Development Of Adolescent Idiopathic Scoliosis Research Articles

Identification of Epigenetic Biomarkers of Adolescent Idiopathic Scoliosis Progression: A Workflow to Assess Local Gene Expression.

Adolescent idiopathic scoliosis (AIS) is a three-dimensional structural deformity of the spine that affects 2-3% of adolescents under the age of 16. AIS etiopathogenesis is not completely understood; however, the disease phenotype is correlated to multiple genetic loci and results from genetic-environmental interactions. One of the primary, still unresolved issues is the implementation of reliable diagnostic and prognostic markers. For clinical management improvement, predictors of curve progression are particularly needed. Recently, an epigenetic contribution to AIS development and progression was proposed; nevertheless, validation of data obtained in peripheral tissues and identification of the specific mechanisms and genes under epigenetic control remain limited. In this study, we propose a methodological approach for the identification of epigenetic markers of AIS progression through an original workflow based on the preliminary characterization of local expression of candidate genes in tissues directly involved in the pathology. The feasibility of the proposed methodological protocol has been originally tested here in terms of identification of the putative epigenetic markers of AIS progression, collection of the different tissues, retrieval of an appropriate amount and quality of RNA and DNA, and identification of suitable reference genes.

A Genetic Variant of FAM46A is Associated With the Development of Adolescent Idiopathic Scoliosis in the Chinese Population.

A genetic case-control study. To replicate recently reported genetic loci associated with adolescent idiopathic scoliosis (AIS) in the Chinese Han population, and to determine the relationship between gene expression and the clinical features of the patients. A recent study conducted in the Japanese population identified several novel susceptible loci, which might provide new insights into the etiology of AIS. However, the association of these genes with AIS in other populations remains unclear. A total of 1210 AIS and 2500 healthy controls were recruited for the genotyping of 12 susceptibility loci. Paraspinal muscles used for gene expression analysis were obtained from 36 AIS and 36 patients with congenital scoliosis. The difference regarding genotype and allele frequency between patients and controls was analyzed by χ 2 analysis. The t test was performed to compare the target gene expression level between controls and AIS patients. Correlation analysis was performed between gene expression and phenotypic data, including Cobb angle, bone mineral density, lean mass, height, and body mass index. Four SNPs, including rs141903557, rs2467146, rs658839, and rs482012, were successfully validated. Allele C of rs141903557, allele A of rs2467146, allele G of rs658839, and allele T of single nucleotide polymorphism rs482012 showed significantly higher frequency in patients. Allele C of rs141903557, allele A of rs2467146, allele G of rs658839, and allele T of rs482012 could notably increase the risk of AIS patients, with an odds ratio of 1.49, 1.16, 1.11, and 1.25, respectively. Moreover, tissue expression of FAM46A was significantly lower in AIS patients as compared with controls. Moreover, FAM46A expression was remarkably correlated with bone mineral density of patients. Four SNPs were successfully validated as novel susceptibility loci associated with AIS in the Chinese population. Moreover, FAM46A expression was associated with the phenotype of AIS patients.

Circulatory Adipokines and Incretins in Adolescent Idiopathic Scoliosis: A Pilot Study.

Adolescent idiopathic scoliosis (AIS) is a three-dimensional malformation of the spine of unknown cause that develops between 10 and 18 years old and affects 2-3% of adolescents, mostly girls. It has been reported that girls with AIS have a taller stature, lower body mass index (BMI), and bone mineral density (BMD) than their peers, but the causes remain unexplained. Energy metabolism discrepancies, including alterations in adipokine and incretin circulatory levels, could influence these parameters and contribute to disease pathophysiology. This pilot study aims to compare the anthropometry, BMD, and metabolic profile of 19 AIS girls to 19 age-matched healthy controls. Collected data include participants' fasting metabolic profile, anthropometry (measurements and DXA scan), nutritional intake, and physical activity level. AIS girls (14.8 ± 1.7 years, Cobb angle 27 ± 10°), compared to controls (14.8 ± 2.1 years), were leaner (BMI-for-age z-score ± SD: -0.59 ± 0.81 vs. 0.09 ± 1.11, p = 0.016; fat percentage: 24.4 ± 5.9 vs. 29.2 ± 7.2%, p = 0.036), had lower BMD (total body without head z-score ± SD: -0.6 ± 0.83 vs. 0.23 ± 0.98, p = 0.038; femoral neck z-score: -0.54 ± 1.20 vs. 0.59 ± 1.59, p = 0.043), but their height was similar. AIS girls had higher adiponectin levels [56 (9-287) vs. 32 (7-74) μg/mL, p = 0.005] and lower leptin/adiponectin ratio [0.042 (0.005-0.320) vs. 0.258 (0.024-1.053), p = 0.005]. AIS participants with a Cobb angle superior to 25° had higher resistin levels compared to controls [98.2 (12.8-287.2) vs. 32.1 (6.6-73.8), p = 0.0013]. This pilot study suggests that adipokines are implicated in AIS development and/or progression, but more work is needed to confirm their role in the disease.

A Genetic Variant of the ROBO3 Gene is Associated With Adolescent Idiopathic Scoliosis in the Chinese Population.

A case-control association study. This study aimed to reveal whether mutations within roundabout receptor 3 ( ROBO3 ) gene were related to adolescent idiopathic scoliosis (AIS) in Chinese Han population and to investigate the functional role of ROBO3 in the pathogenesis and progression of AIS. ROBO3 is essential for the regulation of hindbrain axonal cell migration and midline crossing. Studies have demonstrated that ROBO3 homozygous mutations are associated with horizontal gaze palsy with progressive scoliosis. However, whether and how ROBO3 contributed to the development of scoliosis remains unclear. Whole exome sequencing was performed in 135 AIS patients and 267 healthy controls to evaluate the differences of single nucleotide polymorphism variants within ROBO3 . Then the identified variant of ROBO3 was genotyped in another cohort included 1140 AIS patients and 1580 controls. Moreover, paraspinal muscles were collected from 39 AIS patients and 45 lumbar disk herniation patients for the measurement of ROBO3 mRNA expression. The χ 2 test, Fisher exact test or the Student t test were used to compare intergroup data. Pearson correlation was used to determine the association between ROBO3 expression and clinical phenotypes. A significant association was identified between the gene variant (rs74787566) of ROBO3 and the development of AIS through exome sequencing. The genotyping cohort demonstrated a higher frequency of allele A in AIS patients compared to controls (7.89% vs . 4.30%, P <0.001, odds ratio=1.87). In addition, the expression of ROBO3 in paraspinal muscles was inversely correlated with the Cobb angle ( P =0.043, r2 =0.1059). A significant association was identified between the gene variant (rs74787566) of ROBO3 and the development of AIS. The reduced expression of ROBO3 could result in the progression of curve magnitude in patients with AIS. Further studies are needed to verify the functional role of ROBO3 in the development of AIS. Level III.

Association of FBN1 polymorphism with susceptibility of adolescent idiopathic scoliosis: a case-control study

BackgroundFibrillin-1 (FBN1) is an extracellular matrix glycoprotein essential to the structural component of microfibrils and FBN1 gene polymorphisms can be associated with adolescent idiopathic scoliosis (AIS) susceptibility. This study aimed to evaluate the potential role of the FBN1 rs12916536 polymorphism in AIS development or severity and the variation in Cobb angle in relation to patient’s characteristics.MethodsDNA from 563 subjects (185 AIS patients and 378 controls) were genotyped using a validated TaqMan allelic discrimination assay. A multivariate logistic regression model evaluated the association between polymorphism and AIS, using the adjusted odds ratios (OR) with their respective 95% confidence intervals (95% CI). A linear regression analysis evaluated the variation in Cobb angle according to the patient’s age and body mass index (BMI).ResultsAmong the AIS group there was a predominance of females (12:1), low or normal BMI (90%), 58% had a Cobb angle greater than 45° and 74% were skeletally mature. Age was a risk factor (4-fold) for curve progression higher than BMI (P < 0.001). The allelic frequency of the rs12916536 G > A polymorphism was 40% in controls and 31% in AIS cases; and this difference was statistically significant (P = 0.004). FBN1 rs12916536 GA + AA genotypes were associated with a lower risk of AIS susceptibility (OR = 0.58 and 95% CI = 0.35–0.98), after adjustment for age, sex and BMI. However, no significant differences were detected in polymorphism distribution with the severity of the disease (Cobb < 45° or ≥ 45°).ConclusionAge was a risk factor for progression of the scoliotic curve and FBN1 rs12916536 polymorphism a protective factor for AIS susceptibility.

Polymorphisms in paired box 1 gene were associated with susceptibility of adolescent idiopathic scoliosis: A case–control study

BackgroundAssociation of genetic polymorphisms in paired box 1 (PAX-1) gene can influence the development of adolescent idiopathic scoliosis (AIS). PAX-1 is mainly expressed in the region of the vertebral bodies and intervertebral discs, being important for the proper formation of spinal structures.Objectives:The objective of this study was to evaluate the association of polymorphisms in PAX-1 gene with the susceptibility of AIS.Settings and Design:This was an analytical observational case–control study.Materials and Methods:Samples of 59 AIS indicated for surgical treatment, and 119 controls, without spinal disease were genotyped for PAX-1 rs6137473 and rs169311 polymorphisms.Statistical Analysis:The association of the polymorphisms with AIS was evaluated by a multivariable logistic regression model, using odds ratios (OR) and 95% confidence intervals (CI).Results:According to Lenke's classification, 89.8% had Type I and 10.2% II curves. The mean value of the Cobb angle of the proximal thoracic curve was 30.8°, 58.7° thoracic, and 30.4° for the lumbar and on the bending films 14.6°, 40.7°, and 11°, respectively. Among the AIS group, there was a predominance of females (8.8:1). The PAX-1 rs169311 and rs6137473 polymorphisms were positively associated with developing the AIS (OR = 1.98; 95% CI = 1.2–3.3 and OR = 3.16; 95% CI = 1.4–7.3, respectively). The rs6137473 polymorphism was associated with the lumbar modifier B and C compared to A (OR = 2.52; 95% CI = 1.1–5.8).Conclusions:PAX-1 polymorphisms were associated with an increased risk of developing the AIS and with curve severity and can be used as a biomarker to map the risk of developing surgical-grade AIS, guiding the treatment of patients.

A Functional SNP in the Promoter of LBX1 Is Associated With the Development of Adolescent Idiopathic Scoliosis Through Involvement in the Myogenesis of Paraspinal Muscles.

Previous studies have shown that LBX1 is associated with adolescent idiopathic scoliosis (AIS) in multiple populations. For the first time, rs1322330 located in the putative promoter region of LBX1 was found significantly associated with AIS in the Chinese population [p = 6.08 × 10–14, odds ratio (OR) = 1.42, 95% confidence interval of 1.03–1.55]. Moreover, the luciferase assay and electrophoretic mobility shift assay supported that the allele A of rs1322330 could down-regulate the expression of LBX1 in the paraspinal muscles of AIS. In addition, silencing LBX1 in the myosatellite cells resulted in significantly inhibited cell viability and myotube formation, which supported an essential role of LBX1 in muscle development of AIS. To summarize, rs1322330 may be a novel functional SNP regulating the expression of LBX1, which was involved in the etiology of AIS possibly via regulation of myogenesis in the paraspinal muscles.

Risk Factors, Lifestyle and Prevention among Adolescents with Idiopathic Juvenile Scoliosis: A Cross Sectional Study in Eleven First-Grade Secondary Schools of Palermo Province, Italy.

Adolescent idiopathic scoliosis (AIS) has an incidence of 2–3% in the general population and a multifactorial etiology. The present study aims to analyze modifiable risk factors and their interactions in the development of AIS in order to increase knowledge about the disease and to prevent the evolution of AIS in young students with tailored public health strategies. A cross-sectional study was conducted over two consecutive school years among students attending 11 first-grade secondary schools in the province of Palermo, Italy. A self-administered questionnaire that investigated socio-demographical, physical and anamnestic characteristics and habits, focusing on possible risk factors associated with idiopathic scoliosis, was administered. In addition, a clinical evaluation was performed with Adams’ test and Bunnel’s inclinometer. Suspected AIS cases were associated with the practice of high-risk sports (p < 0.05), weekly physical activity lasting ≥3 h (p < 0.05), lower back pain (p < 0.001), posture disorders (p < 0.01) and having had no contact with a physician (p < 0.01). Practice of high-risk sports (adj OR = 1.83; CI 95% 1.11–4.76) and suffering of posture disorders (adj OR = 1.67; CI 95% 1.12–3.60) showed a significant association with a confirmed diagnosis of AIS (Cobb angle ≥ 10° at X-ray). The risk factors associated with AIS are still unclear. Therefore, it is crucial to identify early modifiable and multiple risk factors to prevent the evolution of scoliosis in school-age children.

The role of endocrine hormones in the pathogenesis of adolescent idiopathic scoliosis.

Adolescent idiopathic scoliosis (AIS) is a common spinal deformity characterized by changes in the three-dimensional structure of the spine. It usually initiates during puberty, the peak period of human growth when the secretion of numerous hormones is changing, and it is more common in females than in males. Accumulating evidence shows that the abnormal levels of many hormones including estrogen, melatonin, growth hormone, leptin, adiponectin and ghrelin, may be related to the occurrence and development of AIS. The purpose of this review is to provide a summary and critique of the research published on each hormone over the past 20years, and to highlight areas for future study. It is hoped that the presentation will help provide a better understanding of the role of endocrine hormones in the pathogenesis of AIS.

Genetic variant of MIR4300HG is associated with progression of adolescent idiopathic scoliosis in a Chinese population

BackgroundA recent genome-wide association study identified a susceptible locus in MIR4300HG gene that was associated with curve progression of adolescent idiopathic scoliosis (AIS) in the Japanese population. However, the association between the gene and curve progression in other populations remains unclear.MethodsA cohort of 1952 AIS patients and 2495 healthy controls were included in the case-control analysis. In the case-only analysis, 747 patients were assigned to the progression group and 520 patients were assigned to the non-progression group, respectively. Rs35333564 was genotyped for all the subjects. Paraspinal muscles of 76 patients were collected for the analysis of gene expression. Chi-square test and ANOVA test were used for the intergroup comparison. Pearson correlation analysis was performed to investigate the relationship between the gene expression and curve magnitude.ResultsVariant rs35333564 was significantly associated with the curve severity of AIS (p = 0.025), but not the development of AIS (p = 0.418). Genotype GG was indicated by remarkably lower expression of MIR4300 (p = 0.020) which was significantly correlated with curve magnitude (p = 0.010). As a predicted target gene of MIR4300, the expression of CRTC1 was negatively correlated with MIR4300 expression (p = 0.012, r = −0.287) and positively correlated with curve severity (p = 0.025, r = 0.257).ConclusionsThe association between rs35333564 and curve progression was successfully replicated in a Chinese AIS population. CRTC1 may be the target gene of MIR4300 that plays a role in the curve progression of AIS.

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

A genetic case-control study. 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. 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. 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. 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. 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.

Lower androgen levels promote abnormal cartilage development in female patients with adolescent idiopathic scoliosis

BackgroundAdolescent idiopathic scoliosis (AIS) is a disease characterized by changes in the three-dimensional structure of the spine. Studies have shown that the development of AIS might be associated with genetic, biomechanics, endocrine factors and abnormal bone or cartilage development.MethodsBlood samples collected from 301 female patients (161 females with AIS and 140 females without AIS) were used for genotyping. Forty-eight serum samples from 161 females with AIS and 40 serum samples from 140 females without AIS were subjected to enzyme-linked immunosorbent assays (ELISAs). We also evaluated 32 facet joints (18 females with AIS and 14 females without AIS from the 301 female patients) using immunohistochemistry, Western blotting, and isolation of human primary chondrocytes, among other methods. We treated the AIS primary chondrocytes with dihydrotestosterone (DHT) to verify the relationship among androgen, the androgen receptor (AR), and its downstream pathway proteins.ResultsThe serum androgen level in the AIS group was significantly decreased (1.94±0.09 vs. 2.284±0.103) compared with that in the non-AIS (control) group. The single nucleotide polymorphism genotyping results showed that the mutation rates of rs6259 between the AIS and control groups were significantly different (G/G genotype: 48.4% vs. 42.1%, G/A genotype: 40.4% vs. 35.7%, P<0.05). The levels of interleukin (IL)-6 and metalloproteinase (MMP)-13 were increased in the cartilage of AIS patients, and these patients also exhibited decreased AR levels. The cell experiment results showed that androgen reduced the degree of abnormal cartilage development in female AIS patients through the AR/IL-6/signal transducer and activator of transcription 3 (STAT3) signaling pathway.ConclusionsOur study provides a new perspective on the pathogenesis of AIS and indicates that decreased androgen levels in female AIS patients play a potential role in the development of AIS via the AR/IL-6/STAT3 signaling pathway.

Etiopathogenesis of adolescent idiopathic scoliosis: Review of the literature and new epigenetic hypothesis on altered neural crest cells migration in early embryogenesis as the key event

Adolescent idiopathic scoliosis (AIS) affects 2–3% of children. Numerous hypotheses on etiologic/causal factors of AIS were investigated, but all failed to identify therapeutic targets and hence failed to offer a cure. Therefore, currently there are only two options to minimize morbidity of the patients suffering AIS: bracing and spinal surgery. From the beginning of 1960th, spinal surgery, both fusion and rod placement, became the standard of management for progressive adolescent idiopathic spine deformity. However, spinal surgery is often associated with complications. These circumstances motivate AIS scientific community to continue the search for new etiologic and causal factors of AIS. While the role of the genetic factors in AIS pathogenesis was investigated intensively and universally recognized, these studies failed to nominate mutation of a particular gene or genes combination responsible for AIS development. More recently epigenetic factors were suggested to play causal role in AIS pathogenesis. Sharing this new approach, we investigated scoliotic vertebral growth plates removed during vertebral fusion (anterior surgery) for AIS correction. In recent publications we showed that cells from the convex side of human scoliotic deformities undergo normal chondrogenic/osteogenic differentiation, while cells from the concave side acquire a neuronal phenotype. Based on these facts we hypothesized that altered neural crest cell migration in early embryogenesis can be the etiological factor of AIS. In particular, we suggested that neural crest cells failed to migrate through the anterior half of somites and became deposited in sclerotome, which in turn produced chondrogenic/osteogenic-insufficient vertebral growth plates. To test this hypothesis we conducted experiments on chicken embryos with arrest neural crest cell migration by inhibiting expression of Paired-box 3 (Pax3) gene, a known enhancer and promoter of neural crest cells migration and differentiation. The results showed that chicken embryos treated with Pax3 siRNA (microinjection into the neural tube, 44 h post-fertilization) progressively developed scoliotic deformity during maturation. Therefore, this analysis suggests that although adolescent idiopathic scoliosis manifests in children around puberty, the real onset of the disease is of epigenetic nature and takes place in early embryogenesis and involves altered neural crest cells migration. If these results confirmed and further elaborated, the hypothesis may shed new light on the etiology and pathogenesis of AIS.

Genetic Variants of CHD7 Are Associated with Adolescent Idiopathic Scoliosis.

A case-control association study. The aim of this study was to investigate whether CHD7 was associated with adolescent idiopathic scoliosis in Chinese Han population and to further explore the functional role of CHD7 in the development of adolescent idiopathic scoliosis (AIS). Several studies have explored the association of CHD7 with scoliosis in patients of European descent, while the results were inconsistent. There was a lack of study investigating the association of CHD7 with AIS in Chinese Han population. Variants within CHD7 were genotyped in 965 AIS patients and 976 healthy controls. Whole exome sequencing was performed in 96 AIS patients. Paraspinal muscles of 43AIS patients and 38 lumbar disc herniation patients were collected for the evaluation of the gene expression. Intergroup comparison was performed with the χ2 test for genotyping data or Student t test for tissue expression. The relationship of CHD7 expression with clinical phenotypes was determined by the Pearson correlation. Variant rs121434341 of CHD7 was significantly associated with AIS. AIS patients were found to have a remarkable higher frequency of allele G when compared with healthy controls (2.89% vs. 1.57%, P = 0.0018), with an odds ratio value of 1.89. A pathogenic mutation affecting normal splicing was identified in a patient. Moreover, the expression level of CHD7 in AIS patients was significantly lower than in the controls (0.0008437 ± 0.00004583 vs. 0.001129 ± 0.00003773, P < 0.001), and CHD7 expression was positively correlated with bone mineral contents (P = 0.036, r = 0.32). Genetic variants of CHD7 were significantly associated with AIS. Moreover, the decreased expression of CHD7 may be involved in the abnormal bone mass of AIS patients. Further studies are warranted to investigate the functional role of CHD7 in the pathogenesis of AIS.Level of Evidence: 3.

Polygenic Risk Score of Adolescent Idiopathic Scoliosis for Potential Clinical Use.

Adolescent idiopathic scoliosis (AIS) is a common disease causing three-dimensional spinal deformity in as many as 3% of adolescents. Development of a method that can accurately predict the onset and progression of AIS is an immediate need for clinical practice. Because the heritability of AIS is estimated as high as 87.5% in twin studies, prediction of its onset and progression based on genetic data is a promising option. We show the usefulness of polygenic risk score (PRS) for the prediction of onset and progression of AIS. We used AIS genomewide association study (GWAS) data comprising 79,211 subjects in three cohorts and constructed a PRS based on association statistics in a discovery set including 31,999 female subjects. After calibration using a validation data set, we applied the PRS to a test data set. By integrating functional annotations showing heritability enrichment in the selection of variants, the PRS demonstrated an association with AIS susceptibility (p=3.5 × 10-40 with area under the receiver-operating characteristic [AUROC]=0.674, sensitivity=0.644, and specificity=0.622). The decile with the highest PRS showed an odds ratio of as high as 3.36 (p=1.4 × 10-10 ) to develop AIS compared with the fifth in decile. The addition of a predictive model with only a single clinical parameter (body mass index) improved predictive ability for development of AIS (AUROC=0.722, net reclassification improvement [NRI] 0.505 ± 0.054, p=1.6 × 10-8 ), potentiating clinical use of the prediction model. Furthermore, we found the Cobb angle (CA), the severity measurement of AIS, to be a polygenic trait that showed a significant genetic correlation with AIS susceptibility (rg=0.6, p=3.0 × 10-4 ). The AIS PRS demonstrated a significant association with CA. These results indicate a shared polygenic architecture between onset and progression of AIS and the potential usefulness of PRS in clinical settings as a predictor to promote early intervention of AIS and avoid invasive surgery. © 2021 American Society for Bone and Mineral Research (ASBMR).

Novel Mutations in UTS2R are Associated with Adolescent Idiopathic Scoliosis in the Chinese Population.

A case-control study. To investigate the association of urotensin II (UTS2) signals with the susceptibility of adolescent idiopathic scoliosis (AIS) in the Chinese Han population. Dysregulated UTS2 signals induced by impaired cerebrospinal fluid flow have been implicated in the development of idiopathic scoliosis through studies on zebrafish. Furthermore, mutations in urotensin II receptor (UTS2R) were reported to cause severe scoliosis in zebrafish. In spite of the evidence presented in animal models, there is still a lack of knowledge concerning the role of UTS2 signaling related genes in AIS. In the discovery stage, exons of UTS2, UTS2R, and UTS2D were sequenced for 200 AIS patients and 200 healthy controls. Newly identified mutations were further genotyped in another independent cohort of 1000 AIS patients and 1000 controls by allelic-specific multiple ligase detection reactions. Gene expression analysis was performed in 36 AIS patients and 36 age-matched congenital scoliosis patients. The Chi-square test was used to compare the genotyping data between the groups. Gene expression analysis was compared with the Student t test. Association between two novel mutations (rs11654140, c.51T > C; rs568196624, c.1146C > G) and the development of AIS was identified. Allele C of rs11654140 and allele G of rs568196624 were significantly associated with the risk of AIS (1.5% vs. 0.5%, odds ratio = 3.02, P = 0.01 for rs11654140; 1.41% vs. 0.58%, odds ratio = 2.29, P = 0.04 for rs568196624). The mRNA expression of UTS2R in the AIS group was significantly higher as compared with that in the control group (0.059 ± 0.015 vs. 0.035 ± 0.013, P < 0.01). Rare mutations in UTS2R were significantly associated with AIS. Expression of UTS2R was significantly increased in AIS patients. The role of UTS2 signaling in the development of AIS is worthy of further investigation.Level of Evidence: N/A.

Dysregulated Bone Metabolism Is Related to High Expression of miR-151a-3p in Severe Adolescent Idiopathic Scoliosis.

Adolescent idiopathic scoliosis (AIS) is a common complex disease, and bone homeostasis plays an important role in its pathogenesis. Recent advances in epigenetic research show that dysregulated miRNAs may participate in the development of orthopedic diseases and AIS. The aim of this study was to detect differentially expressed miRNAs in severe AIS and elucidate the mechanism of miRNA deregulation in the pathogenesis of AIS. In the present study, miRNA expression profiles were detected in severe and mild AIS patients as well as healthy controls by miRNA sequencing. Candidate miRNAs were validated in a larger cohort. Primary osteoblasts from severe AIS patients were extracted and isolated to determine the effect of the candidate miRNAs on bone metabolism. Finally, we determined the methylation level in primary osteoblasts from severe AIS patients. The result showed that miR-151a-3p was overexpressed in severe AIS patients. Reduced GREM1 expression was observed in primary osteoblasts from severe AIS patients. miR-151a-3p directly inhibited GREM1 in primary osteoblasts. Relatively lower methylation levels were detected in primary osteoblasts from severe AIS patients. In conclusion, our study revealed that plasma miR-151a-3p levels may serve as a biomarker for severe AIS. Overexpression of miR-151a-3p may interrupt bone homeostasis via inhibiting GREM1 expression. Our result may provide a new biomarker for the early detection of AIS and increase our understanding of the pathogenesis of AIS.

Genetic Variant of TBX1 Gene Is Functionally Associated With Adolescent Idiopathic Scoliosis in the Chinese Population.

A genetic association study. The aim of this study was to investigate whether rs1978060 of TBX1 gene was a susceptible locus of adolescent idiopathic scoliosis (AIS) in the Chinese Han population and to better define the functional role of TBX1 in the development and progression of AIS. A recent genome-wide association study reported a novel susceptible locus in TBX1 gene, which was associated with the development of AIS in the Japanese population. However, there is a paucity of knowledge concerning the functional role of TBX1 in the Chinese AIS population. The SNP rs1978060 was genotyped in 1725 female AIS patients and 2600 healthy controls. Paraspinal muscle samples were collected from 30 AIS patients and 26 age-matched congenital scoliosis (CS) patients for the analysis of tissue expression. The differences of genotype and allele distributions between the patients and the controls were calculated using the χ test. The Pearson correlation analysis was carried out to investigate the relation between the expression of the PAX1 gene and the curve severity. SNP rs1978060 was significantly associated with the susceptibility of AIS. Allele G of rs1978060 could significantly add to the risk of AIS with an odds ratio of 1.12. The tissue expression of TBX1 was obviously decreased in AIS patients. There was a remarkable correlation between the curve magnitude and the TBX1 expression (r = -0.519, P = 0.003). The association between TBX1 and the susceptibility of AIS was successfully replicated in the Chinese population. Moreover, rs1978060 may be a functional variant regulating the expression of TBX1. More studies were warranted to explore the functional role of TBX1 in the onset and progression of AIS. 3.

Upregulation of cGMP-dependent Protein Kinase (PRKG1) in the Development of Adolescent Idiopathic Scoliosis.

ObjectiveTo explore the molecular regulatory mechanisms underlying fibroblast differentiation and dysfunction in the development of adolescent idiopathic scoliosis (AIS) in an effort to identify candidate therapeutic targets for AIS.MethodsThe GSE110359 dataset, obtained from the bone marrow stromal cells of 12 AIS patients and five healthy controls, was retrieved from the GEO database. The data were preprocessed and differentially expressed genes (DEGs) were identified. KEGG pathway and Gene Ontology (GO)‐Biological Process (BP) enrichment analyses were performed to identify the function of the DEGs. A protein–protein interaction (PPI) and a microRNA‐transcription factor (TF)‐target co‐regulatory network were constructed to identify hub genes in the development of AIS. In addition, hub DEGs were evaluated by quantitative PCR (qPCR) and immunohistochemical staining.ResultsA total of 188 DEGs including 100 up‐regulated and 88 down‐regulated genes were obtained. The up‐regulated DEGs were related to “p53 signaling pathway”, “FoxO signaling pathway”, and “cGMP‐PKG signaling pathway” terms, while the down‐regulated DEGs were significantly enriched in seven terms including “protein processing in endoplasmic reticulum”. The key up‐regulated genes, PRKG1, CCNG2, and KAT2B, and the key down‐regulated genes, MAP2K1 and DUSP6, were identified by the PPI and miRNA‐TF‐Target regulatory network analyses. mRNA expression patterns for PRKG1, DUSP6, and KAT2B were successfully verified by qPCR. In addition, PRKG1 protein levels were found to be elevated during the immunohistochemical analysis.ConclusionIncreased expression of PRKG1 in AIS patients might be an attractive therapeutic target for AIS. However, further gain or loss‐of‐function studies should be conducted.

A Novel Coding Variant in SLC39A8 Is Associated With Adolescent Idiopathic Scoliosis in Chinese Han Population.

A genetic case-control association study. The aim of this study was to investigate the association of SLC39A8 with the susceptibility of adolescent idiopathic scoliosis (AIS) in Chinese Han population. A recent exome-wide association study identified a missense variant rs13107325 in SLC39A8 that was associated with AIS. However, there was a lack of study validating the association of this novel mutation with AIS in other populations. The variant rs13107325 was genotyped in 965 AIS patients and 976 healthy controls by allelic specific multiple ligase detection reactions. Variants located in the coding region of SLC39A8 were identified by exon sequencing for 192 AIS patients and 192 controls. Paraspinal muscles from 36 AIS patients and 36 age-matched congenital scoliosis patients were collected for the gene expression analysis. Comparison between the cases and controls was performed with the χ test for genotyping data or with Student t test for gene expression analysis. For the missense variant rs13107325, there was no case of mutation detected in the patients or the controls. All the subjects had homozygous genotype CC. Exon sequencing revealed that a coding variant rs11097773 of SLC39A8 had a significantly different distribution of minor allele frequency between patients and controls (7.81% vs. 14.8%, P = 0.002). The mRNA expression of SLC39A8 in the patients was remarkably lower than that in the controls (0.0015 ± 0.00026 vs. 0.0021 ± 0.00033, P < 0.001). The association of previously reported novel mutation (rs13107325 in SLC39A8) with AIS was not replicated in the Chinese population. Interestingly, a novel coding variant rs11097773 of SLC39A8 is found significantly associated with AIS. Moreover, the expression of SLC39A8 was obviously decreased in AIS patients. Further study is warranted to clarify the functional role of rs11097773 in the development of AIS. 3.