Yu-juan Liu

Exome sequencing identifies SLCO2A1 mutations as a cause of primary hypertrophic osteoarthropathy.

By using whole-exome sequencing, we identified a homozygous guanine-to-adenine transition at the invariant -1 position of the acceptor site of intron 1 (c.97-1G>A) in solute carrier organic anion transporter family member 2A1 (SLCO2A1), which encodes a prostaglandin transporter protein, as the causative mutation in a single individual with primary hypertrophic osteoarthropathy (PHO) from a consanguineous family. In two other affected individuals with PHO from two unrelated nonconsanguineous families, we identified two different compound heterozygous mutations by using Sanger sequencing. These findings confirm that SLCO2A1 mutations inactivate prostaglandin E(2) (PGE(2)) transport, and they indicate that mutations in SLCO2A1 are the pathogenic cause of PHO. Moreover, this study might also help to explain the cause of secondary hypertrophic osteoarthropathy.

Association of polymorphisms in low-density lipoprotein receptor-related protein 5 gene with bone mineral density in postmenopausal Chinese women

AbstractAim:To investigate the possible association of Q89R, N740N and A1330V polymorphisms in low-density lipoprotein receptor-related protein 5 (LRP5) gene with bone mineral density (BMD) in postmenopausal Chinese women.Methods:Q89R, N740N and A1330V genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 647 unrelated healthy postmenopausal Han Chinese women aged 43–76 years in Shanghai. BMD at lumbar spine 1–4 and the left proximal femur including the femoral neck, trochanter and Wards triangle were measured by dual-energy X-ray absorptionmetry in all subjects.Results:The distribution of the Q89R, N740N and A1330V genotypes in this population was as follows: QQ 80.5%, QR 18.7%, and RR 0.8%; TT 66.9%, TC31.1%, and CC2.0%; AA 68.0%, AV 29.7%, and VV 2.3%. The frequencies of the Q89R, N740N and A1330V genotypes and alleles did not deviate from the Hardy-Weinberg equilibrium. We found that the Q89R and A1330V polymorphisms were in linkage disequilibrium in our population (χ2=13.50, P<0.01). Both before and after adjusting for age, years since menopause, height, and weight, the Q89R or N740N genotypes were significantly associated with BMD at the femoral neck (P<0.05). Subjects with the Q89R QQ genotype or the N740N TT genotype had a significantly higher BMD at the femoral neck, compared with those with the QR/RR or TC/CC genotypes, respectively. No significant association was found between A1330V polymorphism and BMD at any site.Conclusion:Our findings suggest that the LRP5 gene is a candidate for the genetic determination of BMD in postmenopausal Chinese women.

No association of the polymorphisms of the frizzled-related protein gene with peak bone mineral density in Chinese nuclear families

BackgroundThe Wnt/beta-catenin signaling pathway plays an important role in skeletal development. Polymorphisms of frizzled-related protein (FRZB), an antagonist of this pathway, may generate variations in bone mineral density (BMD). In this study, we analyzed the association between FRZB genotypes and peak BMD variation in the spines and hips of two relatively large samples of Chinese female-offspring and male-offspring nuclear families.MethodsWe recruited 1,260 subjects from 401 female-offspring nuclear families and 1,296 subjects from 427 male-offspring nuclear families and genotyped four tagging single nucleotide polymorphisms (tagSNPs) (rs6433993, rs409238, rs288324, and rs4666865) spanning the entire FRZB gene. The SNPs rs288326 and rs7775, which are associated with hip osteoarthritis, were not selected in this study because of their low minor allele frequencies (MAFs) in Chinese people. The quantitative transmission disequilibrium test (QTDT) was used to analyze the association between each SNP and haplotype with peak BMD in female- and male-offspring nuclear families.ResultsIn the female-offspring nuclear families, we found no evidence of an association between either single SNPs or haplotypes and peak BMD in the spine or hip. In the male-offspring nuclear families, no within-family association was observed for either SNPs or haplotypes, although a significant total association was found between rs4666865 and spine BMD (P = 0.0299).ConclusionOur results suggest that natural variation in FRZB is not a major contributor to the observed variability in peak BMD in either Chinese females or males. Because ethnic differences in the FRZB genotypes may exist, other studies in different population are required to confirm such results.

Hip axis length changes in 10,554 males and females and the association with femoral neck fracture.

Hip axis length (HAL) has been proposed as an independent predictor of hip fracture risk in Caucasian females. There are, however, few data concerning its predictive risk in Chinese. The aim of this study was to investigate the changes of HAL in healthy Chinese population and the relationship between HAL and femoral neck fracture. The study population included 10,554 healthy Chinese people (8665 females, 1889 males) aged 20-97 yrs living in Shanghai. Cases were 106 patients (82 females, 24 males) aged 52 yrs old and over with femoral neck fracture. Controls were 106 age-matched healthy persons. All subjects were measured bone mineral density (BMD) at any site of proximal femur and HAL using dual-energy X-ray absorptiometry. HAL had significantly positive correlations with height and weight. After the adjustment of height and weight, HAL increased with age at 50 yrs of age and over in females, and no difference was found among the age groups in males. Males had longer HAL than females in all age groups. The peak BMD appeared in 30-44 yrs for females and 20-24 yrs for males and decreased thereafter, especially for females at 50 yrs old and over. HAL was similar in both fracture and control groups, whereas the BMD values at proximal femur were significantly lower in fracture group than in controls. There was no evidence that subjects with femoral neck fracture had longer HAL. Because of the limitations of retrospective study and relatively small fracture sample, prospective studies are required to determine the conclusions.

Association between VDR and ESR1 gene polymorphisms with bone and obesity phenotypes in Chinese male nuclear families.

AbstractAim:The goal of this study was to determine whether polymorphisms in the vitamin D receptor (VDR) and estrogen receptor alpha (ESR1) genes are associated with variations of peak bone mineral density (BMD) and obesity phenotypes in young Chinese men.Methods:A total of 1215 subjects from 400 Chinese nuclear families were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and allele-specific multiple PCR (ASM-PCR) analysis at the ApaI, FokI, and CDX2 sites in the VDR gene and the PvuII and XbaI sites in the ESR1 gene. BMD at the lumbar spine and hip, total fat mass, and total lean mass were measured using dual energy X-ray absorptiometry. The associations between VDR and ESR1 gene polymorphisms with peak BMD, body mass index (BMI), total fat mass, total lean mass, and percentage fat mass (PFM) were determined using quantitative transmission disequilibrium tests (QTDTs).Results:Using QTDTs, no significant within-family associations were obtained between genotypes or haplotypes of the VDR and ESR1 genes and peak BMD. For the obesity phenotypes, the within-family associations were significant between CDX2 genotypes and BMI (P=0.046), fat mass (P=0.004), and PFM (P=0.020). Further, PvuII was significantly associated with the variation of fat mass and PFM (P=0.002 and P=0.039, respectively). A subsequent 1000 permutations were in agreement with these within-family association results.Conclusion:Our findings showed that VDR and ESR1 polymorphisms were associated with total fat mass in young Chinese men, but we failed to find a significant association between VDR and ESR1 genotypes and peak BMD. These findings suggested that the VDR and ESR1 genes are quantitative trait loci (QTL) underlying fat mass variation in young Chinese men.

Susceptibility genes for osteoporotic fracture in postmenopausal chinese women

To identify the susceptibility genes for osteoporotic fracture in postmenopausal Chinese women, a two‐stage case‐control association study using joint analysis was conducted in 1046 patients with nontraumatic vertebra, hip, or distal radius fractures and 2303 healthy controls. First, 113 single‐nucleotide polymorphisms (SNPs) in 16 potential osteoporosis candidate genes reported in recent genomewide association studies, meta‐analyses studies, large‐scale association studies, and functional studies were genotyped in a small‐sample‐size subgroup consisting of 541 patients with osteoporotic fractures and 554 healthy controls. Variants and haplotypes in SPTBN1, TNFRSF11B, CNR2, LRP4, and ESR1 that have been identified as being associated with osteoporotic fractures were further reanalyzed in the entire case‐control group. We identified one SNP in TNFRSF11B (rs3102734), three SNPs in ESR1 (rs9397448, rs2234693, and rs1643821), two SNPs in LRP4 (rs17790156 and rs898604), and four SNPs in SPTBN1 (rs2971886, rs2941583, rs2941584, and rs12475342) were associated with all of the broadly defined osteoporotic fractures. The most significant polymorphism was rs3102734, with increased risk of osteoporotic fractures (odds ratio, 1.35; 95% confidence interval [CI], 1.17–1.55, Bonferroni p = 2.6 × 10−4). Furthermore, rs3102734, rs2941584, rs12475342, rs9397448, rs2234693, and rs898604 exhibited significant allelic, genotypic, and/or haplotypic associations with vertebral fractures. SNPs rs12475342, rs9397448, and rs2234693 showed significant genotypic associations with hip fractures, whereas rs3102734, rs2073617, rs1643821, rs12475342, and rs2971886 exhibited significant genotypic and/or haplotypic associations with distal radius fractures. Accordingly, we suggest that in addition to the clinical risk factors, the variants in TNFRSF11B, SPTBN1, ESR1, and LRP4 are susceptibility genetic loci for osteoporotic fracture in postmenopausal Chinese women.

Identification of the CLCN7 gene mutations in two Chinese families with autosomal dominant osteopetrosis (type II)

Here we report the identification of two different mutations in chloride channel 7 gene in two unrelated patients with autosomal dominant osteopetrosis type II. We determined that one patient (a 32-year-old woman) carried a heterozygous gene for a R767W mutation in exon 24, and another patient (a 17-year-old boy) carried a heterozygous gene for a novel frameshift mutation (Glu798FS) in exon 25. Recent studies have reported loss-of-function mutations in the chloride channel 7 (CLCN7) gene as a cause of autosomal dominant osteopetrosis type II (ADO-II). The identification of gene mutations in Chinese with ADO has not been reported previously. In this study, we identified mutations of the CLCN7 gene in two unrelated Chinese families with ADO-II. Two probands with ADO-II were diagnosed based on their bone characteristics on X-rays and their laboratory results. All 25 exons of the CLCN7 gene, including the exon–intron boundaries, were sequenced. We found in family 1 that the proband (a 32-year-old woman) was heterozygous for a CLCN7 mutation. The nonsynonymous mutation consisted of a heterozygous C/T transition at codon 2327 in exon 24, which resulted in an arginine (CGG)-to tryptophan (TGG) substitution at position 767 (R767W). The same heterozygous mutation (C/T) was determined in her father and son, who were asymptomatic with normal skeleton radiography. In family 2, we found that the proband (a 17-year-old boy) carried a novel frameshift mutation (Glu798FS) resulting from a G insertion between codon 60 and codon 61 in exon 25. The heterozygous –/G insertion is predicted to elongate the peptide of CLCN7 by 120 amino acids after position 797 amino acids. Similarly, some individuals of this family carried the same heterozygous mutation, but they are all asymptomatic. Furthermore, the R767W and Glu798FS mutations were not found in 100 unrelated controls. Our present findings suggest that the novel Glu798FS mutation in exon 25 and R767W in exon 24 in the CLCN7 gene were responsible for ADO-II in these Chinese patients.

Serum osteocalcin levels are inversely associated with plasma glucose and body mass index in healthy Chinese women

Aim:Osteocalcin, a biochemical marker of bone formation, has been suggested to be involved in the regulation of energy metabolism. The aim of this study was to investigate the possible association between serum osteocalcin and markers of glucose and lipid metabolism in a large sample of healthy Chinese women.Methods:A total of 2032 healthy Chinese women in Shanghai, aged 20–94 (including 1396 discovery-study subjects and 636 postmenopausal women for a reduplication analysis) were recruited. Their serum osteocalcin, calcium and the relevant measurements were analyzed. A Spearman correlation analysis was performed between osteocalcin and the other markers of energy metabolism including triglyceride, total cholesterol, fasting plasma glucose (FPG), serum insulin, body mass index and homeostasis model assessment-insulin resistance. Separate multiple regression analyses were performed with data from the discovery and reduplication subjects to determine whether serum osteocalcin concentration was an independent predictor of the glucose or lipid metabolism markers.Results:For the discovery-study subjects, serum osteocalcin was found to be negatively associated with weight (r=−0.08, P=0.002), BMI (−0.13, P<0.001) and FPG (r=−0.13, P=0.001). Similar results were also found in the reduplication subjects (weight: r=−0.19, P=0.016; BMI: r=−0.23, P=0.003; FPG: r=−0.28, P<0.001). In the multiple regression analysis, serum osteocalcin was revealed as a potential independent predictor for FPG (β=−0.07 and –0.210 for discovery and reduplication, respectively, P<0.01) and BMI (β=−0.127 and –0.299 for discovery and reduplication, respectively, P<0.01).Conclusion:Serum osteocalcin is negatively associated with weight BMI and FPG in healthy Chinese women. Therefore, osteocalcin might contribute to obesity and diabetes.

Association between SNP and haplotypes in PPARGC1 and adiponectin genes and bone mineral density in Chinese nuclear families

AbstractAim:To assess the contribution of single nucleotide polymorphisms (SNP) and haplotypes in the peroxisome proliferator-activated receptor-γ co-activator-1 (PPARGC1) and adiponectin genes to normal bone mineral density (BMD) variation in healthy Chinese women and men.Methods:We performed population-based (ANOVA) and family-based (quantitative trait locus transmission disequilibrium test) association studies of PPARGC1 and adiponectin genes. SNP in the 2 genes were genotyped. BMD was measured using dual-energy X-ray absorptiometry in the lumbar spine and hip in 401 nuclear families with a total of 1260 subjects, including 458 premenopausal women, 20–40 years of age; 401 post-menopausal women (mothers), 43–74 years of age; and 401 men (fathers), 49–76 years of age.Results:Significant within-family association was found between the Thr394Thr polymorphism in the PPGAGC1 gene and peak BMD in the femoral neck (P=0.026). Subsequent permutations were in agreement with this significant within-family association result (P=0.016), but Thr394Thr SNP only accounted for 0.7% of the variation in femoral neck peak BMD. However, no significant within-family association was detected between each SNP in the adiponectin gene and peak BMD. Although no significant association was found between BMD and SNP in the PPARGC1 and adiponectin genes in both men and postmenopausal women, haplotype 2 (T-T) in the adiponectin gene was associated with lumbar spine BMD in postmenopausal women (P=0.019).Conclusion:Our findings suggest that Thr394Thr SNP in the PPARGC1 gene was associated with peak BMD in the femoral neck in Chinese women. Confirmation of our results is needed in other populations and with more functional markers within and flanking the PPARGC1 or adiponectin genes region.

A novel VCP mutation as the cause of atypical IBMPFD in a Chinese family

INTRODUCTION Inclusion-body myopathy (IBM) with Pagets disease of bone (PDB) and frontotemporal dementia (FTD), designated as IBMPFD, is a rare, autosomal dominant disorder (MIM 605382). IBMPFD is caused by mutations in the gene that encode valosin-containing protein (VCP). We investigated a Chinese family in which multiple members were diagnosed with PDB and suffered from weakness of the limbs. However, no members of this family were diagnosed with FTD. We made a preliminary diagnosis of PDB, but failed to identify an SQSTM1 mutation in any of the patients. We used whole-exome sequencing to identify the pathogenic gene mutation affecting the Chinese male proband. MATERIALS AND METHODS Altogether, 254 subjects, including one 56-year-old male proband, four affected, related individuals and additional nine family members from a non-consanguineous Chinese family, and 240 healthy donors were recruited and genomic DNA was extracted. All eight exons and the exon-intron boundaries of the SQSTM1 gene were amplified by polymerase chain reaction (PCR) and directly sequenced in five patients (II13, II4, II5, II8, II9). Using whole-exome sequencing, we identified a novel mutation in VCP as the disease-causing mutation. We confirmed the result by sequencing a 500-bp region of the promoter and the coding region of VCP in all 254 of the participants using Sanger sequencing. RESULTS No mutation in the SQSTM1 gene was identified in the five patients examined using direct Sanger sequencing. However, through whole-exome sequencing we were able to identify a novel missense mutation in exon 3 of the VCP gene (p.Gly97Glu) in the Chinese male proband. This mutation was confirmed using Sanger sequencing. The proband, four affected individuals and three unaffected individuals carried this mutation. We were able to correctly diagnose the patients with atypical IBMPFD. Structural analysis of the p.Gly97Glu mutation in the VCP protein showed that the affected amino-acid is located in the interface of the protein. This abnormality may therefore interfere with protein function. CONCLUSIONS This is the first report of a family from China with IBMPFD. A novel VCP mutation was found as the cause of atypical IBMPFD in a Chinese family. Our findings confirm that VCP gene mutations can be a pathogenic cause of IBMPFD.