Ossification In Humans Research Articles

A rare case of deformity: Fibrodysplasia ossificans progressiva

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder and the most debilitating condition of heterotopic ossification in humans. Misdiagnosis is common and they suffer from complications of incorrect diagnosis and subsequent inappropriate treatment hence, to create awareness, we are reporting a genetically confirmed case of FOP. A 10-year-old boy, born to a non-consanguineously married couple, presented with progressive stiffness of the neck, back, and limbs with restriction of movements. Examination showed two firm non-tender immobile swellings over the back with scoliosis towards the left side. A bilateral hallux valgus deformity with restricted movements of the great toe was noted. Outside treated as mixed connective tissue disorder without any improvement. The X-ray showed calcifications in the swellings. Genetic testing showed the pathogenic variant in the activin A receptor type 1 gene. The FOP should be considered in children presenting with progressive stiffness, deformity of the feet, and ectopic ossifications to avoid misdiagnosis and inappropriate treatment.

The Distinct Role of the Extra-Large G Protein ɑ-Subunit XLɑs.

GNAS is one of the most complex gene loci in the human genome and encodes multiple gene products including Gsα, XLαs, NESP55, A/B, and AS transcripts. XLαs, the extra-large G protein ɑ-subunit, is paternally expressed. XLɑs and Gsɑ share the common 2-13 exons with different promoters and first exons. Therefore, XLɑs contains most of the functional domains of Gsα including receptor and effector binding sites. In vitro studies suggest a "Gsɑ"-like function of XLɑs regarding the stimulation of cAMP generation in response to receptor activation with different cellular actions. However, it is unclear whether XLαs has an important physiological function in humans. Pseudopseudohypoparathyroidism (PPHP) and progressive osseous heteroplasia (POH) are caused by paternally inherited mutations of GNAS. Maternal uniparental disomy of chromosome 20 [UPD(20)mat] lacks paternal chromosome 20. Therefore, the phenotypes of these diseases may be secondary to the abnormal functions of XLɑs, at least partly. From the phenotypes of human diseases like PPHP, POH, and UPD(20)mat, as well as some animal models with deficient XLɑs functions, it could be seen that XLɑs is involved in the growth and development of the mammalian fetus, plays a different role in glucose, lipid, and energy metabolism when compared with Gsɑ, and could prevent heterotopic ossification in humans and mice. More in vivo and in vitro studies, especially the development of conditional XLɑs knockout mice, are needed to clarify the physiopathologic roles and related signal pathways of XLɑs.

A case of fibrodysplasia ossificans progressiva with sternocleidomastoid muscle calcification

Fibrodysplasia ossificans progressiva (FOP) is a rare and disabling genetic condition characterized by congenital skeletal malformations and progressive heterotopic ossification in humans with no ethnic, racial, gender, or geographic predilection. Diagnosis of this condition can be made clinically in the presence of radiographic evidence of heterotopic ossification along with symmetrical malformations of the great toes. The course of the disease is unpredictable and often progresses in the early childhood and patients become immobile and confined to a wheelchair by their twenties. Survival beyond the third decade is uncommon. We hereby report a case of FOP in a 7½-year-old girl.

Is “Fibrodysplasia Ossificans Progressiva” a Vascular Disease? A Groundbreaking Pathogenic Model

Fibrodysplasia ossificans progressiva is the most severe and disabling disorder of ectopic ossification in humans. It is characterized by congenital skeletal abnormalities in association with extraskeletal widespread endochondral osteogenesis. Virtually all patients show the same mutation in the “activin A type-I/activin-like kinase-2” receptor encoding gene. As a result of this discovery there have been significant advances in the knowledge of the cellular and molecular basis of the disease. Besides allowing a better understanding of ossification process, recent evidence indicates that the primary disturbance lies within basic mechanisms of cell differentiation that are key in several physiological pathways and in the genesis of diseases with a major impact on health. In this article we summarize these breakthroughs, with implications that go beyond the limits of this devastating disease to insinuate a new model of human pathophysiology.

¿Es la «fibrodisplasia osificante progresiva» una enfermedad de origen vascular? Un modelo patogénico innovador

Fibrodysplasia ossificans progressiva is the most severe and disabling disorder of ectopic ossification in humans. It is characterized by congenital skeletal abnormalities in association with extraskeletal widespread endochondral osteogenesis. Virtually all patients show the same mutation in the “Activin A type-I/activin-like kinase-2” receptor encoding gene. As a result of this discovery there have been significant advances in the knowledge of the cellular and molecular basis of the disease. Besides allowing a better understanding of ossification process, recent evidence indicates that the primary disturbance lies within basic mechanisms of cell differentiation that are key in several physiological pathways and in the genesis of diseases with a major impact on health. In this article we summarize these breakthroughs, with implications that go beyond the limits of this devastating disease to insinuate a new model of human pathophysiology.

Principles of cranial base ossification in humans and rats

Conclusions: 1. The principle of bilateral symmetry depends on the chordal cartilage that is the keystone in cranial base ossification in rats and humans, due to its anatomical situation and for the production of the chordin protein that regulates the bone morphogenetic protein BMP-7. 2. In humans and in rats, foramen lacerum closure follows a line of intramembranous ossification that depends on BMP-7, regulated by the first branchial pouch. 3. The cranial base ossification patterns and centres are similar in humans and in rats, except in the otic capsule, palate and the lateral pterygoid plate. 4. The neural crest may induce cranial ossification through the cranial nerves. Objectives: To study the patterns of cranial base ossification in humans and in rats, considering the chordal cartilage, and the otic, nasal and orbit capsules, as well as the participation of the branchial arches and pouches. Methods: This was a light microscopy study of human fetal specimens obtained from spontaneous abortions with the following crown-rump-lengths (crl) 45, 74, 90, 134, 145 and 270 mm, and a 1-day-old neonate (360 mm crl), who had died of sudden death syndrome. We also examined Webster albino rat embryos of 16, 18 and 20 days of gestation and a postnatal series of rats 8 h and 1, 3, 4, 6, 7, 10 and 13 days old, as well as adult animals. Results: In the 45 mm human fetus, the chordal cartilage with the nasal, otic and orbit capsules initiates cranial base ossification. Foramen lacerum closure begins in the 16-day-old rat embryo, following a line of membranous ossification between the external pterygoid process and the lateral alisphenoidal wing at ovalis foramen level. This is not a timing symmetrical process, which may persist until the 10th postnatal day in the rat. In the human fetus of 74 mm, the foramen lacerum space is closed by a membranous fusion ossification between the chordal cartilage and otic capsule, finishing at the 270 mm specimen. Endochondral ossification of the human otic capsule first appeared in the 145 mm (18 weeks) fetal specimen with four ossifying centres. The rat otic cartilaginous capsule showed rapid endochondral ossification, in the third and fourth postnatal day specimens.

Fibrodysplasia ossificans progressiva (FOP): watch the great toes!

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder and the most disabling condition of heterotopic (extraskeletal) ossification in humans. Extraskeletal bone formation associated with inflammation preceding the osseous conversion usually begins in the first decade, predominantly in the head, neck, and shoulders. All patients have malformed great toes. Most patients have a spontaneous mutation of the ACVR1 gene. We report a 17-year-old girl with malformed great toes who had her first episode of heterotopic ossification and impaired mobility of the left hip at the age of 13years. No inflammatory fibroproliferative masses preceded the onset of heterotopic ossification. Radiographic studies demonstrated myositis ossificans, but failure to associate the great toe malformation with heterotopic ossification led to a failure to diagnose FOP. She underwent repeated and unnecessary operative procedures to remove a recurrent lesion. FOP was finally suspected when the great toe malformation was correlated with the trauma-induced heterotopic ossification. Genetic analysis confirmed the presence of the classic FOP mutation (ACVR1 c.617G>A; R206H). This case highlights the importance of examining the great toes in anyone with heterotopic ossification. The association of malformations of the great toe with heterotopic ossification in all cases of classic FOP will lead to prompt clinical diagnosis and the prevention of iatrogenic harm.

Early Mortality and Cardiorespiratory Failure in Patients with Fibrodysplasia Ossificans Progressiva

Fibrodysplasia ossificans progressiva, a rare genetic disorder of progressive extraskeletal ossification, is the most disabling form of heterotopic ossification in humans. However, little is known about the lifespan or causes of mortality in these patients. We undertook this study to determine the lifespan and causes of mortality in individuals who had fibrodysplasia ossificans progressiva. We reviewed comprehensive mortality reports from two large registries of patients with fibrodysplasia ossificans progressiva. Together, these registries comprise >90% of all known patients with this condition in the world. We noted the sex, dates of birth and death, and the cause of death for each individual. We verified the cause of death with extensive medical records, when available. We also collected date of birth, current age, and sex information for each living patient member of the International Fibrodysplasia Ossificans Progressiva Association. Sixty deaths (thirty male and thirty female patients) were reported in the fibrodysplasia ossificans progressiva community during a thirty-three-year-period. For all sixty patients, the median age at the time of death was forty years (range, three to seventy-seven years). Data were sufficient to establish the cause of death in forty-eight (80%) of the sixty individuals. The median age at the time of death for the forty-eight patients (twenty-four male and twenty-four female patients) with an established cause of death was also forty years. The median lifespan estimated from the 371 individuals in the international fibrodysplasia ossificans progressiva community who were alive and the sixty who had died was fifty-six years (95% confidence interval, fifty-one to sixty years). The most common causes of death in patients with fibrodysplasia ossificans progressiva were cardiorespiratory failure from thoracic insufficiency syndrome (54%; median age, forty-two years) and pneumonia (15%; median age, forty years). Fibrodysplasia ossificans progressiva is not only an extremely disabling disease but also a condition of considerably shortened lifespan. The most common cause of death in patients with fibrodysplasia ossificans progressiva is cardiorespiratory failure from thoracic insufficiency syndrome.

Pharmacological Inhibition of 5-Lipoxygenase Accelerates and Enhances Fracture-Healing

Loss of cyclooxygenase-2 activity is known to impair fracture-healing in animal models and to inhibit heterotopic ossification in humans. Cyclooxygenase-2 is the rate-limiting enzyme involved in the conversion of arachidonic acid into prostaglandins. Arachidonic acid also is a substrate for 5-lipoxygenase, which catalyzes the initial steps in leukotriene synthesis. In contrast to cyclooxygenase-2, genetic ablation of 5-lipoxygenase accelerates and enhances fracture-healing in mice. The goal of this study was to determine if systemic inhibition of 5-lipoxygenase with an orally delivered drug could accelerate fracture-healing. Closed femoral fractures were made in Sprague-Dawley rats. The rats were treated with oral doses of vehicle (ninety-five rats), celecoxib (fifty-nine rats), or AA-861 (a 5-lipoxygenase inhibitor; eighty-nine rats). Fracture-healing was measured with use of radiographs, histomorphometry, and biomechanical testing. Effects of drug treatments on callus cell proliferation and gene expression were determined by incorporation of bromodeoxyuridine and quantitative polymerase chain reactions, respectively. AA-861 treatment decreased fracture-bridging time, significantly increased early callus cartilage (5.6-fold; p < 0.001) and bone formation (4.2-fold; p = 0.015), and significantly increased callus mechanical properties compared with the vehicle-treated rat fractures. Callus cell proliferation rate was increased by AA-861 treatment, compared with vehicle, at day 2 after fracture (3.68% compared with 2.08%; p < 0.001; 95% confidence interval, -2.81 to -0.039) but was reduced by celecoxib treatment at day 4 after fracture (4.22% compared with 1.84%; p < 0.001; 95% confidence interval, 2.27 to 4.07). At day 10 after fracture, AA-861 and celecoxib treatment increased Type-II collagen mRNA levels (16.0-fold and 6.1-fold, respectively; p < 0.001 for both), but only AA-861 treatment caused an increase in Type-X collagen mRNA (6.3-fold; p < 0.001). AA-861 treatment significantly increased cyclooxygenase-2 (4.0-fold at day 10; p < 0.001) and osteopontin mRNA levels (3.6-fold at day 7; p = 0.024), while decreasing 5-lipoxygenase mRNA levels (5.6-fold at day 4; p < 0.001). Systemic inhibition of 5-lipoxygenase with an orally delivered drug significantly accelerated and enhanced fracture-healing in this rat model. Gene expression analysis indicates that cyclooxygenase-2 is necessary for callus chondrocytes to progress into hypertrophy so as to complete endochondral ossification. Conversely, inhibition of 5-lipoxygenase alters the inflammatory response, which enhances callus chondrocyte hypertrophy and accelerates endochondral ossification.

Ossification of a Collagen Membrane Cross‐Linked by Sugar: A Human Case Series

Collagen membranes cross-linked by glycation (GLYM) for guided bone regeneration (GBR) and guided tissue regeneration (GTR) are used extensively with proven safety and efficacy. Complete GLYM ossification, when placed in contact with bone, was described in a canine jaw model, suggesting that GLYM may serve as an ossification substrate. The purpose of this case series was to histologically evaluate GLYM in GBR procedures in humans. We retrospectively selected seven consecutive patients with implant-related bony defects who underwent GBR with GLYM. Six defects had bone grafts, and one had a barrier alone. Selection criteria were primary closure upon post-surgical examination and tissue that was 2- to 3-mm thick over the implant's cover screw. Tissue was removed when the implants were uncovered after 20 to 29 weeks. Decalcified sections were stained and analyzed under light microscopy. In five of seven specimens, GLYM was identified and preserved its barrier effect. The mean membrane thickness was 0.17 +/- 0.054 mm. In two cases, the bone grafts under the membrane were embedded in new bone, whereas in five cases, they were embedded in fibrous connective tissue. Formation of new dense bone was observed along the side of the membrane facing the original bone, and various degrees of membrane ossification were evident in all five cases. GLYM maintained its barrier effect in five of seven cases for 25 weeks and induced dense new bone along its interface with underlying tissues. To the best of our knowledge, this is the first report on GLYM ossification in humans with direct mineral apposition on glycated collagen and suggests a new concept of tissue-integrated active barriers.

Dysregulated BMP Signaling and Enhanced Osteogenic Differentiation of Connective Tissue Progenitor Cells From Patients With Fibrodysplasia Ossificans Progressiva (FOP)

The study of FOP, a disabling genetic disorder of progressive heterotopic ossification, is hampered by the lack of readily available connective tissue progenitor cells. We isolated such cells from discarded primary teeth of patients with FOP and controls and discovered dysregulation of BMP signaling and rapid osteoblast differentiation in FOP cells compared with control cells. Fibrodysplasia ossificans progressiva (FOP), the most disabling condition of progressive heterotopic ossification in humans, is caused by a recurrent heterozygous missense mutation in activin receptor IA (ACVR1), a bone morphogenetic protein (BMP) type I receptor, in all classically affected individuals. A comprehensive understanding of FOP has been limited, in part, by a lack of readily available connective tissue progenitor cells in which to study the molecular pathology of this disorder. We derived connective tissue progenitor cells from discarded primary teeth (SHED cells) of patients with FOP and controls and examined BMP signaling and osteogenic differentiation in these cells. SHED cells transmitted BMP signals through both the SMAD and p38 mitogen-activated protein kinase (MAPK) pathways and responded to BMP4 treatment by inducing BMP responsive genes. FOP cells showed ligand-independent BMP signaling and ligand-dependent hyper-responsiveness to BMP stimulation. Furthermore, FOP cells showed more rapid differentiation to an osteogenic phenotype than control cells. This is the first study of BMP signaling and osteogenic differentiation in connective tissue progenitor cells from patients with FOP. Our data strongly support both basal and ligand-stimulated dysregulation of BMP signaling consistent with in silico studies of the mutant ACVR1 receptor in this condition. This study substantially extends our understanding of dysregulated BMP signaling in a progenitor cell population relevant to the pathogenesis of this catastrophic disorder of progressive ectopic ossification.

The Effectiveness of Two COX-2 Inhibitors in the Prophylaxis Against Heterotopic New Bone Formation: An Experimental Study in Rabbits

The purpose of this study was to present the effectiveness of 2 cyclooxygenase-2 inhibitors, meloxicam and parecoxib, in the prevention of heterotopic ossification. Eighteen white mature male rabbits were divided into 3 groups of 6 animals. One was the control group; 1 group was administered meloxicam subcutaneously, and 1 group was administered parecoxib subcutaneously. For the induction of heterotopic new bone, the Michelsson model was used. The extent of heterotopic new bone development was assessed according to Scott's classification. The meloxicam and parecoxib groups developed significantly less heterotopic bone when compared to the control group. There was no significant difference between the meloxicam and parecoxib group. We concluded that meloxicam and parecoxib are capable of preventing the development of heterotopic new bone in rabbits. The clinical relevance of this study is that meloxicam and parecoxib may be helpful in reducing heterotopic ossification in humans.

The effect of ketoprophen on lumbar spinal fusion healing in a rabbit model

Several reports have shown that nonsteroidal antiinflammatory drugs (NSAIDs) have an inhibitory effect in osteogenesis and reduce heterotopic ossification in humans. A deleterious effect of NSAIDs in posterolateral intertransverse process fusion has also been suggested. The authors used a validated rabbit model to try to determine the influence of the NSAID ketoprophen on the fusion rate in lumbar spinal arthrodesis. Thirty New Zealand male rabbits underwent posterolateral (intertransverse process) bilateral spinal fusions at a single level, using autologous bone graft obtained from both iliac crests. The animals were randomized after the operation, so that 15 rabbits received ketoprophen as a postoperative analgesic and the other 15 received the postoperative analgesic tramadol. The animals were killed 8 weeks after surgery, and fusion status was determined by inspection, palpation, anteroposterior radiographs, and histological analysis. A solid fusion was obtained in eight rabbits (53%), and pseudarthrosis in seven rabbits (47%) in each group. These findings suggest that the use of ketoprophen after intertransverse spinal fusion at a single level does not decrease the fusion rate, compared with tramadol.

The effect of ketoprophen on lumbar spinal fusion healing in a rabbit model

Object Several reports have shown that nonsteroidal antiinflammatory drugs (NSAIDs) have an inhibitory effect in osteogenesis and reduce heterotopic ossification in humans. A deleterious effect of NSAIDs in posterolateral intertransverse process fusion has also been suggested. The authors used a validated rabbit model to try to determine the influence of the NSAID ketoprophen on the fusion rate in lumbar spinal arthrodesis. Methods Thirty New Zealand male rabbits underwent posterolateral (intertransverse process) bilateral spinal fusions at a single level, using autologous bone graft obtained from both iliac crests. The animals were randomized after the operation, so that 15 rabbits received ketoprophen as a postoperative analgesic and the other 15 received the postoperative analgesic tramadol. The animals were killed 8 weeks after surgery, and fusion status was determined by inspection, palpation, anteroposterior radiographs, and histological analysis. Results A solid fusion was obtained in eight rabbits (53%), and pseudarthrosis in seven rabbits (47%) in each group. Conclusions These findings suggest that the use of ketoprophen after intertransverse spinal fusion at a single level does not decrease the fusion rate, compared with tramadol.

A new era for fibrodysplasia ossificans progressiva: a druggable target for the second skeleton

Fibrodysplasia ossificans progressiva (FOP) is a disabling genetic condition that leads to the formation of a second (heterotopic) skeleton, and is the most catastrophic disorder of heterotopic ossification in humans. Throughout childhood and early adult life, FOP progressively immobilizes all of the joints of the normotopic skeleton, rendering movement impossible. At present, there is no effective prevention or treatment. Recently, a recurrent mutation in the glycine-serine activation domain of the activin receptor IA/activin-like kinase-2, a bone morphogenetic protein type I receptor, was reported in all sporadic and familial cases of classic FOP, making this one of the most highly specific disease-causing mutations in the human genome. The discovery of the FOP gene establishes a critical milestone in understanding FOP, reveals a highly conserved druggable target in the TGF-β/bone morphogenetic protein signaling pathway and compels therapeutic approaches for the development of small molecule signal transduction inhibitors for activin-like kinase-2. Effective therapies for FOP, and possibly for a vast array of more common conditions of heterotopic ossification, will be based on blocking activin-like kinase-2, a critical node in the BMP signaling pathway.

1138. In Vivo Somatic Cell Gene Transfer of an Engineered Noggin Mutein Prevents BMP4-Induced Heterotopic Ossification in the Mouse: Implications for the Treatment of Fibrodysplasia Ossificans Progressiva and Other Disorders of Heterotopic Ossification in Humans

1138. In Vivo Somatic Cell Gene Transfer of an Engineered Noggin Mutein Prevents BMP4-Induced Heterotopic Ossification in the Mouse: Implications for the Treatment of Fibrodysplasia Ossificans Progressiva and Other Disorders of Heterotopic Ossification in Humans

Regulation of human cranial osteoblast phenotype by FGF-2, FGFR-2 and BMP-2 signaling.

The formation of cranial bone requires the differentiation of osteoblasts from undifferentiated mesenchymal cells. The balance between osteoblast recruitment, proliferation, differentiation and apoptosis in sutures between cranial bones is essential for calvarial bone formation. The mechanisms that control human osteoblasts during normal calvarial bone formation and premature suture ossification (craniosynostosis) begin to be understood. Our studies of the human calvaria osteoblast phenotype and calvarial bone formation showed that premature fusion of the sutures in non-syndromic and syndromic (Apert syndrome) craniosynostoses results from precocious osteoblast differentiation. We showed that Fibroblast Growth Factor-2 (FGF-2), FGF receptor-2 (FGFR-2) and Bone Morphogenetic Protein-2 (BMP-2), three essential factors involved in skeletal development, regulate the proliferation, differentiation and apoptosis in human calvaria osteoblasts. Mechanisms that induce the differentiated osteoblast phenotype have also been identified in human calvaria osteoblasts. We demonstrated the implication of molecules (N-cadherin, Il-1) and signaling pathways (src, PKC) by which these local factors modulate human calvaria osteoblast differentiation and apoptosis. The identification of these essential signaling molecules provides new insights into the pathways controlling the differentiated osteoblast phenotype, and leads to a more comprehensive view in the mechanisms that control normal and premature cranial ossification in humans.

Heterotopic Ossification: Two Rare Forms and What They Can Teach Us.

Heterotopic ossification is characterized by the formation of normal bone at ectopic soft-tissue locations. Regardless of the etiology of heterotopic ossification, requisite pathogenetic conditions include an inductive signal capable of stimulating morphogenesis, a population of inducible osteoprogenitor cells, and a heterotopic environment conducive to osteogenesis. Two rare heritable and developmental forms of heterotopic ossification, fibrodysplasia ossificans progressiva and progressive osseous heteroplasia, provide valuable clinical and pathogenetic insights into heterotopic ossification in humans. A fundamental understanding of the developmental and molecular pathology of these disorders may lead to more effective strategies for preventing and treating heterotopic ossification in humans.

Extracellular matrix alterations during endochondral ossification in humans

Immunohistochemical methods were employed to examine alterations in the cartilage extracellular matrix constituents associated with endochondral ossification in humans. The distributions of chondroitin 4- and 6-sulfate and keratan sulfate proteoglycan (PG) determinants, cartilage PG link protein, collagen types I and II, and fibronectin were determined in iliac crest growth-plate specimens using the avidin-biotin-horseradish peroxidase system. Collagen type II was distributed throughout the growth plate, providing a framework within which chondrocytes divided and formed clusters of differentiating (hypertrophic) cells. The septa between these clusters and their subchondral extensions into underlying bone trabeculae were rich in PG, PG link protein, and collagen type II and resembled the extracellular matrix of reserve cartilage. The territorial matrix associated with the differentiating cells within the clusters contained reduced amounts of collagen type II, PG link protein, and possibly cartilage PG. Collagen type I and fibronectin were detected within the cytoplasm of the maturing and degenerating cells, and fibronectin localized intensely to the pericellular matrix envelopes of these cells. These alterations presumably facilitate the degradation of the matrix associated with the cell clusters by invading vascular tissue, while the septa, which retain the characteristics of more typical cartilage matrix, are not degraded and firmly anchor the cartilage to the subchondral bone.