Interaction among Calcium Diet Content, PTH (1-34) Treatment and Balance of Bone Homeostasis in Rat Model: The Trabecular Bone as Keystone.

The present study is the second step (concerning normal diet restoration) of the our previous study (concerning the calcium-free diet) to determine whether normal diet restoration, with/without concomitant PTH (1-34) administration, can influence amounts and deposition sites of the total bone mass. Histomorphometric evaluations and immunohistochemical analysis for Sclerostin expression were conducted on the vertebral bodies and femurs in the rat model. The final goals are (i) to define timing and manners of bone mass changes when calcium is restored to the diet, (ii) to analyze the different involvement of the two bony architectures having different metabolism (i.e., trabecular versus cortical bone), and (iii) to verify the eventual role of PTH (1-34) administration. Results evidenced the greater involvement of the trabecular bone with respect to the cortical bone, in response to different levels of calcium content in the diet, and the effect of PTH, mostly in the recovery of trabecular bony architecture. The main findings emerged from the present study are (i) the importance of the interplay between mineral homeostasis and skeletal homeostasis in modulating and guiding bone's response to dietary/metabolic alterations and (ii) the evidence that the more involved bony architecture is the trabecular bone, the most susceptible to the dynamical balance of the two homeostases.

PTH(1-34) effects on repairing experimentally drilled holes in rat femur: novel aspects - qualitative vs. quantitative improvement of osteogenesis.

The timetable of effects on bone repair of the active fraction-parathyroid hormone, PTH(1-34), was analytically investigated from the morphometric viewpoint in 3-month-old male Sprague-Dawley rats, whose femurs were drilled at mid-diaphyseal level (transcortical holes). The animals were divided into groups with/without PTH(1-34) administration, and sacrificed at different times (10, 28, 45 days after surgery). The observations reported here need to be framed in the context of our previous investigations regarding bone histogenesis (Ferretti et al. Anat Embryol. 2002; 206: 21-29) in which we demonstrated the occurrence of two successive bone-forming processes during both skeletal organogenesis and bone repair, i.e. static and dynamic osteogenesis: the former (due to stationary osteoblasts, haphazardly grouped in cords) producing preliminary bad quality trabecular bone, the latter (due to typical polarized osteoblasts organized in ordered movable laminae) producing mechanically valid bone tissue. The primary function of static osteogenesis is to provide a rigid scaffold containing osteocytes (i.e. mechano-sensors) for osteoblast laminae acting in dynamic osteogenesis. In the present work, histomorphometric analysis revealed that, already 10 days after drilling, despite the holes being temporarily filled by the same amount of newly formed trabecular bone by static osteogenesis independently of the treatment, the extent of the surface of movable osteoblast-laminae (covering the trabecular surface) was statistically higher in animals submitted to PTH(1-34) administration than in control ones; this datum strongly suggests the effect of PTH(1-34) alone in anticipating the occurrence of dynamic osteogenesis involved in the production of good quality bone (with more ordered collagen texture) more suitable for loading. This study could be crucial in further translational clinical research in humans for defining the best therapeutic strategies to be applied in recovering severe skeletal lesions, particularly as regards the time of PTH(1-34) administration.

Expression and functional proteomic analyses of osteocytes from Xenopus laevis tested under mechanical stress conditions: preliminary observations on an appropriate new animal model.

Hitherto, the role of the osteocyte as transducer of mechanical stimuli into biological signals is far from settled. In this study, we used an appropriate model represented by the cortex of Xenopus laevis long bone diaphysis lacking (unlike the mammalian one) of vascular structures and containing only osteocytes inside the bone matrix. These structural features allow any change of protein profile that might be observed upon different experimental conditions, such as bone adaptation to stress/mechanical loading, to be ascribed specifically to osteocytes. The study was conducted by combining ultrastructural observations and two-dimensional electrophoresis for proteomic analysis. The osteocyte population was extracted from long bones of lower limbs of amphibian skeletons after different protocols (free and forced swimming). The experiments were performed on 210 frogs subdivided into five trials, each including free swimming frogs (controls) and frogs submitted to forced swimming (stressed). The stressed groups were obliged to swim (on movable spheres covering the bottom of a pool on a vibrating plate) continuously for 8 h, and killed 24 h later along with the control groups. Long bones free of soft tissues (periosteum, endosteum and bone marrow), as well as muscles of posterior limbs, were processed and analyzed for proteins differentially expressed or phosphorylated between the two sample groups. The comparative analysis showed that protein phosphorylation profiles differ between control and stressed groups. In particular, we found in long bones of stressed samples that both Erk1/2 and Akt are hyperphosphorylated; moreover, the different phosphorylation of putative Akt substrates (recognized by specific Akt phosphosubstrates-antibody) in stressed vs. control samples clearly demonstrated that Akt signaling is boosted by forced swimming (leading to an increase of mechanical stress) of amphibian long bones. In parallel, we found in posterior limb muscles that the expression of heat shock protein HSP27 and HSP70 stress markers increased upon the forced swimming condition. Because the cortexes of frog long bones are characterized by the presence of only osteocytes, all our results establish the suitability of the X. laevis animal model to study the bone response to stress conditions mediated by this cell type and pave the way for further analysis of the signaling pathways involved in these signal transduction mechanisms.

Morphology and cell biology - two sides of the same coin: Importance of morphology in choosing Cre experimental models for targeting osteoblasts vs osteocytes.

The present letter to editor comments the manuscript entitled "Targeting osteocytes vs osteoblasts" by Y. Kitase and M. Prideaux, where we underlie the importance morphology in choosing Cre experimental models for targeting osteoblasts vs osteocytes.

In Vitro and in Vivo Bone Remodeling Models: Complementary Pieces of the same Puzzle.

The present letter to editor comments on the manuscript entitled "Assembling the Puzzle Pieces. Insights for in Vitro Bone Remodeling" by O. Krasnova & I. Neganova; in this context, we underlie the importance of in vivo models to corroborate in vitro bone remodeling systems.

Unexpected Absence of Skeletal Responses to Dietary Magnesium Depletion: Basis for Future Perspectives?

It's known that a magnesium (Mg)-deficient diet is associated with an increased risk of osteoporosis. The aim of this work is to investigate, by a histological approach, the effects of a Mg-deprived diet on the bone of 8-weeks-old C57BL/6J male mice. Treated and control mice were supplied with a Mg-deprived or normal diet for 8 weeks, respectively. Body weight, serum Mg concentration, expression of kidney magnesiotropic genes, and histomorphometry on L5 vertebrae, femurs, and tibiae were evaluated. Body weight gain and serum Mg concentration were significantly reduced, while a trend toward increase was found in gene expression in mice receiving the Mg-deficient diet, suggesting the onset of an adaptive response to Mg depletion. Histomorphometric parameters on the amount of trabecular and cortical bone, number of osteoclasts, and thickness of the growth plate in femoral distal and tibial proximal metaphyses did not differ between groups; these findings partially differ from most data present in the literature showing that animals fed a Mg-deprived diet develop bone loss and may be only in part explained by differences among the experimental protocols. However, the unexpected findings we recorded on bones could be attributed to genetic differences that may have developed after multiple generations of inbreeding.

Striated muscle fiber apoptosis after experimental tendon lesion in a rat model.

Tendon lesions induce muscular atrophy, the nature of which has not yet been clearly related to lesion etiology and entity. In the present study, tendon and muscle alterations were assessed after experimental tendon lesion of the Infraspinatus muscle in young rats. The consequences of lesions differed on the basis of both extension and injured tissue vascularization, that is apoptosis and/or degeneration, differing mainly by energy demands: apoptosis requires high energy levels (proportional to vascular supply), but degeneration does not. It is well known that tendons are poorly supplied with blood compared with muscular masses, which are abundantly vascularized. Five weeks after tendon surgical section, tendon/muscle samples were taken for TUNEL and transmission electron microscopy. The structural results reported here identified different tendon/muscle alterations: degeneration of tendon without signs of apoptosis, and atrophy of muscle fibers due only to apoptosis. This led to the formulation of the following hypothetical sequence of events: a tendon lesion, not recovering quickly due to the poor tendon blood supply, results in degeneration of the injured tendon, which, in turn, induces a partial disuse of the muscle mass, which consequently atrophies (proportionally to the severity of tendon lesion) by striated muscular fiber apoptosis. The authors suggest that the different behavior of the two tissues depends on the marked difference in their vascularization.

Osteocyte apoptosis and absence of bone remodeling in human auditory ossicles and scleral ossicles of lower vertebrates: a mere coincidence or linked processes?

Considering the pivotal role as bone mechanosensors ascribed to osteocytes in bone adaptation to mechanical strains, the present study analyzed whether a correlation exists between osteocyte apoptosis and bone remodeling in peculiar bones, such as human auditory ossicles and scleral ossicles of lower vertebrates, which have been shown to undergo substantial osteocyte death and trivial or no bone turnover after cessation of growth. The investigation was performed with a morphological approach under LM (by means of an in situ end-labeling technique) and TEM. The results show that a large amount of osteocyte apoptosis takes place in both auditory and scleral ossicles after they reach their final size. Additionally, no morphological signs of bone remodeling were observed. These facts suggest that (1) bone remodeling is not necessarily triggered by osteocyte death, at least in these ossicles, and (2) bone remodeling does not need to mechanically adapt auditory and scleral ossicles since they appear to be continuously submitted to stereotyped stresses and strains; on the contrary, during the resorption phase, bone remodeling might severely impair the mechanical resistance of extremely small bony segments. Thus, osteocyte apoptosis could represent a programmed process devoted to make stable, when needed, bone structure and mechanical resistance.

Effects of different doses of ferutinin on bone formation/resorption in ovariectomized rats.

This study analyzes the effects of different doses of ferutinin on bone loss caused by estrogen deficiency in ovariectomized rats, in comparison with estradiol benzoate. Thirty female Sprague-Dawley rats were ovariectomized and treated for 30 days from the day after ovariectomy. Static/dynamic histomorphometric analyses were performed on trabecular and cortical bone of lumbar vertebrae and femurs. Very low weight increments were recorded only in all F-OVX groups, with respect to the others. Although the great differences in weight, that could imply a decrease of bone mass in F-OVX groups compared to the control ovariectomized group (C-OVX), trabecular bone in lumbar vertebrae did not show significant differences, suggesting that ferutinin, opposing estrogen deficiency, inhibits bone resorption. Newly formed cortical bone was always low in all F-OVX groups and high in C-OVX, suggesting that it is mainly devoted in answering mechanical demands. In contrast, in distal femoral metaphyses, trabecular bone was reduced and the number of osteoclasts was increased in C-OVX with respect to all other groups, suggesting that it is mainly devoted in answering metabolic demands; moreover, ferutinin dose of 2 mg/kg seemed to be more effective than the lower doses used and estrogens, particularly in those skeletal regions with higher metabolic activity. Our results suggest that the role of ferutinin in preventing osteoporosis caused by estrogen deficiency is expressed in decreasing bone erosion; moreover, in all F-OVX groups bone turnover is very low and seems correlated to the trivial body weight increase, which, in turn, depends on ferutinin treatment.

The Osteocyte: From "Prisoner" to "Orchestrator".

Osteocytes are the most abundant bone cells, entrapped inside the mineralized bone matrix. They derive from osteoblasts through a complex series of morpho-functional modifications; such modifications not only concern the cell shape (from prismatic to dendritic) and location (along the vascular bone surfaces or enclosed inside the lacuno-canalicular cavities, respectively) but also their role in bone processes (secretion/mineralization of preosseous matrix and/or regulation of bone remodeling). Osteocytes are connected with each other by means of different types of junctions, among which the gap junctions enable osteocytes inside the matrix to act in a neuronal-like manner, as a functional syncytium together with the cells placed on the vascular bone surfaces (osteoblasts or bone lining cells), the stromal cells and the endothelial cells, i.e., the bone basic cellular system (BBCS). Within the BBCS, osteocytes can communicate in two ways: by means of volume transmission and wiring transmission, depending on the type of signals (metabolic or mechanical, respectively) received and/or to be forwarded. The capability of osteocytes in maintaining skeletal and mineral homeostasis is due to the fact that it acts as a mechano-sensor, able to transduce mechanical strains into biological signals and to trigger/modulate the bone remodeling, also because of the relevant role of sclerostin secreted by osteocytes, thus regulating different bone cell signaling pathways. The authors want to emphasize that the present review is centered on the morphological aspects of the osteocytes that clearly explain their functional implications and their role as bone orchestrators.

Influence of ferutinin on bone metabolism in ovariectomized rats. II: role in recovering osteoporosis.

The aim of the present investigation, which represents an extension of a previous study, was to investigate the effect of ferutinin in recovering severe osteoporosis due to estrogen deficiency after rat ovariectomy and to compare phytoestrogen effects with those of estrogens commonly used in hormone replacement therapy (HRT) by women with postmenopausal osteoporosis. The animal model used was the Sprague-Dawley ovariectomized rat. Ferutinin was orally administered (2 mg kg(-1) per day) for 30 or 60 days starting from 2 months after ovariectomy (i.e. when osteoporosis was clearly evident) and its effects were compared with those of estradiol benzoate (1.5 microg per rat twice a week, subcutaneously injected) vs. vehicle-treated ovariectomized (OVX) and sham-operated (SHAM) rats. Histomorphometric analyses were performed on trabecular bone of lumbar vertebrae (4th and 5th) and distal femoral epiphysis, as well as on cortical bone of femoral diaphysis. Bone histomorphometric analyses showed that ferutinin seems to display the same effects on bone mass recorded with estradiol benzoate, thus suggesting that it could enhance the recovery of bone loss due to severe estrogen deficiency in OVX rats. On this basis, the authors propose listing ferutinin among the substances representing a potential alternative for the treatment of postmenopausal osteoporosis, which occurs as a result of estrogen deficiency.

WISP-2 expression induced by Teriparatide treatment affects in vitro osteoblast differentiation and improves in vivo osteogenesis.

The Osteocyte, recognized as a major orchestrator of osteoblast and osteoclast activity, is the most important key player during bone remodeling processes. Imbalances occurring during bone remodeling, caused by hormone perturbations or by mechanical loading alterations, can induce bone pathologies such as osteoporosis. Recently, the active fraction of parathormone, PTH (1-34) or Teriparatide (TPTD), was chosen as election treatment for osteoporosis. The effect of such therapy is dependent on the temporal manner of administration. The molecular reasons why the type of administration regimen is so critical for the fate of bone remodeling are numerous and not yet well known. Our study attempts to analyze diverse signaling pathways directly activated in osteocytes upon TPTD treatment. By means of gene array analysis, we found many molecules upregulated or downregulated in osteocytes. Later, we paid attention to Wisp-2, a protein involved in the Wnt pathway, that is secreted by MLO-Y4 cells and increases upon TPTD treatment and that is able to positively influence the early phases of osteogenic differentiation. We also confirmed the pro osteogenic property of Wisp-2 during mesenchymal stem cell differentiation into the preliminary osteoblast phenotype. The same results were confirmed with an in vivo approach confirming a remarkable Wisp-2 expression in metaphyseal trabecular bone. These results highlighted the anabolic roles unrolled by osteocytes in controlling the action of neighboring cells, suggesting that the perturbation of certain signaling cascades, such as the Wnt pathway, is crucial for the positive regulation of bone formation.

Leptin increases growth of primary ossification centers in fetal mice.

The effect of peripheral leptin on fetal primary ossification centers during the early phases of bone histogenesis was investigated by administration of leptin to pregnant mice. Fourteen pregnant mice were divided into two groups. The treated pregnant group was subcutaneously injected in the intrascapular region with supraphysiologic doses (2 mg kg(-1)) of leptin (Vinci Biochem, Firenze, Italy) in a volume of 0.1 mL per 10 g body weight, at the 7th, 9th and 11th day of gestation. The control group was treated with physiological solution in the same manner and same times as the treated group. The new-born mice were killed 1 day after birth and the primary ossification centers were stained with Alizarin Red S after diaphanizing the soft tissues in 1% potassium hydroxide. The development of both endochondral and intramembranous ossification centers was morphometrically analysed in long bones. The results showed that the ossification centers of mice born by mothers treated with leptin grow more rapidly in both length and cross-sectional area compared with mice born by the untreated mothers. As the development of long bones depends on endochondral ossification occurring at proximal and distal epiphyseal plates as well as on intramembranous ossification along the periosteal surface, it appears that leptin activates the differentiation and proliferation of both chondrocytes and osteoblasts. The role of leptin as a growth factor of cartilage and bone is discussed in the light of the data reported in the literature.

Influence of ferutinin on bone metabolism in ovariectomized rats. I: role in preventing osteoporosis.

Phytoestrogens play a role in maintaining bone mass in the post-menopausal period for their putative function as osteoprotective agents. The aim of the present study was to investigate the influence of Ferutinin, a phytoestrogen found in the plants of Ferula genus, on bone loss in ovariectomized rats. Such an animal model can simulate the various clinical syndromes deriving from osteoporosis. The effect of the daily oral administration of ferutinin to ovariectomized rats (dosed at 2 mg/kg per day for 30 and 60 days) was compared to that of estradiol benzoate (subcutaneously administered at the dose of 1.5 microg/rat twice a week). After the sacrifice, histomorphometrical analyses were performed on trabecular bone of L4-L5 vertebrae and distal femoral metaphysis, as well as on cortical bone of femoral diaphysis; biochemical parameters (bone mineral components and markers) were also evaluated from the rat serum. The histomorphometrical analyses of trabecular and cortical bone from lumbar vertebrae and femur showed that ferutinin has the same antiosteoporotic effect of estradiol benzoate on bone mass, and in some cases is even stronger. This fact suggests that it could prevent osteoporosis caused by severe estrogen deficiency in ovariectomized rats. The possibility of using ferutinin as an alternative to the commonly employed hormonal replacing therapy in post-menopausal women is discussed.

Two peculiar conditions following a coma: a clinical case of heterotopic ossification concomitant with keloid formation.

The etiology and formation pattern of heterotopic ossifications (HO) are still unknown. They occur in soft tissues in which bone does not normally form, near one or more proximal joints. In this article, the authors report a peculiar case of a 31-year-old patient affected by scapulo-humeral ankylosis that occurred about 6 months after a coma, in which two unusual concomitant conditions were observed: HO formation in the scapulo-humeral region and the development of keloids during wound repair. The scapulo-humeral ankylosis was resolved surgically with the removal of the HO, which was then studied morphologically to understand its formation pattern. By light microscopy and transmission electron microscopy, it was observed that heterotopic bone displays the normal microscopic structure of primary bone, in which two types of bone tissue were recognized, i.e., woven-fibered bone, deeply located and produced first, and lamellar bone. This suggests that the pattern of HO formation retraces the ontogenetic steps that normally occur during intramembranous ossification. The authors also discuss the peculiar concomitance of HO formation and keloid development, speculating that, although they are different conditions localized in dissimilar regions, they might be hypothetically triggered by a common event, such as the release of factors likely issued during the coma status.

Role of Osteocytes in Myeloma Bone Disease: Anti-sclerostin Antibody as New Therapeutic Strategy.

Osteocytes are terminally differentiated cells of the osteoblast lineage. They are involved in the regulation of bone remodeling by increasing osteoclast formation or decreasing bone formation by the secretion of the osteoblast inhibitor sclerostin. Monoclonal antibody anti-sclerostin, Romosozumab, has been developed and tested in clinical trials in patients with osteoporosis. In the last years, the role of osteocytes in the development of osteolytic bone lesions that occurs in multiple myeloma, have been underlined. Myeloma cells increase osteocyte death through the up-regulation of both apoptosis and autophagy that, in turn, triggers osteoclast formation, and activity. When compared to healthy controls, myeloma patients with bone disease have higher osteocyte cell death, but the treatment with proteasome inhibitor bortezomib has been shown to maintain osteocyte viability. In preclinical mouse models of multiple myeloma, treatment with blocking anti-sclerostin antibody increased osteoblast numbers and bone formation rate reducing osteolytic bone lesions. Moreover, the combination of anti-sclerostin antibody and the osteoclast inhibitor zoledronic acid increased bone mass and fracture resistance synergistically. However, anti-sclerostin antibody did not affect tumor burden in vivo or the efficacy of anti-myeloma drugs in vitro. Nevertheless, the combination therapy of anti-sclerostin antibody and the proteasome inhibitor carfilzomib, displayed potent anti-myeloma activity as well as positive effects on bone disease in vivo. In conclusion, all these data suggest that osteocytes are involved in myeloma bone disease and may be considered a novel target for the use of antibody-mediated anti-sclerostin therapy also in multiple myeloma patients.

Structural and ultrastructural analyses of bone regeneration in rabbit cranial osteotomy: Piezosurgery versus traditional osteotomes.

Clinical advantages of piezosurgery have been already proved. However, few investigations have focused on the dynamics of bone healing. The aim of this study was to evaluate, in adult rabbits, bone regeneration after cranial linear osteotomies with two piezoelectrical devices (Piezosurgery® Medical - PM and Piezosurgery® Plus - PP), comparing them with conventional rotary osteotomes (RO). PP was characterized by an output power three times higher than PM. Fifteen days after surgery, histomorphometric analyses showed that the osteotomy gap produced with PM and PP was about half the size of that produced by RO, and in a more advanced stage of recovery. Values of regenerated bone area with respect to the total osteotomy area were about double in PM and PP samples compared with RO ones, while the number of TRAP-positive (tartrate-resistant acid phosphatase positive) osteoclasts per linear surface showed a significant increase, suggesting greater bone remodelling. Under scanning electron microscopy, regenerated bone displayed higher cell density and less mineralized matrix compared with pre-existent bone for all devices used. Nanoindentation tests showed no changes in elastic modulus. In conclusion, PM/PP osteotomies can be considered equivalent to each other, and result in more rapid healing compared with those using RO.

Biocompatibility Analyses of Al2O3-Treated Titanium Plates Tested with Osteocyte and Fibroblast Cell Lines.

Osseointegration of a titanium implant is still an issue in dental/orthopedic implants durable over time. The good integration of these implants is mainly due to their surface and topography. We obtained an innovative titanium surface by shooting different-in-size particles of Al2O3 against the titanium scaffolds which seems to be ideal for bone integration. To corroborate that, we used two different cell lines: MLO-Y4 (murine osteocytes) and 293 (human fibroblasts) and tested the titanium scaffolds untreated and treated (i.e., Al2O3 shot-peened titanium surfaces). Distribution, density, and expression of adhesion molecules (fibronectin and vitronectin) were evaluated under scanning electron microscope (SEM) and confocal microscope (CM). DAPI and fluorochrome-conjugated antibodies were used to highlight nuclei, fibronectin, and vitronectin, under CM; cell distribution was analyzed after gold-palladium sputtering of samples by SEM. The engineered biomaterial surfaces showed under SEM irregular morphology displaying variously-shaped spicules. Both SEM and CM observations showed better outcome in terms of cell adhesion and distribution in treated titanium surfaces with respect to the untreated ones. The results obtained clearly showed that this kind of surface-treated titanium, used to manufacture devices for dental implantology: (i) is very suitable for cell colonization, essential prerequisite for the best osseointegration, and (ii) represents an excellent solution for the development of further engineered implants with the target to obtain recovery of stable dental function over time.

Morphological Study: Ultrastructural Aspects of Articular Cartilage and Subchondral Bone in Patients Affected by Post-Traumatic Shoulder Instability.

Post-traumatic shoulder instability is a frequent condition in active population, representing one of most disabling pathologies, due to altered balance involving joints. No data are so far available on early ultrastructural osteo-chondral damages, associated with the onset of invalidating pathologies, like osteoarthritis-OA. Biopsies of glenoid articular cartilage and sub-chondral bone were taken from 10 adult patients underwent arthroscopic stabilization. Observations were performed under Transmission Electron Microscopy-TEM in tangential, arcuate and radial layers of the articular cartilage and in the sub-chondral bone. In tangential and arcuate layers chondrocytes display normal and very well preserved ultrastructure, probably due to the synovial liquid supply; otherwise, throughout the radial layer (un-calcified and calcified) chondrocytes show various degrees of degeneration; occasionally, in the radial layer evidences of apoptosis/autophagy were also observed. Concerning sub-chondral bone, osteocytes next to the calcified cartilage also show signs of degeneration, while osteocytes farther from the osteo-chondral border display normal ultrastructure, probably due to the bone vascular supply. The ultrastructural features of the osteo-chondral complex are not age-dependent. This study represents the first complete ultrastructural investigation of the articular osteo-chondral complex in shoulder instability, evaluating the state of preservation/viability of both chondrocytes and osteocytes throughout the successive layers of articular cartilage and sub-chondral bone. Preliminary observations here collected represent the morphological basis for further deepening of pathogenesis related to shoulder instability, enhancing the relationship between cell shape and microenvironment; in particular, they could be useful in understanding if the early surgical treatment in shoulder instability could avoid the onset of OA. Anat Rec, 300:1208-1218, 2017. © 2016 Wiley Periodicals, Inc.

Does static precede dynamic osteogenesis in endochondral ossification as occurs in intramembranous ossification?

Endochondral ossification takes place with calcified cartilage cores providing a rigid scaffold for new bone formation. Intramembranous ossification begins in connective tissue and new bone formed by a process of static ossification (SO) followed by dynamic ossification (DO) as previously described. The aim of the present study was to determine if the process of endochondral ossification is similar to that of intramembranous ossification with both a static and a dynamic phase of osteogenesis. Endochondral ossification centers of the tibiae and humeri of newborn and young growing rabbits were studied by light and transmission electron microscopy. The observations clearly showed that in endochondral ossification, the calcified trabeculae appeared to be lined first by osteoclasts. The osteoclasts were then replaced by flattened cells (likely cells of the reversal phase) and finally by irregularly arranged osteoblastic laminae, typical of DO. This cellular sequence did not include osteoblasts seen in the phase of SO. These findings clearly support our working hypothesis that SO only forms in soft tissues to provide a rigid framework for DO, and that DO requires a rigid mineralized surface. The presence of osteocytes in contact with the calcified cartilage also suggests the existence of stationary osteoblasts in endochondral ossification. Stationary osteoblasts did not appear to be a unique feature of SO. The presence of stationary osteoblasts may appear to provide the initial osteocytes during osteogenesis that may function as mechanosensors throughout the bone tissue. If this is the case, then bone would be capable of sensing mechanical strains from its inception.