Activated FGFR3 suppresses bone regeneration and bone mineralization in an ovariectomized mouse model.

BACKGROUND: Postmenopausal osteoporosis is a widespread health concern due to its prevalence among older adults and an associated high risk of fracture. The downregulation of bone regeneration delays fracture healing. Activated fibroblast growth factor receptor 3 (FGFR3) accelerates bone regeneration at juvenile age and downregulates bone mineralization at all ages. However, the impact of FGFR3 signaling on bone regeneration and bone mineralization post-menopause is still unknown. This study aimed to evaluate the impact of FGFR3 signaling on bone regeneration and bone mineralization during menopause by developing a distraction osteogenesis (DO) mouse model after ovariectomy (OVX) using transgenic mice with activated FGFR3 driven by Col2a1 promoter (Fgfr3 mice). METHODS: The OVX or sham operations were performed in 8-week-old female Fgfr3 and wild-type mice. After 8 weeks of OVX surgery, DO surgery in the lower limb was performed. The 5-day-latency period followed by performing distraction for 9 days. Bone mineral density (BMD) and bone regeneration was assessed by micro-computed tomography (micro-CT) scan and soft X-ray. Bone volume in the distraction area was also evaluated by histological analysis after 7 days at the end of distraction. Osteogenic differentiation and mineralization of bone marrow-derived mesenchymal stem cells (BMSCs) derived from each mouse after 8 weeks of the OVX or sham operations were also evaluated with and without an inhibitor for FGFR3 signaling (meclozine). RESULTS: BMD decreased after OVX in both groups, and it further deteriorated in Fgfr3 mice. Poor callus formation after DO was also observed in both groups with OVX, and the amount of regenerated bone was further decreased in Fgfr3 mice. Similarly, histological analysis revealed that Fgfr3 OVX mice showed lower bone volume. Osteogenic differentiation and mineralization of BMSCs were also deteriorated in Fgfr3 OVX mice. An inhibitor for FGFR3 signaling dramatically reversed the inhibitory effect of OVX and FGFR3 signaling on BMSC mineralization. CONCLUSION: Upregulated FGFR3 decreased newly regenerated bone after DO and BMD in OVX mice. FGFR3 signaling can be a potential therapeutic target in patients with postmenopausal osteoporosis.

Gain-of-Function of FGFR3 Accelerates Bone Repair Following Ischemic Osteonecrosis in Juvenile Mice.

Bone collapse, bone deformity, and a long treatment period are major clinical problems associated with juvenile ischemic osteonecrosis (JIO). Accelerating the process of bone repair in JIO is expected to shorten the treatment duration and better maintain morphology. We previously indicated that both bone formation and resorption were accelerated following distraction osteogenesis-mediated limb lengthening in genetically engineered mutant mice with a gain-of-function mutation in fibroblast growth factor receptor 3 (FGFR3) gene (i.e., Fgfr3 mice). The purpose of this study was to investigate the role of FGFR3 in the bone repair process following surgically induced ischemic osteonecrosis in the mutant mice. Epiphyseal deformity was less in the Fgfr3 mice compared to the wild-type mice at 6 weeks following ischemic osteonecrosis in skeletally immature age. Assessment of the morphology by micro-computed tomography (CT) revealed that the trabecular bone volume was increased in the Fgfr3 mice. Dynamic bone histomorphometry revealed increased rates of bone formation and mineral apposition in the Fgfr3 mice at 4 weeks post-surgery. The number of tartrate-resistant acid phosphatase (TRAP)-positive cells rapidly increased, and the numbers of TdT-mediated dUTP nick-end labeling (TUNEL)-positive cells rapidly decreased in the Fgfr3 mice. Vascular endothelial growth factor (VEGF) expression was increased at the earlier phase post-surgery in the Fgfr3 mice. The activation of FGFR3 signaling shortens the time needed for bone repair after ischemic osteonecrosis by accelerating revascularization, bone resorption, and new bone formation. Our findings are clinically relevant as a new potential strategy for the treatment of JIO.

Disease-specific complications and multidisciplinary interventions in achondroplasia.

Achondroplasia (ACH) is the most common skeletal dysplasia and characterized by a disproportionate short stature, macrocephaly with frontal bossing, exaggerated lumbar lordosis, and trident hands. It is induced by activated mutations in the fibroblast growth factor receptor 3 (FGFR3) gene. In addition to short stature, patients with ACH have a high prevalence of medical complications, including upper airway obstructive apnea, increased mortality, foramen magnum stenosis, hydrocephalus, developmental delay, recurrent ear infections, genu varum, obesity, and spinal canal stenosis, throughout their whole life. Several investigational drugs that modulate abnormal FGFR3 signaling have recently emerged, vosoritide being the most developed. This review presents the different disease-specific complications of ACH occurring in neonates, infants, childhood, adolescent, and adults and reports the current multidisciplinary interventions for these various complications. Moreover, we propose treatment strategies for children with ACH from the perspective of quality of life in adulthood.

Appropriate Surgical Timing of Salter Innominate Osteotomy for Residual Acetabular Dysplasia in Children.

BACKGROUND: Salter innominate osteotomy (SIO) provides favorable results for treating residual acetabular dysplasia in young children. In this study, we examined the midterm results of SIO according to the age at surgery to determine the optimal timing of this procedure. METHODS: We retrospectively examined 50 hips of 42 patients (8 boys and 34 girls) with acetabular dysplasia who underwent SIO and were followed up until skeletal maturity. The center-edge angle (CEA) was measured based on the anteroposterior radiographs of the hip obtained before surgery, 5 weeks after surgery, and at the latest follow-up. Severin classification was evaluated at the latest follow-up. Patients were categorized into 3 groups according to age at surgery: younger than 7 years of age (group A), 7 to 8 years of age (group B), and 9 years of age or older (group C). RESULTS: The mean preoperative CEA level of 0.9 degrees improved to 17.1 degrees postoperatively, which was increased to 28.1 degrees at the latest examination. Overall, 45 hips (90%) were classified as Severin I or II, with 96% in group A, 94% in group B, and 57% in group C. In group C, postoperative acetabular coverage was similar to that in the other groups (16.6 degrees in group A, 14.8 degrees in group B, and 18.1 degrees in group C), although the final outcome was unsatisfactory. The average improvement in CEA from postoperative to skeletal maturity was significantly smaller in group C than in the other groups (12.7 degrees in group A, 11.3 degrees in group B, and 3.0 degrees in group C). CONCLUSIONS: SIO showed favorable outcomes with satisfactory acetabular coverage at skeletal maturity. However, satisfactory acetabular coverage could not be obtained in some older patients because of limited postoperative remodeling capacity and smaller secondary improvement of CEA. We recommend that SIO should be performed in patients aged 8 years or younger. LEVEL OF EVIDENCE: Level III-retrospective comparative study.

Bypass of Epiphyseal Fragmentation Following Early Salter Innominate Osteotomy and Its Clinical Relevance in Legg-Calvé-Perthes Disease.

BACKGROUND: It has been demonstrated that early femoral varus osteotomy (FVO) produces a greater probability of skipping or interruption of epiphyseal fragmentation, thereby shortening the length of fragmentation stage for hips in the active stage of Legg-Calvé-Perthes disease. This "bypassing phenomenon" is thought to effect less disease severity or outcome, whereas it remains to be elucidated whether this phenomenon is specific to early FVO. We sought to investigate the presence and characteristics of the "bypassing phenomenon" following pelvic osteotomy performed in the avascular necrosis or early fragmentation stage as well as its correlation with disease severity and radiographic outcomes. METHODS: A retrospective review of data was conducted for 79 patients with unilateral Legg-Calvé-Perthes disease who had been diagnosed from 1987 to 2015, undergone the Salter innominate osteotomy (SIO) during the stage of avascular necrosis or in the early part of the fragmentation stage between 6.0 and 12.0 years of age, and followed up until skeletal maturity. Epiphyseal fragmentation was classified into 4 patterns according to a previous study. We compared lateral pillar groups and Stulberg grades between patients with and without bypass of the fragmentation stage. RESULTS: The mean age at surgery and follow-up period was 8.1 and 7.9 years, respectively. Sixty hips were in the Waldenström stage I and 19 hips in stage IIa at the surgery. In hips receiving SIO during stage I, the mean duration of the fragmentation stage was 276 days. The fragmentation pattern was typical for 40 hips, abortive for 17 hips, and atypical with horizontal fissure for 3 hips. Patients whose fragmentation was aborted experienced significantly less severe lateral pillar involvement and more favorable Stulberg outcomes at skeletal maturity. CONCLUSIONS: Incomplete bypass of epiphyseal fragmentation was observed in 28% of patients following early SIO performed in the avascular necrosis stage. In contrast to FVO, no patient bypassed fragmentation completely. Patients with incomplete bypass had a significantly higher proportion of less severe hips and a significantly greater probability of being associated with favorable radiographic outcomes compared with those without bypass. LEVEL OF EVIDENCE: Level IV-therapeutic study.

A cross-sectional nationwide survey of osteosclerotic skeletal dysplasias in Japan.

BACKGROUND: The osteosclerotic skeletal dysplasias (OSSDs) are a heterogeneous group of disorders characterized by systemic bone sclerosis. Little is known about OSSDs because of their rarity. We conducted a cross-sectional nationwide survey of OSSDs and examined the incidence, epidemiology, and therapeutic interventions on these disorders. METHODS: This study consisted of a two-step survey. The number of patients with OSSDs who had visited medical institutions between April 2017 and March 2018 was reported from a total of 341 facilities (1364 departments from pediatrics, orthopaedic surgery, neurosurgery, and otolaryngology in each facility) by the first questionnaire. In the secondary survey, their clinical features were assessed by collecting demographic data, diagnostic details, current status, family histories, therapeutic interventions, histories of bone fracture and osteomyelitis, severity assessed by the modified Rankin Scale (mRS) and recent lifestyle conditions of the patient by the EQ-5D. RESULTS: In the first survey, 51 facilities (56 departments) reported one or more OSSDs patients, including 50 patients with osteopetrosis and 57 patients of other OSSDs. Among 87 patients eligible for inclusion in the analysis in the secondary survey, we investigated detailed information on the 42 patients with osteopetrosis. The number of initial visits of osteopetrosis patients during the surveillance period was five per year, indicating that the estimated incidence of osteopetrosis seemed to be 0.6 per 100,000 live births. Eighty-six bone fractures were reported in 22 patients (52%), and interventions of pseudarthrosis were conducted in five patients. Nine patients (23%) showed significant disabilities with the mRS of grade 3 or higher. Neurological complications and severe anemia were the factors that deteriorate patients' quality of life. CONCLUSIONS: This is the first study to examine the detailed epidemiology of OSSDs in Japan. We demonstrated that the incidence of OSSDs is extremely rare. Bone fragility and delayed fracture healing seem to be important orthopaedic problems for patients with osteopetrosis.

Treatment approaches for congenital transverse limb deficiency: Data analysis from an epidemiological national survey in Japan.

BACKGROUND: Congenital limb deficiency is a rare anomaly that impairs limb function. Transverse deficiency accounts for approximately half of congenital limb deficiency cases. In Japan, there have been no detailed data of clinical features, especially treatment approaches, of this disorder. The present study aimed to investigate the status of treatment approaches of congenital transverse limb deficiency in Japan. METHODS: From the national epidemiological survey of congenital limb deficiency undertaken in Japan in 2016, all the data of 200 patients with congenital transverse limb deficiencies were extracted. These data were analysed to reveal the treatment approaches of congenital transverse limb deficiency and its basic clinical features. RESULTS: Surgical treatments and prosthetic or orthotic intervention had been implemented or planned for about one-quarter of patients, respectively. In the upper limb deficiencies, prosthetic or orthotic intervention was likely chosen in cases of deficiency at the metacarpal or proximal to metacarpal level. Surgical treatment was chosen only in cases of deficiency at the carpal or distal to carpal level. Although the number of patients with transverse lower limb deficiencies was small, prosthetic or orthotic intervention was likely chosen in proximal deficiencies, and surgical treatment was likely chosen in distal deficiencies. CONCLUSIONS: Herein, we revealed the status of treatment approaches for congenital transverse limb deficiency in Japan. Approximately half of the patients had no history of-and no plans for-surgical, prosthetic, or orthotic interventions. Further treatment advances may enable patients with congenital limb deficiencies to increase their participation in daily activities. STUDY DESIGN: Cross-sectional survey.

Impact of fracture characteristics and disease-specific complications on health-related quality of life in osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a connective tissue disease with bone fragility. Several studies have indicated that physical function in adult OI was correlated to the disease severity, but there have been no reports delineating the impact of the fracture characteristics and disease-specific complications on health-related quality of life (HRQoL). The purpose of this study is to clarify the factors impacted on HRQoL in adult OI patients. We conducted a cross-sectional study between July 2016 and March 2018 and sent a questionnaire regarding HRQoL using Short Form-36 (SF-36) to the OI patients at the age of 20 years or older who had a medical history of the investigators' institutions. The 40 patients completely answered the SF-36. Mental component summary and role/social component summary were unremarkable. Physical component summary (PCS) was significantly associated with z-score for height, teeth abnormality, and cardiopulmonary insufficiency (partial regression coefficient, 3.04, - 9.70, and - 11.35; p, < 0.001, 0.047, and 0.025, respectively). PCS was also significantly lower in the patients who had an initial fracture before the age of 2 years than those without occurrence of fractures until 2 years old (25.80 ± 17.15 versus 44.20 ± 16.54; p = 0.002), or those who had lower extremity fractures more than five times as compared with normal populations. Physical function was decreased in OI patients who had fractures before 2 years old, especially in lower extremity. Appropriate medical managements for cardiopulmonary insufficiency are required not only to maintain physical function but also to decrease mortality.

Fibrodysplasia ossificans progressiva: Review and research activities in Japan.

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic skeletal disorder manifesting progressive heterotopic ossification (HO) and congenital malformation of the great toes. Since 2007, we have conducted research on FOP. Here, we review the findings on FOP published to date, including the results of our research. Epidemiological studies in Japan have indicated that FOP has nearly the same prevalence in Japan as in the rest of the world. Basic research on its pathoetiology has progressed rapidly since the identification of the causal gene in 2006. Clinical and radiological findings have been thoroughly researched, including early radiological signs, and diagnostic criteria were established, designating FOP as an intractable disease in Japan. In patients with FOP, the progression of HO is associated with numerous disabilities, often manifesting in vicious cycles that can lead to early mortality. Through cross-sectional and short-term longitudinal studies, we have explored patient education, quality of life, and activities of daily living among Japanese patients. The management of FOP requires education of patients and caregivers, the use of medications to settle inflammation and flare-ups, instructions to ensure proper oral care, and other compensatory approaches that aid in rehabilitation. An avoidance of medical intervention, which may cause HO to progress, is also important. The advent of new drugs to prevent HO could have clinical benefit.

Lower limb pain following allogeneic hematopoietic stem cell transplantation in Japanese children.

BACKGROUND: Patients who undergo allogeneic hematopoietic stem cell transplantation (allo-HSCT) can experience musculoskeletal pains in their lower limbs in the early and late post-transplantation period. This study investigated demographics and clinical characteristics of the lower limb pain (LLP) among Japanese survivors of pediatric allo-HSCT. METHODS: A total of 143 consecutive Japanese patients who had undergone allo-HSCT less than 18 years of age in a single institute between 2005 and 2015 were reviewed. Patients referred for in-house orthopedic evaluation of their sustained LLPs that impaired ambulation were defined as LLP group. Illness/transplantation-related parameters were compared between LLP group and non-LLP group. RESULTS: Ninety children with a mean age of 8.5 years at transplantation were enrolled. Their median follow-up period following transplantation was 6.3 years (range, 2.1-12.5). There were four patients in LLP group, whose etiologies were AVN of the femoral head and insufficiency fracture (ISF) of the tibia or the medial cuneiform bone. Cumulative dose of steroids that administered from six months before transplantation to six months after discharge from hospitalization for transplantation was significantly higher in LLP group than non-LLP group. Additionally, the two groups differed significantly in terms of hospitalization period after transplantation. LLP caused by AVN of the femoral head manifested between six months and two years, whereas that caused by ISF within the first six months after transplantation. CONCLUSIONS: The incidence of sustained LLP that impairs ambulation following contemporary allo-HSCT is not common in Japanese pediatric survivors. The risk of developing musculoskeletal LLP may increase with a higher steroid dosage in the peri-transplant period. LLP caused by AVN of the femoral head is likely to manifest later than that caused by ISF.

Physical, Mental, and Social Problems of Adolescent and Adult Patients with Achondroplasia.

Patients with achondroplasia (ACH) require various medical interventions throughout the lifetime. Survey of health-related quality of life (HRQoL) in adult ACH patients is essential for the evaluation of treatment outcomes performed during childhood such as growth hormone administration and limb lengthening surgeries, but no study focused on the treatment strategy by analyzing HRQoL of ACH patients. The purpose of this study was to assess whether final height impacted on HRQoL and to evaluate what kinds of medical interventions were positively or negatively associated with HRQoL. We included 184 ACH patients (10-67 years old) who were registered in the patients' associations or who had a medical history of the investigators' institutions, and analyzed HRQoL by using Short Form-36 and patient demographics. Physical component summary (PCS) was significantly lower than the standard values in each age, especially in elderly populations, while mental component summary (MCS) was similar to the standard values. Role/social component summary was deteriorated only in elderly populations. The PCS was improved in the patients who had a height of 140 cm or taller (p < 0.001). The PCS and MCS were strongly associated with the past medical history of spine surgeries (p < 0.001 and p = 0.028, respectively). A treatment strategy would be planned to gain a final height of 140 cm or taller during childhood in combination with growth hormone administration and limb lengthening surgeries. Appropriate medical management for neurological complications of adult ACH patients is required to maintain physical and mental function.

Congenital limb deficiency in Japan: a cross-sectional nationwide survey on its epidemiology.

BACKGROUND: Congenital limb deficiency is a rare and intractable disease, which impairs both function and appearance of the limbs. To establish adequate medical care, it is necessary to reveal the actual conditions and problems associated with this disease. However, there have been no extensive epidemiological surveys in Japan addressing this disease. This is the first nationwide epidemiological survey of congenital limb deficiency in this country. METHODS: With the cooperation of epidemiology experts, we performed a two-stage nationwide survey to estimate the number of patients with congenital limb deficiency and reveal basic patient features. We targeted orthopaedic surgery, paediatric, and plastic surgery departments. Hospitals were categorized according to the institution type and the number of hospital beds; hospitals were randomly selected from these categories. We selected 2283 departments from a total 7825 departments throughout Japan. In this study, we defined congenital limb deficiency as partial or total absence of the limbs, proximal to the proximal interphalangeal joint of the fingers/lesser toes or interphalangeal joint of the thumb/great toe. We distributed the first survey querying the number of initial patient visits from January 2014 to December 2015. Targets of the second survey were departments that reported one or more initial patient visits in the first survey. RESULTS: In the first survey, 1767 departments responded (response rate: 77.4%). Among them, 161 departments reported one or more initial patient visits. We conducted the second survey among these 161 departments, of which 96 departments responded (response rate: 59.6%). The estimated number of initial visits by patients with congenital limb deficiency was 417 (95% confidence interval: 339-495) per year in 2014 and 2015. The estimated prevalence of congenital limb deficiency in Japan was 4.15 (95% confidence interval: 3.37-4.93) per 10,000 live births. The sex ratio was 1.40. Upper limbs were more affected than lower limbs. CONCLUSIONS: We revealed the estimated number of initial patient visits per year and birth prevalence of congenital limb deficiency in Japan. Our results will contribute to establishing the disease concept and grades of severity of congenital limb deficiency.

Early radiographic risk factors for rigid relapse in idiopathic clubfoot treated with the Ponseti method.

BACKGROUND: This study aimed at identifying early risk factors for rigid relapse in idiopathic clubfoot using foot radiographs. METHODS: Thirty-four patients with 43 idiopathic clubfeet treated with the Ponseti method were retrospectively reviewed. RESULTS: There were seven rigid relapse recalcitrant to manipulation and requiring extensive soft-tissue release. Three radiograabphic measurements on the maximum dorsiflexion lateral (MD-Lat) radiograph, talocalcaneal (TaloCalc-Lat), tibiocalcaneal (TibCalc-Lat), and calcaneus-first metatarsal (CalcMT1-Lat) angles, showed significant differences between patients with and without rigid relapse. The TaloCalc-Lat and CalcMT1-Lat angles showed significant hazard ratio for rigid relapse by multivariate survival analysis. Clubfeet demonstrating TibCalc-Lat>90° and CalcMT1-Lat<5° have a 24.9-fold odds ratio to develop rigid relapse compared to those demonstrating TibCalc-Lat≤90° or CalcMT1-Lat≥5°. CONCLUSIONS: The TaloCalc-Lat, TibCalc-Lat, and CalcMT1-Lat angles on the MD-Lat radiograph immediately before the tenotomy, probably representing intrinsic tightness of the midfoot and/or hindfoot, are significant risk factors for rigid relapse in patients treated with the Ponseti method.

Is the timing of surgery associated with avascular necrosis after unstable slipped capital femoral epiphysis? A multicenter study.

BACKGROUND: An unstable slipped capital femoral epiphysis (SCFE) is associated with a high rate of avascular necrosis (AVN). The etiology of AVN seems to be multifactorial, although it is not thoroughly known. The aims of our study were to determine the rate of AVN after an unstable SCFE and to investigate the risk factors for AVN, specifically evaluating the notion of an "unsafe window", during which medical interventions would increase the risk for AVN. METHODS: This retrospective multicenter study included 60 patients with an unstable SCFE diagnosed between 1985 and 2014. Timing of surgery was evaluated for three time periods, from acute onset of symptoms to surgery: period I, <24 h; period II, between 24 h and 7 days; and period III, >7 days. Multivariate logistic regression analysis was used to identify risk factors for AVN. RESULTS: Closed reduction and pinning was performed in 43 patients and in situ pinning in 17. Among these cases, 16 patients (27%) developed AVN. The rate of AVN was significantly higher in patients treated by closed reduction and pinning (15/43, 35%) than in those treated by in situ pinning (1/17, 5.9%) (p = 0.022). In patients treated by closed reduction and pinning, the incidence of AVN was 2/11 (18%) in period I, 10/13 (77%) in period II and 3/15 (20%) in period III, showing the significantly higher rate in period II (p = 0.002). The surgery provided in period II was identified as an independent risk factor for the development of AVN. CONCLUSIONS: Our rate of AVN was 27% using two classical treatment methods. Time-to-surgery, between 24 h and 7 days, was independently associated with AVN, supporting the possible existence of an "unsafe window" in patients with unstable SCFE treated by closed reduction and pinning.

Mutations in B3GALT6, which encodes a glycosaminoglycan linker region enzyme, cause a spectrum of skeletal and connective tissue disorders.

Proteoglycans (PGs) are a major component of the extracellular matrix in many tissues and function as structural and regulatory molecules. PGs are composed of core proteins and glycosaminoglycan (GAG) side chains. The biosynthesis of GAGs starts with the linker region that consists of four sugar residues and is followed by repeating disaccharide units. By exome sequencing, we found that B3GALT6 encoding an enzyme involved in the biosynthesis of the GAG linker region is responsible for a severe skeletal dysplasia, spondyloepimetaphyseal dysplasia with joint laxity type 1 (SEMD-JL1). B3GALT6 loss-of-function mutations were found in individuals with SEMD-JL1 from seven families. In a subsequent candidate gene study based on the phenotypic similarity, we found that B3GALT6 is also responsible for a connective tissue disease, Ehlers-Danlos syndrome (progeroid form). Recessive loss-of-function mutations in B3GALT6 result in a spectrum of disorders affecting a broad range of skeletal and connective tissues characterized by lax skin, muscle hypotonia, joint dislocation, and spinal deformity. The pleiotropic phenotypes of the disorders indicate that B3GALT6 plays a critical role in a wide range of biological processes in various tissues, including skin, bone, cartilage, tendon, and ligament.

Tranilast stimulates endochondral ossification by upregulating SOX9 and RUNX2 promoters.

Endochondral ossification is an essential process for reparative phase of fracture healing, which starts with the differentiation of mesenchymal cells into chondrocytes followed by substitution of bone tissue. It is strictly controlled by the expression of crucial transcriptional factors: SOX9 in the early phase and RUNX2 in the late phase. Screening of FDA-approved compounds revealed that an anti-allergic drug, tranilast, that has been used for more than 30 years in clinical practice, enhanced the SOX9 promoter in chondrogenic cells and the RUNX2 promoter in osteoblastic cells. We observed that tranilast increased mRNA expression of both Sox9 and Runx2 in differentiating ATDC5 chondrogenic progenitor cells. Tranilast upregulated mRNA expression of chondrogenic marker genes (Col2a1, Acan, Col10a1, and Mmp13) in differentiating ATDC5 cells. Moreover, tranilast upregulated mRNA expression of essential signaling molecules involved in endochondral ossification (Pthrp, Ihh, and Axin2). In the later phase of differentiation of ATDC5 cells, tranilast increased synthesis of matrix proteoglycans, induced the alkaline phosphatase activity, and tended to accelerate mineralization. Tranilast is a potential agent that accelerates fracture repair by promoting the regulatory steps of endochondral ossification.

Characterisation of novel RUNX2 mutation with alanine tract expansion from Japanese cleidocranial dysplasia patient.

Cleidocranial dysplasia (CCD; MIM 119600) is an autosomal dominant skeletal dysplasia characterised by hypopalstic and/or aplastic clavicles, midface hypoplasia, absent or delayed closure of cranial sutures, moderately short stature, delayed eruption of permanent dentition and supernumerary teeth. The molecular pathogenesis can be explained in about two-thirds of CCD patients by haploinsufficiency of the RUNX2 gene. In our current study, we identified a novel and rare variant of the RUNX2 gene (c.181_189dupGCGGCGGCT) in a Japanese patient with phenotypic features of CCD. The insertion led an alanine tripeptide expansion (+3Ala) in the polyalanine tract. To date, a RUNX2 variant with alanine decapeptide expansion (+10Ala) is the only example of a causative variant of RUNX2 with polyalanine tract expansion to be reported, whilst RUNX2 (+1Ala) has been isolated from the healthy population. Thus, precise analyses of the RUNX2 (+3Ala) variant were needed to clarify whether the tripeptide expanded RUNX2 is a second disease-causing mutant with alanine tract expansion. We therefore investigated the biochemical properties of the mutant RUNX2 (+3Ala), which contains 20 alanine residues in the polyalanine tract. When transfected in COS7 cells, RUNX2 (+3Ala) formed intracellular ubiquitinated aggregates after 24h, and exerted a dominant negative effect in vitro. At 24h after gene transfection, whereas slight reduction was observed in RUNX2 (+10Ala), all of these mutants significantly activated osteoblast-specific element-2, a cis-acting sequence in the promoter of the RUNX2 target gene osteocalcin. The aggregation growth of RUNX2 (+3Ala) was clearly lower and slower than that of RUNX2 (+10Ala). Furthermore, we investigated several other RUNX2 variants with various alanine tract lengths, and found that the threshold for aggregation may be RUNX2 (+3Ala). We conclude that RUNX2 (+3Ala) is the cause of CCD in our current case, and that the accumulation of intracellular aggregates in vitro is related to the length of the alanine tract.

Characteristic calcaneal ossification: an additional early radiographic finding in infants with fibrodysplasia ossificans progressiva.

BACKGROUND: We have clinically encountered children with fibrodysplasia ossificans progressiva who had abnormal calcaneal ossification. OBJECTIVE: To evaluate whether calcaneal ossification variants are significant radiographic findings in children with fibrodysplasia ossificans progressiva. MATERIALS AND METHODS: Lateral feet radiographs in nine children who fulfilled the diagnostic criteria of fibrodysplasia ossificans progressiva were reviewed. The studies were obtained during infancy or early childhood. RESULTS: Fourteen lateral foot radiographs of fibrodysplasia ossificans progressiva were available for this study (ages at examination: 1-104 months). Four children ages 2 months to 11 months showed double calcaneal ossification centers; 7 children had plantar calcaneal spurs that decreased in size with age. Overall, eight of nine children with fibrodysplasia ossificans progressiva demonstrated double calcaneal ossifications and/or plantar calcaneal spurs in infancy or childhood. CONCLUSION: Double calcaneal ossification centers in early infancy and plantar calcaneal spurs in childhood are frequently seen in children with fibrodysplasia ossificans progressiva and may be a useful radiologic indicator for early diagnosis.

Low bone mineral density in achondroplasia and hypochondroplasia.

BACKGROUND: Achondroplasia (ACH) and hypochondroplasia (HCH) are the most common form of short-limb skeletal dysplasias caused by activated fibroblast growth factor receptor 3 (FGFR3) signaling. Although decreased bone mass was reported in gain-of-function mutation in Fgfr3 mice, both disorders have never been described as osteoporotic. In the present study, we evaluated bone mineral density (BMD) in ACH and HCH patients. METHODS: We measured spinal BMD (L1-L4) in 18 ACH and four HCH patients with an average age of 19.8 ± 7.5 years (range, 10-33 years). BMD Z-score in each individual was calculated for normalizing age and gender. Correlation between body mass index (BMI) and BMD was analyzed. Moreover, BMD and Z-score were compared between ACH patients and HCH patients. RESULTS: The average BMD of ACH/HCH patients was 0.805 ± 0.141 g/cm(2) (range, 0.554-1.056 g/cm(2) ), resulting in an average Z-score of -1.1 ± 0.8 (range, -2.4 to 0.6) of the standard value. A slightly positive correlation was observed between BMI and BMD (r = 0.45; P = 0.13). There was no significant difference in BMD and Z-score between ACH and HCH patients. CONCLUSION: Spinal BMD was reduced in ACH/HCH patients, and was mildly correlated with individual BMI. We should carefully monitor BMD and examine osteoporosis-related symptoms in adolescent and adult ACH/HCH patients. © 2016 Japan Pediatric Society.