The Deceiving Appearances of Osteoclasts

Osteoclasts are multinucleated cells responsible for the resorption of bone matrix. During the development and growth of a human, they ensure proper contouring of the bones and extension of the medullary spaces to provide for hematopoiesis. In the adult skeleton, osteoclasts ensure remodeling of the bone matrix by means of a process of internal turnover. Skeletal homeostasis is achieved by focal osteoclast-mediated degradation of the bone matrix and osteoblast-mediated formation of new bone matrix without compromising the overall architecture and anatomy of bones. Under ideal conditions, the amount of new bone equals the amount resorbed, with no net change in the volume of bone. As a consequence of remodeling, the molecular composition of the adult skeleton is not static but changes as new bone fills each resorption site. Thus, new carbonato–calcium phosphate crystals replace the highly crystalline, less soluble mature mineral component of bone.1 The renewal of bone matrix is central for the important role bone has in mineral exchange and homeostasis.

Remodeling does not occur in other mineralized tissues, such as dentin and enamel, or in the bone of all species. In marine fishes, for example, osteoclastic bone resorption is limited to tooth-bearing bones or to rare hyperostotic sites.2

The mechanisms of coupling the resorptive and anabolic processes in time and space are incompletely understood but are fundamental to pathologic states in which they are uncoupled. It is lamentable that the rejuvenation of bone matrix is imperfect and that remodeling does not prevent skeletal aging. Peak bone mass is achieved during the third decade of life. Subsequently, bone mass declines and can be associated with an increased risk of fracture. Pathologic bone loss occurs in osteoporosis, certain malignant conditions, inflammation, and immobilization, increasing the risk of fragility fractures.

Osteoclasts attach firmly to bone with a membrane specialization that encloses an acidified zone of resorption (Figure 1Figure 1Schematic Representation of the Structure and Actions of the Osteoclast.). In histologic sections of cortical bone, osteoclasts are found at the apex of tunnels, or cutting cones, which are the areas within which the mineral and organic components of the bone matrix are resorbed. In histologic sections of cancellous bone, osteoclasts are found in lacunae, or erosions, on the surfaces of the trabeculae, which are the so-called rods and plates of bone tissue.

Bone-biopsy specimens are not taken routinely but are useful for monitoring bone physiology in clinical trials. The prevailing system of bone histomorphometrics defines standards of nomenclature for measurements of features of bone tissue in biopsy specimens of transiliac bone.3 The specimens examined in the study reported by Weinstein et al. in this issue of the Journal 4 were available from a previous trial (1994 to 1997) involving alendronate in healthy postmenopausal women.5 In that trial, women were randomly assigned to placebo or oral alendronate at 1, 5, or 10 mg per day for 3 years or 20 mg per day for 2 years followed by placebo for 1 year. Incidentally, in 2008, we would not consider 2 or 3 years to be long-term use of alendronate; many physicians now prescribe alendronate indefinitely for patients who have or are at risk for osteoporosis. In the original trial, a subgroup of women volunteered to undergo transiliac bone biopsies at the end of 3 years of the study. Weinstein et al. resectioned, processed, and examined those valuable specimens for histomorphometry of cancellous osteoclasts and a possible association with the amount of alendronate that was administered.

Several maneuvers were applied to improve the quality of the slides and to avoid processing artifacts. Rigorous criteria were used to identify and enumerate profiles of osteoclasts in 5-mm sections. Weinstein et al. calculated a dose-dependent increase in the number of osteoclasts. Many of these osteoclasts appeared normal, but many others were giant, detached from the bone, or apoptotic. The article by Weinstein et al. provides a unique and detailed description of osteoclasts in women who had been treated with alendronate for 2 or 3 years, as well as new information about the effects of alendronate on the number of osteoclasts and their morphology in human bone. It is known that the drug blocks bone resorption through the inhibition of geranyl pyrophosphate synthase or farnesyl pyrophosphate synthase in osteoclasts and subsequent interference with the osteoclast-membrane specializations that are required for bone resorption.6

Studies in mice, mouse or rabbit cells, and osteoclast surrogates such as murine macrophage-like J774.2 cells in vitro indicate that bisphosphonates reduce the number of osteoclasts by means of apoptosis.7,8 In contrast to those studies that show a reduced number of osteoclasts, the study by Weinstein et al. and other studies referenced by the authors, in which biopsy specimens of human bone were obtained after the administration of bisphosphonates, describe an increase in numbers of seemingly normal and apoptotic osteoclasts. It is possible that the effects of bisphosphonates in humans may not be accurately reproduced in those experimental models because of differences among species in achieved levels of the drug, the sensitivity of the macrophage-like cells, confounding interactions with other cells, or differences among species in the fate of apoptotic cells.

Weinstein and colleagues also raise questions about the seemingly normal appearances of osteoclasts and whether it is possible to estimate bone resorption on the basis of the number of normal-appearing osteoclasts in a bone-biopsy specimen. Treatment with alendronate resulted in decreased biochemical markers of bone turnover in the original trial from which these biopsy specimens were obtained, but the new data suggest discordance between bone resorption and the number of normal-appearing osteoclasts. Accordingly, Weinstein et al. deduced that the normal-appearing osteoclasts were likely to resorb bone poorly. Perhaps new tests or criteria will be developed to identify osteoclasts that have normal function. Is a foamy, vacuole-laden cytoplasm a good index of function? It would be tedious but perhaps worthwhile to determine whether serial 5-mm sections of the osteoclasts in bone-biopsy specimens such as those examined in the present study would reveal abnormalities in other portions of the osteoclasts that appear normal in one random section. Weinstein et al. propose that alendronate acts to promote the accumulation and endurance of apoptotic cells. This is a testable hypothesis. The presence of cells with large numbers of nuclei, even a year after cessation of treatment with alendronate, raises questions about their importance and whether they are not cleared from the bone because of the bisphosphonate that has been stored in the microscopic milieu.

A major and riveting finding is that not all samples from subjects treated with higher doses of alendronate showed the abnormalities in osteoclasts. Only slightly more than half the specimens revealed the abnormalities, and just 30% of the osteoclasts showed evidence of apoptosis. This may be due to the method of sampling, but one wonders about individual clinical features or responses to alendronate that may account for the ostensible absence of abnormal osteoclasts in so many (10 of 17) of the biopsy specimens from two groups with the highest cumulative dose.

The report by Weinstein and colleagues describes dose-dependent increases in the number of normal-appearing and abnormal osteoclasts in biopsy specimens from women who were treated with alendronate. The study was small, with just a few samples generating the conclusions, and with an apparent lack of consistency in those findings within treatment groups. Nevertheless, the observations reported raise testable hypotheses about the effects of bisphosphonates on human osteoclasts. The observations also underscore the need for more information about how these widely used drugs inhibit bone resorption and about the process of apoptosis.

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