Identification of FAT3 as a new candidate gene for adolescent idiopathic scoliosis.

In an effort to identify rare alleles associated with adolescent idiopathic scoliosis (AIS) whole-exome sequencing was performed on a discovery cohort of 73 unrelated patients and 70 age-and sex matched controls, all of French-Canadian ancestry. A collapsing gene burden test was performed to analyze rare protein-altering variants using case-control statistics. Since no single gene achieved statistical significance, targeted exon sequencing was performed for 24 genes with the smallest p values, in an independent replication cohort of unrelated severely affected females with AIS and sex-matched controls (N = 96 each). An excess of rare, potentially protein-altering variants was noted in one particular gene, FAT3, although it did not achieve statistical significance. Independently, we sequenced the exomes of all members of a rare multiplex family of three affected sisters and unaffected parents. All three sisters were compound heterozygous for two rare protein-altering variants in FAT3. The parents were single heterozygotes for each variant. The two variants in the family were also present in our discovery cohort. A second validation step was done, using another independent replication cohort of 258 unrelated AIS patients having reach their skeletal maturity and 143 healthy controls to genotype nine FAT3 gene variants, including the two variants previously identified in the multiplex family: p.L517S (rs139595720) and p.L4544F (rs187159256). Interestingly, two FAT3 variants, rs139595720 (genotype A/G) and rs80293525 (genotype C/T), were enriched in severe scoliosis cases (4.5% and 2.7% respectively) compared to milder cases (1.4% and 0.7%) and healthy controls (1.6% and 0.8%). Our results implicate FAT3 as a new candidate gene in the etiology of AIS.

Spinal malformation - A biochemical analysis using congenital kyphosis rats.

Spinal kyphosis involves the vertebrae curving excessively backward, beyond their physiological curvature. Although the normal structure of the spinal vertebrae is extremely important for maintaining posture, the normal function of the thoracic and abdominal organs, and cosmetics, our knowledge concerning the pathogenesis of this disease is lacking. Furthermore, the responsible gene has not yet been identified. In this short review, we summarize the current state of kyphosis research and introduce the molecular and cellular mechanisms associated with the pathogenesis of this disease, based on findings obtained using rats that develop kyphosis.

Paraspinal myopathy-induced intervertebral disc degeneration and thoracolumbar kyphosis in TSC1mKO mice model-a preliminary study.

BACKGROUND: Increasing kyphosis of the spine in a human is a well-recognized clinical phenomenon that has been associated with back pain, poor physical performance and disability. The pathophysiology of age-related kyphosis is complex and has been associated with physiological changes in vertebrae, intervertebral disc (IVD) and paraspinal musculature, which current cross-sectional studies are unable to demonstrate. Creating an in vivo, paraspinal myopathic animal model for longitudinal study of these changes under controlled conditions is thus warranted. PURPOSE: To confirm the TSC1 gene knockout effect on paraspinal muscle musculature; to analyze the development of spinal kyphosis, IVD degeneration and vertebra structural changes in a longitudinal manner to gain insights into the relationship between these processes. STUDY DESIGN: A prospective cohort study of 28 female mice, divided into 4 groups-9-month-old TSC1mKO (n=7), 9-month-old control (n=4), 12-month-old TSC1mKO (n=8), and 12-month-old controls (n=9). METHODS: High resolution micro-computed tomography was used to measure sagittal spinal alignment (Cobb's angle), vertebral height, vertebral body wedging, disc height index (DHI), disc wedge index (DWI), histomorphometry of trabecular bone and erector spinae muscle cross-sectional area. Paraspinal muscle specimens were harvested to assess for myopathic features with H&E stain, muscle fiber size, density of triangular fiber and central nucleus with WGA/DAPI stain, and percentage of fibers with PGC-1α stain. Intervertebral discs were evaluated for disc score using FAST stain. RESULTS: Compared to controls, paraspinal muscle sections revealed features of myopathy in TSC1mKO mice similar to human sarcopenic paraspinal muscle. While there was significantly greater presence of small triangular fiber and density of central nucleus in 9-and 12-month-old TSC1mKO mice, significantly larger muscle fibers and decreased erector spinae muscle cross-sectional area were only found in 12-month-old TSC1mKO mice compared to controls. TSC1mKO mice developed accelerated thoracolumbar kyphosis, with significantly larger Cobb angles found only at 12 months old. Structural changes to the trabecular bone in terms of higher bone volume fraction and quality, as well as vertebral body wedging were observed only in 12-month-old TSC1mKO mice when compared to controls. Disc degeneration was observed as early as 9 months in TSC1mKO mice and corresponded with disc wedging. However, significant disc height loss was only observed when comparing 12-month-old TSC1mKO mice with controls. CONCLUSIONS: This study successfully shows the TSC1 gene knockout effect on the development of paraspinal muscle myopathy in a mouse which is characteristic of sarcopenia. The TSC1mKO mice is by far the best model available to study the pathological consequence of sarcopenia on mice spine. With paraspinal muscle myopathy established as early as 9 months, TSC1mKO mice developed disc degeneration and disc wedging. This is followed by kyphosis of the spine at 12 months with concomitant disc height loss and vertebral body wedging due to bone remodeling. Age-related bone loss was not found in our study, suggesting osteoporosis and myopathy-induced vertebral body wedging are likely two independent processes. CLINICAL SIGNIFICANCE: This is the first study to provide key insights on the early and late consequences of paraspinal myopathy on intervertebral disc degeneration, spinal kyphosis, and vertebral body changes. With this new understanding, future studies evaluating therapies for spinal degeneration may be performed to develop time-sensitive interventions.

Identical twins with progressive kyphoscoliosis and ophthalmoplegia.

The possible differential diagnoses for children presenting with kyphoscoliosis, skeletal deformities and ophthalmoplegia are diverse. We present 11-year-old identical twins with these symptoms, with interesting etiological concern for those practicing in the fields of neurology, pediatrics, spine surgery and related specialties. A new presentation for a rare genetic condition was the final diagnosis for our patients. In this movement disorder round we describe our approach to this clinical constellation and discuss clinical significance of this genetic condition.

Two novel variants in PLOD1 causing hydrocephalus in female newborn with kyphoscoliotic Ehlers-Danlos syndrome.

The kyphoscoliotic Ehlers-Danlos syndrome (kEDS) is a rare autosomal recessive connective tissue disorder characterized by hyperextensible skin and joints, kyphoscoliosis, and severe muscle hypotonia at birth. Causal variants have been identified in PLOD1 resulting in lysyl hydroxylase deficiency responsible for kEDS. However, the detailed phenotype of kEDS during the perinatal period is still poorly recognized. Here, we describe a case of a female newborn presenting with prenatal hydrocephalus and severe hypotonia after birth with two novel compound heterozygous variants, c.2T > C (p.?) and c.1462del (p. Arg488Glyfs*9) in the PLOD1 gene. Our case suggests that in addition to the reported phenotype during the neonatal period, prenatal hydrocephalus should also be differentially diagnosed to exclude the potential of kEDS.

Female-Specific Susceptibility Locus in BOC and SEC16B are Associated with Adolescent Idiopathic Scoliosis.

STUDY DESIGN: A genetic case-control study. OBJECTIVES: To investigate whether the variants in BOC, SEC16B, and SH2D1B are sex-specifically and functionally associated with the susceptibility of adolescent idiopathic scoliosis (AIS) in Chinese Han population. SUMMARY OF BACKGROUND DATA: A recent genome-wide association study identified three female-specific susceptibility loci of AIS in Japanese population. However, the association of these genes with AIS in other populations remains unclear. Further investigation of the functional role of the three genes was warranted. METHODS: SNPs rs73235136, rs545608, and rs142502288 were genotyped in 1599 AIS patients and 2985 controls. Paraspinal muscle collected from 40 AIS and 30 lumber disc herniation patients during surgical interventions was used for gene expression analysis. The difference regarding genotype and allele frequency between patients and controls was analyzed by chi-square analysis. Expression of BOC and SEC16B was compared between AIS and lumber disc herniation patients by the Student t test. Pearson correlation analysis was performed to evaluate the relationship between gene expression level and clinical phenotypes. RESULTS: SNPs rs73235136 of BOC and rs545608 of SEC16B were found to be remarkably associated with AIS only in females. Allele C of rs73235136 and allele G of rs545608 could significantly add to the risk of female AIS patients, with an odds ratio of 1.087 and 1.033, respectively. However, there was no significant difference between the male patients and controls regarding genotype or allele frequency of rs73235136 and rs545608. No polymorphism at rs142502288 was detected in either patients or controls, and all the subjects had genotype of AA. Moreover, tissue expression of BOC and SEC16B was significantly lower in AIS patients compared with controls. BOC expression was positively associated with bone mineral contents, and expression of SEC16B was negatively correlated with curve severity in AIS patients. CONCLUSION: Female-specific variants in BOC and SEC16B were associated with AIS. Expression of BOC and SEC16B was significantly lower in AIS patients. The role of BOC and SEC16B in the development of AIS is worthy of further investigation.Level of Evidence: 3.

Recurrence of an early postzygotic rescue of an inherited unbalanced translocation resulting in mosaic segmental uniparental isodisomy of chromosome 11q in siblings.

Balanced translocations are associated with a risk of transmission of unbalanced chromosomal rearrangements in the offspring. Such inherited chromosomal abnormalities are typically non-mosaic as they are present in the germline. We report the recurrence in two siblings of a mosaicism for a chromosomal rearrangement inherited from their asymptomatic father who carried a balanced t(2;11)(q35;q25) translocation. Both siblings exhibited a similar phenotype including intellectual disability, dysmorphic features, kyphoscoliosis, and cervical spinal stenosis. Karyotyping, fluorescence in situ hybridization and SNP array analysis of blood lymphocytes of both siblings identified two cell lines: one carrying a 2q35q37.3 duplication and a 11q25qter deletion (~90% cells), and one carrying an 11q uniparental isodisomy of maternal origin (~10% cells). We hypothesize that these mosaics were related to a postzygotic rescue mechanism which unexpectedly recurred in both siblings.

Musculoskeletal abnormalities in a large international cohort of boys with 49,XXXXY.

49,XXXXY is the rarest X and Y chromosomal variation, with an incidence of 1 in 80,000-100,000 live male births and has been associated with numerous musculoskeletal abnormalities. Data was collected from an international cohort of boys with 49,XXXXY over 10 years. Children were evaluated by a multidisciplinary team consisting of a pediatric orthopedist, a neurogeneticist, a neurodevelopmentalist, and two physical therapists. Increased rates of torticollis (32.4%), hamstring tightness (42%), radioulnar synostosis (67.6%), pes planus (65.2%), and other foot abnormalities (86.9%) were observed. Several anomalies increased with age, specifically hamstring tightness, kyphosis, and scoliosis. The elucidation of the orthopedic profile of this population is necessary in order to provide healthcare providers with current medical information. This research further supports the necessity for the comprehensive multidisciplinary treatment of boys with 49,XXXXY.

The first case report of Kyphoscoliotic Ehlers-Danlos syndrome of chinese origin with a novel PLOD1 gene mutation.

BACKGROUND: Kyphoscoliotic Ehlers-Danlos syndrome (kEDS) is a rare autosomal recessive connective tissue disorder characterized by progressive kyphoscoliosis, congenital muscular hypotonia, marked joint hypermobility, and severe skin hyperextensibility and fragility. Deficiency of lysyl hydroxylase 1 (LH1) due to mutations of PLOD1 (procollagen-lysine, 2-oxoglutarate 5-dioxygenase 1) gene has been identified as the pathogenic cause of kEDS (kEDS-PLOD1). Up to now, kEDS-PLOD1 has not been reported among Chinese population. CASE PRESENTATION: A 17-year-old Chinese male patient presenting with hypotonia, joint hypermobility and scoliosis was referred to our hospital. After birth, he was found to have severe hypotonia leading to delayed motor development. Subsequently, joint hypermobility, kyphoscoliosis and amblyopia were found. Inguinal hernia was found at age 5 years and closed by surgery. At the same time, he presented with hyperextensible and bruisable velvety skin with widened atrophic scarring after minor trauma. Dislocation of elbow joint was noted at age of 6 years. Orthopedic surgery for correction of kyphoscoliosis was performed at age 10 years. His family history was unremarkable. Physical examination revealed elevated blood pressure. Slight facial dysmorphologies including high palate, epicanthal folds, and down-slanting palpebral fissures were found. He also had blue sclerae with normal hearing. X-rays revealed severe degree of scoliosis and osteopenia. The Echocardiography findings were normal. Laboratory examination revealed a slightly elevated bone turnover. Based on the clinical manifestations presented by our patient, kEDS was suspected. Genetic analysis revealed a novel homozygous missense mutation of PLOD1 (c.1697 G > A, p.C566Y), confirming the diagnosis of kEDS-PLOD1. The patient was treated with alfacalcidol and nifedipine. Improved physical strength and normal blood pressure were reported after 12-month follow-up. CONCLUSIONS: This is the first case of kEDS-PLOD1 of Chinese origin. We identified one novel mutation of PLOD1, extending the mutation spectrum of PLOD1. Diagnosis of kEDS-PLOD1 should be considered in patients with congenital hypotonia, progressive kyphoscoliosis, joint hypermobility, and skin hyperextensibility and confirmed by mutation analysis of PLOD1.

Natural history of achondroplasia: A retrospective review of longitudinal clinical data.

There are limited data on the longitudinal frequency and severity of the symptoms and complications of achondroplasia. We undertook a retrospective electronic chart review of 114 patients to develop a more thorough understanding of the lifetime impact of achondroplasia. Craniocervical stenosis (involving the foramen magnum with or without cervical vertebrae C1 and/or C2) was noted in nearly 50% of patients with craniovertebral junction imaging; however, corrective decompression surgery was only needed in 6% of patients. No children in our cohort died at 4 years of age or under. Kyphosis was present in most patients but usually resolved in early childhood. Cervical and lumbar stenosis were diagnosed in children and adults while, genu varum, elbow contractures, and radial head dislocations were identified during childhood. Central sleep apnea and obstructive sleep apnea were present in children, while the diagnosis of obstructive sleep apnea was shown to recur in adulthood. Cardiovascular risk factors were present in only 7% of patients. A range of mental health disorders were identified, with most diagnoses being made before 18 years of age. Our data show that achondroplasia has a significant impact on patients' physical health, and complications continue to be reported and require intervention throughout patients' lifetimes. This highlights the need for continuous support beyond pediatric care, by adult care clinicians experienced with managing the long-term complications of achondroplasia.

Upregulated miR-224-5p suppresses osteoblast differentiation by increasing the expression of Pai-1 in the lumbar spine of a rat model of congenital kyphoscoliosis.

Congenital scoliosis is defined by the presence of structural anatomical malformations that arise from failures of vertebral formation or segmentation before and after birth. The understanding of genetic background and key genes for congenital scoliosis is still poor. We herein report that the excess expression of plasminogen activator inhibitor-1 (Pai-1) induced by the upregulation of miR-224-5p is involved in the pathogenesis of congenital kyphoscoliosis through impaired osteoblast differentiation. We first investigated the variety and progression of abnormalities of the lumbar spines in Ishibashi (IS) rats, a rat model of congenital kyphoscoliosis. The rats had already shown fusion and division of the primary ossification center at postnatal day 4. Over time, the rats showed various abnormalities of the lumbar spine, including the fusion of the annular epiphyseal nucleus. At postnatal day 42, spinal curvature was clearly observed due to the fusion of the vertebral bodies. Using a microRNA array, we found that the expression of miR-224-5p was increased in the lumbar spine of the rats at postnatal day 4. The expression of Pai-1, which is involved in osteoblast differentiation regulated by miR-224-5p, was also increased, while the levels of type I collagen, a marker of osteoblast differentiation, were decreased in the lumbar spine. These results indicate that the aberrant expression of miRNA-224-5p and its target genes is involved in the impaired osteoblast differentiation and may provide a partial molecular explanation for the pathogenesis of congenital scoliosis.

Posterior Spinal Fusion for Severe Spinal Deformities in Musculocontractural Ehlers-Danlos Syndrome: Detailed Observation of a Novel Case and Review of 2 Reported Cases.

BACKGROUND: Musculocontractural Ehlers-Danlos syndrome caused by pathogenic variants in CHST14 (mcEDS-CHST14) is a recently delineated connective tissue disorder characterized by multisystem congenital malformations and progressive connective tissue fragility-related manifestations. With only 2 cases of mcEDS-CHST14 containing precise information on surgical spinal correction being reported to date, there remains no consensus on treatment standards. This study describes the detailed clinical and radiologic outcomes of the third known patient with mcEDS-CHST14 who successfully underwent surgery for severe kyphoscoliosis. CASE DESCRIPTION: The patient was a 19-year-old girl with mcEDS-CHST14 who suffered from low back pain and decreased daily activities caused by progressive kyphoscoliosis. She underwent posterior spinal fusion with an all-pedicle screw construct from T4 to L4 for a preoperative main curve Cobb angle of 69 degrees and kyphotic angle of 27 degrees. Postoperative Cobb angle of the main curve and kyphotic angle were 26 and 6 degrees, respectively. Although sufficient correction was achieved without disseminated intravascular coagulation or other serious sequelae, a large amount of blood (2600 g) was lost due to tissue fragility. Her low back pain was decreased at 1 year after surgery. CONCLUSIONS: On the basis of the present and 2 earlier reported cases, posterior spinal fusion may be a reasonable surgical option for severe progressive spinal deformities in patients with mcEDS-CHST14. However, careful attention is needed for possible massive blood loss from tissue fragility.

Clinical Observations and Treatment Approaches for Scoliosis in Prader-Willi Syndrome.

Prader-Willi syndrome (PWS) is recognized as the first example of genomic imprinting, generally due to a de novo paternal 15q11-q13 deletion. PWS is considered the most common genetic cause of marked obesity in humans. Scoliosis, kyphosis, and kyphoscoliosis are commonly seen in children and adolescents with PWS with a prevalence of spinal deformities cited between 15% to 86%. Childhood risk is 70% or higher, until skeletal maturity, with a bimodal age distribution with one peak before 4 years of age and the other nearing adolescence. As few reports are available on treating scoliosis in PWS, we described clinical observations, risk factors, therapeutic approaches and opinions regarding orthopedic care based on 20 years of clinical experience. Treatments include diligent radiographic screening, starting once a child can sit independently, ongoing physical therapy, and options for spine casting, bracing and surgery, depending on the size of the curve, and the child's age. Similarly, there are different surgical choices including a spinal fusion at or near skeletal maturity, versus a construct that allows continued growth while controlling the curve for younger patients. A clear understanding of the risks involved in surgically treating children with PWS is important and will be discussed.

STIM1 R304W in mice causes subgingival hair growth and an increased fraction of trabecular bone.

Calcium signaling plays a central role in bone development and homeostasis. Store operated calcium entry (SOCE) is an important calcium influx pathway mediated by calcium release activated calcium (CRAC) channels in the plasma membrane. Stromal interaction molecule 1 (STIM1) is an endoplasmic reticulum calcium sensing protein important for SOCE. We generated a mouse model expressing the STIM1 R304W mutation, causing Stormorken syndrome in humans. Stim1R304W/R304W mice showed perinatal lethality, and the only three animals that survived into adulthood presented with reduced growth, low body weight, and thoracic kyphosis. Radiographs revealed a reduced number of ribs in the Stim1R304W/R304W mice. Microcomputed tomography data revealed decreased cortical bone thickness and increased trabecular bone volume fraction in Stim1R304W/R304W mice, which had thinner and more compact bone compared to wild type mice. The Stim1R304W/+ mice showed an intermediate phenotype. Histological analyses showed that the Stim1R304W/R304W mice had abnormal bone architecture, with markedly increased number of trabeculae and reduced bone marrow cavity. Homozygous mice showed STIM1 positive osteocytes and osteoblasts. These findings highlight the critical role of the gain-of-function (GoF) STIM1 R304W protein in skeletal development and homeostasis in mice. Furthermore, the novel feature of bilateral subgingival hair growth on the lower incisors in the Stim1R304W/R304W mice and 25 % of the heterozygous mice indicate that the GoF STIM1 R304W protein also induces an abnormal epithelial cell fate.

A novel de novo mutation in MYH7 gene in a patient with early onset muscular weakness and severe kyphoscoliosis: A case report.

INTRODUCTION: Various phenotypes have been identified for MYH7 gene mutation-related myopathy. Here, we describe a patient with severe muscular weakness and skeletal deformity with de novo heterozygous MYH7 gene mutation. PATIENT CONCERNS: A 33-year-old woman presented with early onset of muscular weakness, with delayed motor development during infancy. At age 8 years, she was unable to walk, with signs of skeletal deformity, including the progression of kyphoscoliosis. At age 31 years, she developed dyspnea. DIAGNOSIS: She diagnosed with esophageal hiatal hernia with abdominal CT. In electromyography, short duration, small amplitude motor unit action potential (MUAP), and early recruitment patterns were observed in the involved proximal muscles, suggesting myopathy. Muscle histopathology showed fiber-type disproportion. INTERVENTIONS: Next-generation sequencing study revealed a heterozygous in-frame deletion variation in the exon 14 of the MYH7 gene (c.1498_1500del/p.Glu500del), which is a novel variation confirmed by conventional Sanger sequencing. Compared with the parental test, this variant was concluded as de novo. OUTCOMES: She received laparoscopic hiatal hernia repair and Nissen fundoplication for esophageal hiatal hernia. After surgery, her postural dyspnea improved. As there is no fundamental treatment for MYH7-related myopathies, she continued conservative treatment for her symptoms. CONCLUSION: Here, we presented a rare case of de novo mutation of the myosin head domain in the MYH7 gene. This report broadens both the phenotypic and genotypic spectra of MYH7-related myopathies.

The retinol-retinoic acid metabolic pathway is impaired in the lumbar spine of a rat model of congenital kyphoscoliosis.

Although congenital scoliosis is defined as a genetic disease characterized by a congenital and abnormal curvature of the spinal vertebrae, our knowledge of the genetic underpinnings of the disease is insufficient. We herein show that the downregulation of the retinol-retinoic acid metabolism pathway is involved in the pathogenesis of congenital scoliosis. By analyzing DNA microarray data, we found that the expression levels of genes associated with the retinol metabolism pathway were decreased in the lumbar spine of Ishibashi rats (IS), a rat model of congenital kyphoscoliosis. The expression of Adh1 and Aldh1a2 (alcohol dehydrogenase), two enzymes that convert retinol to retinoic acid in this pathway, were decreased at both the gene and protein levels. Rarα, a receptor of retinoic acid and bone morphogenetic protein 2, which play a central role in bone formation and are located downstream of this pathway, were also downregulated. Interestingly, the serum retinol levels of IS rats were higher than those of wild-type control rats. These results indicate that the adequate conversion from retinol to retinoic acid is extremely important in the regulation of normal bone formation and it may also be a key factor for understanding the pathogenesis of congenital scoliosis.

Identifying pathogenic variants in the Follistatin-like 1 gene (FSTL1) in patients with skeletal and atrioventricular valve disorders.

BACKGROUND: Follistatin-like 1 (Fstl1) is a glycoprotein expressed throughout embryonic development. Homozygous loss of Fstl1 in mice results in skeletal and respiratory defects, leading to neonatal death due to a collapse of the trachea. Furthermore, Fstl1 conditional deletion from the endocardial/endothelial lineage results in postnatal death due to heart failure and profound atrioventricular valve defects. Here, we investigated patients with phenotypes similar to the phenotypes observed in the transgenic mice, for variants in FSTL1. METHODS: In total, 69 genetically unresolved patients were selected with the following phenotypes: campomelic dysplasia (12), small patella syndrome (2), BILU (1), and congenital heart disease patients (54), of which 16 also had kyphoscoliosis, and 38 had valve abnormalities as their main diagnosis. Using qPCR, none of 69 patients showed copy number variations in FSTL1. The entire gene body, including microRNA-198 and three validated microRNA-binding sites, were analyzed using Sanger sequencing. RESULTS: No variants were found in the coding region. However, 8 intronic variants were identified that differed significantly in their minor allele frequency compared to controls. Variant rs2272515 was found to significantly correlate (p < 0.05) with kyphoscoliosis. CONCLUSION: We conclude that pathogenic variants in FSTL1 are unlikely to be responsible for skeletal or atrioventricular valve anomalies in humans.