- Email: [email protected]
Morphometry of bone resorption — introduction and overview
Abstracts from Bone Morphology
A26
101 ELECTRONMICROSCOPYEXA~WNATIONOFOSTEOBLAST-LIKE CELLS(Ros 17/2.8) AFTER COLORIMETRIC EVALUATION OF THEPROLIFERATION. S. PALLE C. GENTY, C. ALEXANDRE. -7 LBTO-GIP EXERCICE.faculte de mtdecine, 15 rue A. Part. 42023 Saint-Etienne cedex 2, Prance studies were performed after evaluation in by calorimetric method. The staining and destaining procedure presents the advantage of preserving the integrity of the cells and offers the possibility of evaluating growth rate and ultrastructural analysis on the same culture. ROS 17/2.8 were grown on ThermanoxTM coverslips, fixed in Situ for 30 minutes in 2.5% glutaraldehyde, stained with methylene blue (I%, w/v in borate buffer O.lM at pH 8.4). The dye fixed by the cells was released with hydrochloric acid (O.lM) and the obtained blue acid solution was measured in a spectrophotometer to evaluate the number of cells. The cells were. then prepared for routine electron microscopy. They were embedded in Situ by reversing gelatin capsules filled with spurr resin on the top of the coverslip. After performing calorimetric procedure, electron microscopic observations show normal structural aspect of destained cells when compared to control cells. The cell membranes and intracellular organites are well preserved and collagen fibrils are not removed after treatment. The main advantage of such a technic is the possibility to evaluate the cell proliferation and to perform an ultrastructural study on the same culture. In this way, osteoblast cells could be studied throughout the different stages of their differentiation during the growth. Electron microscopic
Situ of cells proliferation
1992, Lexington,
Kentucky
102 HISTOl4ORPHOl4ETRIC OSTEOCYTR-OSTEOBLASTS
Institute
STUDY ON RELATIONSHIPS
C.Palumbo, G. Marotti and M. Ferretti of HumanAnatomy, University of Modena, Italy
Our previous TEM studies showed that preosteocytes and osteocytes have an asymmetrical arborization, the cytoplasmic processes radiating from the vascular side of the cells (osteocyte-VP) being much longer than those radiating from the opposite mineral side (osteocyte-MP). We also found an unexpectedly large number of osteocyteVP coming into contact with the osteoblastic laminae by means of adherens and gap junctions. In the present investigation an attempt has been made to estimate the number of osteocyte-VP which contacts each osteoblast. Analyses were performed with SEM on growing endosteal surfaces of the medullary canal of the tibia in 4 rabbits of different age. The area of each osteoblast was measured on the SEH micrographs by means of an Image Analyzer. The number of osteocyte’ cytoplasmic processes was estimated indirectly by counting the canalicular openings present on the same microscopic fields after the removal of the osteoblasts and the osteoid seam. In all four animals, the surface area of the osteoblasts (OA) was found to vary a great deal, while the density of canalicular openings was fairly uniform. Moreover, while the OA mean value increased significantly with the age of the animals, the density of canalicular openings did not; it would therefore appear that the older the animal and the more flattened the osteoblasts, the greater the number of canaliculi beneath them. Since osteoblast activity has previously been shown to be inversely proportional to their secretory surface, it appears that the less active osteoblasts should contact a greater number of osteocyte cytoplasmic processes. These findings suggest that osteocytes might play an important role in modulating osteoblast activity and in recruiting osteoblasts that transform into osteocytes, possibly by means of inhibitory signals transmitted via gap junctions.
103
104
STRUCTURAL VARIABLES IN POSTMENOPAUSAL WOMEN: COMPARISON AMONG NORMAL WOMEN AND PATIENTS WITH OSTEOPOROSIS OR PRIMARY HYPERPARATHYROIDISM. M&&&B RW Mellish, M Schnitaer. J Nirves, F Corn-ran, RR Recker, D Kimmel, SJ Silverberg, E Shane, JP Bilerikian, R Lindsay, DW Oempster. Helen Hayes Hospital, W Haverstraw, NY, Oepts Med. Pherm and Pathol, Columbia UniversltY, NY, snd Creighton University, Omaha, NE. This study compares the microarchitecture of bone in two groups of postmenopausal (PM) women with osteoporosis fOPI and primary hyperperathyroidism (PHPT) to that of normal PM women (N) matched by years PM. Histomorphometric variables of cancellous bone structure : bone volume @V/TV), trabacular thickness fTb.Th), number lTb.Nl and separation fTb.Sp), node fN.Nd) and terminus lN.Tm) number, total strut length fTSL1, node to node lNd.Nd), node to terminus lNd.Tm), terminus to terminus lTm.Tml strut lengths and node to terminus rstio fN.Nd/N.Tm) in 17 women with untreated primary OP (65.1~1.4 yrs, 15.8~23 yrsPM), and 15 women with PHPT (573~2.1 yrs, 10.721.8 yrsPM1 were compared to those of 31 N 159.821.4 yrs, 134~1.5 yrsPM). Variabies expressing the smount of bone (TSL, M//TV end Tb.N) were lower in OP fpsO.0031 but were higher in PHPT fNS) as compared to N. TbSp was higher fp-zO.001) in OP. and lower in F’HPT (NM, while Tb.Th was unchangedin both groups. Variables expressing connectivity fN.Nd, Nd.Nd, N.NdM.Tm) were lower in OP (p~O.003) but hisher in PHPT INS to p=O.O4). Vsrisbles expressing loss of connectivity (N.Tm, Nd.Tm, Tm.Tm), were unchanged or higher in OP fNS to p=O.Ol) but lower in PHPT fp=O.O9-G.O1). BV/TV and Tb.N correlated positively with varisbles of connectivity fN.Nd, Nd.Nd. and N.NdM.Tm) in all three Oroups although the strongest correlations were in N. In conclusion: (1) in 3 groups of PM women, structursl variables reflecting the amount of bone were lower in OP and higher in PHPT than N (21 variables reflecting connectivity were lower in OP. higher in PHPT than N whereas those reflecting loss of connactivfty were unchanged or hipher in OP but lower in PHPT (3) these findincts contrast bone loss and architectural deterioration in OP with conservation of cancallous bone structure in PHPT.
MORPHOMETRY OF BONE RESORPTION - INTRODUCTION AND OVERVIEW A. M. Parfitt, Henry Ford Hospital, Detroit Bone resorption is a more difficult process to study than bone formation for many reasons. It is more rapid, so that its traces are less prevalent and sampling problems are more serious. Resorption must be inferred from what has been removed rather than from what has been added; there is no marker analogous to tetracycline, and small erosions may be difficult to distinguish from minor surface irregularities. There is disagreement concerning mononuclear osteoclasts -their existence, identification, prevalence and contribution to bone remodeling. Consequently, both within and between reader variability are greater than for indices of formation. Erosion and resorption rates can only be measured indirectly. Osteoclasts are motile, and their domain of activity is larger than their current area of contact with bone, but little is known about their pathways in three-dimensions. The usual fate of an erosion cavity is to be refilled with new bone; if the osteoblasts needed for this are late in arrival, the life span of the untilled cavity (reversal phase) is prolonged, and the surface extent of such cavities will increase. In both cortical and cancellous bone, the frequency distribution of completed cavity depth is bimodal. The shallower cavities may never be refilled, but remain as quasipermanent defects on the surfaces of trabeculae and haversian canals. The deeper cavities will eventually be at least partly refilled. Completed cavity depth is the most important aspect of resorption from several standpoints - qualitative diagnosis, focal bone balance, and the indirect measurement of erosion and resorption rates. A sustained increase in erosion depth, with intratrabecular tunneling, is the most distinctive feature of osteitis fibrosa, and a transient increase during early menopause is the most likely explanation for trabecular plate perforation and consequent loss of connectivity. Unfortunately, there are substantial differences between methods of measurement, large enough to alter conclusions about the cellular mechanisms of bone loss.
A26
101 ELECTRONMICROSCOPYEXA~WNATIONOFOSTEOBLAST-LIKE CELLS(Ros 17/2.8) AFTER COLORIMETRIC EVALUATION OF THEPROLIFERATION. S. PALLE C. GENTY, C. ALEXANDRE. -7 LBTO-GIP EXERCICE.faculte de mtdecine, 15 rue A. Part. 42023 Saint-Etienne cedex 2, Prance studies were performed after evaluation in by calorimetric method. The staining and destaining procedure presents the advantage of preserving the integrity of the cells and offers the possibility of evaluating growth rate and ultrastructural analysis on the same culture. ROS 17/2.8 were grown on ThermanoxTM coverslips, fixed in Situ for 30 minutes in 2.5% glutaraldehyde, stained with methylene blue (I%, w/v in borate buffer O.lM at pH 8.4). The dye fixed by the cells was released with hydrochloric acid (O.lM) and the obtained blue acid solution was measured in a spectrophotometer to evaluate the number of cells. The cells were. then prepared for routine electron microscopy. They were embedded in Situ by reversing gelatin capsules filled with spurr resin on the top of the coverslip. After performing calorimetric procedure, electron microscopic observations show normal structural aspect of destained cells when compared to control cells. The cell membranes and intracellular organites are well preserved and collagen fibrils are not removed after treatment. The main advantage of such a technic is the possibility to evaluate the cell proliferation and to perform an ultrastructural study on the same culture. In this way, osteoblast cells could be studied throughout the different stages of their differentiation during the growth. Electron microscopic
Situ of cells proliferation
1992, Lexington,
Kentucky
102 HISTOl4ORPHOl4ETRIC OSTEOCYTR-OSTEOBLASTS
Institute
STUDY ON RELATIONSHIPS
C.Palumbo, G. Marotti and M. Ferretti of HumanAnatomy, University of Modena, Italy
Our previous TEM studies showed that preosteocytes and osteocytes have an asymmetrical arborization, the cytoplasmic processes radiating from the vascular side of the cells (osteocyte-VP) being much longer than those radiating from the opposite mineral side (osteocyte-MP). We also found an unexpectedly large number of osteocyteVP coming into contact with the osteoblastic laminae by means of adherens and gap junctions. In the present investigation an attempt has been made to estimate the number of osteocyte-VP which contacts each osteoblast. Analyses were performed with SEM on growing endosteal surfaces of the medullary canal of the tibia in 4 rabbits of different age. The area of each osteoblast was measured on the SEH micrographs by means of an Image Analyzer. The number of osteocyte’ cytoplasmic processes was estimated indirectly by counting the canalicular openings present on the same microscopic fields after the removal of the osteoblasts and the osteoid seam. In all four animals, the surface area of the osteoblasts (OA) was found to vary a great deal, while the density of canalicular openings was fairly uniform. Moreover, while the OA mean value increased significantly with the age of the animals, the density of canalicular openings did not; it would therefore appear that the older the animal and the more flattened the osteoblasts, the greater the number of canaliculi beneath them. Since osteoblast activity has previously been shown to be inversely proportional to their secretory surface, it appears that the less active osteoblasts should contact a greater number of osteocyte cytoplasmic processes. These findings suggest that osteocytes might play an important role in modulating osteoblast activity and in recruiting osteoblasts that transform into osteocytes, possibly by means of inhibitory signals transmitted via gap junctions.
103
104
STRUCTURAL VARIABLES IN POSTMENOPAUSAL WOMEN: COMPARISON AMONG NORMAL WOMEN AND PATIENTS WITH OSTEOPOROSIS OR PRIMARY HYPERPARATHYROIDISM. M&&&B RW Mellish, M Schnitaer. J Nirves, F Corn-ran, RR Recker, D Kimmel, SJ Silverberg, E Shane, JP Bilerikian, R Lindsay, DW Oempster. Helen Hayes Hospital, W Haverstraw, NY, Oepts Med. Pherm and Pathol, Columbia UniversltY, NY, snd Creighton University, Omaha, NE. This study compares the microarchitecture of bone in two groups of postmenopausal (PM) women with osteoporosis fOPI and primary hyperperathyroidism (PHPT) to that of normal PM women (N) matched by years PM. Histomorphometric variables of cancellous bone structure : bone volume @V/TV), trabacular thickness fTb.Th), number lTb.Nl and separation fTb.Sp), node fN.Nd) and terminus lN.Tm) number, total strut length fTSL1, node to node lNd.Nd), node to terminus lNd.Tm), terminus to terminus lTm.Tml strut lengths and node to terminus rstio fN.Nd/N.Tm) in 17 women with untreated primary OP (65.1~1.4 yrs, 15.8~23 yrsPM), and 15 women with PHPT (573~2.1 yrs, 10.721.8 yrsPM1 were compared to those of 31 N 159.821.4 yrs, 134~1.5 yrsPM). Variabies expressing the smount of bone (TSL, M//TV end Tb.N) were lower in OP fpsO.0031 but were higher in PHPT fNS) as compared to N. TbSp was higher fp-zO.001) in OP. and lower in F’HPT (NM, while Tb.Th was unchangedin both groups. Variables expressing connectivity fN.Nd, Nd.Nd, N.NdM.Tm) were lower in OP (p~O.003) but hisher in PHPT INS to p=O.O4). Vsrisbles expressing loss of connectivity (N.Tm, Nd.Tm, Tm.Tm), were unchanged or higher in OP fNS to p=O.Ol) but lower in PHPT fp=O.O9-G.O1). BV/TV and Tb.N correlated positively with varisbles of connectivity fN.Nd, Nd.Nd. and N.NdM.Tm) in all three Oroups although the strongest correlations were in N. In conclusion: (1) in 3 groups of PM women, structursl variables reflecting the amount of bone were lower in OP and higher in PHPT than N (21 variables reflecting connectivity were lower in OP. higher in PHPT than N whereas those reflecting loss of connactivfty were unchanged or hipher in OP but lower in PHPT (3) these findincts contrast bone loss and architectural deterioration in OP with conservation of cancallous bone structure in PHPT.
MORPHOMETRY OF BONE RESORPTION - INTRODUCTION AND OVERVIEW A. M. Parfitt, Henry Ford Hospital, Detroit Bone resorption is a more difficult process to study than bone formation for many reasons. It is more rapid, so that its traces are less prevalent and sampling problems are more serious. Resorption must be inferred from what has been removed rather than from what has been added; there is no marker analogous to tetracycline, and small erosions may be difficult to distinguish from minor surface irregularities. There is disagreement concerning mononuclear osteoclasts -their existence, identification, prevalence and contribution to bone remodeling. Consequently, both within and between reader variability are greater than for indices of formation. Erosion and resorption rates can only be measured indirectly. Osteoclasts are motile, and their domain of activity is larger than their current area of contact with bone, but little is known about their pathways in three-dimensions. The usual fate of an erosion cavity is to be refilled with new bone; if the osteoblasts needed for this are late in arrival, the life span of the untilled cavity (reversal phase) is prolonged, and the surface extent of such cavities will increase. In both cortical and cancellous bone, the frequency distribution of completed cavity depth is bimodal. The shallower cavities may never be refilled, but remain as quasipermanent defects on the surfaces of trabeculae and haversian canals. The deeper cavities will eventually be at least partly refilled. Completed cavity depth is the most important aspect of resorption from several standpoints - qualitative diagnosis, focal bone balance, and the indirect measurement of erosion and resorption rates. A sustained increase in erosion depth, with intratrabecular tunneling, is the most distinctive feature of osteitis fibrosa, and a transient increase during early menopause is the most likely explanation for trabecular plate perforation and consequent loss of connectivity. Unfortunately, there are substantial differences between methods of measurement, large enough to alter conclusions about the cellular mechanisms of bone loss.