and OC and expression of CD61 in vivo and in vitro. In OP rats, CST5 overexpression increased trabecular bones and bone mineral density of bone tissues, but decreased trabecular separation, fat within the bone marrow cavities and the number of osteoclasts through inhibiting the NF‐κB pathway. In vivo experiments showed that CST5 elevation inhibited growth in number and area of osteoclastic resorption pits and restrained osteoclastic bone absorption by inhibiting the NF‐κB pathway. In summary, overexpression of CST5 suppresses the activation and bone resorption of osteoclasts by inhibiting the activation of the NF‐κB pathway.     

Tectorigenin inhibits RANKL‐induced osteoclastogenesis via suppression of NF‐κB signalling and decreases bone loss in ovariectomized C57BL/6

Metabolism of bone is regulated by the balance between osteoblast‐mediated bone formation and osteoclast‐mediated bone resorption. Activation of osteoclasts could lead to osteoporosis. Thus, inhibiting the activity of osteoclasts becomes an available strategy for the treatment of osteoporosis. Tectorigenin is an extract of Belamcanda chinensis In the present study, the anti‐osteoclastogenesis effects of tectorigenin were investigated in vitro and in vivo. The results showed preventive and therapeutic effects of tectorigenin at concentrations of 0, 10, 40, and 80 μmol/L in the maturation and activation of osteoclasts. A signalling study also indicated that tectorigenin treatment reduces activation of NF‐κB signalling in osteoclastogenesis. Animal experiment demonstrated that tectorigenin treatment (1‐10 mg/kg, abdominal injection every 3 days) significantly inhibits bone loss in ovariectomized C57BL/6. Our data suggest that tectorigenin is a potential pharmacological choice for osteoporosis.     

Sphingosine‐1‐phosphate (S1P) receptors: Promising drug targets for treating bone‐related diseases

Sphingosine‐1‐phosphate (S1P) is a natural bioactive lipid molecule and a common first or second messenger in the cardiovascular and immune systems. By binding with its receptors, S1P can serve as mediator of signalling during cell migration, differentiation, proliferation and apoptosis. Although the predominant role of S1P in bone regeneration has been noted in many studies, this role is not as well‐known as its roles in the cardiovascular and immune systems. In this review, we summarize previous research on the role of S1P receptors (S1PRs) in osteoblasts and osteoclasts. In addition, S1P is regarded as a bridge between bone resorption and formation, which brings hope to patients with bone‐related diseases. Finally, we discuss S1P and its receptors as therapeutic targets for treating osteoporosis, inflammatory osteolysis and bone metastasis based on the biological effects of S1P in osteoclastic/osteoblastic cells, immune cells and tumour cells.     

Suppression of bone formation by osteoclastic expression of semaphorin 4D

Most of the currently available drugs for osteoporosis inhibit osteoclastic bone resorption; only a few drugs promote osteoblastic bone formation. It is thus becoming increasingly necessary to identify the factors that regulate bone formation. We found that osteoclasts express semaphorin 4D (Sema4D), previously shown to be an axon guidance molecule, which potently inhibits bone formation. The binding of Sema4D to its receptor Plexin-B1 on osteoblasts resulted in the activation of the small GTPase RhoA, which inhibits bone formation by suppressing insulin-like growth factor-1 (IGF-1) signaling and by modulating osteoblast motility. Sema4d-/- mice, Plxnb1-/- mice and mice expressing a dominant-negative RhoA specifically in osteoblasts showed an osteosclerotic phenotype due to augmented bone formation. Notably, Sema4D-specific antibody treatment markedly prevented bone loss in a model of postmenopausal osteoporosis. Thus, Sema4D has emerged as a new therapeutic target for the discovery and development of bone-increasing drugs.     

Mechanisms influencing bone metabolism in chronic illness

Bone is permanently renewed by the coordinated actions of bone-resorbing osteoclasts and bone-forming osteoblasts, which model and remodel bone structure during growth and adult life. The origin of osteoblastic cells (osteoblasts, osteocytes and bone-lining cells) differs from that of osteoclasts, but both cell groups communicate with each other using cytokines and cell-cell contact as to optimally maintain bone homeostasis. This communication in many ways uses the same players as the communication between cells in the immune system. During acute life-threatening illness massive bone resorption is the rule, while bone formation is suppressed. During chronic illness, the balance between bone formation and bone resorption also shifts, frequently resulting in decreased bone mass and density. Several factors may contribute to the osteopenia that accompanies chronic illness, the most important being undernutrition and low body weight, inflammatory cytokines, disorders of the neuroendocrine axis (growth hormone/IGF-1 disturbances, thyroid and gonadal deficiency), immobilization, and the long-term use of glucocorticoids. Their combined effects not only influence the generation and activity of all bone cells involved, but probably also regulate their life span by apoptotic mechanisms. Osteopenia or even osteoporosis and bone fragility, and before puberty also decreased linear growth and lower peak bone mass are therefore frequent consequences of chronic illnesses.     

Arbutin ameliorates glucocorticoid-induced osteoporosis through activating autophagy in osteoblasts

Chronic long-term glucocorticoid use causes osteoporosis partly by interrupting osteoblast homeostasis and exacerbating bone loss. Arbutin, a natural hydroquinone glycoside, has been reported to have biological activities related to the differentiation of osteoblasts and osteoclasts. However, the role and underlying mechanism of arbutin in glucocorticoid-induced osteoporosis are elusive. In this study, we demonstrated that arbutin administration ameliorated osteoporotic disorders in glucocorticoid dexamethasone (Dex)-induced mouse model, including attenuating the loss of bone mass and trabecular microstructure, promoting bone formation, suppressing bone resorption, and activating autophagy in bone tissues. Furthermore, Dex-stimulated mouse osteoblastic MC3T3-E1 cells were treated with arbutin. Arbutin treatment rescued Dex-induced repression of osteoblast differentiation and mineralization, the downregulation of osteogenic gene expression, reduced autophagic marker expression, and decreased autophagic puncta formation. The application of autophagy inhibitor 3-MA decreased autophagy, differentiation, and mineralization of MC3T3-E1 cells triggered by arbutin. Taken together, our findings suggest that arbutin treatment fends off glucocorticoid-induced osteoporosis, partly through promoting differentiation and mineralization of osteoblasts by autophagy activation.     

The HIV proteins Tat and Nef promote human bone marrow mesenchymal stem cell senescence and alter osteoblastic differentiation

To maintain bone mass turnover and bone mineral density (BMD), bone marrow (BM) mesenchymal stem cells (MSCs) are constantly recruited and subsequently differentiated into osteoblasts. HIV‐infected patients present lower BMD than non‐HIV infected individuals and a higher prevalence of osteopenia/osteoporosis. In antiretroviral treatment (ART)‐naive patients, encoded HIV proteins represent pathogenic candidates. They are released by infected cells within BM and can impact on neighbouring cells. In this study, we tested whether HIV proteins Tat and/or Nef could induce senescence of human BM‐MSCs and reduce their capacity to differentiate into osteoblasts. When compared to nontreated cells, MSCs chronically treated with Tat and/or Nef up to 30 days reduced their proliferative activity and underwent early senescence, associated with increased oxidative stress and mitochondrial dysfunction. The antioxidant molecule N‐acetyl‐ cysteine had no or minimal effects on Tat‐ or Nef‐induced senescence. Tat but not Nef induced an early increase in NF‐κB activity and cytokine/chemokine secretion. Tat‐induced effects were prevented by the NF‐κB inhibitor parthenolide, indicating that Tat triggered senescence via NF‐κB activation leading to oxidative stress. Otherwise, Nef‐ but not Tat‐treated cells displayed early inhibition of autophagy. Rapamycin, an autophagy inducer, reversed Nef‐induced senescence and oxidative stress. Moreover, Tat+Nef had cumulative effects. Finally, Tat and/or Nef decreased the MSC potential of osteoblastic differentiation. In conclusion, our in vitro data show that Tat and Nef could reduce the number of available precursors by inducing MSC senescence, through either enhanced inflammation or reduced autophagy. These results offer new insights into the pathophysiological mechanisms of decreased BMD in HIV‐infected patients.     

Prostate cancer bone metastases promote both osteolytic and osteoblastic activity

Advanced prostate cancer is frequently accompanied by the development of metastasis to bone. In the past, prostate cancer bone metastases were characterized as being osteoblastic (i.e., increasing bone density) based on radiographs. However, emerging evidence suggests that development of prostate cancer bone metastases requires osteoclastic activity in addition to osteoblastic activity. The complexities of how prostate tumor cells influence bone remodeling are just beginning to be elucidated. Prostate cancer cells produce a variety of pro-osteoblastic factors that promote bone mineralization. For example, both bone morphogenetic proteins and endothelin-1 have well recognized pro-osteoblastic activities and are produced by prostate cancer cells. In addition to factors that enhance bone mineralization prostate cancer cells produced factors that promote osteoclast activity. Perhaps the most critical pro-osteoclastogenic factor produced by prostate cancer cells is receptor activator of NFkappaB ligand (RANKL), which has been shown to be required for the development of osteoclasts. Blocking RANKL results in inhibiting prostate cancer-induced osteoclastogenesis and inhibits development and progression of prostate tumor growth in bone. These findings suggest that targeting osteoclast activity may be of therapeutic benefit. However, it remains to be defined how prostate cancer cells synchronize the combination of osteoclastic and osteoblastic activity. We propose that as the bone microenvironment is changed by the developing cancer, this in turn influences the prostate cancer cells' balance between pro-osteoclastic and pro-osteoblastic activity. Accordingly, the determination of how the prostate cancer cells and bone microenvironment crosstalk are important to elucidate how prostate cancer cells modulate bone remodeling.     

The skeleton in primary hyperparathyroidism: a review focusing on bone remodeling, structure, mass, and fracture.

The mechanisms behind the influence of PHPT on the skeleton are closely connected with bone turnover. Throughout life, the skeleton is continuously renewed by bone remodeling, a process which serves the purpose of repairing damaged bone and adapting the skeleton to changes in physical load. In this process, old bone is removed by osteoclastic resorption and new bone is laid down by osteoblastic formation. Bone mass increases with growth in the first decades of life, and around the age of 30 years the peak bone mass is reached. Thereafter, as a result of mechanisms involving bone remodeling, a net bone loss is seen: 1) A reversible bone loss because of increase in the remodeling space, i.e., the amount of bone resorped but not yet reformed during the remodeling cycle. This mechanism leads to decrease in average trabecular thickness and cortical width, and to increase in cortical porosity. 2) An irreversible bone loss caused by negative bone balance, where the amount of bone formed by the osteoblasts is exceeded by the amount of bone resorbed by the osteoclasts at the same remodeling site. Consequently, progressive thinning of trabecular elements, reduced cortical width and increased cortical porosity is seen. 3) Finally, perforation of trabecular plates by deep resorption lacunae leads to complete irreversible removal of structural bone components. Parathyroid hormone, together with vitamin D, are the principal modulators in calcium homeostasis. The main actions of PTH are executed in bone and kidneys. In the kidneys, PTH increases the tubular re-absorption of calcium, thereby tending to increase serum calcium. PTH also induces increased conversion of 25(OH)-D to 1,25(OH)2-D. This last action, enhances intestinal calcium absorption and increased skeletal calcium mobilization, which further adds to the circulating calcium pool. In bone, the "acute" regulatory actions of PTH on serum calcium are probably accompliced via activation of osteocytes and lining cells. A second mechanism of PTH in bone is the regulation of bone remodeling. The action seems to be an increased recruitment from osteoblastic precursor cells and activation of mature osteoclasts. It is supposed that these responses are predominantly mediated indirectly through actions on osteoblast-like or nonosteoblast-like stromal cells, as osteoclasts themselves to not have PTH receptors. Bone metabolism and bone mass are studied by biochemical bone markers, bone histomorphometry, and densitometry. As bone markers and bone histomorphometry give information on bone metabolism from different points of view, these methods are preferably combined. Histomorphometry gives detailed information about bone turnover on cellular level, the whole remodeling sequence is described, and the bone balance can be calculated. However, they focus on a small volume, and may, therefore, not be representative for the whole skeleton. On the other hand, studies of bone markers supply general information about turnover in the whole skeleton, but they do not give facts on the bone turnover on the cellular or tissue level and bone balance. Bone densitometry is the principal method in studying bone mass, but valuable information concerning bone structure also comes from histomorphometry. Bone remodeling is considerably increased in PHPT. Studies of bone markers show increase in both resorptive and formative markers, and the increases seem to be of equivalent size. This is in agreement with histomorphometric findings and shows that the coupling between resorption and formation is preserved. By histomorphometry on iliac crest biopsies, trabecular bone remodeling is found increased by 50%, judged by the increase in activation frequency; a measure of how often new remodeling is initiated on the trabecular bone surface. In PHPT, such remodeling activity is repeated about once every year. Reconstruction of the whole remodeling sequence does not show major deviations in lengths of the resorptive and formative periods compared to normal. Furthermore, the amount of bone removed by the osteoclasts during the resorptive phase is matched by the amount of new bone formed by the osteoblasts leading to a bone balance very close to zero. Compared with trabecular bone, the turnover rate in cortical bone is considerably lower, around 10%. Remodeling of the cortical bone takes place at the endocortical, the pericortical, and the Haversian surfaces. Endocortical bone remodeling activities are very similar to trabecular remodeling activities with good correlation between individual parameters. Periosteal remodeling activity is negligible in PHPT, as it is in the normal state. Cortical porosity, which reflects the remodeling activity on the Haversian surface, is increased by 30-65% in PHPT. (ABSTRACT TRUNCATED)     

Role of vitamin D in bone resorption.

The idea that vitamin D must function at the bone site to promote bone mineralization has long existed since its discovery as an anti-rachitic agent. However, the definite evidence for this is still lacking. In contrast, much evidence has accumulated that 1 alpha,25(OH)2D3 in involved in bone resorption. 1 alpha,25(OH)2D3 tightly regulates differentiation of osteoclast progenitors into osteoclasts. Osteoclast progenitors have been thought to belong to the monocyte-macrophage lineage. 1 alpha,25(OH)2D3 greatly stimulates differentiation and activation of mononuclear phagocytes. Recent reports have indicated that differentiation of mononuclear phagocytes into osteoclasts is strictly regulated by osteoblastic cells, the process of which is also stimulated by 1 alpha,25(OH)2D3. In the differentiation of mononuclear phagocytes into osteoclasts, the target cells for 1 alpha,25(OH)2D3 appear to be osteoblastic stromal cells. Osteoblastic cells produce several proteins such as BGP, MGP, osteopontin and the third component of complement (C3) in response to the vitamin. They appear to be somehow involved in osteoclast differentiation and functions. Thus, 1 alpha,25(OH)2D3 seems to be involved in the differentiation of osteoclast progenitors into osteoclasts directly and also by an indirect mechanism involving osteoblastic cells. The precise role of osteoblastic cells in osteoclast development has to be elucidated in the future.     

Targeting bone remodeling by isoflavone and 3,3'-diindolylmethane in the context of prostate cancer bone metastasis

Prostate cancer (PCa) bone metastases have long been believed to be osteoblastic because of bone remodeling leading to the formation of new bone. However, recent studies have shown increased osteolytic activity in the beginning stages of PCa bone metastases, suggesting that targeting both osteolytic and osteoblastic mediators would likely inhibit bone remodeling and PCa bone metastasis. In this study, we found that PCa cells could stimulate differentiation of osteoclasts and osteoblasts through the up-regulation of RANKL, RUNX2 and osteopontin, promoting bone remodeling. Interestingly, we found that formulated isoflavone and 3,3'-diindolylmethane (BR-DIM) were able to inhibit the differentiation of osteoclasts and osteoblasts through the inhibition of cell signal transduction in RANKL, osteoblastic, and PCa cell signaling. Moreover, we found that isoflavone and BR-DIM down-regulated the expression of miR-92a, which is known to be associated with RANKL signaling, EMT and cancer progression. By pathway and network analysis, we also observed the regulatory effects of isoflavone and BR-DIM on multiple signaling pathways such as AR/PSA, NKX3-1/Akt/p27, MITF, etc. Therefore, isoflavone and BR-DIM with their multi-targeted effects could be useful for the prevention of PCa progression, especially by attenuating bone metastasis mechanisms.     

Extracellular matrix networks in bone remodeling

Bones are constantly remodeled throughout life to maintain robust structure and function. Dysfunctional remodeling can result in pathological conditions such as osteoporosis (bone loss) or osteosclerosis (bone gain). Bone contains 100s of extracellular matrix (ECM) proteins and the ECM of the various bone tissue compartments plays essential roles directing the remodeling of bone through the coupled activity of osteoclasts (which resorb bone) and osteoblasts (which produce new bone). One important role for the ECM is to serve as a scaffold upon which mineral is deposited. This scaffold is primarily type I collagen, but other ECM components are involved in binding of mineral components. In addition to providing a mineral scaffolding role, the ECM components provide structural flexibility for a tissue that would otherwise be overly rigid. Although primarily secreted by osteoblast-lineage cells, the ECM regulates cells of both the osteoblast-lineage (such as progenitors, mature osteoblasts, and osteocytes) and osteoclast-lineage (including precursors and mature osteoclasts), and it also influences the cross-talk that occurs between these two oppositional cells. ECM influences the differentiation process of mesenchymal stem cells to become osteoblasts by both direct cell-ECM interactions as well as by modulating growth factor activity. Similarly, the ECM can influence the development of osteoclasts from undifferentiated macrophage precursor cells, and influence osteoclast function through direct osteoclast cell binding to matrix components. This comprehensive review will focus on how networks of ECM proteins function to regulate osteoclast- and osteoblast-mediated bone remodeling. The clinical significance of these networks on normal bone and as they relate to pathologies of bone mass and geometry will be considered. A better understanding of the dynamic role of ECM networks in regulating tissue function and cell behavior is essential for the development of new treatment approaches for bone loss.     

Matrix IGF-1 maintains bone mass by activation of mTOR in mesenchymal stem cells

Insulin-like growth factor 1 (IGF-1), the most abundant growth factor in the bone matrix, maintains bone mass in adulthood. We now report that IGF-1 released from the bone matrix during bone remodeling stimulates osteoblastic differentiation of recruited mesenchymal stem cells (MSCs) by activation of mammalian target of rapamycin (mTOR), thus maintaining proper bone microarchitecture and mass. Mice with knockout of the IGF-1 receptor (Igf1r) in their pre-osteoblastic cells showed lower bone mass and mineral deposition rates than wild-type mice. Further, MSCs from Igf1rflox/flox mice with Igf1r deleted by a Cre adenovirus in vitro, although recruited to the bone surface after implantation, were unable to differentiate into osteoblasts. We also found that the concentrations of IGF-1 in the bone matrix and marrow of aged rats were lower than in those of young rats and directly correlated with the age-related decrease in bone mass. Likewise, in age-related osteoporosis in humans, we found that bone marrow IGF-1 concentrations were 40% lower in individuals with osteoporosis than in individuals without osteoporosis. Notably, injection of IGF-1 plus IGF binding protein 3 (IGFBP3), but not injection of IGF-1 alone, increased the concentration of IGF-1 in the bone matrix and stimulated new bone formation in aged rats. Together, these results provide mechanistic insight into how IGF-1 maintains adult bone mass, while also providing a further rationale for its therapeutic targeting to treat age-related osteoporosis.     

Lactoferrin and bone; structure-activity relationships.

The maintenance of the mechanical integrity of the skeleton depends on bone remodeling, the well-coordinated balance between bone formation by osteoblasts and bone resorption by osteoclasts. The coupled action of osteoblasts and osteoclasts is regulated by the action of many local and circulating hormones and factors as well as central regulation by a neurological mechanism. We have previously shown that lactoferrin can promote bone growth. At physiological concentrations, lactoferrin potently stimulates the proliferation and differentiation of primary osteoblasts and acts as a survival factor. Lactoferrin also affects osteoclasts, potently inhibiting their formation. In vivo, local injection of lactoferrin results in substantial increases in bone formation and bone area. In a critical bone-defect model in vivo, lactoferrin was also seen to promote bone growth. The mitogenic effect of lactoferrin in osteoblast-like cells is mediated mainly through low-density lipoprotein-receptor protein-1 (LRP1), a member of the low-density lipoprotein-receptor-related proteins that are primarily known as endocytic receptors; however, LRP1 is not necessary for the anti-apoptotic actions of lactoferrin. Lactoferrin also induces the activation of p42/44 mitogen-activated protein kinase (MAPK) signalling and the PI3-kinase-dependent phosphorylation of Akt in osteoblasts. In this study, we examined other properties of lactoferrin and the way they affect osteogenic activity. The degree of glycosylation, iron-binding, and the structure-activity relationships indicate that lactoferrin maintains osteogenic activity in deglycosylated, holo, and apo forms, and in with various small fragments of the molecule. These data suggest that lactoferrin signals through more than 1 membrane-bound receptor to produce its anabolic skeletal effects, and that it signals through diverse pathways. We conclude that lactoferrin might have a physiological role in bone growth and healing and a potential therapeutic role as an anabolic factor in osteoporosis.     

Nutritional factors and bone homeostasis: synergistic effect with zinc and genistein in osteogenesis

Bone homeostasis is regulated through osteoclasts and osteoblasts. Osteoporosis, which is induced with its accompanying decrease in bone mass with increasing age, is widely recognized as a major public health problem. Bone loss may be due to decreased osteoblastic bone formation and increased osteoclastic bone resorption. There is growing evidence that nutritional and food factors may play a part in the prevention of bone loss with aging and have been to be worthy of notice in the prevention of osteoporosis. Zinc, an essential trace element, or genistein, which are contained in soybeans, has been shown to have a stimulatory effect on osteoblastic bone formation and an inhibitory effect on osteoclastic bone resorption, thereby increasing bone mass. These factors have an effect on protein synthesis and gene expression, which are related to bone formation in osteoblastic cells and bone resorption in osteoclastic cells. The combination of zinc and genistein is found to reveal the synergistic effect on bone anabolic effect. The oral administration of those factors has been shown to prevent on bone loss in ovariectomized rats, an animal model for osteoporosis, indicating a role in the prevention of osteoporosis. Supplemental intake of ingredient with the combination of zinc and genistein has been shown to have a preventive effect on osteoporosis in human subjects, suggesting a role in the prevention of bone loss.