The nature of the multinucleated cells involved in the resorption processes occurring inside macroporous calcium-phosphate biomaterials grafted into rabbit bone was studied using light microscopy, histomorphometric analysis, enzymatic detection of tartrate-resistant acid phosphatase (TRAP) activity, scanning, and electron microscopy. Samples were taken at days 7, 14, and 21 after implantation. As early as day 7, osteogenesis and resorption were observed at the surface of the biomaterials, inside the macropores. Resorption of both newly formed bone and calcium-phosphate biomaterials was associated with two types of multinucleated cells. Giant multinucleated cells were found only at the surface of the biomaterials; they showed a large number of nuclei, were TRAP negative, developed no ruffled border, and contained numerous vacuoles with large accumulation of mineral crystals from the biomaterials. Osteoclasts exhibited TRAP positivity and well-defined ruffled border. They were observed at the surface of both newly formed bone and biomaterials, around the implant, and inside the macropores. In contract with the biomaterials, infoldings of their ruffled border were observed between the mineral crystals, deeply inside the microporosity. The microporosity of the biomaterials (i.e., the noncrystalline spaces inside the biomaterials) increased underneath this type of cell as compared with underneath giant cells or to the depth of the biomaterials. These observations demonstrate that macroporous calcium-phosphate biomaterials implanted in bone elicit osteogenesis and the recruitment of a double multinucleated cell population having resorbing activity: giant multinucleated cells that resorb biomaterials and osteoclasts that resorb newly formed bone and biomaterials.
Prolonged corticosteroid (CS) therapy induces osteoporosis and fractures. Osteoporosis is characterized at the histomorphometric level by reduced bone volume (BV/TV) and disruption of the three-dimensional (3D) trabecular architecture. Several stereological methods have been proposed to characterize these alterations: measurements of trabecular thickness and trabecular number, star volumes, interconnectivity index (ICI) of the bone marrow spaces, and trabecular bone pattern factor (TBP(f)). These methods were computerized with a single program running on an image analyzer to evaluate the bone changes in a series of iliac biopsies performed on 31 male patients. All of them were asthmatic and had received CS for a long period of time. BV/TV was reduced when compared with age-matched controls. In the CS-treated population, exponential relationships were obtained between bone volume and the different connectivity parameters. The various methods used to measure connectivity were well correlated. When the population was divided into two groups (BV/TV greater or less than an 11% threshold), the architectural disturbances were found to imply two mechanisms. A progressive decline in trabecular thickness was noted in both groups versus controls. Trabecular perforations were not established in the group with BV/TV> 11% with the star volume or ICI, although some alterations were detected by trabecular bone pattern factor measurement. However, perforations were revealed in the group with BV/TV < 11% by all the different methods. Perforations seemed to occur when the trabecular thickness was below 70 mu m. This strongly suggests that bone histomorphometry should take into consideration bone volume in combination with detailed 3D descriptors of the trabecular architecture. Several histological methods need to be used in combination to appreciate the 3D architecture of trabecular bone.
SUMMARYMorphological and immunocytological studies have demonstrated the presence of paramyxovirus antigens in Paget's bone disease tissue and in particular antigens related to measles virus and respiratory syncytial virus. To examine the relationship between measles virus and Paget's bone disease we used in situ hybridization and a cloned measles virus DNA probe specific for the nucleocapsid protein to detect and locate measles virus RNA sequences in Paget's bone tissue. In five patients with the disease, measles virus RNA sequences were detected not only in 80 to 90% of the multinucleated osteoclasts where there is morphological and immunocytological evidence of measles virus activity but also in 30 to 40% of mononucleated bone cells, mainly osteoblasts, osteocytes, fibroblasts and lympho-monocytes. In contrast, no hybridization was observed in bone tissue from three control patients without signs of Paget's bone disease. These results indicate that the host cell range for measles virus in Paget's disease is more widespread than has been supposed. They also demonstrate the usefulness of the in situ hybridization method to detect viral genetic information in cells where viral antigenic activity is not detectable. These observations further support the hypothesis that measles virus is involved in the pathogenesis of Paget's bone disease.
Ultrastructural observations in Paget's disease of bone clarify aspects of bone cells in bone tissue and demonstrate the presence of specific intranuclear inclusions composed of microcylinders in the osteoclasts. The morphologic analysis of these structures suggests an analogy with virus material of the measles group. Results obtained using indirect immunofluorescence and immunoperoxidase techniques lend further support to the hypothesis of a viral etiology in Paget's disease of bone.
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