SGMS2 in primary osteoporosis with facial nerve palsy.

Pathogenic heterozygous variants in SGMS2 cause a rare monogenic form of osteoporosis known as calvarial doughnut lesions with bone fragility (CDL). The clinical presentations of SGMS2-related bone pathology range from childhood-onset osteoporosis with low bone mineral density and sclerotic doughnut-shaped lesions in the skull to a severe spondylometaphyseal dysplasia with neonatal fractures, long-bone deformities, and short stature. In addition, neurological manifestations occur in some patients. SGMS2 encodes sphingomyelin synthase 2 (SMS2), an enzyme involved in the production of sphingomyelin (SM). This review describes the biochemical structure of SM, SM metabolism, and their molecular actions in skeletal and neural tissue. We postulate how disrupted SM gradient can influence bone formation and how animal models may facilitate a better understanding of SGMS2-related osteoporosis.

Variation in the fibroblast growth factor 23 (FGF23) gene associates with serum FGF23 and bone strength in infants.

Introduction: The effects of genetic variation in fibroblast growth factor 23 (FGF23) are unclear. This study explores the associations of single-nucleotide polymorphisms (SNPs) of FGF23 with phosphate and vitamin D metabolism and bone strength in early childhood. Methods: The study is part of the vitamin D intervention in infant (VIDI) trial (2013-2016), in which healthy term infants born to mothers of Northern European origin received vitamin D3 supplementation of 10 or 30 µg/day from 2 weeks to 24 months of age (ClinicalTrials.gov NCT01723852). Intact and C-terminal FGF23 (cFGF23), 25-hydroxyvitamin D (25-OHD), parathyroid hormone, phosphate, and peripheral quantitative computed tomography (pQCT)-derived bone strength parameters were analyzed at 12 and 24 months. The study included 622 VIDI participants with genotyping data on FGF23 SNPs rs7955866, rs11063112, and rs13312770. Results: Rs7955866 minor allele homozygotes had lowest cFGF23 at both time-points (mixed model for repeated measurements, pvariant = 0.009). Minor alleles of rs11063112 were associated with a greater age-related decrease in phosphate concentration (pinteraction = 0.038) from 12 to 24 months. Heterozygotes of rs13312770 had the greatest total bone mineral content (total BMC), cross-sectional area (total CSA), and polar moment of inertia (PMI) at 24 months (ANOVA p = 0.005, 0.037, and 0.036, respectively). Rs13312770 minor alleles were associated with a greater increase of total BMC, but a smaller increase of total CSA and PMI, during follow-up (pinteraction <0.001, 0.043, and 0.012, respectively). Genotype of FGF23 did not modify 25-OHD. Conclusion: The study finds that genetic variation in FGF23 modifies cFGF23, phosphate, and pQCT-derived bone strength parameters from 12 to 24 months of age. These findings potentially promote an understanding of the regulation of FGF23 and its role in bone metabolism and temporal changes thereof during early childhood.

A multi-omics study to characterize the transdifferentiation of human dermal fibroblasts to osteoblast-like cells.

Background: Various skeletal disorders display defects in osteoblast development and function. An in vitro model can help to understand underlying disease mechanisms. Currently, access to appropriate starting material for in vitro osteoblastic studies is limited. Native osteoblasts and their progenitors, the bone marrow mesenchymal stem cells, (MSCs) are problematic to isolate from affected patients and challenging to expand in vitro. Human dermal fibroblasts in vitro are a promising substitute source of cells. Method: We developed an in vitro culturing technique to transdifferentiate fibroblasts into osteoblast-like cells. We obtained human fibroblasts from forearm skin biopsy and differentiated them into osteoblast-like cells with ß-glycerophosphate, ascorbic acid, and dexamethasone treatment. Osteoblastic phenotype was confirmed by staining for alkaline phosphatase (ALP), calcium and phosphate deposits (Alizarin Red, Von Kossa) and by a multi-omics approach (transcriptomic, proteomic, and phosphoproteomic analyses). Result: After 14 days of treatment, both fibroblasts and MSCs (reference cells) stained positive for ALP together with a significant increase in bone specific ALP (p = 0.04 and 0.004, respectively) compared to untreated cells. At a later time point, both cell types deposited minerals, indicating mineralization. In addition, fibroblasts and MSCs showed elevated expression of several osteogenic genes (e.g. ALPL, RUNX2, BMPs and SMADs), and decreased expression of SOX9. Ingenuity Pathways Analysis of RNA sequencing data from fibroblasts and MSCs showed that the osteoarthritis pathway was activated in both cell types (p_adj. = 0.003 and 0.004, respectively). Discussion: These data indicate that our in vitro treatment induces osteoblast-like differentiation in fibroblasts and MSCs, producing an in vitro osteoblastic cell system. This culturing system provides an alternative tool for bone biology research and skeletal tissue engineering.

Biallelic KIF24 Variants Are Responsible for a Spectrum of Skeletal Disorders Ranging From Lethal Skeletal Ciliopathy to Severe Acromesomelic Dysplasia.

Skeletal dysplasias comprise a large spectrum of mostly monogenic disorders affecting bone growth, patterning, and homeostasis, and ranging in severity from lethal to mild phenotypes. This study aimed to underpin the genetic cause of skeletal dysplasia in three unrelated families with variable skeletal manifestations. The six affected individuals from three families had severe short stature with extreme shortening of forelimbs, short long-bones, and metatarsals, and brachydactyly (family 1); mild short stature, platyspondyly, and metaphyseal irregularities (family 2); or a prenatally lethal skeletal dysplasia with kidney features suggestive of a ciliopathy (family 3). Genetic studies by whole genome, whole exome, and ciliome panel sequencing identified in all affected individuals biallelic missense variants in KIF24, which encodes a kinesin family member controlling ciliogenesis. In families 1 and 3, with the more severe phenotype, the affected subjects harbored homozygous variants (c.1457A>G; p.(Ile486Val) and c.1565A>G; p.(Asn522Ser), respectively) in the motor domain which plays a crucial role in KIF24 function. In family 2, compound heterozygous variants (c.1697C>T; p.(Ser566Phe)/c.1811C>T; p.(Thr604Met)) were found C-terminal to the motor domain, in agreement with a genotype-phenotype correlation. In vitro experiments performed on amnioblasts of one affected fetus from family 3 showed that primary cilia assembly was severely impaired, and that cytokinesis was also affected. In conclusion, our study describes novel forms of skeletal dysplasia associated with biallelic variants in KIF24. To our knowledge this is the first report implicating KIF24 variants as the cause of a skeletal dysplasia, thereby extending the genetic heterogeneity and the phenotypic spectrum of rare bone disorders and underscoring the wide range of monogenetic skeletal ciliopathies. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Targeted Exome Sequencing of Genes Involved in Rare CNVs in Early-Onset Severe Obesity.

Context: Rare copy number variants (CNVs) have been associated with the development of severe obesity. However, the potential disease-causing contribution of individual genes within the region of CNVs is often not known. Objective: Screening of rare variants in genes involved in CNVs in Finnish patients with severe early-onset obesity to find candidate genes linked to severe obesity. Methods: Custom-made targeted exome sequencing panel to search for rare (minor allele frequency <0.1%) variants in genes affected by previously identified CNVs in 92 subjects (median age 14 years) with early-onset severe obesity (median body mass index (BMI) Z-score + 4.0). Results: We identified thirteen rare heterozygous variants of unknown significance in eleven subjects in twelve of the CNV genes. Two rare missense variants (p.Pro405Arg and p.Tyr232Cys) were found in SORCS1, a gene highly expressed in the brain and previously linked to diabetes risk. Four rare variants were in genes in the proximal 16p11.2 region (a frameshift variant in TAOK2 and missense variants in SEZ6L2, ALDOA and KIF22) and three rare missense variants were in genes in the 22q11.21 region (AIFM3, ARVCF and KLHL22). Conclusion: We report several rare variants in CNV genes in subjects with childhood obesity. However, the role of the individual genes in the previously identified rare CNVs to development of obesity remains uncertain. More studies are needed to understand the potential role of the specific genes within obesity associated CNVs.

Abnormal Bone Tissue Organization and Osteocyte Lacunocanalicular Network in Early-Onset Osteoporosis Due to SGMS2 Mutations.

Pathological variants in SGMS2, encoding sphingomyelin synthase 2 (SMS2), result in a rare autosomal dominant skeletal disorder with cranial doughnut lesions. The disease manifests as early-onset osteoporosis or a more severe skeletal dysplasia with low bone mineral density, frequent fractures, long-bone deformities, and multiple sclerotic cranial lesions. The exact underlying molecular features and skeletal consequences, however, remain elusive. This study investigated bone tissue characteristics in two adult males with a heterozygous SGMS2 mutation p.Arg50* and significant bone fragility. Transiliac bone biopsy samples from both (patient 1: 61 years; patient 2: 29 years) were analyzed by bone histomorphometry, confocal laser scanning microscopy, and quantitative backscattered electron imaging (qBEI). Bone histomorphometry portrayed largely normal values for structural and turnover parameters, but in both patient 1 and patient 2, respectively, osteoid thickness (-1.80 SD, -1.37 SD) and mineralizing surface (-1.03 SD, -2.73 SD) were reduced and osteoid surface increased (+9.03 SD, +0.98 SD), leading to elevated mineralization lag time (+8.16 SD, +4.10 SD). qBEI showed low and heterogeneous matrix mineralization (CaPeak -2.41 SD, -3.72 SD; CaWidth +7.47 SD, +4.41 SD) with a chaotic arrangement of collagenous fibrils under polarized light. Last, osteocyte lacunae appeared abnormally large and round in shape and the canalicular network severely disturbed with short-spanned canaliculi lacking any orderliness or continuity. Taken together, these data underline a central role for functional SMS2 in bone matrix organization and mineralization, lacunocanalicular network, and in maintaining skeletal strength and integrity. These data bring new knowledge on changes in bone histology resulting from abnormal sphingomyelin metabolism and aid en route to better understanding of sphingolipid-related skeletal disorders. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Phosphate Concentrations and Modifying Factors in Healthy Children From 12 to 24 Months of Age.

CONTEXT: Phosphate homeostasis and its modifiers in early childhood are inadequately characterized. OBJECTIVE: To determine physiological plasma phosphate concentration and modifying factors in healthy infants at 12 to 24 months of age. DESIGN: This study included 525 healthy infants (53% girls), who participated in a randomized vitamin D intervention trial and received daily vitamin D3 supplementation of either 10 or 30 µg from age 2 weeks to 24 months. Biochemical parameters were measured at 12 and 24 months. Dietary phosphate intake was determined at 12 months. MAIN OUTCOME MEASURES: Plasma phosphate concentrations at 12 and 24 months of age. RESULTS: Mean (SD) phosphate concentration decreased from 12 months (1.9 ±â€…0.15 mmol/L) to 24 months (1.6 ±â€…0.17 mmol/L) of age (P < 0.001 for repeated measurements). When adjusted by covariates, such as body size, creatinine, serum 25-hydroxyvitamin D, intact and C-terminal fibroblast growth factor 23, mean plasma phosphate was higher in boys than girls during follow-up (P = 0.019). Phosphate concentrations were similar in the vitamin D intervention groups (P > 0.472 for all). Plasma iron was associated positively with plasma phosphate at both time points (B, 0.006 and 0.005; 95% CI, 0.004-0.009 and 0.002-0.008; P < 0.001 at both time points, respectively). At 24 months of age, the main modifier of phosphate concentration was plasma creatinine (B, 0.007; 95% CI 0.003-0.011, P < 0.001). CONCLUSION: Plasma phosphate concentration decreased from age 12 to 24 months. In infants and toddlers, the strongest plasma phosphate modifiers were sex, iron, and creatinine, whereas vitamin D supplementation did not modify phosphate concentrations.

An ARHGAP25 variant links aberrant Rac1 function to early-onset skeletal fragility.

Ras homologous guanosine triphosphatases (RhoGTPases) control several cellular functions, including cytoskeletal actin remodeling and cell migration. Their activities are downregulated by GTPase-activating proteins (GAPs). Although RhoGTPases are implicated in bone remodeling and osteoclast and osteoblast function, their significance in human bone health and disease remains elusive. Here, we report defective RhoGTPase regulation as a cause of severe, early-onset, autosomal-dominant skeletal fragility in a three-generation Finnish family. Affected individuals (n = 13) presented with multiple low-energy peripheral and vertebral fractures despite normal bone mineral density (BMD). Bone histomorphometry suggested reduced bone volume, low surface area covered by osteoblasts and osteoclasts, and low bone turnover. Exome sequencing identified a novel heterozygous missense variant c.652G>A (p.G218R) in ARHGAP25, encoding a GAP for Rho-family GTPase Rac1. Variants in the ARHGAP25 5' untranslated region (UTR) also associated with BMD and fracture risk in the general population, across multiple genomewide association study (GWAS) meta-analyses (lead variant rs10048745). ARHGAP25 messenger RNA (mRNA) was expressed in macrophage colony-stimulating factor (M-CSF)-stimulated human monocytes and mouse osteoblasts, indicating a possible role for ARHGAP25 in osteoclast and osteoblast differentiation and activity. Studies on subject-derived osteoclasts from peripheral blood mononuclear cells did not reveal robust defects in mature osteoclast formation or resorptive activity. However, analysis of osteosarcoma cells overexpressing the ARHGAP25 G218R-mutant, combined with structural modeling, confirmed that the mutant protein had decreased GAP-activity against Rac1, resulting in elevated Rac1 activity, increased cell spreading, and membrane ruffling. Our findings indicate that mutated ARHGAP25 causes aberrant Rac1 function and consequently abnormal bone metabolism, highlighting the importance of RhoGAP signaling in bone metabolism in familial forms of skeletal fragility and in the general population, and expanding our understanding of the molecular pathways underlying skeletal fragility. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

A new family with epiphyseal chondrodysplasia type Miura.

Epiphyseal chondrodysplasia, Miura type (ECDM) is a skeletal dysplasia with tall stature and distinctive skeletal features caused by heterozygous NPR2 pathogenic variants. Only four families have been reported. We present a family with five affected individuals (mother, three sons, and daughter). The mother's phenotype was relatively mild: borderline tall stature and elongated halluces operated during childhood. The children were remarkably more severely affected with tall stature, scoliosis, and elongated toes and fingers leading to suspicion of Marfan syndrome. Progressive valgus deformities (at the hips, knees, and ankles) were the main complaints and necessitated orthopedic investigations and surgery. Radiographs showed coxa valga, scoliosis, multiple pseudoepiphyses of the fingers and toes with uneven elongation of the digits and ankle valgus. The two older brothers underwent osteotomies and guided growth for axial deformities and arthrodesis for elongated halluces. Genetic testing confirmed the clinical diagnosis of ECDM: all affected individuals had a heterozygous c.2647G>A (p.Val883Met) NPR2 variant in a highly conserved region in the carboxyl-terminal guanylyl cyclase domain. This two-generation family elucidates the clinical and radiological variability of the disease. These rare cases are important to gain further understanding of the fundamental processes of growth regulation.

GNAS, PDE4D, and PRKAR1A Mutations and GNAS Methylation Changes Are Not a Common Cause of Isolated Early-Onset Severe Obesity Among Finnish Children.

Context: Pseudohypoparathyroidism type Ia (PHP1A) is caused by inactivating mutations involving GNAS exons 1-13, encoding the alpha-subunit of the stimulatory G protein (Gsα). Particularly PHP1A, but also other disorders involving the Gsα-cAMP-signaling pathway, have been associated with early-onset obesity. Thus, patients with mutations in the genes encoding PDE4D and PRKAR1A can also be obese. Furthermore, epigenetic GNAS changes, as in pseudohypoparathyroidism type Ib (PHP1B), can lead to excessive weight. Objective: Search for genetic variants in GNAS, PDE4D, and PRKAR1A and for methylation alterations at the GNAS locus in Finnish subjects with isolated severe obesity before age 10 years. Methods: Next generation sequencing to identify pathogenic variants in the coding exons of GNAS, PDE4D, and PRKAR1A; Multiplex Ligation-dependent Probe Amplification (MLPA) and methylation-sensitive MLPA (MS-MLPA) to search for deletions in GNAS and STX16, and for epigenetic changes at the four differentially methylated regions (DMR) within GNAS. Results: Among the 88 subjects (median age 13.8 years, median body mass index Z-score +3.9), we identified one rare heterozygous missense variant of uncertain significance in the XL exon of GNAS in a single patient. We did not identify clearly pathogenic variants in PDE4D and PRKAR1A, and no GNAS methylation changes were detected by MS-MLPA. Conclusions: Our results suggest that coding GNAS mutations or methylation changes at the GNAS DMRs, or coding mutations in PDE4D and PRKAR1A are not common causes of isolated childhood obesity in Finland.

Severe Phenotype of APECED (APS1) Increases Risk for Structural Bone Alterations.

Objective: Immunological abnormalities, the resulting endocrinopathies and their treatments may impact bone health in patients with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED, APS1). The aim of the present study was to describe skeletal characteristics in patients with APECED and the prevalence and risk factors of compromised bone health. Patients and methods: We performed a cross-sectional study on 44 patients (27 females) with APECED and 82 age-, gender- and ethnicity-matched control subjects (54 females). We determined the prevalence of osteoporosis by dual-energy X-ray absorptiometry and skeletal characteristics by peripheral quantitative computed tomography at radius and tibia. Results: Patients were examined at the median age of 37.8 years (range, 7.0-70.1). Dual-energy X-ray absorptiometry indicated osteoporosis in four adult patients (9%); radiographs showed vertebral fractures in three patients. The prevalence of multiple non-spinal fractures was higher in patients than in controls. On peripheral quantitative computed tomography, bone characteristics at distal and proximal radius did not differ between the groups. At distal tibia, patients had lower total (p = 0.009) and trabecular (p = 0.033) volumetric bone mineral density. At the proximal tibia, patients had lower cortical thickness (p < 0.001) than controls. Severity of APECED phenotype influenced both radial and tibial characteristics: cortical thickness and total and trabecular volumetric bone mineral density were lower in patients with ≥7 disease manifestations as compared with more mildly affected patients, whose values were similar to controls. Conclusions: APECED associated with bone structural alterations, especially in patients with a high number of disease manifestations. This may increase the risk of fractures with aging, but symptomatic osteoporosis was rare.

Rare Variants in Genes Linked to Appetite Control and Hypothalamic Development in Early-Onset Severe Obesity.

Context: The hypothalamic circuit has an essential role in the regulation of appetite and energy expenditure. Pathogenic variants in genes involved in the hypothalamic leptin-melanocortin pathway, including melanocortin-4-receptor (MC4R), have been associated with monogenic obesity. Objective: To determine the rate and spectrum of rare variants in genes involved in melanocortin pathway or hypothalamic development in patients with severe early-onset obesity (height-adjusted weight >60% before age 10 years). Methods: We used a custom-made targeted exome sequencing panel to assess peripheral blood DNA samples for rare (minor allele frequency <0.5%), pathogenic/likely pathogenic variants in 24 genes related to the hypothalamic circuit in 92 subjects (51% males, median age 13.7 years) with early-onset severe obesity (median body mass index (BMI) Z-score + 4.0). Results: We identified a novel frameshift deletion in MC4R (p.V103Afs5*) in two unrelated patients and a previously reported MC4R variant (p.T112M) in one patient. In addition, we identified rare heterozygous missense variants in ADCY3 (p.G1110R), MYT1L (p.R807Q), ISL1 (p.I347F), LRP2 (p.R2479I, and p.N3315S) and a hemizygous missense variant in GRPR (p.L87M) (each in one patient), possibly contributing to the obesity phenotype in these patients. Altogether 8 % (7/92) of the subjects had rare pathogenic/likely pathogenic variants in the studied genes. Conclusions: Rare genetic variants within the hypothalamic circuit are prevalent and contribute to the development of severe early-onset obesity. Targeted exome sequencing is useful in identifying affected subjects. Further studies are needed to evaluate the variants' clinical significance and to define optimal treatment.

Genetic variation in GC and CYP2R1 affects 25-hydroxyvitamin D concentration and skeletal parameters: A genome-wide association study in 24-month-old Finnish children.

Vitamin D is important for normal skeletal homeostasis, especially in growing children. There are no previous genome-wide association (GWA) studies exploring genetic factors that influence vitamin D metabolism in early childhood. We performed a GWA study on serum 25-hydroxyvitamin D (25(OH)D) and response to supplementation in 761 healthy term-born Finnish 24-month-old children, who participated in a randomized clinical trial comparing effects of 10 µg and 30 µg of daily vitamin D supplementation from age 2 weeks to 24 months. Using the Illumina Infinium Global Screening Array, which has been optimized for imputation, a total of 686085 markers were genotyped across the genome. Serum 25(OH)D was measured at the end of the intervention at 24 months of age. Skeletal parameters reflecting bone strength were determined at the distal tibia at 24 months using peripheral quantitative computed tomography (pQCT) (data available for 648 children). For 25(OH)D, two strong GWA signals were identified, localizing to GC (Vitamin D binding protein) and CYP2R1 (Vitamin D 25-hydroxylase) genes. The GWA locus comprising the GC gene also associated with response to supplementation. Further evidence for the importance of these two genes was obtained by comparing association signals to gene expression data from the Genotype-Tissue Expression project and performing colocalization analyses. Through the identification of haplotypes associated with low or high 25(OH)D concentrations we used a Mendelian randomization approach to show that haplotypes associating with low 25(OH)D were also associated with low pQCT parameters in the 24-month-old children. In this first GWA study on 25(OH)D in this age group we show that already at the age of 24 months genetic variation influences 25(OH)D concentrations and determines response to supplementation, with genome-wide significant associations with GC and CYP2R1. Also, the dual association between haplotypes, 25(OH)D and pQCT parameters gives support for vertical pleiotropy mediated by 25(OH)D.

The human long non-coding RNA gene RMRP has pleiotropic effects and regulates cell-cycle progression at G2.

RMRP was the first non-coding nuclear RNA gene implicated in a disease. Its mutations cause cartilage-hair hypoplasia (CHH), an autosomal recessive skeletal dysplasia with growth failure, immunodeficiency, and a high risk for malignancies. This study aimed to gain further insight into the role of RNA Component of Mitochondrial RNA Processing Endoribonuclease (RMRP) in cellular physiology and disease pathogenesis. We combined transcriptome analysis with single-cell analysis using fibroblasts from CHH patients and healthy controls. To directly assess cell cycle progression, we followed CHH fibroblasts by pulse-labeling and time-lapse microscopy. Transcriptome analysis identified 35 significantly upregulated and 130 downregulated genes in CHH fibroblasts. The downregulated genes were significantly connected to the cell cycle. Multiple other pathways, involving regulation of apoptosis, bone and cartilage formation, and lymphocyte function, were also affected, as well as PI3K-Akt signaling. Cell-cycle studies indicated that the CHH cells were delayed specifically in the passage from G2 phase to mitosis. Our findings expand the mechanistic understanding of CHH, indicate possible pathways for therapeutic intervention and add to the limited understanding of the functions of RMRP.

Genetic Variation of the Vitamin D Binding Protein Affects Vitamin D Status and Response to Supplementation in Infants.

CONTEXT: Single nucleotide polymorphisms (SNPs) of the vitamin D binding protein encoding the GC (group component) gene affect 25-hydroxyvitamin D (25OHD) concentrations, but their influence on vitamin D status and response to vitamin D supplementation in infants is unknown. OBJECTIVE: To study GC genotype-related differences in 25OHD concentrations and the response to supplementation during a vitamin D intervention study in infants. DESIGN: In this randomized controlled trial, healthy term infants received vitamin D3 (10 or 30 µg/d) from 2 weeks to 24 months of age. GC SNPs rs2282679, rs4588, rs7041, and rs1155563 were genotyped. rs4588/7041 diplotype and haplotypes of rs2282679, rs4588, and rs7041 (Haplo3SNP) and of all four SNPs (Haplo4SNP) were determined. MAIN OUTCOME MEASURES: 25OHD measured in cord blood at birth and at 12 and 24 months during intervention. RESULTS: A total of 913 infants were included. Minor allele homozygosity of all studied GC SNPs, their combined haplotypes, and rs4588/rs7041 diplotype 2/2 were associated with lower 25OHD concentrations at all time points in one or both intervention groups [analysis of covariance (ANCOVA) P < 0.043], with the exception of rs7041, which did not affect 25OHD at birth. In the high-dose supplementation group receiving 30 µg/d vitamin D3, but not in those receiving 10 µg/d, genotype of rs2282679, rs4588, and rs7041; diplotype; and Haplo3SNP significantly affected intervention response (repeated measurement ANCOVA Pinteraction < 0.019). Minor allele homozygotes had lower 25OHD concentrations and smaller increases in 25OHD throughout the intervention. CONCLUSIONS: In infants, vitamin D binding protein genotype affects 25OHD concentration and efficiency of high-dose vitamin D3 supplementation.

A preliminary transcriptome analysis suggests a transitory effect of vitamin D on mitochondrial function in obese young Finnish subjects.

OBJECTIVE: The effect of vitamin D at the transcriptome level is poorly understood, and furthermore, it is unclear if it differs between obese and normal-weight subjects. The objective of the study was to explore the transcriptome effects of vitamin D supplementation. DESIGN AND METHODS: We analysed peripheral blood gene expression using GlobinLock oligonucleotides followed by RNA sequencing in individuals participating in a 12-week randomised double-blinded placebo-controlled vitamin D intervention study. The study involved 18 obese and 18 normal-weight subjects (of which 20 males) with mean (±s.d.) age 20.4 (±2.5) years and BMIs 36 (±10) and 23 (±4) kg/m2, respectively. The supplemental daily vitamin D dose was 50 µg (2000 IU). Data were available at baseline, 6- and 12-week time points and comparisons were performed between the vitamin D and placebo groups separately in obese and normal-weight subjects. RESULTS: Significant transcriptomic changes were observed at 6 weeks, and only in the obese subjects: 1724 genes were significantly upregulated and 186 genes were downregulated in the vitamin D group compared with placebo. Further analyses showed several enriched gene categories connected to mitochondrial function and metabolism, and the most significantly enriched pathway was related to oxidative phosphorylation (adjusted P value 3.08 × 10-14). Taken together, our data suggest an effect of vitamin D supplementation on mitochondrial function in obese subjects. CONCLUSIONS: Vitamin D supplementation affects gene expression in obese, but not in normal-weight subjects. The altered genes are enriched in pathways related to mitochondrial function. The present study increases the understanding of the effects of vitamin D at the transcriptome level.

Osteoporosis and skeletal dysplasia caused by pathogenic variants in SGMS2.

Mechanisms leading to osteoporosis are incompletely understood. Genetic disorders with skeletal fragility provide insight into metabolic pathways contributing to bone strength. We evaluated 6 families with rare skeletal phenotypes and osteoporosis by next-generation sequencing. In all the families, we identified a heterozygous variant in SGMS2, a gene prominently expressed in cortical bone and encoding the plasma membrane-resident sphingomyelin synthase SMS2. Four unrelated families shared the same nonsense variant, c.148C>T (p.Arg50*), whereas the other families had a missense variant, c.185T>G (p.Ile62Ser) or c.191T>G (p.Met64Arg). Subjects with p.Arg50* presented with childhood-onset osteoporosis with or without cranial sclerosis. Patients with p.Ile62Ser or p.Met64Arg had a more severe presentation, with neonatal fractures, severe short stature, and spondylometaphyseal dysplasia. Several subjects had experienced peripheral facial nerve palsy or other neurological manifestations. Bone biopsies showed markedly altered bone material characteristics, including defective bone mineralization. Osteoclast formation and function in vitro was normal. While the p.Arg50* mutation yielded a catalytically inactive enzyme, p.Ile62Ser and p.Met64Arg each enhanced the rate of de novo sphingomyelin production by blocking export of a functional enzyme from the endoplasmic reticulum. SGMS2 pathogenic variants underlie a spectrum of skeletal conditions, ranging from isolated osteoporosis to complex skeletal dysplasia, suggesting a critical role for plasma membrane-bound sphingomyelin metabolism in skeletal homeostasis.

A novel MYT1L mutation in a patient with severe early-onset obesity and intellectual disability.

The genetic background of severe early-onset obesity is still incompletely understood. Deletions at 2p25.3 associate with early-onset obesity and variable intellectual disability. Myelin-transcriptor-factor-1-like (MYT1L) gene in this locus has been proposed a candidate gene for obesity. We report on a 13-year-old boy presenting with overweight already at 1 year of age (body mass index [BMI] Z-score +2.3) and obesity at 2 years of age (BMI Z-score +3.8). The patient had hyperphagia and delayed neurological, cognitive and motor development. He also had speech delay, strabismus, hyperactivity and intellectual disability. Brain MRI was normal. The parents and sister had normal BMI. Whole-genome sequencing identified in the index patient a novel de novo frameshift deletion that introduces a premature termination of translation NM_015025.2(MYT1L): c.2215_2224delACGCGCTGCC, p.(Thr739Alafs*7) in MYT1L. The frameshift variant was confirmed by Sanger sequencing. Our finding supports the association of MYT1L mutations with early-onset syndromic obesity. The identification of novel monogenic forms of childhood-onset obesity will provide insights to the involved genetic and biologic pathways.

Low free 25-hydroxyvitamin D and high vitamin D binding protein and parathyroid hormone in obese Caucasians. A complex association with bone?

BACKGROUND: Studies have shown altered vitamin D metabolism in obesity. We assessed differences between obese and normal-weight subjects in total, free, and bioavailable 25-hydroxyvitamin D (25(OH)D, 25(OH)DFree, and 25(OH)DBio, respectively), vitamin D binding protein (DBP), parathyroid hormone (PTH) and bone traits. METHODS: 595 37-47-year-old healthy Finnish men and women stratified by BMI were examined in this cross-sectional study. Background characteristic and intakes of vitamin D and calcium were collected. The concentrations of 25(OH)D, PTH, DBP, albumin and bone turnover markers were determined from blood. 25(OH)DFree and 25(OH)DBio were calculated. pQCT was performed at radius and tibia. RESULTS: Mean±SE (ANCOVA) 25(OH)DFree (10.8±0.6 vs 12.9±0.4 nmol/L; P = 0.008) and 25(OH)DBio (4.1±0.3 vs 5.1±0.1 nmol/L; P = 0.003) were lower in obese than in normal-weight women. In men, 25(OH)D (48.0±2.4 vs 56.4±2.0 nmol/L, P = 0.003), 25(OH)DFree (10.3±0.7 vs 12.5±0.6 pmol/L; P = 0.044) and 25(OH)DBio (4.2±0.3 vs 5.1±0.2 nmol/L; P = 0.032) were lower in obese. Similarly in all subjects, 25(OH)D, 25(OH)DFree and 25(OH)DBio were lower in obese (P<0.001). DBP (399±12 vs 356±7mg/L, P = 0.008) and PTH (62.2±3.0 vs 53.3±1.9 ng/L; P = 0.045) were higher in obese than in normal-weight women. In all subjects, PTH and DBP were higher in obese (P = 0.047and P = 0.004, respectively). In obese women, 25(OH)D was negatively associated with distal radius trabecular density (R2 = 0.089, P = 0.009) and tibial shaft cortical strength index (CSI) (R2 = 0.146, P = 0.004). 25(OH)DFree was negatively associated with distal radius CSI (R2 = 0.070, P = 0.049), radial shaft cortical density (CorD) (R2 = 0.050, P = 0.045), and tibial shaft CSI (R2 = 0.113, P = 0.012). 25(OH)DBio was negatively associated with distal radius CSI (R2 = 0.072, P = 0.045), radial shaft CorD (R2 = 0.059, P = 0.032), and tibial shaft CSI (R2 = 0.093, P = 0.024). CONCLUSIONS: The associations between BMI and 25(OH)D, 25(OH)DFree, and 25(OH)DBio, DBP, and PTH suggest that obese subjects may differ from normal-weight subjects in vitamin D metabolism. BMI associated positively with trabecular bone traits and CSI in our study, and slightly negatively with cortical bone traits. Surprisingly, there was a negative association of free and bioavailable 25(OH)D and some of the bone traits in obese women.