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M Amling

Publications and source records attributed to M Amling.

At least 37 records · Page 2Linked to original sources
PubMed2000

Osteopenia and decreased bone formation in osteonectin-deficient mice.

Bone continuously remodels in response to mechanical and physiological stresses, allowing vertebrates to renew bone as adults. Bone remodeling consists of the cycled synthesis and resorption of collagenous and noncollagenous extracellular matrix proteins, and an imbalance in this process can lead to disease states such as osteoporosis, or more rarely, osteopetrosis. There is evidence that the extracellular matrix glycoprotein osteonectin or secreted protein acidic and rich in cysteine (BM-40) may be important in bone remodeling. Osteonectin is abundant in bone and is expressed in areas of active remodeling outside the skeleton. In vitro studies indicate that osteonectin can bind collagen and regulate angiogenesis, metalloproteinase expression, cell proliferation, and cell-matrix interactions. In some osteopenic states, such as osteogenesis imperfecta and selected animal models for bone fragility, osteonectin expression is decreased. To determine the function of osteonectin in bone, we used contact x-ray, histomorphometry, and Northern blot analysis to characterize the skeletal phenotype of osteonectin-null mice. We found that osteonectin-null mice have decreased bone formation and decreased osteoblast and osteoclast surface and number, leading to decreased bone remodeling with a negative bone balance and causing profound osteopenia. These data indicate that osteonectin supports bone remodeling and the maintenance of bone mass in vertebrates.

Animals↗
PubMed1999

A Cbfa1-dependent genetic pathway controls bone formation beyond embryonic development.

The molecular mechanisms controlling bone extracellular matrix (ECM) deposition by differentiated osteoblasts in postnatal life, called hereafter bone formation, are unknown. This contrasts with the growing knowledge about the genetic control of osteoblast differentiation during embryonic development. Cbfa1, a transcriptional activator of osteoblast differentiation during embryonic development, is also expressed in differentiated osteoblasts postnatally. The perinatal lethality occurring in Cbfa1-deficient mice has prevented so far the study of its function after birth. To determine if Cbfa1 plays a role during bone formation we generated transgenic mice overexpressing Cbfa1 DNA-binding domain (DeltaCbfa1) in differentiated osteoblasts only postnatally. DeltaCbfa1 has a higher affinity for DNA than Cbfa1 itself, has no transcriptional activity on its own, and can act in a dominant-negative manner in DNA cotransfection assays. DeltaCbfa1-expressing mice have a normal skeleton at birth but develop an osteopenic phenotype thereafter. Dynamic histomorphometric studies show that this phenotype is caused by a major decrease in the bone formation rate in the face of a normal number of osteoblasts thus indicating that once osteoblasts are differentiated Cbfa1 regulates their function. Molecular analyses reveal that the expression of the genes expressed in osteoblasts and encoding bone ECM proteins is nearly abolished in transgenic mice, and ex vivo assays demonstrated that DeltaCbfa1-expressing osteoblasts were less active than wild-type osteoblasts. We also show that Cbfa1 regulates positively the activity of its own promoter, which has the highest affinity Cbfa1-binding sites characterized. This study demonstrates that beyond its differentiation function Cbfa1 is the first transcriptional activator of bone formation identified to date and illustrates that developmentally important genes control physiological processes postnatally.

Amino Acid Sequence↗
PubMed1999

The tyrosine phosphatase SHP-1 is a negative regulator of osteoclastogenesis and osteoclast resorbing activity: increased resorption and osteopenia in me(v)/me(v) mutant mice.

Naturally occuring inactivating mutations of the Src homology 2 (SH2) domain-containing tyrosine phosphatase 1 (SHP-1) in mice give rise to the motheaten (me) phenotype. me/me mice have multiple hematopoietic abnormalities, suggesting that this phosphatase plays an important role in hematopoiesis. SHP-1 binds to and is activated by several hematopoietic surface receptors, including the colony-stimulating factor type 1 receptor. We have examined the role of SHP-1 in osteoclastogenesis and osteoclast function using mice with the viable motheaten (me(v)/me(v)) mutation, which has markedly decreased SHP-1 activity. Histomorphometric analysis of 6-week-old me(v)/me(v) mice and control littermates showed a marked osteopenia with an increase in bone resorption indices. The number of formed osteoclast-like cells (OCLs) in cocultures of me(v)/me(v) hematopoietic cells with normal osteoblasts was significantly increased. In contrast, the number of OCLs formed in the coculture of normal bone marrow cells with the me(v)/me(v) osteoblasts was not significantly different from controls. The bone-resorbing activity of me(v)me(v) OCLs and authentic osteoclasts was also found to be increased. Finally, Western blotting of proteins from me(v)/me(v) and control OCLs revealed an overall increase in tyrosine phosphorylation in the me(v)/me(v) lysates. These in vivo and in vitro results suggest that SHP-1 is a negative regulator of bone resorption, affecting both the formation and the function of osteoclasts.

Animals↗
PubMed1999

Ablation of the PTHrP gene or the PTH/PTHrP receptor gene leads to distinct abnormalities in bone development.

Parathyroid hormone (PTH) and parathyroid hormone-related peptide (PTHrP) bind to and activate the same PTH/PTHrP receptor. Deletion of either the PTHrP gene or the PTH/PTHrP receptor gene leads to acceleration of differentiation of growth plate chondrocytes. To explore further the functional relationships of PTHrP and the PTH/PTHrP receptor, bones of knockout mice were analyzed early in development, and the phenotypes of double-knockout mice were characterized. One early phenotype is shared by both knockouts. Normally, the first chondrocytes to become hypertrophic are located in the centers of long bones; this polarity is greatly diminished in both these knockouts. The PTH/PTHrP receptor-deficient (PTH/PTHrP-R(-/-)) mice exhibited 2 unique phenotypes not shared by the PTHrP(-/-) mice. During intramembranous bone formation in the shafts of long bones, only the PTH/PTHrP-R(-/-) bones exhibit a striking increase in osteoblast number and matrix accumulation. Furthermore, the PTH/PTHrP-R(-/-) mice showed a dramatic decrease in trabecular bone formation in the primary spongiosa and a delay in vascular invasion of the early cartilage model. In the double-homozygous knockout mice, the delay in vascular invasion did not occur. Thus, PTHrP must slow vascular invasion by a mechanism independent of the PTH/PTHrP receptor.

Animals↗
PubMed1999

Osteoblasts are a new target for prolactin: analysis of bone formation in prolactin receptor knockout mice.

Bone development is a multistep process that includes patterning of skeletal elements, commitment of hematopoietic and/or mesenchymental cells to chondrogenic and osteogenic lineages, and further differentiation into three specialized cell types: chondrocytes in cartilage and osteoblasts and osteoclasts in bone. Although PRL has a multitude of biological actions in addition to its role in the mammary gland, very little is known about its effect on bone. Mice carrying a germline null mutation for the PRL receptor gene have been produced in our laboratory and used to study the role of PRL in bone formation. In -/- embryos, we observed an alteration in bone development of calvaria. In adults, histomorphometric analysis showed that the absence of PRL receptors leads to a decrease in bone formation rate using double calcein labeling and a reduction of bone mineral density, measured by dual energy x-ray absorptiometry. In addition, serum estradiol, progesterone, testosterone, and PTH levels were analyzed. We also established that osteoblasts, but not osteoclasts, express PRL receptors. This suggests that an effect of PRL on osteoblasts could be required for normal bone formation and maintenance of bone mass. Thus, the PRL receptor knockout mouse model provides a new tool to investigate the involvement of PRL in bone metabolism.

Absorptiometry, Photon↗
PubMed1999

Rescue of the skeletal phenotype of vitamin D receptor-ablated mice in the setting of normal mineral ion homeostasis: formal histomorphometric and biomechanical analyses.

1,25-Dihydroxyvitamin D3 has been shown to play an important role in vitro in regulating osteoblast gene transcription and promoting osteoclast differentiation. To address the role of the vitamin D receptor (VDR) in skeletal homeostasis, formal histomorphometric analyses were performed in VDR null mice in the setting of impaired mineral ion homeostasis as well as in VDR null mice in whom normal mineral ion homeostasis had been preserved. In hypocalcemic VDR null mice, there was an increase in bone volume as a result of a dramatic increase in osteoid. There was also an increase in the number of osteoblasts without a significant change in the number of osteoclasts. Examination of the growth plate revealed marked disorganization, with an increase in vascularity and matrix. Biomechanical parameters demonstrated increased bone fragility in the hypocalcemic VDR null mice. In the VDR ablated mice in whom normal mineral ion homeostasis had been preserved, none of these measurements was significantly different from those in wild-type littermates raised under identical conditions. Notably, the morphology and width of the growth plate were indistinguishable from those in wild-type controls, demonstrating that a calcium/phosphorus/lactose-enriched diet started at 16 days of age in the VDR null mice permits the development of both normal morphology in the growth cartilage and adjacent metaphysis and normal biomechanical competence of cortical bone. Thus, the principle action of the VDR in skeletal growth, maturation, and remodeling is its role in intestinal calcium absorption. The skeletal consequences of VDR ablation are a result of impaired intestinal calcium absorption and/or the resultant secondary hyperparathyroidism and hypophosphatemia.

Animals↗
PubMed1999

The Gorham-Stout syndrome (Gorham's massive osteolysis). A report of six cases with histopathological findings.

The Gorham-Stout Syndrome (Gorham's massive osteolysis) is a rare condition in which spontaneous, progressive resorption of bone occurs. The aetiology is poorly understood. We report six cases of the condition and present evidence that osteolysis is due to an increased number of stimulated osteoclasts. This suggests that early potent antiresorptive therapy such as with calcitonin or bisphosphonates may prevent local progressive osteolysis.

Adult↗
PubMed1999

Oligodeoxynucleotide targeted to the alphav gene inhibits alphav integrin synthesis, impairs osteoclast function, and activates intracellular signals to apoptosis.

The alphav integrin subunit is highly expressed in osteoclasts where it dimerizes with beta1 and beta3 subunits to form receptors for vitronectin and bone sialoproteins. Inhibition of osteoclast adhesion and function has previously been achieved by alphavbeta3 antibodies or Arg-Gly-Asp-containing peptides which have the disadvantages of blocking a single receptor type, or of being rather nonspecific, respectively. Here we show that alphav integrin expression in rabbit osteoclasts can be inhibited by partially phosphorothioated antisense oligodeoxynucleotide (ODN) spanning the adenine-uracil-guanine (AUG) translational start site of the human/rabbit alphav gene, a procedure which offers the advantage of affecting all the alphav receptors with high efficiency. The alphav antisense ODN caused a dose-dependent, substrate-specific reduction of osteoclast adhesion and bone resorption. Control ODNs, such as sense, inverted, and mismatch, were without effect, providing evidence of specificity of the antisense reagent. It is likely as a consequence of loss of substrate interaction, the antisense ODN induced osteoclast retraction and apoptosis, increase of the cyclin/cyclin-dependent kinase complex inhibitor p21WAF1/CIP1, and inhibition of the cell survival gene, bcl-2. Although the expression of the cell death-promoting gene, bax, remained unchanged, a reduction of the bcl-2/bax ratio, known to underlie the intracellular signal to apoptosis, was observed. This finding led us to hypothesize that these changes could provide a link between reduction of alphav synthesis and osteoclast programmed death. In conclusion, this study provides novel insights into the use of alphav antisense ODN as an efficacious mechanism for blocking osteoclast function and underscores for the first time the involvement of integrins in bone cell apoptosis. In vivo studies may verify potential application of this ODN as alternative therapy for bone diseases.

Animals↗
PubMed1998

Dissociation between bone resorption and bone formation in osteopenic transgenic mice.

Bone mass is maintained constant in vertebrates through bone remodeling (BR). BR is characterized by osteoclastic resorption of preexisting bone followed by de novo bone formation by osteoblasts. This sequence of events and the fact that bone mass remains constant in physiological situation lead to the assumption that resorption and formation are regulated by each other during BR. Recent evidence shows that cells of the osteoblastic lineage are involved in osteoclast differentiation. However, the existence of a functional link between the two activities, formation and resorption, has never been shown in vivo. To define the role of bone formation in the control of bone resorption, we generated an inducible osteoblast ablation mouse model. These mice developed a reversible osteopenia. Functional analyses showed that in the absence of bone formation, bone resorption continued to occur normally, leading to an osteoporosis of controllable severity, whose appearance could be prevented by an antiresorptive agent. This study establishes that bone formation and/or bone mass do not control the extent of bone resorption in vivo.

Animals↗
PubMed1998

[Quantitative morphology of vertebral body cortical bone. Building block for noninvasive calculation of fracture threshold in osteoporosis].

The vertebral bodies consist of two main structures, trabecular and cortical bone. The histological changes within the spine, especially in cortical bone, leading to osteoporotic fractures remain, however, poorly understood. Therefore, the complete front column of the spine was removed in 26 autopsy cases without skeletal diseases and in 11 cases with proven osteoporosis. A sagittal segment prepared through the center of all vertebral bodies was undecalcified embedded in plastic, ground to a 1-mm-thick block and stained using a modification of the von Kossa method. The analysis included measurement of the mean cortical thickness of both ventral and dorsal shell (from C3 to L5). The qualitative investigation of the structure of the cortical ring completed the analysis. The skeletally intact specimens had high cortical thickness values in the cervical spine (285 +/- 22 microns), a decrease in the thoracic spine (244 +/- 14 microns) and an increase in the lumbar spine (290 +/- 15 microns). The mean thickness of the ventral shell is in general higher than the thickness of the dorsal shell. The cortical thickness of the spine showed no gender-specific differences (P = n.s.). There was a slight decrease in the cortical thickness with age; however, this decrease and the correlation of cortical thickness to age was only significant below vertebral body T8 (r = 0.225 to 0.574; Pr < 0.05 to Pr < 0.005). Most interestingly, osteoporosis is characterized by a significant decrease in cortical thickness throughout the whole spine. This decrease in cortical thickness was more marked in the dorsal shell (P < 0.05) than in the ventral shell (ventral from C3 to T6 (P < 0.05) below T6 (P = n.s.). We therefore conclude that in osteoporosis, biomechanical competence is affected by both trabecular bone loss and decrease of cortical thickness. This suggests that, in addition to trabecular bone measurements, the cortical thickness is of special interest for diagnostic radiological examinations (CT) to yield clues about the risk of vertebral fractures.

Adolescent↗
PubMed1998

[Biology of osteoclasts; their role in bone metastases].

Bone is a common site for metastasis of malignant tumors. These can be recognized radiologically as either lytic or sclerotic lesions since the tumor cells stimulate resident bone cells to cause excessive local resorption or new bone formation. The osteoclast, as the only cell being capable of resorbing bone, is of major importance for the homing of tumor cells in bone and progression of metastasis due to bone destruction. Thus, the improvement of our means of therapeutic intervention towards prevention of tumor progression and pathological fractures will depend on our better understanding of both the molecular basis of bone resorption and the cellbiology of the osteoclast. This article presents our current opinion of the molecular mechanisms of bone resorption by the osteoclast. After describing the morphological features of the osteoclast, aspects such as cell mobility, attachment, enzymes synthesis, transmembrane transport, osteoclast differentiation and function, as well as the protooncogenes c-src and c-cbl and their role in bone resorption are presented in detail.

Bone Neoplasms↗
PubMed1998

Heterogeneity of trabecular bone structure in the calcaneus using magnetic resonance imaging.

The purpose of this study was to quantify the heterogeneity in the trabecular bone structure in the calcaneus. Magnetic resonance (MR) images of the calcaneus were obtained in the sagittal plane at an in-plane resolution of 195 microns and a slice thickness of 1000 microns in 12 young normal subjects. Regions of interest (ROI) were selected to cover the calcaneus using a grid of square boxes (10 mm per side). A thresholding technique based on the regional intensity histogram was used to segment the images into trabecular bone and marrow phases and to calculate measures such as apparent trabecular bone area fraction, apparent trabecular spacing, apparent trabecular thickness and apparent trabecular number. Bone mineral density (BMD) of the calcaneus was assessed using dual-energy X-ray absorptiometry (DXA). Histological sections of three calcanei were also analyzed using transmission light illumination, and the results used to calibrate our computational software. For a relatively narrow inter-subject variation in posterior BMD, a significant inter-subject variation was seen in MRI-derived structural parameters. Furthermore, the spatial heterogeneity of the structural parameters in the posterior region was as high as 40%. Thus, the posterior tuberosity of the calcaneus, a typical site for BMD and single-point ultrasound assessments, can demonstrate significant regional variation in trabecular bone structure.

Absorptiometry, Photon↗
PubMed1998

Oligonephronia, not exuberant apoptosis, accounts for the development of glomerulosclerosis in the bcl-2 knockout mouse.

BACKGROUND: A main function attributed to B cell leukaemia/lymphoma 2 gene (bcL-2) is its ability to confer resistance against apoptosis. In bcl-2 deficient mice, extensive apoptosis occurs during abnormal nephrogenesis, and renal failure is found very quickly after birth. However, the underlying mechanisms remain poorly understood. The aim of the present study was to clarify whether the degenerative process in the kidneys seen after birth is based either on increased apoptosis of glomerular cells or on mechanisms independent from the genetic defect. MATERIALS AND METHODS: Kidneys from 7-56-day-old bcl-2 knockout mice and wild-type litter mates were studied. Glomerular number, glomerular tuft volume, cell counts in 'non-sclerotic' glomeruli as well as the glomerular damage score were determined by histomorphometrical studies. Apoptosis was evaluated by morphological criteria and the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL)-technique. RESULTS: The number of nephrons at birth was severely decreased in bcl-2 knockout mice compared to controls (<20%; P < 0.001). These nephrons undergo dramatic hypertrophy with an approximately 4-fold increase in volume (P < 0.001). In hypertrophic, but 'non-sclerotic' glomeruli, the number density of glomerular cells progressively declined with time (P<0.001). Starting with day 20, enlarged glomeruli developed sclerosis beginning with a segmental distribution, but quickly progressing to global sclerosis. Apoptosis was neither detected in non-sclerotic glomeruli nor in stages prior to fully established sclerosis. As shown by the glomerular damage score, post-natal degeneration of kidneys from bcl-2 knockout animals proceeded rapidly. CONCLUSIONS: Bcl-2 knockout mice exhibit deficient nephrogenesis resulting in severe oligonephronia at birth. Post-partum development of glomerulosclerosis does not seem to be due to augmented apoptosis. The degenerative process appears to be based on a glomerular overload with increased mechanical stress to the filtration barrier, leading via glomerular hypertrophy, podocyte damage and formation of tuft adhesions to glomerulosclerosis.

Animals↗
PubMed1998

Normalization of mineral ion homeostasis by dietary means prevents hyperparathyroidism, rickets, and osteomalacia, but not alopecia in vitamin D receptor-ablated mice.

1,25-Dihydroxyvitamin D3 plays a major role in intestinal calcium transport. To determine what phenotypic abnormalities observed in vitamin D receptor (VDR)-ablated mice are secondary to impaired intestinal calcium absorption rather than receptor deficiency, mineral ion levels were normalized by dietary means. VDR-ablated mice and control littermates were fed a diet that has been shown to prevent secondary hyperparathyroidism in vitamin D-deficient rats. This diet normalized growth and random serum ionized calcium levels in the VDR-ablated mice. The correction of ionized calcium levels prevented the development of parathyroid hyperplasia and the increases in PTH messenger RNA synthesis and in serum PTH levels. VDR-ablated animals fed this diet did not develop rickets or osteomalacia. However, alopecia was still observed in the VDR-ablated mice with normal mineral ions, suggesting that the VDR is required for normal hair growth. This study demonstrates that normalization of mineral ion homeostasis can prevent the development of hyperparathyroidism, osteomalacia, and rickets in the absence of the genomic actions of 1,25-dihydroxyvitamin D3.

Alopecia↗
PubMed1998

PTHrP and Bcl-2: essential regulatory molecules in chondrocyte differentiation and chondrogenic tumors.

Human chondrosarcomas (CS) are a frequent form of malignant bone tumors. The accurate distinction between benign solitary enchondroma and conventional CS of bones is a major diagnostic goal. Although the histological characteristics of chondrogenic tumors and the grading of CS (G1 to G3) have been defined by several authors, immunohistochemical markers for the different entities and grades are still missing and the mechanisms of tumorigenesis remain poorly understood. In addition to the emerging evidence that parathyroid hormone-related peptide (PTHrP) plays a critical role in endochondral bone formation we have recently reported that Bcl-2 lies downstream of PTHrP in the regulation of chondrocyte differentiation. To further characterize chondrogenic tumors and to determine whether PTHrP and the regulation of Bcl-2-expression is of relevance to tumorigenesis, we analyzed the expression of both PTHrP and Bcl-2 on a series of 23 cases of solitary enchondroma (9 cases) and primary CS (14 cases) using light and confocal microscopy. While all 9 enchondromas exhibited a detectable level of PTHrP-expression only, 2 showed low levels of immunoreactivity for Bcl-2. In sharp contrast, strong coexpression of Bcl-2 and PTHrP was found in 11 (composed of 3 CS G3, 7 CS G2, and one dedifferentiated CS) out of 14 CS, while the expression level of these proteins was below the detection limit in two CS G1 and one dedifferentiated CS. To verify this data 3 cases each of enchondroma, CS G2, and CS G3 respectively, were subjected to quantitative confocal analysis, after double labeling for PTHrP and Bcl-2. The results showed a significant increase in the expression of both PTHrP and Bcl-2, in malignant CS versus the benign enchondromas. Most interestingly, the levels of expression of both PTHrP and Bcl-2 correlated with the degree of malignancy of the chondrogenic tumors. These results therefore suggest that both PTHrP and Bcl-2 play a role in the tumorigenesis of chondrogenic tumors and further indicate that both proteins may participate in the same pathway regulating chondrocyte differentiation.

Adult↗