摘要:

AbstractThis cross‐sectional twin study examined the influence of constitutional, lifestyle, and genetic factors on bone mineral density (BMD) in elderly women. BMD, at the lumbar spine, femoral neck, Ward's triangle, total hip, and total forearm, total body bone mineral content (BMC), and lean mass and fat mass were measured using dual energy X‐ray absorptiometry in 69 volunteer female twin pairs (37 monozygotic [MZ], 32 dizygotic [DZ]) aged 60–89 years. Height and weight were measured. Medical history and lifetime tobacco and alcohol use were determined by questionnaire. In terms of within‐pair differences, lean mass was independently associated with BMD at all sites. In contrast, fat mass was not associated with BMD at any site once allowance had been made for lean mass. Lifetime tobacco use was independently associated with BMD at the lumbar spine, total hip, and forearm. Total body BMC was independently predicted by lean mass, fat mass, tobacco use, and alcohol consumption. Age and the above independently predictive body composition and lifestyle factors accounted for 20–33% of variation in BMD. After allowing for these covariates, MZ and DZ correlations were consistent with about 75% of residual variation in BMD at the nonforearm sites being determined by genetic factors. For total body BMC, the covariates explained 75% of total variation, and genetic factors 76% of the residual variation. Therefore, at the proximal femur and lumbar spine, after taking into account the relation of BMD with lean mass and smoking, genetic factors appear to play a substantial role in explaining variation in BMD in elde

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101102

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractAlkaline phosphatase is the marker enzyme for matrix vesicles, extracellular organelles that play a major role in primary bone formation and calcification. Recently, we developed osteosarcoma × fibrosarcoma hybrids in which alkaline phosphatase expression was greatly reduced, a phenomenon known as extinction. In the present study, we used two cell hybrids, LTA‐1 and LTA‐5, constructed from a human osteoblast‐like osteosarcoma, TE85, and a mouse fibrosarcoma, La−t−, to examine the differential distribution of alkaline phosphatase between matrix vesicles and the plasma membrane, postulated to be the parent membrane from which matrix vesicles are derived. While alkaline phosphatase in plasma membranes was extinguished, enzyme activity in matrix vesicles from LTA‐1 hybrid cells was 34.2% of that present in matrix vesicles from the TE85 parent cells and 200 times that found in La−t−matrix vesicles. Matrix vesicles from LTA‐5 had alkaline phosphatase levels similar to La−t−. When other membrane enzymes (phospholipase A2, 5′‐nucleotidase, and Na+/K+ATPase) were examined, hybrid matrix vesicle and plasma membrane levels were similar to those of TE85 and significantly higher than in La−t−membrane fractions. Northern analysis detected mRNA for alkaline phosphatase in TE85 cells, but not in the hybrids or La−t−cells. In contrast, reverse transcription‐polymerase chain reaction (RT‐PCR) revealed alkaline phosphatase mRNA in the hybrid cells, but at very low levels. Taken together, the data indicate that regulation of plasma membrane and matrix vesicle alkaline phosphatase is independent and suggest that matrix vesicle biogenesis is independent and distinct from that of plasma membrane biogenesis. Analysis of 1B‐ and 1L‐type alkaline phosphatase mRNA by RT‐PCR showed that alternate promoter usage of the alkaline phosphatase gene was not responsible for the differential localization of this enzyme in matrix vesicle. Thus, it is likely that matrix vesicle and plasma membrane alkaline phosphatase are regulated

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101103

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractAvian tibial dyschondroplasia (ATD), a disease characterized by an almost total lack of mineralization in affected areas of growth plate cartilage, may involve defective matrix vesicle (MV) mineralization. To explore the biochemical defect in ATD, both normal and diseased tissue were analyzed for the amount of isolatable MVs, their chemical composition, and their ability to induce mineral formation. We found significantly fewer MVs in ATD tissue, and in contrast to normal MVs, which rapidly mineralized when incubated in synthetic cartilage lymph, those isolated from ATD lesions induced only limited mineralization even after prolonged incubation. Analysis by detergent extraction revealed a nearly dysfunctional nucleational core in ATD MVs. Thus, in ATD tissue, there is a defect in the formation of MVs, and those that form are nearly inactive. There were also alterations in the lipid‐dependent Ca2+‐binding proteins (annexins) in ATD MVs. There were lower levels of annexins II and VI in endogenously produced collagenase‐released matrix vesicles (CRMVs), but not in matrix vesicle–enriched microsomes (MVEMs) produced by tissue homogenization. These findings indicate that there is insufficient Ca2+in ATD cells to enable incorporation of the annexins into MVs. Finally, there was evidence of phospholipid breakdown in ATD MVs, as well as in ATD tissue generally. This indicated that the ATD lesions were becoming necrotic. Taken together, these findings indicate that there is a defect in tissue vascularization such that the supply of mineral ions and nutrients to ATD cartilage is inadequate to support normal MV formation and subsequent mineral

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101104

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractWhen grown in medium containing ascorbic acid and β‐glycerol phosphate, mouse MC3T3‐E1 cells express an osteoblast phenotype and produce a highly mineralized extracellular matrix. The purpose of this study was to independently examine the role of the collagenous matrix and functional osteoblasts on the mineralization process. Cultures with and without an extensive collagenous matrix were prepared by growing MC3T3‐E1 cells in the presence and absence of ascorbic acid. Matrix‐rich cultures mineralized at much lower calcium phosphate ion products than nonmatrix cultures. At higher ion products, spontaneous precipitation in the medium and cell layers of nonmatrix cultures were observed. In contrast, mineral in matrix‐rich cultures was still exclusively associated with collagen fibrils and not with ectopic sites in the cell layer or medium. To examine the effect of cell viability on matrix mineralization, cells were grown 8 or 16 days in the presence of ascorbic acid, then killed and incubated in a mineralizing medium. Significant mineralization was not observed in the collagenous matrix of 8‐day killed cultures or age‐matched controls. At 16 days mineral was associated with collagen fibrils at specific foci in the matrix of both viable and killed cultures. This observation is consistent with the concept that collagenous matrices must undergo a maturation process before they can support mineral induction and growth. It further shows that osteoblast‐like cells are not required for mineralization of mature matrices, but are required for m

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101105

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractA number of cytokines have been shown to exert their effects via a recently discovered signaling cascade. One step in this pathway is mediated by a family of nonreceptor protein tyrosine kinases, the Janus kinases or JAK kinases, which become phosphorylated upon ligand‐receptor binding and receptor phosphorylation. This in turn is followed by phosphorylation of certain members of a family of latent transcription factors, called signal transducers and activators of transcription (STATs), which subsequently enter the nucleus, bind to DNA in a sequence‐specific fashion, and modulate transcription. In view of the apparent role of leukemia inhibitory factor (LIF) in bone remodeling, we sought to determine which, if any, of the JAK/STAT family members are involved in mediating the actions of LIF using the MC3T3‐E1 cell line (a spontaneously immortalized osteoblast) and normal murine calvarial osteoblasts. We report here rapid and transient phosphorylation of the LIF receptor, and similarly, we detect phosphorylation of predominantly JAK1 and to a minor extent JAK2 in response to LIF treatment in MC3T3‐E1 cells. In these experiments we also detect phosphorylation of STAT1 and to a much lesser degree STAT3 upon addition of LIF. Phosphorylation of the STAT proteins correlates directly with their ability to bind DNA in a gel mobility shift assay in MC3T3‐E1 cells and in normal calvarial osteoblasts. These studies suggest that LIF action in these cells, as in other cell types, is mediated in part via specific members of the JAK/STA

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101106

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractType I receptors for bone morphogenetic proteins (BMPs), i.e., BMPR‐IA and BMPR‐IB, are transmembrane serine/threonine kinases, that bind osteogenic protein‐1 (OP‐1, also termed BMP‐7) and BMP‐4. Using antibodies specific to BMPR‐IA and ‐IB, we have studied the expression of BMP type I receptors in the bone formation process during embryonic development and fracture healing. In the mouse embryo, both BMPR‐IA and ‐IB were expressed in condensing mesenchymal cells at 13.5 dayspost coitum(p.c.). At 15.5 days p.c., expression of BMPR‐IB, but not of BMPR‐IA, was observed in the cells in perichondrium of developing cartilage. At 17.5 and 19.5 days p.c., expression of both receptors was observed in chondrocytes and in osteoblasts. In normal rat adult bone, expression of BMPR‐IA, but not of BMPR‐IB, was observed in osteoblasts in the periosteum. Three days after the femoral fracture, expression of BMPR‐IA and ‐IB was up‐regulated in cells at the proliferating osteogenic layer of the periosteum. On day 7, both receptors were found in fibroblast‐like spindle cells and chondrocytes in the endochondral ossification sites, and osteoblasts in the newly formed trabecular bone. Expression of BMPR‐IA was higher than that of BMPR‐IB in osteogenic layer on day 3 and in osteoblasts in the trabecular bone on day 7. On day 14, expression of BMP type I receptors was observed at similar sites, albeit with lower expression levels than were observed on day 7. The present data suggest that expression of BMP type I receptors is up‐regulated during bone formation, and that they may p

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101107

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractExtracellular calcium concentration is critically important for normal function of the body. Recently, reports have shown that cells derived from parathyroid glands contain an extracellular calcium receptor that is responsive to changes in extracellular calcium. Bone is also intimately involved in calcium homeostasis; therefore, we sought to test the hypothesis that extracellular calcium has direct effects on bone cells. Extracellular calcium was increased by the addition of varying concentrations of CaCl2(0.4–2.0 mM) to the control medium. An increase in extracellular calcium increased cell proliferation, as assessed by3H‐thymidine incorporation, in a number of cell types including normal human bone cells derived from vertebrae (HBV155) and a number of human osteosarcoma cell lines. The increase in cell proliferation by elevated CaCl2was dose dependent, whereas MgCl2was not effective at the doses tested (up to 2 mM added MgCl2). To test the hypothesis that the mitogenic activity of elevated extracellular calcium involved a growth factor, levels of insulin‐like growth factor II (IGF‐II) were measured in the conditioned medium of HBV155 cells by radioimmunoassay after removal of binding proteins by size exclusion chromatography. The effects of an increase in extracellular calcium by 1 mM were: 1) increased culture media levels of IGF‐II within 1 h of treatment, 2) the increase in IGF‐II levels reached a maximum after 8 h of treatment, and 3) IGF‐II levels were still elevated after 24 h of treatment. Furthermore, a blocking monoclonal antibody against IGF‐II abolished the increased cell proliferation in HBV155 cells following elevation of extracellular calcium. Taken together, these findings suggest that an increase in extracellular calcium results in an increase in IGF‐II which is required for the subsequent increase in c

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101108

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractOsteopontin is a phosphorylated glycoprotein believed to be secreted by osteoblasts and deposited into the bone matrix to facilitate osteoclast adhesion or to initiate osteoid mineralization. Previously we have presented contradictory evidence that osteoclasts express osteopontin mRNA in human remodeling bone. The aim of this study was to ascertain whether osteoclasts synthesize and deposit osteopontin in resorption lacunae. We characterized expression of osteopontin mRNA and protein expression in both intramembranous and endochondral ossification, as well as remodeling bone, in the human osteophyte. Osteopontin mRNA was expressed in osteoclasts with tartrate‐resistant acid phosphatase (TRAP) positivity within resorption lacunae. The osteoclasts and immediate resorption surfaces also expressed osteopontin. However, osteopontin mRNA and protein were weak (transient) or undetectable in osteoblasts at adjacent bone formation sites; no osteopontin expression was observed in the osteoid, although occasional reactivity was observed in osteocytes and the mineral‐osteoid interface. In contrast, osteopontin was highly expressed in the osteoblasts and matrix of woven bone during intramembranous and endochondral ossification. The matrix expression correlated with mineralization; however, in some instances osteopontin deposition was observed prior to mineralization. Similarly, osteopontin expression was evident in cartilage matrix, solely at foci of mineralization. Chondroclasts expressed osteopontin mRNA and protein; the surfaces of resorbed calcified cartilage also expressed osteopontin. Abnormal, unmineralized matrices apparently lacked deposited osteopontin, but were nevertheless resorbed by osteoclasts; the osteoclasts and resorbed surfaces expressed no osteopontin protein. That osteoclasts are responsible for the deposition of osteopontin was confirmed in vitro, whereby resorption pits in whale dentine and bovine bone slices, produced by isolated human osteoclasts, contained deposited osteopontin. Osteopontin may facilitate the adhesion (or detachment) of the osteoclast to the bone surface. Alternatively, the possibility that osteopontin may act as a postresorptive signal to recruit osteoblasts, or to polarize and direct the mineralization of the formed osteoid, is discus

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101109

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractAlthough the action of bone morphogenetic protein (BMP) on osteoblast differentiation has been extensively investigated, its effect on osteoclast differentiation remains unknown. In the present study, in vitro effects of BMP‐2 on osteoclast‐like cell formation and bone resorption were examined. BMP‐2 (1–100 ng/ml) significantly stimulated bone resorption by preexistent osteoclast‐like cells in mouse bone cell cultures containing stromal cells, whereas it did not affect the bone‐resorbing activity of isolated rabbit osteoclast‐like cells. When BMP‐2 was added to unfractionated bone cells after degeneration of preexistent osteoclast‐like cells, BMP‐2 dose‐dependently stimulated osteoclast‐like cell formation at a minimal effective concentration of 10 pg/ml. BMP‐2 also enhanced the osteoclast‐like cell formation induced by 1,25‐dihydroxyvitamin D3(1,25(OH)2D3). Moreover, osteoclast‐like cells newly formed by BMP‐2 from unfractionated bone cells possessed the ability to form pits on dentine slices. Because these results indicated that BMP‐2 directly or indirectly stimulated osteoclast differentiation and activity, we next examined the direct effect of BMP‐2 on osteoclast precursors in the absence of stromal cells using hemopoietic blast cells derived from spleen cells. The mRNA for BMP‐2/4 receptor was detected in hemopoietic blast cells supported by granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) as well as osteoblastic MC3T3‐E1 cells and MC3T3‐G2/PA6 stromal cells by RNase protection assay. BMP‐2 dose‐dependently stimulated osteoclast‐like cell formation from hemopoietic blast cells supported by GM‐CSF at a minimal effective concentration of 10 pg/ml. BMP‐2 also enhanced 1,25(OH)2D3‐induced osteoclast‐like cell formation from hemopoietic blast cells. The present data are the first to indicate that BMP‐2 stimulates bone resorption through both direct stimulation of osteoclast formation and activ

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101110

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY

摘要:

AbstractGene expression patterns have been investigated in prolonged cultures of rat mandibular condylar cartilage (MCC) cells to examine the possibility of culture‐induced phenotypic changes. MCC cells were isolated from newborn rats and grown in the presence of 10% fetal bovine serum (FBS) and basic fibroblast growth factor (bFGF). MCC cells were passaged and cultured in the presence of 10% FBS and bFGF until confluent. After confluence, the medium was changed to that supplemented with 10% FBS, ascorbate, and β‐glycerophosphate (day 1). Mineralization and gene expression of MCC cells have been investigated. Mineralization, visualized by staining with Alizarin red, was observed to begin at day 13 in culture, and increased up to day 22 in culture, which was the length of this study. Type II collagen and aggrecan mRNAs were highly expressed at the start of culture (day 1–4) and decreased in a time‐dependent manner. Type I collagen, alkaline phosphatase, and osteopontin mRNAs expressed biphasic patterns that peaked at the start of culture and the beginning of mineralization (day 13–16). Osteonectin mRNA was expressed throughout the culture period. Osteocalcin mRNA was expressed before the beginning of mineralization peaking at day 7. These observations suggest that the gene expression patterns of MCC cells can be categorized into two different periods in prolonged culture: maturation (day 1–10) and mineralization (day 13–22). The cell culture system of MCC represents a new model system in which the differentiation of embryonic MCC cells c

ISSN:0884-0431    

DOI:10.1002/jbmr.5650101111

出版商:John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)    

年代:1995

数据来源: WILEY