BACKGROUND: Congenital scoliosis (CS) is a complex spinal malformation of unknown etiology with abnormal bone metabolism. Fibroblast growth factor 23 (FGF23), secreted by osteoblasts and osteocytes, can inhibit bone formation and mineralization. This research aims to investigate the relationship between CS and FGF23. METHODS: We collected peripheral blood from two pairs of identical twins for methylation sequencing of the target region. FGF23 mRNA levels in the peripheral blood of CS patients and age-matched controls were measured. Receiver operator characteristic (ROC) curve analyses were conducted to evaluate the specificity and sensitivity of FGF23. The expression levels of FGF23 and its downstream factors fibroblast growth factor receptor 3 (FGFr3)/tissue non-specific alkaline phosphatase (TNAP)/osteopontin (OPN) in primary osteoblasts from CS patients (CS-Ob) and controls (CT-Ob) were detected. In addition, the osteogenic abilities of FGF23-knockdown or FGF23-overexpressing Ob were examined. RESULTS: DNA methylation of the FGF23 gene in CS patients was decreased compared to that of their identical twins, accompanied by increased mRNA levels. CS patients had increased peripheral blood FGF23 mRNA levels and decreased computed tomography (CT) values compared with controls. The FGF23 mRNA levels were negatively correlated with the CT value of the spine, and ROCs of FGF23 mRNA levels showed high sensitivity and specificity for CS. Additionally, significantly increased levels of FGF23, FGFr3, OPN, impaired osteogenic mineralization and lower TNAP levels were observed in CS-Ob. Moreover, FGF23 overexpression in CT-Ob increased FGFr3 and OPN levels and decreased TNAP levels, while FGF23 knockdown induced downregulation of FGFr3 and OPN but upregulation of TNAP in CS-Ob. Mineralization of CS-Ob was rescued after FGF23 knockdown. CONCLUSIONS Our results suggested increased peripheral blood FGF23 levels, decreased bone mineral density in CS patients, and a good predictive ability of CS by peripheral blood FGF23 levels. FGF23 may contribute to osteopenia in CS patients through FGFr3/TNAP / OPN pathway.
Humans ; Osteopontin/genetics* ; Alkaline Phosphatase/metabolism* ; Receptor, Fibroblast Growth Factor, Type 3/metabolism* ; Scoliosis/genetics* ; Osteoblasts/metabolism* ; Calcinosis ; RNA, Messenger/metabolism* ; Bone Diseases, Metabolic/metabolism* ; Fibroblast Growth Factors/genetics*

BACKGROUND: Osteopenia has been well documented in adolescent idiopathic scoliosis (AIS). Bone marrow stem cells (BMSCs) are a crucial regulator of bone homeostasis. Our previous study revealed a decreased osteogenic ability of BMSCs in AIS-related osteopenia, but the underlying mechanism of this phenomenon remains unclear. METHODS: A total of 22 AIS patients and 18 age-matched controls were recruited for this study. Anthropometry and bone mass were measured in all participants. Bone marrow blood was collected for BMSC isolation and culture. Osteogenic and adipogenic induction were performed to observe the differences in the differentiation of BMSCs between the AIS-related osteopenia group and the control group. Furthermore, a total RNA was extracted from isolated BMSCs to perform RNA sequencing and subsequent analysis. RESULTS: A lower osteogenic capacity and increased adipogenic capacity of BMSCs in AIS-related osteopenia were revealed. Differences in mRNA expression levels between the AIS-related osteopenia group and the control group were identified, including differences in the expression of LRRC17 , DCLK1 , PCDH7 , TSPAN5 , NHSL2 , and CPT1B . Kyoto Encyclopedia of Genes and Genomes enrichment analyses revealed several biological processes involved in the regulation of autophagy and mitophagy. The Western blotting results of autophagy markers in BMSCs suggested impaired autophagic activity in BMSCs in the AIS-related osteopenia group. CONCLUSION Our study revealed that BMSCs from AIS-related osteopenia patients have lower autophagic activity, which may be related to the lower osteogenic capacity and higher adipogenic capacity of BMSCs and consequently lead to the lower bone mass in AIS patients.
Humans ; Adolescent ; Scoliosis/genetics* ; Cell Differentiation/physiology* ; Osteogenesis/genetics* ; Bone Diseases, Metabolic/genetics* ; Kyphosis ; Autophagy/genetics* ; Bone Marrow Cells ; Cells, Cultured ; Doublecortin-Like Kinases

Identification of genetic risk factors is the hotspot of adolescent idiopathic scoliosis (AIS). Through candidate gene approach and genome-wide association studies (GWAS), some genes were preliminary identified. To review AIS related genes,and construct the gene network map of AIS gene. We searched on NCBI PubMed and Web of Science database using search terms "adolescent idiopathic scoliosis" and "gene", to classify induction genes. We then constructed gene diagram using string-db. We found 35 AIS genes relating to connective tissue, nervous system active substances, melatonin synthesis and metabolism, puberty and growth, and genes whose function is unknown. Gene diagram shows that a network relationship between gene and other genes,in which IL6, ESR1, ESR2, VDR, TGFB1, IGF1 gene may as the key gene about AIS' genetic mechanism. Two sites of 3 GWAS results outside the network, it is suggesting new pathway that need to be explored. The study about AIS susceptibility gene is still preliminary, requiring in-depth research to identify the new networks.
Adolescent ; Genetic Predisposition to Disease ; Genome-Wide Association Study ; Humans ; Insulin-Like Growth Factor I ; genetics ; Matrilin Proteins ; genetics ; Scoliosis ; genetics ; Transforming Growth Factor beta1 ; genetics

BACKGROUND: Degenerative lumbar scoliosis (DLS) progresses with aging after 50-60 years, and the genetic association of DLS remains largely unclear. In this study, the genetic association between collagen type II alpha 1 (COL2A1) gene and DLS was investigated. METHODS: COL2A1 gene polymorphism was investigated in DLS subjects compared to healthy controls to investigate the possibility of its association with COL2A1 gene. Based on a single nucleotide polymorphism (SNP) database, SNP (rs2276454) in COL2A1 were selected and genotyped using direct sequencing in 51 patients with DLS and 235 healthy controls. The SNP effects were analyzed using three models of codominant, dominant, and recessive. Logistic regression models were calculated for odds ratios (ORs) with 95% confidence intervals (CIs) and corresponding p-values, controlling age and gender as co-variables. RESULTS: SNP (rs2276454) in COL2A1 was significantly associated with the degenerative lumbar scoliosis in the codominant (OR, 1.90; 95% CI, 1.17 to 3.10; p = 0.008) and dominant models (OR, 3.58; 95% CI, 1.59 to 9.29; p = 0.001). CONCLUSIONS: The results suggest that COL2A1 is associated with the risk of DLS in Korean population.
Aged ; Asian Continental Ancestry Group ; Collagen Type II/*genetics ; Female ; Humans ; *Lumbar Vertebrae ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; Scoliosis/ethnology/*genetics

PURPOSE: The incidence of adolescent idiopathic scoliosis (AIS) has rapidly increased, and with it, physician consultations and expenditures (about one and a half times) in the last 5 years. Recent etiological studies reveal that AIS is a complex genetic disorder that results from the interaction of multiple gene loci and the environment. For personalized treatment of AIS, a tool that can accurately measure the progression of Cobb's angle would be of great use. Gene analysis utilizing single nucleotide polymorphism (SNP) has been developed as a diagnostic tool for use in Caucasians but not Koreans. Therefore, we attempted to reveal AIS-related genes and their relevance in Koreans, exploring the potential use of gene analysis as a diagnostic tool for personalized treatment of AIS therein. MATERIALS AND METHODS: A total of 68 Korean AIS and 35 age- and sex-matched, healthy adolescents were enrolled in this study and were examined for 10 candidate scoliosis gene SNPs. RESULTS: This study revealed that the SNPs of rs2449539 in lysosomal-associated transmembrane protein 4 beta (LAPTM4B) and rs5742612 in upstream and insulin-like growth factor 1 (IGF1) were associated with both susceptibility to and curve severity in AIS. The results suggested that both LAPTM4B and IGF1 genes were important in AIS predisposition and progression. CONCLUSION: Thus, on the basis of this study, if more SNPs or candidate genes are studied in a larger population in Korea, personalized treatment of Korean AIS patients might become a possibility.
Adolescent ; Disease Progression ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Insulin-Like Growth Factor I/genetics ; Korea ; Male ; Membrane Proteins/genetics ; Oncogene Proteins/genetics ; *Polymorphism, Single Nucleotide ; Scoliosis/*genetics/pathology/radiography

OBJECTIVETo investigate whether the titrate-resistant acid phosphatase 5 (ACP5) gene polymorphisms were associated with the occurrence or curve severity of adolescent idiopathic scoliosis (AIS).

METHODSThere were 372 AIS patients from January 2006 to December 2008 and 239 normal controls from March 2005 to August 2006 were recruited. The Cobb angles were ≥ 10° in all AIS patients. Using the haplotype data of Han population from the Hapmap Project, two tag SNPs (rs2229531, rs2071484) were defined for ACP5 gene. PCR-restriction fragment length polymorphism was used for the genotyping.

RESULTSNo polymorphism in rs2229531 was found in this study. The genotype and allele frequency distribution in rs2071484 were similar between AIS patients and normal controls (χ(2) = 3.336 and 1.438, P > 0.05). The mean maximum Cobb angles of different genotypes of rs2071484 in ACP5 gene were 38° ± 19° in AA, 34° ± 14° in AG and 38° ± 21° in GG, which were similar with each other among AIS patients who reached skeletal maturity or received surgery treatment (P = 0.157).

CONCLUSIONThe ACP5 gene is neither associated with the occurrence nor the curve severity of AIS.

Acid Phosphatase ; genetics ; Adolescent ; Child ; Female ; Humans ; Isoenzymes ; genetics ; Male ; Polymorphism, Genetic ; Scoliosis ; genetics ; Tartrate-Resistant Acid Phosphatase

Persistent generalized low bone mineral density (BMD) has been reported in patients with adolescent idiopathic scoliosis (AIS). However, the exact mechanisms and causes of the low BMD in AIS patients are largely unknown. The purpose of this study was to examine the relationship between the receptor activator of NF-κB ligand (RANKL)/osteoprotegerin (OPG) levels in osteoblasts (OBs) from AIS patients with low BMD and with comparison made between the patients and controls. Twenty AIS patients and eight age-matched controls were included in the present study. The BMD of lumbar spine and proximal femur was measured in all subjects. OBs from the cancellous bone of each subject was harvested and primarily cultured. The mRNA and protein expression of RANKL and OPG in OBs was detected by RT-PCR and Western blotting. The results showed BMD was lower in AIS patients than in controls. A significantly higher mRNA and protein expression of RANKL was observed in OBs from AIS patients, while no significant difference was found in the expression of OPG between AIS patients and controls. As a result, RANKL/OPG ratio in patients with AIS was remarkably higher than controls. Our study preliminarily demonstrated expression of RANKL was higher in OBs from AIS patients with low BMD as compared with controls, suggesting the unbalanced RANKL/OPG ratio caused by an over-expression of RANKL in OBs may be responsible for the low BMD in AIS patients.
Adolescent ; Bone Density ; genetics ; Child ; Humans ; Osteoblasts ; metabolism ; RANK Ligand ; genetics ; Receptor Activator of Nuclear Factor-kappa B ; genetics ; metabolism ; Scoliosis ; genetics ; Young Adult

OBJECTIVETo determine whether the matrix metalloproteinase 9 gene (MMP9) polymorphism is associated with the onset or progression of adolescent idiopathic scoliosis (AIS) in Chinese Han female.

METHODSThree single nucleotide polymorphisms (SNPs) (rs17576, rs2250889, rs1805088) were genotyped through TaqMan-based real-time PCR assay in 190 AIS patients and 190 controls, all of whom were females from Chinese Han population with matched age. Analyses performed included Hardy Weinberg equilibrium test, Pearson chi-square test, Logistic regression analysis, linkage disequilibrium analysis and haplotype analysis. The mean maximum Cobb angles with different genotypes in case-only dataset were also compared.

RESULTSAll 3 SNPs have reached Hardy-Weinberg equilibrium in the controls. Genotype and allele frequencies of all SNPs were found similar between cases and controls by Pearson chi-square test and Logistic regression. Genotype-phenotype analysis showed that patients with CC genotype in rs2250889 featured larger maximum Cobb angles.

CONCLUSIONMMP9 may not be a predisposition gene of AIS in Han female. However, homozygous mutation in rs2250889 can render scoliosis more severe, implying that MMP9 defect may result in deterioration of AIS.

Adolescent ; Asian Continental Ancestry Group ; genetics ; Child ; China ; Female ; Genetic Association Studies ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Matrix Metalloproteinase 9 ; genetics ; Phenotype ; Polymorphism, Single Nucleotide ; genetics ; Scoliosis ; genetics

OBJECTIVETo identify whether SH3GL1 gene serves as a disease associated gene of adolescent idiopathic scoliosis (AIS).

METHODSPositioning candidate cloning: "case-sibling or case-family control design" research scheme based on family constellation was designed. Fifty-six AIS patients (15 male and 41 female, mean age 15 years old, ranged from 8 to 22 years old, Cobb angle from 25 degrees to 110 degrees , average Cobb angle of 67.5 degrees ) from November 2007 to December 2008 were recruited. In all patients, blood preparation was collected, and genome DNA was extracted. According to nucleotide sequence of gene SH3GL1, primer pair for PCR amplification, cloning, and sequencing with 10 exons as emphasis was designed. Sequence comparative analysis for exon sequencing result between sib pairs or family pairs, and that between sib pair or family pairs and NCBI (National Center for Biotechnology Information) were conducted through Vector NTI Advance 10.3 software to judge whether basic group mutation occurred or not. Amino acid sequence comparative analysis for prediction was made.

RESULTSTen exons of the candidate gene SH3GL1 were successfully amplified and cloned in genome DNA of an AIS sib pair and family pairs, and the sequencing obtained positive results. Twelve basic group mutations were found in 10 exons of the candidate gene SH3GL1 of patients with AIS. These mutations were located in the second exon (3 mutations), the fourth exon (1 mutations), the fifth exon (4 mutations), the sixth exon (1 mutations), the eighth exon (1 mutations), and the tenth exon (2 mutations, noncoding region). If basic group in 515 of mRNA was mutated to T, termination codon(TAG) came into being and open reading frame was altered. The sequence of protein showed brachytmema protein was encoded, which could cause changes of primary structure.

CONCLUSIONSH3GL1 is possibly one of the disease associated genes of AIS.

Adolescent ; Base Sequence ; Child ; Exons ; genetics ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; Male ; Scoliosis ; genetics ; Sequence Alignment ; Young Adult

OBJECTIVETo investigate the association between the tissue inhibitor of metalloproteinase-2 (TIMP-2) gene polymorphisms with the predisposition and disease severity of thoracic adolescent idiopathic scoliosis (AIS).

METHODSThree hundred and fifty-four female thoracic AIS patients treated from January 2007 to March 2009 and 210 healthy female who underwent health examination during March 2005 to June 2006 as normal controls were recruited in this study. One SNP-418G/C (rs8179090) in the promoter region were selected for TIMP-2 gene. PCR- RFLP was used for genotyping.

RESULTSNo significant differences of genotype and allele frequency distribution were found between AIS patients and normal controls (P > 0.05). In AIS patients, the frequency of C allele of the patients with the body mass index (BMI) < 17 kg/m(2) was significantly higher than those with the BMI > or = 17 kg/m(2) (P < 0.05), and the frequency of C allele of cases with the major Cobb angle > or = 40 degrees was significantly higher than that with Cobb angle < 40 degrees (P < 0.05). Among the patients who reached skeletal maturity without any interference of natural history, significantly higher average maximum Cobb angle was found in patients with GC and CC genotype compare with those with GG genotype.

CONCLUSIONSThe SNP-418G/C (rs8179090) in the promoter region of TIMP-2 gene may be associated with abnormal growth pattern and curve progression of thoracic AIS. TIMP-2 gene is a disease-modifier gene of thoracic AIS.

Adolescent ; Case-Control Studies ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Polymorphism, Genetic ; Polymorphism, Single Nucleotide ; Scoliosis ; genetics ; Tissue Inhibitor of Metalloproteinase-2 ; genetics