The gelatinases, matrix metalloproteinases 2 and 9, play individual roles in skeleton development.

The gelatinases, a subgroup of the matrix metalloproteinases (MMPs) superfamily are composed of two members; MMP2 and MMP9. They are known to degrade gelatin among other components of the extracellular matrix. Recently, the two gelatinases were found to be necessary for neural crest cell migration and to compensate for each other loss in these cells. To characterize their involvement in the skeletal system, and to better reveal their individual or common roles, we have generated double knockout (dKO) mice, lacking both MMP2 and MMP9. Comprehensive analysis of the skeleton morphological and mechanical parameters at postnatal day (P) 0, P21, 3 months (M) and 8M of age, revealed an unexpected distinct role for each gelatinase; MMP2 was found to be involved merely in intramembranous ossification which led to a smaller skull and inferior cortical parameters upon its loss, while MMP9 was found to affect only the endochondral ossification process, which led to shorter long-bones in its absence. Importantly, the dKO mice demonstrated a combination of both the skull and long bone phenotypes as found in the single-KOs, and not a severer additive phenotype. Transcriptome analysis on the cortical bone, the growth plate and the skull frontal bone, found many genes that were differentially expressed as a direct or indirect result of MMP-loss, and reinforced the specific and distinct role of each gelatinase in each bone type. Altogether, these results suggest that although both gelatinases share the same substrates and are highly expressed in flat and long bones, they are indispensable and control separately the development of different bones.

BMP3 Affects Cortical and Trabecular Long Bone Development in Mice.

Bone morphogenetic proteins (BMPs) have a major role in tissue development. BMP3 is synthesized in osteocytes and mature osteoblasts and has an antagonistic effect on other BMPs in bone tissue. The main aim of this study was to fully characterize cortical bone and trabecular bone of long bones in both male and female Bmp3-/- mice. To investigate the effect of Bmp3 from birth to maturity, we compared Bmp3-/- mice with wild-type littermates at the following stages of postnatal development: 1 day (P0), 2 weeks (P14), 8 weeks and 16 weeks of age. Bmp3 deletion was confirmed using X-gal staining in P0 animals. Cartilage and bone tissue were examined in P14 animals using Alcian Blue/Alizarin Red staining. Detailed long bone analysis was performed in 8-week-old and 16-week-old animals using micro-CT. The Bmp3 reporter signal was localized in bone tissue, hair follicles, and lungs. Bone mineralization at 2 weeks of age was increased in long bones of Bmp3-/- mice. Bmp3 deletion was shown to affect the skeleton until adulthood, where increased cortical and trabecular bone parameters were found in young and adult mice of both sexes, while delayed mineralization of the epiphyseal growth plate was found in adult Bmp3-/- mice.

Cellular and Molecular Alterations Underlying Abnormal Bone Growth in X-Linked Hypophosphatemia.

X-linked hypophosphatemia (XLH), the most common form of hereditary hypophosphatemic rickets, is caused by inactivating mutations of the phosphate-regulating endopeptidase gene (PHEX). XLH is mainly characterized by short stature, bone deformities and rickets, while in hypophosphatemia, normal or low vitamin D levels and low renal phosphate reabsorption are the principal biochemical aspects. The cause of growth impairment in patients with XLH is not completely understood yet, thus making the study of the growth plate (GP) alterations necessary. New treatment strategies targeting FGF23 have shown promising results in normalizing the growth velocity and improving the skeletal effects of XLH patients. However, further studies are necessary to evaluate how this treatment affects the GP as well as its long-term effects and the impact on adult height.

C-type Natriuretic Peptide-induced PKA Activation Promotes Endochondral Bone Formation in Hypertrophic Chondrocytes.

Longitudinal bone growth is achieved by a tightly controlled process termed endochondral bone formation. C-type natriuretic peptide (CNP) stimulates endochondral bone formation through binding to its specific receptor, guanylyl cyclase (GC)-B. However, CNP/GC-B signaling dynamics in different stages of endochondral bone formation have not been fully clarified, especially in terms of the interaction between the cyclic guanine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) pathways. Here, we demonstrated that CNP activates the cAMP/protein kinase A (PKA) pathway and that this activation contributed to the elongation of the hypertrophic zone in the growth plate. Cells of the chondrogenic line ATDC5 were transfected with Förster resonance energy transfer (FRET)-based cGMP and PKA biosensors. Dual-FRET imaging revealed that CNP increased intracellular cGMP levels and PKA activities in chondrocytes. Further, CNP-induced PKA activation was enhanced following differentiation of ATDC5 cells. Live imaging of the fetal growth plate of transgenic mice, expressing a FRET biosensor for PKA, PKAchu mice, showed that CNP predominantly activates the PKA in the hypertrophic chondrocytes. Additionally, histological analysis of the growth plate of PKAchu mice demonstrated that CNP increased the length of the growth plate, but coadministration of a PKA inhibitor, H89, inhibited the growth-promoting effect of CNP only in the hypertrophic zone. In summary, we revealed that CNP-induced cGMP elevation activated the cAMP/PKA pathway, and clarified that this PKA activation contributed to the bone growth-promoting effect of CNP in hypertrophic chondrocytes. These results provide insights regarding the cross-talk between cGMP and cAMP signaling in endochondral bone formation and in the physiological role of the CNP/GC-B system.

Application of 3D MAPs pipeline identifies the morphological sequence chondrocytes undergo and the regulatory role of GDF5 in this process.

The activity of epiphyseal growth plates, which drives long bone elongation, depends on extensive changes in chondrocyte size and shape during differentiation. Here, we develop a pipeline called 3D Morphometric Analysis for Phenotypic significance (3D MAPs), which combines light-sheet microscopy, segmentation algorithms and 3D morphometric analysis to characterize morphogenetic cellular behaviors while maintaining the spatial context of the growth plate. Using 3D MAPs, we create a 3D image database of hundreds of thousands of chondrocytes. Analysis reveals broad repertoire of morphological changes, growth strategies and cell organizations during differentiation. Moreover, identifying a reduction in Smad 1/5/9 activity together with multiple abnormalities in cell growth, shape and organization provides an explanation for the shortening of Gdf5 KO tibias. Overall, our findings provide insight into the morphological sequence that chondrocytes undergo during differentiation and highlight the ability of 3D MAPs to uncover cellular mechanisms that may regulate this process.

Characterization of the growth plate-bone interphase region using cryo-FIB SEM 3D volume imaging.

The interphase region at the base of the growth plate includes blood vessels, cells and mineralized tissues. In this region, cartilage is mineralized and replaced with bone. Blood vessel extremities permeate this space providing nutrients, oxygen and signaling factors. All these different components form a complex intertwined 3D structure. Here we use cryo-FIB SEM to elaborate this 3D structure without removing the water. As it is challenging to image mineralized and unmineralized tissues in a hydrated state, we provide technical details of the parameters used. We obtained two FIB SEM image stacks that show that the blood vessels are in intimate contact not only with cells, but in some locations also with mineralized tissues. There are abundant red blood cells at the extremities of the vessels. We also documented large multinucleated cells in contact with mineralized cartilage and possibly also with bone. We observed membrane bound mineralized particles in these cells, as well as in blood serum, but not in the hypertrophic chondrocytes. We confirm that there is an open pathway from the blood vessel extremities to the mineralizing cartilage. Based on the sparsity of the mineralized particles, we conclude that mainly ions in solution are used for mineralizing cartilage and bone, but these are augmented by the supply of mineralized particles.

Sex-specific effects of 17ß-estradiol and dihydrotestosterone (DHT) on growth plate chondrocytes are dependent on both ERα and ERß and require palmitoylation to translocate the receptors to the plasma membrane.

Rat costochondral cartilage growth plate chondrocytes exhibit cell sex-specific responses to 17ß-estradiol (E2), testosterone, and dihydrotestosterone (DHT). Mechanistically, E2 and DHT stimulate proliferation and extracellular matrix synthesis in chondrocytes from female and male rats, respectively, by signaling through protein kinase C (PKC) and phospholipase C (PLC). Estrogen receptors (ERα; ERß) and androgen receptors (ARs) are present in both male and female cells, but it is not known whether they interact to elicit sex-specific signaling. We used specific agonists and antagonists of these receptors to examine the relative contributions of ERs and ARs in membrane-mediated E2 signaling in female chondrocytes and DHT signaling in male chondrocytes. PKC activity in female chondrocytes was stimulated by agonists of ERα and ERß and required intact caveolae; PKC activity was inhibited by the E2 enantiomer and by an inhibitor of ERß. Western blots of cell lysates co-immunoprecipitated for ERα suggested the formation of a complex containing both ERα and ERß with E2 treatment. DHT and DHT agonists activated PKC in male cells, while AR inhibition blocked the stimulatory effect of DHT on PKC. Inhibition of ERα and ERß also blocked PKC activation by DHT. Western blots of whole-cell lysates, plasma membranes, and caveolae indicated the translocation of AR to the plasma membrane and specifically to caveolae with DHT treatment. These results suggest that E2 and DHT promote chondrocyte differentiation via the ability of ARs and ERs to form a complex. The results also indicate that intact caveolae and palmitoylation of the membrane receptor(s) or membrane receptor complex containing ERα and ERß is required for E2 and DHT membrane-associated PKC activity in costochondral cartilage cells.

Rhizomelia and Impaired Linear Growth in a Girl with Juvenile Paget Disease: The Natural History of the Condition.

In ultra-rare bone diseases, information on growth during childhood is sparse. Juvenile Paget disease (JPD) is an ultra-rare disease, characterized by loss of function of osteoprotegerin (OPG). OPG inhibits osteoclast activation via the receptor activator of nuclear factor-κB (RANK) pathway. In JPD, overactive osteoclasts result in inflammatory-like bone disease due to grossly elevated bone resorption. Knowledge on the natural history of JPD, including final height and growth, is limited. Most affected children receive long-term antiresorptive treatment, mostly with bisphosphonates, to contain bone resorption, which may affect growth. In this study, we report the follow-up of height, growth velocity, and skeletal maturation in a 16-year-old female patient with JPD. The patient was treated with cyclic doses of pamidronate starting at 2.5 years of age and with 2 doses of denosumab at the age of 8 years, when pamidronate was paused. In the following years, a sustainable decline in a height z-score and a stunted pubertal growth spurt; despite appropriate maturation of the epiphyseal plates of the left hand, the proximal right humerus and both femora were observed. Whether this reflects the growth pattern in JPD or might be associated to the antiresorptive treatments is unclear, since there is very limited information available on the effect of bisphosphonates and denosumab on growth and the growth plate in pediatric patients. Studies are needed to understand the natural history of an ultra-rare bone disease and to assess the effects of antiresorptive treatment on the growing skeleton.

Phosphatase inhibition by LB-100 enhances BMN-111 stimulation of bone growth.

Activating mutations in fibroblast growth factor receptor 3 (FGFR3) and inactivating mutations in the natriuretic peptide receptor 2 (NPR2) guanylyl cyclase both result in decreased production of cyclic GMP in chondrocytes and severe short stature, causing achondroplasia (ACH) and acromesomelic dysplasia, type Maroteaux, respectively. Previously, we showed that an NPR2 agonist BMN-111 (vosoritide) increases bone growth in mice mimicking ACH (Fgfr3Y367C/+). Here, because FGFR3 signaling decreases NPR2 activity by dephosphorylating the NPR2 protein, we tested whether a phosphatase inhibitor (LB-100) could enhance BMN-111-stimulated bone growth in ACH. Measurements of cGMP production in chondrocytes of living tibias, and of NPR2 phosphorylation in primary chondrocytes, showed that LB-100 counteracted FGF-induced dephosphorylation and inactivation of NPR2. In ex vivo experiments with Fgfr3Y367C/+ mice, the combination of BMN-111 and LB-100 increased bone length and cartilage area, restored chondrocyte terminal differentiation, and increased the proliferative growth plate area, more than BMN-111 alone. The combination treatment also reduced the abnormal elevation of MAP kinase activity in the growth plate of Fgfr3Y367C/+ mice and improved the skull base anomalies. Our results provide a proof of concept that a phosphatase inhibitor could be used together with an NPR2 agonist to enhance cGMP production as a therapy for ACH.

Growth Modulation by Stimulating the Growth Plate: A Pilot Study.

This study investigates the ability of low-intensity pulsed ultrasound (LIPUS) or direct injection of recombinant growth hormone (rGH) to stimulate local growth of long bones. In a randomized controlled animal trial, healthy immature rabbits were allocated to 1 of the following 4 conditions: epiphyseal rGH periosteal injection, transdermal LIPUS, saline periosteal injection, or no treatment. New bone deposition was labeled with calcein at days 1 and 18, and microscopic measurements of growth were conducted by blinded observers. Statistically significant differences in growth were observed between the LIPUS and rGH stimulated legs compared with contralateral control legs (35% p = 0.04 and 41% p = 0.04, respectively); whereas no difference was observed between the 4 control groups (p = 0.37). There was no evidence of physeal bar formation, suggesting that direct injection of rGH and application of LIPUS around the distal femoral physis in rabbits may have a positive effect on microscopic growth without short-term adverse sequelae.

miR-143 is implicated in growth plate injury by targeting IHH in precartilaginous stem cells.

Precartilaginous stem cells (PCSCs) are able to initiate chondrocyte and bone development. The present study aimed to investigate the role of miR-143 and the underlying mechanisms involved in PCSC proliferation. In a rat growth plate injury model, tissue from the injury site was collected and the expression of miR-143 and its potential targets was determined. PCSCs were isolated from the rabbits' distal epiphyseal growth plate. Cell viability, DNA synthesis, and apoptosis were determined with MTT, BrdU, and flow cytometric analysis, respectively. Real time PCR and western blot were performed to detect the mRNA and protein expression of the indicated genes. Indian hedgehog (IHH) was identified as a target gene for miR-143 with luciferase reporter assay. Decreased expression of miR-143 and increased expression of IHH gene were observed in the growth plate after injury. miR-143 mimics decreased cell viability and DNA synthesis and promoted apoptosis of PCSCs. Conversely, siRNA-mediated inhibition of miR-143 led to increased growth and suppressed apoptosis of PCSCs. Transfection of miR-143 decreased luciferase activity of wild-type IHH but had no effect when the 3'-UTR of IHH was mutated. Furthermore, the effect of miR-143 overexpression was neutralized by overexpression of IHH. Our study showed that miR-143 is involved in growth plate behavior and regulates PCSC growth by targeting IHH, suggesting that miR-143 may serve as a novel target for PCSC-related diseases.

Selection for increased tibia length in mice alters skull shape through parallel changes in developmental mechanisms.

Bones in the vertebrate cranial base and limb skeleton grow by endochondral ossification, under the control of growth plates. Mechanisms of endochondral ossification are conserved across growth plates, which increases covariation in size and shape among bones, and in turn may lead to correlated changes in skeletal traits not under direct selection. We used micro-CT and geometric morphometrics to characterize shape changes in the cranium of the Longshanks mouse, which was selectively bred for longer tibiae. We show that Longshanks skulls became longer, flatter, and narrower in a stepwise process. Moreover, we show that these morphological changes likely resulted from developmental changes in the growth plates of the Longshanks cranial base, mirroring changes observed in its tibia. Thus, indirect and non-adaptive morphological changes can occur due to developmental overlap among distant skeletal elements, with important implications for interpreting the evolutionary history of vertebrate skeletal form.

Prevention of guanylyl cyclase-B dephosphorylation rescues achondroplastic dwarfism.

Activating mutations in the fibroblast growth factor receptor 3 (FGFR3) or inactivating mutations in guanylyl cyclase-B (GC-B), also known as NPR-B or Npr2, cause short-limbed dwarfism. FGFR3 activation causes dephosphorylation and inactivation of GC-B, but the contribution of GC-B dephosphorylation to achondroplasia (ACH) is unknown. GC-B7E/7E mice that express a glutamate-substituted version of GC-B that cannot be inactivated by dephosphorylation were bred with mice expressing FGFR3-G380R, the most common human ACH mutation, to determine if GC-B dephosphorylation is required for ACH. Crossing GC-B7E/7E mice with FGFR3G380R/G380R mice increased naso-anal and long (tibia and femur), but not cranial, bone length twice as much as crossing GC-B7E/7E mice with FGFR3WT/WT mice from 4 to 16 weeks of age. Consistent with increased GC-B activity rescuing ACH, long bones from the GC-B7E/7E/FGFR3G380R/G380R mice were not shorter than those from GC-BWT/WT/FGFR3WT/WT mice. At 2 weeks of age, male but not female FGFR3G380R/G380R mice had shorter long bones and smaller growth plate hypertrophic zones, whereas female but not male GC-B7E/7E mice had longer bones and larger hypertrophic zones. In 2-week-old males, crossing FGFR3G380R/G380R mice with GC-B7E/7E mice increased long bone length and hypertrophic zone area to levels observed in mice expressing WT versions of both receptors. We conclude that preventing GC-B dephosphorylation rescues reduced axial and appendicular skeleton growth in a mouse model of achondroplasia.

Familial Short Stature-A Novel Phenotype of Growth Plate Collagenopathies.

CONTEXT: Collagens are the most abundant proteins in the human body. In a growth plate, collagen types II, IX, X, and XI are present. Defects in collagen genes cause heterogeneous syndromic disorders frequently associated with short stature. Less is known about oligosymptomatic collagenopathies. OBJECTIVE: This work aims to evaluate the frequency of collagenopathies in familial short stature (FSS) children and to describe their phenotype, including growth hormone (GH) treatment response. METHODS: Eighty-seven FSS children (pretreatment height ≤ -2 SD both in the patient and his or her shorter parent) treated with GH were included in the study. Next-generation sequencing was performed to search for variants in the COL2A1, COL9A1, COL9A2, COL9A3, COL10A1, COL11A1, and COL11A2 genes. The results were evaluated using American College of Medical Genetics and Genomics guidelines. The GH treatment response of affected children was retrospectively evaluated. RESULTS: A likely pathogenic variant in the collagen gene was found in 10 of 87 (11.5%) children. Detailed examination described mild asymmetry with shorter limbs and mild bone dysplasia signs in 2 of 10 and 4 of 10 affected children, respectively. Their growth velocity improved from a median of 5.3 cm/year to 8.7 cm/year after 1 year of treatment. Their height improved from a median of -3.1 SD to -2.6 SD and to -2.2 SD after 1 and 3 years of therapy, respectively. The final height reached by 4 of 10 children differed by -0.67 to +1.0 SD and -0.45 to +0.5 SD compared to their pretreatment height and their affected untreated parent's height, respectively. CONCLUSION: Oligosymptomatic collagenopathies are a frequent cause of FSS. The short-term response to GH treatment is promising.

The Formation of the Epiphyseal Bone Plate Occurs via Combined Endochondral and Intramembranous-Like Ossification.

The formation of the epiphyseal bone plate, the flat bony structure that provides strength and firmness to the growth plate cartilage, was studied in the present study by using light, confocal, and scanning electron microscopy. Results obtained evidenced that this bone tissue is generated by the replacement of the lower portion of the epiphyseal cartilage. However, this process differs considerably from the usual bone tissue formation through endochondral ossification. Osteoblasts deposit bone matrix on remnants of mineralized cartilage matrix that serve as a scaffold, but also on non-mineralized cartilage surfaces and as well as within the perivascular space. These processes occur simultaneously at sites located close to each other, so that, a core of the sheet of bone is established very quickly. Subsequently, thickening and reshaping occurs by appositional growth to generate a dense parallel-fibered bone structurally intermediate between woven and lamellar bone. All these processes occur in close relationship with a cartilage but most of the bone tissue is generated in a manner that may be considered as intramembranous-like. Overall, the findings here reported provide for the first time an accurate description of the tissues and events involved in the formation of the epiphyseal bone plate and gives insight into the complex cellular events underlying bone formation at different sites on the skeleton.

Evaluation of impaired growth plate development of long bones in skeletally immature mice by antirheumatic agents.

Restriction of physical growth and development is a known problem in patients with juvenile idiopathic arthritis (JIA). However, the effect of medical treatment for JIA on skeletal growth in affected children has not been properly investigated. We, therefore, hypothesize that naproxen and methotrexate (MTX) affect endochondral ossification and will lead to reduced skeletal development. Treatment of ATDC5 cells, an in vitro model for endochondral ossification, with naproxen or MTX resulted in increased chondrogenic but decreased hypertrophic differentiation. In vivo, healthy growing C57BL/6 mice were treated with naproxen, MTX, or placebo for 10 weeks. At 15 weeks postnatal, both the length of the tibia and the length of the femur were significantly reduced in the naproxen- and MTX-treated mice compared to their controls. Growth plate analysis revealed a significantly thicker proliferative zone, while the hypertrophic zone was significantly thinner in both experimental groups compared to their controls, comparable to the in vitro results. Micro-computed tomography analysis of the subchondral bone region directly below the growth disc showed significantly altered bone microarchitecture in naproxen and MTX groups. In addition, the involvement of the PTHrP-Ihh loop in naproxen- and MTX-treated cells was shown. Overall, these results demonstrate that naproxen and MTX have a profound effect on endochondral ossification during growth plate development, abnormal subchondral bone morphology, and reduced bone length. A better understanding of how medication influences the development of the growth plate will improve understanding of endochondral ossification and reveal possibilities to improve the treatment of pediatric patients.

Histological and molecular characterization of the growing nasal septum in mice.

The nasal septum is a cartilaginous structure that serves as a pacemaker for the development of the midface. The septum is a hyaline cartilage which is surrounded by a perichondrium and epithelium. It remains cartilaginous anteriorly, but posteriorly it undergoes endochondral ossification to form the perpendicular plate of the ethmoid. Understanding of hyaline cartilage differentiation stems predominantly from investigations of growth plate cartilage. It is currently unclear if the morphological and molecular properties of the differentiating nasal septum align with what is known from the growth plate. In this study, we describe growth, molecular, and cellular characteristics of the nasal septum with reference to hyaline cartilage differentiation. The nasal septum grows asynchronous across its length with phases of rapid growth interrupted by more stagnant growth. Growth appears to be driven predominantly by acquisition of chondrocyte hypertrophy. Similarly, cellular differentiation is asynchronous, and differentiation observed in the anterior part precedes posterior differentiation. Overall, the nasal septum is structurally and molecularly heterogeneous. Early and extensive chondrocyte hypertrophy but no ossification is observed in the anterior septum. Onset of hypertrophic chondrocyte differentiation coincided with collagen fiber deposition along the perichondrium. Sox9, Col2, Col10, Mmp13, Sp7, and Runx2 expression was heterogeneous and did not always follow the expected pattern established from chondrocyte differentiation in the growth plate. The presence of hypertrophic chondrocytes expressing bone-related proteins early on in regions where the nasal septum does not ossify displays incongruities with current understanding of hyaline cartilage differentiation. Runx2, Collagen II, Collagen X, and Sp7 commonly used to mark distinct stages of chondrocyte maturation and early bone formation show wider expression than expected and do not align with expected cellular characteristics. Thus, the hyaline cartilage of the nasal septum is quite distinct from growth plate hyaline cartilage, and caution should be taken before assigning cartilage properties to less well-defined cartilage structures using these commonly used markers. Beyond the structural description of the nasal cartilage, this study also provides important information for cartilage tissue engineering when using nasal septal cartilage for tissue regeneration.

Histochemical assessment on the cellular interplay of vascular endothelial cells and septoclasts during endochondral ossification in mice.

This study was aimed to verify the cellular interplay between vascular endothelial cells and surrounding cells in the chondro-osseous junction of murine tibiae. Many CD31-positive endothelial cells accompanied with Dolichos Biflorus Agglutinin lectin-positive septoclasts invaded into the hypertrophic zone of the tibial epiphyseal cartilage. MMP9 immunoreactive cytoplasmic processes of vascular endothelial cells extended into the transverse partitions of cartilage columns. In contrast, septoclasts included several large lysosomes which indicate the incorporation of extracellular matrices despite no immunopositivity for F4/80-a hallmark of macrophage/monocyte lineage. In addition, septoclasts were observed in c-fos-/- mice but not in Rankl-/- mice. Unlike c-fos-/- mice, Rankl-/- mice showed markedly expanded hypertrophic zone and the irregular shape of the chondro-osseous junction. Immunoreactivity of platelet-derived growth factor-bb, which involved in angiogenic roles in the bone, was detected in not only osteoclasts but also septoclasts at the chondro-osseous junction. Therefore, septoclasts appear to assist the synchronous vascular invasion of endothelial cells at the chondro-osseous junction. Vascular endothelial cells adjacent to the chondro-osseous junction possess endomucin but not EphB4, whereas those slightly distant from the chondro-osseous junction were intensely positive for both endomucin and EphB4, while being accompanied with ephrinB2-positive osteoblasts. Taken together, it is likely that vascular endothelial cells adjacent to the chondro-osseous junction would interplay with septoclasts for synchronous invasion into the epiphyseal cartilage, while those slightly distant from the chondro-osseous junction would cooperate with osteoblastic activities presumably by mediating EphB4/ephrinB2. MINI-ABSTRACT: Our original article demonstrated that vascular endothelial cells adjacent to the chondro-osseous junction would interplay with septoclasts for synchronous invasion into the epiphyseal cartilage, while those slightly distant from the chondro-osseous junction would cooperate with osteoblastic activities presumably by mediating EphB4/ephrinB2. (A figure that best represents your paper is Fig. 5c).

Enchondromatosis and Growth Plate Development.

PURPOSE OF REVIEW: Enchondroma is a common cartilage benign tumor that develops from dysregulation of chondrocyte terminal differentiation during growth plate development. Here we provide an overview of recent progress in understanding causative mutations for enchondroma, dysregulated signaling and metabolic pathways in enchondroma, and the progression from enchondroma to malignant chondrosarcoma. RECENT FINDINGS: Several signaling pathways that regulate chondrocyte differentiation are dysregulated in enchondromas. Somatic mutations in the metabolic enzymes isocitrate dehydrogenase 1 and 2 (IDH1/2) are the most common findings in enchondromas. Mechanisms including metabolic regulation, epigenetic regulation, and altered signaling pathways play a role in enchondroma formation and progression. Multiple pathways regulate growth plate development in a coordinated manner. Deregulation of the process can result in chondrocytes failing to undergo differentiation and the development of enchondroma.

Sanders stage 7b: Using the appearance of the ulnar physis improves decision-making for brace weaning in patients with adolescent idiopathic scoliosis.

AIMS: The aim of this study was to investigate whether including the stages of ulnar physeal closure in Sanders stage 7 aids in a more accurate assessment for brace weaning in patients with adolescent idiopathic scoliosis (AIS). METHODS: This was a retrospective analysis of patients who were weaned from their brace and reviewed between June 2016 and December 2018. Patients who weaned from their brace at Risser stage ≥ 4, had static standing height and arm span for at least six months, and were ≥ two years post-menarche were included. Skeletal maturity at weaning was assessed using Sanders staging with stage 7 subclassified into 7a, in which all phalangeal physes are fused and only the distal radial physis is open, with narrowing of the medial physeal plate of the distal ulna, and 7b, in which fusion of > 50% of the medial growth plate of distal ulna exists, as well as the distal radius and ulna (DRU) classification, an established skeletal maturity index which assesses skeletal maturation using finer stages of the distal radial and ulnar physes, from open to complete fusion. The grade of maturity at the time of weaning and any progression of the curve were analyzed using Fisher's exact test, with Cramer's V, and Goodman and Kruskal's tau. RESULTS: We studied a total of 179 patients with AIS, of whom 149 (83.2%) were female. Their mean age was 14.8 years (SD 1.1) and the mean Cobb angle was 34.6° (SD 7.7°) at the time of weaning. The mean follow-up was 3.4 years (SD 1.8). At six months after weaning, the rates of progression of the curve for patients weaning at Sanders stage 7a and 7b were 11.4% and 0%, respectively for those with curves of < 40°. Similarly, the rates of progression of the curve for those being weaned at ulnar grade 7 and 8 using the DRU classification were 13.5% and 0%, respectively. The use of Sanders stages 6, 7a, 7b, and 8 for the assessment of maturity at the time of weaning were strongly and significantly associated (Cramer's V 0.326; p = 0.016) with whether the curve progressed at six months after weaning. Weaning at Sanders stage 7 with subclassification allowed 10.6% reduction of error in predicting the progression of the curve. CONCLUSION: The use of Sanders stages 7a and 7b allows the accurate assessment of skeletal maturity for guiding brace weaning in patients with AIS. Weaning at Sanders stage 7b, or at ulnar grade 8 with the DRU classification, is more appropriate as the curve did not progress in any patient with a curve of < 40° immediately post-weaning. Thus, reaching full fusion in both distal radial and ulnar physes (as at Sanders stage 8) is not necessary and this allows weaning from a brace to be initiated about nine months earlier. Cite this article: Bone Joint J 2021;103-B(1):141-147.