Genetic Skeletal Disorders Research Articles

Heterogeneous spectrum of EXT gene mutations in Chinese patients with hereditary multiple osteochondromas.

Hereditary multiple osteochondroma (HMO) is one of the most common genetic skeletal disorders. It is caused by mutations in either EXT1 or EXT2 resulting in abnormal skeletal growth and morphogenesis. However, the spectrum and frequency of EXT1 and EXT2 mutations in Chinese patients with HMO was not previously investigated.Mutations were identified by performing Sanger sequencing analysis of the complete coding regions and flanking intronic sequences of EXT1 and EXT2, followed by multiplex ligation-dependent probe amplification (MLPA) analysis to detect gene deletions or duplications that could not be identified by the Sanger sequencing method.The present study identified pathogenic mutations in 93% (68/73) of unrelated HMO probands from 73 pedigrees. Mutations in EXT1 and EXT2 were identified in 53% (39/73) and 40% (29/73) of families. We identified 58 distinct mutations in EXT1 and EXT2, including 20 frameshift mutations, 16 nonsense mutations, 7 missense mutations, 9 splice site mutations, 5 large deletions, and 1 in-frame deletion mutation. Twenty-six of these mutations were novel and 32 were previously reported. Most of the mutations in EXT1 were base deletions or insertions (21/33), whereas the majority of those in EXT2 were single base substitution (18/25).Complete sequencing of both the EXT1 and EXT2 followed by MLPA analysis is recommended for genetic analysis of Chinese patients with HMO. This study provides a comprehensive characterization of the genetic aberrations found in Chinese patients with HMO and highlights the diagnostic value of molecular genetic analysis in this particular disease.

How good are we in predicting the risk of infant mortality for newborns undergoing “curative” surgery for structural congenital heart defects? A prospective, population-based cohort (EPICARD) study

Currently accepted birth prevalence for osteochondrodysplasias (OCDs) is about 2 per 10,000 births. Our main goal is to estimate the prevalence of OCDs in Argentina and compare it with other surveillance systems. We examined 1,663,610 births among 160 hospitals of RENAC (Red Nacional de Anomalías Congénitas - National Network of Congenital Anomalies) between November 2009 and December 2016. Cases were detected and registered according to a pre-established protocol, ranked in three diagnostic evidence levels according to available clinical documentation, and categorized according to the 9th edition of the nosology and classification of genetic skeletal disorders. Within our dataset, the most frequent groups were Group-1 (FGFR3, chondrodysplasia) and Group-25 (Osteogenesis Imperfecta and decreased bone density). Birth prevalence per 10,000 for the main OCD types, were: Achondroplasia 0.47 (95% CI: 0.38–0.59), Thanatophoric Dysplasia 0.37 (95% CI: 0.29–0.48), and the Osteogenesis Imperfecta group 0.34 (95% CI: 0.26–0.44). For total OCD, birth prevalence was 2.20 per 10.000 births (95% CI: 1.98–2.44). RENAC prevalence of total OCDs was found to be lower than that reported by the Latin-American Study of Congenital Malformations (ECLAMC) and Utah Birth Defect Network but higher than EUROCAT. Our investigation is the first study of OCD prevalence in Argentina using data from every jurisdiction of the country.

Targeted next-generation sequencing-based molecular diagnosis of congenital hand malformations in Chinese population

Congenital hand malformations is rare and characterized by hand deformities. It is highly heterogeneous, both clinically and genetically, which complicates the identification of causative genes and mutations. Recently, targeted next-generation (NGS) sequencing has been successfully used for the detection of heterogeneous diseases, and the use of NGS also has contributed significantly in evaluating the etiology of heterogeneous disease. Here, we employed targeted NGS to screen 248 genes involved in genetic skeletal disorders, including congenital hand malformations. Three pathogenic mutations located in the GJA1, ROR2 and TBX5 genes were detected in three large Chinese families with congenital hand malformations. Two novel mutations were reported, and a known causative mutation was verified in this Chinese population. This is also the first report that the same panel of targeted NGS was employed to perform molecular diagnosis of different subtypes of congenital hand malformations. Our study supported the application of a targeted NGS panel as an effective tool to detect the genetic cause for heterogeneous diseases in clinical diagnosis.

Mutation of FOP/FGFR1OP in mice recapitulates human short rib-polydactyly ciliopathy.

Skeletal dysplasias are a clinically and genetically heterogeneous group of bone and cartilage disorders. A total of 436 skeletal dysplasias are listed in the 2015 revised version of the nosology and classification of genetic skeletal disorders, of which nearly 20% are still genetically and molecularly uncharacterized. We report the clinical and molecular characterization of a lethal skeletal dysplasia of the short-rib group caused by mutation of the mouse Fop gene. Fop encodes a centrosomal and centriolar satellite (CS) protein. We show that Fop mutation perturbs ciliogenesis in vivo and that this leads to the alteration of the Hedgehog signaling pathway. Fop mutation reduces CSs movements and affects pericentriolar material composition, which probably participates to the ciliogenesis defect. This study highlights the role of a centrosome and CSs protein producing phenotypes in mice that recapitulate a short rib-polydactyly syndrome when mutated.

Diagnosis, follow-up and management of sleep disordered breathing in children with osteogenesis imperfecta

Osteogenesis imperfecta (OI) is the most common genetic skeletal disorder. Extra skeletal findings are common but association with sleep disordered breathing (SDB) have never been described. The aim of this study was to investigate clinical features of children with OI and suspicion of SDB. A retrospective study of patient's clinical records, signs of SDB and polysomnographic recordings was performed. We paid particular attention to symptoms which could be associated with SDB in this population; scoliosis, kyphosis, vertebral arthrodesis, chest wall deformities, basilar impression, autonomy; and data already known to be associated with obstructive sleep apnea such as body mass index and upper airway impairment. We reviewed the clinical charts of 188 patients referred to our genetic skeletal disorders reference center for OI. Among the 15 patients (8%) with polysomnographic recording, 12 (6.4%) had sleep disorder breathing. We found a negative correlation between Brief Assessment of Motor Function (BAMF) and Apnea Hypopnea Index (AHI) ( r = −0.68; P = 0.01) and Desaturation Index ( r = −0.62; P = 0.02). AHI tended to be higher in non-walkers (6.5 ± 3.6 versus 2.4 ± 1.5; P = 0.02), and in type III OI compared to type IV. Two patients were started on continuous positive airway pressure ventilation with clinical improvement. For OI children, symptoms suggestive of obstructive sleep disorders should be searched for systematically, especially when autonomy is compromised, body mass index is high, trunk deformations are identified, and in case of severe type of OI.

Diagnosis, follow-up and management of sleep-disordered breathing in children with osteogenesis imperfecta

ObjectivesOsteogenesis imperfecta (OI) is the most common genetic skeletal disorder. Extraskeletal findings are common but an association with sleep-disordered breathing (SDB) has never been described. The aim of this study was to investigate clinical features of children with OI and suspected SDB. MethodsA retrospective study of clinical records, signs of SDB and polysomnographic recordings of children with OI was performed. We paid particular attention to symptoms that could be associated with SDB in this population – scoliosis, kyphosis, vertebral arthrodesis, chest wall deformities, basilar impression, autonomy – as well as data already known to be associated with obstructive sleep apnea such as body mass index and upper-airway impairment. ResultsWe reviewed the clinical charts of 188 patients referred to our genetic skeletal disorders reference center for OI. Among the 15 patients (8%) with polysomnographic recordings, 12 (6.4%) had sleep-disordered breathing. We found a negative correlation between the Brief Assessment of Motor Function score and Apnea Hypopnea Index (r=−0.68; p=0.01) and Desaturation Index (r=−0.62; p=0.02). The Apnea Hypopnea Index was higher for non-walkers than walkers (mean [SD]: 6.5 [3.6] vs. 2.4 [1.5]; p=0.02) and with type III versus IV OI. Two patients were started on continuous positive airway pressure ventilation, with clinical improvement. ConclusionFor OI children, symptoms suggesting obstructive sleep disorders should be searched for systematically, especially in children with compromised autonomy, high body mass index, trunk deformations, and severe OI type.

Temporal Lobe Malformations in Achondroplasia: Expanding the Brain Imaging Phenotype Associated with FGFR3-Related Skeletal Dysplasias.

Thanatophoric dysplasia, achondroplasia, and hypochondroplasia belong to the fibroblast growth factor receptor 3 (FGFR3) group of genetic skeletal disorders. Temporal lobe abnormalities have been documented in thanatophoric dysplasia and hypochondroplasia, and in 1 case of achondroplasia. We retrospectively identified 13 children with achondroplasia who underwent MR imaging of the brain between 2002 and 2015. All children demonstrated a deep transverse temporal sulcus on MR imaging. Further common neuroimaging findings were incomplete hippocampal rotation (12 children), oversulcation of the mesial temporal lobe (11 children), loss of gray-white matter differentiation of the mesial temporal lobe (5 children), and a triangular shape of the temporal horn (6 children). These appearances are very similar to those described in hypochondroplasia, strengthening the association of temporal lobe malformations in FGFR3-associated skeletal dysplasias.

Overlapping genetic pathways in the skeletal dysplasias of a middle woodland individual: A case study

Studies of interacting/overlapping genetic skeletal disorders are rare for populations today, but even more so for archaeological contexts. The skeletal remains of an adult female (EZ 3-7-1) were excavated in the 1980s from the Middle Woodland (50BC–AD400) context of the Elizabeth site (11PK512) in the lower Illinois Valley (LIV), USA. Reported here are the standard score (z-score) comparisons of the measured skeletal differences of EZ 3-7-1 with a reference sample and a re-analysis of the individual’s pathological changes, with special consideration placed on refining the disease diagnosis. The impressive preservation and meticulous recovery of these skeletal remains have provided the opportunity to identify the first and earliest archaeological example of an individual (EZ 3-7-1) with a combined genetic skeletal dysplasia, Leri-Weill dyschondrosteosis and achondroplasia.

Two novel mutations in the PPIB gene cause a rare pedigree of osteogenesis imperfecta type IX

BackgroundOsteogenesis imperfecta (OI) is a rare genetic skeletal disorder characterized by increased bone fragility and vulnerability to fractures. PPIB is identified as a candidate gene for OI-IX, here we detect two pathogenic mutations in PPIB and analyze the genotype-phenotype correlation in a Chinese family with OI. MethodsNext-generation sequencing (NGS) was used to screen the whole exome of the parents of proband. Screening of variation frequency, evolutionary conservation comparisons, pathogenicity evaluation, and protein structure prediction were conducted to assess the pathogenicity of the novel mutations. Sanger sequencing was used to confirm the candidate variants. RTQ-PCR was used to analyze the PPIB gene expression. ResultsAll mutant genes screened out by NGS were excluded except PPIB. Two novel heterozygous PPIB mutations (father, c.25A>G; mother, c.509G>A) were identified in relation to osteogenesis imperfecta type IX. Both mutations were predicted to be pathogenic by bioinformatics analysis and RTQ-PCR analysis revealed downregulated PPIB expression in the two carriers. ConclusionWe report a rare pedigree with an autosomal recessive osteogenesis imperfecta type IX (OI-IX) caused by two novel PPIB mutations identified for the first time in China. The current study expands our knowledge of PPIB mutations and their associated phenotypes, and provides new information on the genetic defects associated with this disease for clinical diagnosis.

Orthopedic Manifestations of Type I Camurati-Engelmann Disease.

BackgroundCamurati-Engelmann disease (CED) is a rare genetic skeletal disorder characterized by limb pain, muscle emaciation and weakness, and cortical thickening of the diaphysis of long bones. It is caused by mutations in the transforming growth factor beta 1 (TGFB1) (type I) or other unknown gene(s) (type II). We present 8 consecutive patients with type I CED.MethodsWe retrospectively reviewed medical records and radiographs of type I CED patients with special reference to the mode of presentation, process of diagnostic work-up, and disease course. They were 4 sporadic patients, and two pairs of mother and son.ResultsWe categorized the mode of presentation into three groups. Group I had 4 patients who mainly presented with motor disturbances in young age. They drew medical attention for waddling gait, awkward ambulation or running, difficulty in going upstairs, or a positive Gower's sign at age 4 to 6 years. Subsequent development of limb pain and radiographic abnormality led to the diagnosis of CED at age 6 to 29 years. Group II had 3 patients who mainly presented with limb pain at age 15, 20, and 54 years, respectively. Radiographic evaluation and molecular genetic test led to the diagnosis of CED. The remaining 1 patient (group III) was asymptomatic until age 9 years when bony lesions at the tibiae were found incidentally. For the last 10 years, he intermittently complained of leg pain in the morning or after sports activities, which did not interfere with daily life. All the patients in group I showed a body mass index in the underweight range (< 18.4 kg/m2). At the latest follow-up, 4 patients in groups I and II required medication for the limb pain.ConclusionsCED presents with a wide range of severity. Awareness of this rare disease entity may be the key to timely correct diagnosis. This disease entity should be considered in the differential diagnosis of limb pain or motor disturbance in children to avoid unnecessary diagnostic work-up.

Neuroimaging Findings in Pediatric Genetic Skeletal Disorders: A Review.

Genetic skeletal disorders (GSDs) are a heterogeneous group characterized by an intrinsic abnormality in growth and (re-)modeling of cartilage and bone. A large subgroup of GSDs has additional involvement of other structures/organs beside the skeleton, such as the central nervous system (CNS). CNS abnormalities have an important role in long-term prognosis of children with GSDs and should consequently not be missed. Sensitive and specific identification of CNS lesions while evaluating a child with a GSD requires a detailed knowledge of the possible associated CNS abnormalities. Here, we provide a pattern-recognition approach for neuroimaging findings in GSDs guided by the obvious skeletal manifestations of GSD. In particular, we summarize which CNS findings should be ruled out with each GSD. The diseases (n = 180) are classified based on the skeletal involvement (1. abnormal metaphysis or epiphysis, 2. abnormal size/number of bones, 3. abnormal shape of bones and joints, and 4. abnormal dynamic or structural changes). For each disease, skeletal involvement was defined in accordance with Online Mendelian Inheritance in Man. Morphological CNS involvement has been described based on extensive literature search. Selected examples will be shown based on prevalence of the diseases and significance of the CNS involvement. CNS involvement is common in GSDs. A wide spectrum of morphological abnormalities is associated with GSDs. Early diagnosis of CNS involvement is important in the management of children with GSDs. This pattern-recognition approach aims to assist and guide physicians in the diagnostic work-up of CNS involvement in children with GSDs and their management.

Echocardiographic phenotype in osteogenesis imperfecta varies with disease severity

Background and objectivesOur purpose was to investigate cardiovascular abnormalities in children with osteogenesis imperfecta (OI).MethodsTwo hundred children (100 OI, 100 matched volunteers) were prospectively studied. Aortic and left ventricular (LV)...

Enhanced Wnt signaling improves bone mass and strength, but not brittleness, in the Col1a1+/mov13 mouse model of type I Osteogenesis Imperfecta

Osteogenesis Imperfecta (OI) comprises a group of genetic skeletal fragility disorders. The mildest form of OI, Osteogenesis Imperfecta type I, is frequently caused by haploinsufficiency mutations in COL1A1, the gene encoding the α1(I) chain of type 1 collagen. Children with OI type I have a 95-fold higher fracture rate compared to unaffected children. Therapies for OI type I in the pediatric population are limited to anti-catabolic agents. In adults with osteoporosis, anabolic therapies that enhance Wnt signaling in bone improve bone mass, and ongoing clinical trials are determining if these therapies also reduce fracture risk. We performed a proof-of-principle experiment in mice to determine whether enhancing Wnt signaling in bone could benefit children with OI type I. We crossed a mouse model of OI type I (Col1a1+/Mov13) with a high bone mass (HBM) mouse (Lrp5+/p.A214V) that has increased bone strength from enhanced Wnt signaling. Offspring that inherited the OI and HBM alleles had higher bone mass and strength than mice that inherited the OI allele alone. However, OI+HBM and OI mice still had bones with lower ductility compared to wild-type mice. We conclude that enhancing Wnt signaling does not make OI bone normal, but does improve bone properties that could reduce fracture risk. Therefore, agents that enhance Wnt signaling are likely to benefit children and adults with OI type 1.

A novel ACVR1 mutation detected by whole exome sequencing in a family with an unusual skeletal dysplasia

“Disorganized Development of Skeletal Component” (DDSC) is a group of genetic skeletal dysplasia, caused by mutations in 9 genes including ACVR1. The most known ACVR1-related disorder is fibrodysplasia ossificans progressiva (FOP). FOP variants are frequently encountered with diagnostic challenges due to overlapping clinical manifestations and variable severity. Application of high throughput sequencing methods can overcome these limitations by simultaneous investigation of the entire ACVR1 gene together with other genes involved in disorders with similar manifestations. A 33-year-old man with an unusual skeletal dysplasia and no previous clinical diagnosis is presented in this study. Whole exome sequencing detected a novel c.737T>A (p.Phe246Tyr) mutation in ACVR1 gene. Detailed targeted variant analysis in 226 known genes associated with genetic skeletal disorders together with more specific targeted analysis in 9 genes associated with DDSC ruled out the involvement of other investigated genes. Proband’s phenotypically normal father and brother had the same mutation in whom subsequent investigations showed subclinical radiographic findings.The clinical manifestations, the disease course, and the molecular findings of involvement of ACVR1 gene in this family are suggestive of “FOP variant” or an unusual ACVR1-related skeletal dysplasia. Moreover, this report has demonstrated the critical role of the next generation sequencing technique in characterizing such a rare disorder with variable and even no clinical manifestations, providing the opportunity for effective preventive measures such as preimplantation genetic diagnosis.

Nosology and classification of genetic skeletal disorders: 2015 revision

The purpose of the nosology is to serve as a "master" list of the genetic disorders of the skeleton to facilitate diagnosis and to help delineate variant or newly recognized conditions. This is the 9th edition of the nosology and in comparison with its predecessor there are fewer conditions but many new genes. In previous editions, diagnoses that were phenotypically indistinguishable but genetically heterogenous were listed separately but we felt this was an unnecessary distinction. Thus the overall number of disorders has decreased from 456 to 436 but the number of groups has increased to 42 and the number of genes to 364. The nosology may become increasingly important today and tomorrow in the era of big data when the question for the geneticist is often whether a mutation identified by next generation sequencing technology in a particular gene can explain the clinical and radiological phenotype of their patient. This can be particularly difficult to answer conclusively in the prenatal setting. Personalized medicine emphasizes the importance of tailoring diagnosis and therapy to the individual but for our patients with rare skeletal disorders, the importance of tapping into a resource where genetic data can be centralized and made available should not be forgotten or underestimated. The nosology can also serve as a reference for the creation of locus-specific databases that are expected to help in delineating genotype-phenotype correlations and to harbor the information that will be gained by combining clinical observations and next generation sequencing results.

Use of Targeted Exome Sequencing for Molecular Diagnosis of Skeletal Disorders.

Genetic disorders of the skeleton comprise a large group of more than 450 clinically distinct and genetically heterogeneous diseases associated with mutations in more than 300 genes. Achieving a definitive diagnosis is complicated due to the genetic heterogeneity of these disorders, their individual rarity and their diverse radiographic presentations. We used targeted exome sequencing and designed a 1.4Mb panel for simultaneous testing of more than 4,800 exons in 309 genes involved in skeletal disorders. DNA from 69 individuals from 66 families with a known or suspected clinical diagnosis of a skeletal disorder was analyzed. Of 36 cases with a specific clinical hypothesis with a known genetic basis, mutations were identified for eight cases (22%). Of 20 cases with a suspected skeletal disorder but without a specific diagnosis, four causative mutations were identified. Also included were 11 cases with a specific skeletal disorder but for which there was at the time no known associated gene. For these cases, one mutation was identified in a known skeletal disease genes, and re-evaluation of the clinical phenotype in this case changed the diagnoses from osteodysplasia syndrome to Apert syndrome. These results suggest that the NGS panel provides a fast, accurate and cost-effective molecular diagnostic tool for identifying mutations in a highly genetically heterogeneous set of disorders such as genetic skeletal disorders. The data also stress the importance of a thorough clinical evaluation before DNA sequencing. The strategy should be applicable to other groups of disorders in which the molecular basis is largely known.

A pilot study of gene testing of genetic bone dysplasia using targeted next-generation sequencing

Molecular diagnosis of genetic bone dysplasia is challenging for non-expert. A targeted next-generation sequencing technology was applied to identify the underlying molecular mechanism of bone dysplasia and evaluate the contribution of these genes to patients with bone dysplasia encountered in pediatric endocrinology. A group of unrelated patients (n=82), characterized by short stature, dysmorphology and X-ray abnormalities, of which mucopolysacharidoses, GM1 gangliosidosis, mucolipidosis type II/III and achondroplasia owing to FGFR3 G380R mutation had been excluded, were recruited in this study. Probes were designed to 61 genes selected according to the nosology and classification of genetic skeletal disorders of 2010 by Illumina's online DesignStudio software. DNA was hybridized with probes and then a library was established following the standard Illumina protocols. Amplicon library was sequenced on a MiSeq sequencing system and the data were analyzed by MiSeq Reporter. Mutations of 13 different genes were found in 44 of the 82 patients (54%). Mutations of COL2A1 gene and PHEX gene were found in nine patients, respectively (9/44=20%), followed by COMP gene in 8 (18%), TRPV4 gene in 4 (9%), FBN1 gene in 4 (9%), COL1A1 gene in 3 (6%) and COL11A1, TRAPPC2, MATN3, ARSE, TRPS1, SMARCAL1, ENPP1 gene mutations in one patient each (2% each). In conclusion, mutations of COL2A1, PHEX and COMP gene are common for short stature due to bone dysplasia in outpatient clinics in pediatric endocrinology. Targeted next-generation sequencing is an efficient way to identify the underlying molecular mechanism of genetic bone dysplasia.

Of mice and cats (both calico): Mary F Lyon, FRS (1925-2014).

Of mice and cats (both calico): Mary F Lyon, FRS (1925-2014).

Advances in Skeletal Dysplasia Genetics.

Skeletal dysplasias result from disruptions in normal skeletal growth and development and are a major contributor to severe short stature. They occur in approximately 1/5,000 births, and some are lethal. Since the most recent publication of the Nosology and Classification of Genetic Skeletal Disorders, genetic causes of 56 skeletal disorders have been uncovered. This remarkable rate of discovery is largely due to the expanded use of high-throughput genomic technologies. In this review, we discuss these recent discoveries and our understanding of the molecular mechanisms behind these skeletal dysplasia phenotypes. We also cover potential therapies, unusual genetic mechanisms, and novel skeletal syndromes both with and without known genetic causes. The acceleration of skeletal dysplasia genetics is truly spectacular, and these advances hold great promise for diagnostics, risk prediction, and therapeutic design.

Exome sequencing reveals a novel PTHLH mutation in a Chinese pedigree with brachydactyly type E and short stature

Brachydactyly includes shortening of digits due to abnormal development of phalanges, metacarpals, or both. It can occur either as an isolated malformation or with other anomalies as part of many congenital syndromes. It is included as one of the dysostosis groups affecting the limbs in the nosology and classification of genetic skeletal disorders. However, brachydactyly usually shows a high degree of phenotypic variability. In this study, we successfully identified a novel heterozygous mutation of the parathyroid hormone-like hormone (PTHLH) gene by exome sequencing in a Chinese pedigree with brachydactyly and short stature. The PTHLH gene encodes a parathyroid hormone-related protein (PTHrP) that is involved in the regulation of endochondral bone development, and mutations in this gene cause the type E form of brachydactyly. The mutation p.L15R occurs at a hydrophobic core region of the signal peptide, suggesting that this variation probably changes the signal peptide cleavage site at the in silico prediction. Further in vitro functional analysis showed that this mutation can lead to the retention of an N-terminal signal peptide fragment after the nascent proteins are translated.