RESUMEN
Pathogenic variants disrupting the binding between sclerostin (encoded by SOST) and its receptor LRP4 have previously been described to cause sclerosteosis, a rare high bone mass disorder. The sclerostin-LRP4 complex inhibits canonical WNT signaling, a key pathway regulating osteoblastic bone formation and a promising therapeutic target for common bone disorders, such as osteoporosis. In the current study, we crossed mice deficient for Sost (Sost-/-) with our p.Arg1170Gln Lrp4 knock-in (Lrp4KI/KI) mouse model to create double mutant Sost-/-;Lrp4KI/KI mice. We compared the phenotype of Sost-/- mice with that of Sost-/-;Lrp4KI/KI mice, to investigate a possible synergistic effect of the disease-causing p.Arg1170Trp variant in Lrp4 on Sost deficiency. Interestingly, presence of Lrp4KI alleles partially mitigated the Sost-/- phenotype. Cellular and dynamic histomorphometry did not reveal mechanistic insights into the observed phenotypic differences. We therefore determined the molecular effect of the Lrp4KI allele by performing bulk RNA sequencing on Lrp4KI/KI primary osteoblasts. Unexpectedly, mostly genes related to bone resorption or remodeling (Acp5, Rankl, Mmp9) were upregulated in Lrp4KI/KI primary osteoblasts. Verification of these markers in Lrp4KI/KI, Sost-/- and Sost-/-;Lrp4KI/KI mice revealed that sclerostin deficiency counteracts this Lrp4KI/KI effect in Sost-/-;Lrp4KI/KI mice. We therefore hypothesize that models with two inactivating Lrp4KI alleles rather activate bone remodeling, with a net gain in bone mass, whereas sclerostin deficiency has more robust anabolic effects on bone formation. Moreover, these effects of sclerostin and Lrp4 are stronger in female mice, contributing to a more severe phenotype than in males and more detectable phenotypic differences among different genotypes.
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Proteínas Adaptadoras Transductoras de Señales , Remodelación Ósea , Hiperostosis , Sindactilia , Masculino , Femenino , Animales , Ratones , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Ratones Noqueados , Fenotipo , Mutación , Remodelación Ósea/genética , Proteínas Relacionadas con Receptor de LDL/genética , Proteínas Relacionadas con Receptor de LDL/metabolismoRESUMEN
Paget's disease of bone (PDB) is a common, late-onset bone disorder, characterized by focal increases of bone turnover that can result in bone lesions. Heterozygous pathogenic variants in the Sequestosome 1 (SQSTM1) gene are found to be the main genetic cause of PDB. More recently, PFN1 and ZNF687 have been identified as causal genes in patients with a severe, early-onset, polyostotic form of PDB, and an increased likelihood to develop giant cell tumors. In our study, we screened the coding regions of PFN1 and ZNF687 in a Belgian PDB cohort (n = 188). In the PFN1 gene, no variants could be identified, supporting the observation that variants in this gene are extremely rare in PDB. However, we identified 3 non-synonymous coding variants in ZNF687. Interestingly, two of these rare variants (p.Pro937His and p.Arg939Cys) were clustering in the nuclear localization signal of the encoded ZNF687 protein, also harboring the p.Pro937Arg variant, a previously reported disease-causing variant. In conclusion, our findings support the involvement of genetic variation in ZNF687 in the pathogenesis of classical PDB, thereby expanding its mutational spectrum.
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Osteítis Deformante , Humanos , Osteítis Deformante/genética , Osteítis Deformante/patología , Señales de Localización Nuclear/genética , Proteína Sequestosoma-1/genética , Pruebas Genéticas , Factores de Transcripción/genética , Mutación , Profilinas/genéticaRESUMEN
Hyperostosis Cranialis Interna (HCI) is a rare bone disorder characterized by progressive intracranial bone overgrowth at the skull. Here we identified by whole-exome sequencing a dominant mutation (L441R) in SLC39A14 (ZIP14). We show that L441R ZIP14 is no longer trafficked towards the plasma membrane and excessively accumulates intracellular zinc, resulting in hyper-activation of cAMP-CREB and NFAT signaling. Conditional knock-in mice overexpressing L438R Zip14 in osteoblasts have a severe skeletal phenotype marked by a drastic increase in cortical thickness due to an enhanced endosteal bone formation, resembling the underlying pathology in HCI patients. Remarkably, L438R Zip14 also generates an osteoporotic trabecular bone phenotype. The effects of osteoblastic overexpression of L438R Zip14 therefore mimic the disparate actions of estrogen on cortical and trabecular bone through osteoblasts. Collectively, we reveal ZIP14 as a novel regulator of bone homeostasis, and that manipulating ZIP14 might be a therapeutic strategy for bone diseases.
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Proteínas de Transporte de Catión/genética , Homeostasis/genética , Hiperostosis/genética , Mutación , Osteosclerosis/genética , Base del Cráneo/anomalías , Animales , Línea Celular , Células Cultivadas , Modelos Animales de Enfermedad , Células HEK293 , Humanos , Hiperostosis/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoblastos/citología , Osteoblastos/metabolismo , Osteosclerosis/metabolismo , Transducción de Señal/genética , Base del Cráneo/metabolismo , Zinc/metabolismoRESUMEN
A role for WNT4 and WNT5B in bone metabolism was indicated by genome-wide association studies (GWAS) and a Wnt4 knockout mouse model. The aim of this study was therefore to replicate and further investigate the causality between genetic variation in WNT4 and WNT5B and deviating bone mineral density (BMD) values. A WNT4 and WNT5B mutation screening was performed in patients with craniotubular hyperostosis using Sanger sequencing. Here, no putative causal mutations were detected. Moreover, a high and low BMD cohort was selected from the Odense Androgen Study population for re-sequencing. In WNT4 we detected four variants (three rare, one common), while in WNT5B we detected five variants (two rare, three common). For the common variants, no significant difference in genotype frequencies between the high and low BMD cohorts was observed. The SNPs associated with the GWAS were genotyped in these cohorts, but again no significant difference in genotype frequencies was observed. Despite the findings of the GWAS, we were not able to replicate or further verify the genetic association of polymorphisms in WNT4 and WNT5B with BMD. In order to do so, the intronic regions of both genes could be investigated more thoroughly in more extended populations (or extremes) with greater power. Future genetic and functional studies toward adjacent genes of WNT4 and WNT5B can also be interesting to figure out whether the signal from GWAS could possibly be attributed to genetic variation in these genes.
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Densidad Ósea/genética , Predisposición Genética a la Enfermedad , Osteoporosis/genética , Proteínas Wnt/genética , Proteína Wnt4/genética , Estudios de Cohortes , Pruebas Genéticas/métodos , Variación Genética/genética , Estudio de Asociación del Genoma Completo , Genotipo , Humanos , Persona de Mediana Edad , Osteoporosis/fisiopatología , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
Camurati-Engelmann disease (CED, OMIM 131300), or progressive diaphyseal dysplasia, is a rare autosomal dominant skeletal dysplasia, caused by mutations in the transforming growth factor-ß1 (TGFß1) gene. We describe the first Indian CED family with genetic confirmation and presenting manifestations. The proband is a 17-year-old woman who presented with lower limb pain and proximal muscle weakness. Skeletal radiographs of the long bones revealed cortical, periosteal, and endosteal thickenings, predominantly affecting the diaphyses of the long bones. On detailed evaluation, there was a strong family history of bone disorder with similar symptoms of pain and radiological findings in several family members. Exon sequencing of the TGFß1 gene was performed in available family members. Based on clinical and radiographic studies and its familial nature, a diagnosis of CED was made and confirmed by mutation analysis. A heterozygous G to A transition in exon 4 of the TGFß1 gene (R218H) was detected in 5 out of 10 available family members, including 4 affecteds and 1 asymptomatic individual. Many of our affected individuals responded to glucocorticoids and cortical windowing. CED is a rare genetic disease with variable clinical manifestations and incomplete penetrance. CED needs to be considered in the differential diagnosis of nonspecific limb pain and waddling gait in all young individuals.
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Síndrome de Camurati-Engelmann , Adolescente , Pueblo Asiatico , Síndrome de Camurati-Engelmann/diagnóstico por imagen , Femenino , Humanos , Extremidad Inferior , Debilidad Muscular/diagnóstico por imagen , Mialgia/diagnóstico por imagen , Linaje , Radiografía , Cintigrafía , Imagen de Cuerpo EnteroRESUMEN
CONTEXT: Osteopathia striata with cranial sclerosis (OSCS) is a rare bone disorder with X-linked dominant inheritance, characterized by a generalized hyperostosis in the skull and long bones and typical metaphyseal striations in the long bones. So far, loss-of-function variants in AMER1 (also known as WTX or FAM123B), encoding the APC membrane recruitment protein 1 (AMER1), have been described as the only molecular cause for OSCS. AMER1 promotes the degradation of ß-catenin via AXIN stabilization, acting as a negative regulator of the WNT/ß-catenin signaling pathway, a central pathway in bone formation. OBJECTIVE: In this study, we describe a Dutch adult woman with an OSCS-like phenotype, namely, generalized high bone mass and characteristic metaphyseal striations, but no genetic variant affecting AMER1. RESULTS: Whole exome sequencing led to the identification of a mosaic missense variant (c.876A > C; p.Lys292Asn) in CTNNB1, coding for ß-catenin. The variant disrupts an amino acid known to be crucial for interaction with AXIN, a key factor in the ß-catenin destruction complex. Western blotting experiments demonstrate that the p.Lys292Asn variant does not significantly affect the ß-catenin phosphorylation status, and hence stability in the cytoplasm. Additionally, luciferase reporter assays were performed to investigate the effect of p.Lys292Asn ß-catenin on canonical WNT signaling. These studies indicate an average 70-fold increase in canonical WNT signaling activity by p.Lys292Asn ß-catenin. CONCLUSION: In conclusion, this study indicates that somatic variants in the CTNNB1 gene could explain the pathogenesis of unsolved cases of osteopathia striata.
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Mosaicismo , Osteosclerosis , beta Catenina , Humanos , beta Catenina/genética , beta Catenina/metabolismo , Femenino , Osteosclerosis/genética , Osteosclerosis/patología , Mutación Missense , Adulto , Vía de Señalización Wnt/genética , Persona de Mediana Edad , Secuenciación del Exoma , Proteínas Supresoras de Tumor , Proteínas Adaptadoras Transductoras de SeñalesRESUMEN
Introduction: Spondylocostal dysostosis (SCD) is characterized by multiple vertebral abnormalities associated with abnormalities of the ribs. Five genes causative for the disease have been identified. These include DLL3 (OMIM *602768), MESP2 (OMIM #608681), LFNG (OMIM #609813), TBX6 (OMIM *602427), and HES7 (OMIM *608059). Methods: In the current study, we investigated a Pakistani consanguineous family segregating spondylocostal dysotosis. Whole-exome sequencing (WES) followed by Sanger sequencing was performed using DNA of affected and unaffected individuals to identify pathogenic variant(s). The identified variant was interpreted using ACMG classification. Literature review was performed to summarize currently known mutated alleles of DLL3 and the underlying clinical phenotypes. Results: Clinical examination using anthropometric measurements and radiographs diagnosed the patients to be afflicted with SCD. Pedigree analysis of the affected family showed an autosomal recessive inheritance pattern of the disease. WES followed by Sanger sequencing identified a novel homozygous nonsense variant (DLL3(NM_016941.4): c.535G>T; p.Glu179Ter) in the DLL3 gene located on chromosome 19q13.2. Conclusion: The study will be helpful in carrier testing and genetic counseling to prevent segregation of the disease to the next generations within this family. It also provides knowledge for clinicians and researchers in search of a better understanding of SCD anomalies.
RESUMEN
Background: Camurati-Engelmann disease (CED) is a rare bone dysplasia characterized by diffuse diaphyseal osteosclerosis. Skull base involvement in CED can result in hypopituitarism but is seldom reported. Our objective was to report a patient with acquired hypopituitarism due to CED and assess the management challenges. Case Report: A 20-year-old boy presented with lower limb pain. He had walking difficulty in childhood, which was diagnosed as CED and managed with prednisolone. He later discontinued treatment and was lost to follow-up. Current re-evaluation showed short stature (-3.6 standard deviation), low weight (-4.3 standard deviation), and delayed puberty with delayed bone age (13 years). He was found to have secondary hypogonadism (luteinizing hormone level, 0.1 mIU/mL [1.7-8.6 mIU/mL]; follicle-stimulating hormone level, 1.0 mIU/mL [1.5-12.4 mIU/mL]; and testosterone level, 0.087 nmol/L [9-27 nmol/L]), growth hormone deficiency (low insulin-like growth factor I level, 120 ng/mL [226-903 ng/mL] and peak growth hormone level of 7 ng/mL on insulin-induced hypoglycemia), and secondary hypocortisolism (cortisol level, 105 nmol/L [170-550 nmol/L] and adrenocorticotropic hormone level, 6 pg/mL [5-65 pg/mL]). Serum prolactin level was normal (8.3 ng/mL [5-20 ng/mL]), and he was euthyroid on levothyroxine replacement. Magnetic resonance imaging revealed a partially empty sella. Sanger sequencing revealed a missense mutation (p.R218C/c.652C>T) in exon 4 of the TGFß1 gene. The patient was treated with zoledronate, losartan, and oral prednisolone and continued on levothyroxine and testosterone replacement, which resulted in symptomatic improvement. Discussion: The index case manifested severe CED requiring multimodality therapy. Later, he developed combined pituitary hormone deficiencies, which were managed with thyroid and gonadal hormone replacement with the continuation of glucocorticoids. The partial efficacy of bisphosphonates in CED has been reported in the literature. Conclusion: Skull base involvement in CED can lead to structural and functional hypopituitarism as a result of intracranial hypertension.
RESUMEN
We examined the interaction of ECM1 (extracellular matrix protein 1) using yeast two-hybrid screening and identified the type II transmembrane protein, PLSCR1 (phospholipid scramblase 1), as a binding partner. This interaction was then confirmed by in vitro and in vivo co-immunoprecipitation experiments, and additional pull-down experiments with GST-tagged ECM1a fragments localized this interaction to occur within the tandem repeat region of ECM1a. Furthermore, immunohistochemical staining revealed a partial overlap of ECM1 and PLSCR1 in human skin at the basal epidermal cell layer. Moreover, in human skin equivalents, both proteins are expressed at the basal membrane in a dermal fibroblast-dependent manner. Next, immunogold electron microscopy of ultrathin human skin sections showed that ECM1 and PLSCR1 co-localize in the extracellular matrix, and using antibodies against ECM1 or PLSCR1 cross-linked to magnetic immunobeads, we were able to demonstrate PLSCR1-ECM1 interaction in human skin extracts. Furthermore, whereas ECM1 is secreted by the endoplasmic/Golgi-dependent pathway, PLSCR1 release from HaCaT keratinocytes occurs via a lipid raft-dependent mechanism, and is deposited in the extracellular matrix. In summary, we here demonstrate that PLSCR1 interacts with the tandem repeat region of ECM1a in the dermal epidermal junction zone of human skin and provide for the first time experimental evidence that PLSCR1 is secreted by an unconventional secretion pathway. These data suggest that PLSCR1 is a multifunctional protein that can function both inside and outside of the cell and together with ECM1 may play a regulatory role in human skin.
Asunto(s)
Dermis/metabolismo , Epidermis/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Proteínas de Transferencia de Fosfolípidos/metabolismo , Vías Secretoras , Piel/metabolismo , Línea Celular , Células Cultivadas , Dermis/enzimología , Epidermis/enzimología , Matriz Extracelular/enzimología , Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/química , Proteínas de la Matriz Extracelular/genética , Humanos , Uniones Intercelulares/enzimología , Uniones Intercelulares/metabolismo , Queratinocitos/enzimología , Queratinocitos/metabolismo , Metabolismo de los Lípidos , Proteínas de Transferencia de Fosfolípidos/genética , Unión Proteica , Estructura Terciaria de Proteína , Transporte de Proteínas , Piel/enzimologíaRESUMEN
Sclerosteosis is a high bone mass disorder, caused by pathogenic variants in the genes encoding sclerostin or LRP4. Both proteins form a complex that strongly inhibits canonical WNT signaling activity, a pathway of major importance in bone formation. So far, all reported disease-causing variants are located in the third ß-propeller domain of LRP4, which is essential for the interaction with sclerostin. Here, we report the identification of two compound heterozygous variants, a known p.Arg1170Gln and a novel p.Arg632His variant, in a patient with a sclerosteosis phenotype. Interestingly, the novel variant is located in the first ß-propeller domain, which is known to be indispensable for the interaction with agrin. However, using luciferase reporter assays, we demonstrated that both the p.Arg1170Gln and the p.Arg632His variant in LRP4 reduced the inhibitory capacity of sclerostin on canonical WNT signaling activity. In conclusion, this study is the first to demonstrate that a pathogenic variant in the first ß-propeller domain of LRP4 can contribute to the development of sclerosteosis, which broadens the mutational spectrum of the disorder.
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Proteínas Adaptadoras Transductoras de Señales/genética , Hiperostosis/patología , Proteínas Relacionadas con Receptor de LDL/genética , Mutación , Sindactilia/patología , Vía de Señalización Wnt , Humanos , Hiperostosis/etiología , Hiperostosis/metabolismo , Masculino , Persona de Mediana Edad , Pronóstico , Dominios Proteicos , Sindactilia/etiología , Sindactilia/metabolismoRESUMEN
INTRODUCTION: Spondylo-epi-metaphyseal dysplasia (SEMD) represents a group of osteo-chondrodysplasias characterized by vertebral, epiphyseal as well as metaphyseal abnormalities. Several genes have been identified underlying the different forms. METHODOLOGY AND RESULTS: Two relatives (cousins) in a family were found to have disproportionate short stature with clinical and radiological features suggestive of SEMD. Metabolic bone profile was normal including parathyroid hormone and 25(OH)vitamin D3. Exome sequencing revealed a missense mutation (p. T120M) in the von-Willebrand factor A-domain of the Matrilin 3 (MATN3) gene that segregates with the disease in the family. CONCLUSION: We identified a homozygous missense mutation in MATN3, an important structural component of the extracellular matrix of cartilage, as the genetic cause of SEMD in this pedigree. MATN3 mutations have been more commonly associated with multiple epiphyseal dysplasia than SEMD. Recognition of this mutation will aid in enhancing the understanding and expanding the spectrum of this particular skeletal dysplasia.
RESUMEN
Sclerosteosis is a rare autosomal recessive bone disorder marked by hyperostosis of the skull and tubular bones. Initially, we and others reported that sclerosteosis was caused by loss-of-function mutations in SOST, encoding sclerostin. More recently, we identified disease-causing mutations in LRP4, a binding partner of sclerostin, in three sclerosteosis patients. Upon binding to sclerostin, LRP4 can inhibit the canonical WNT signaling that is known to be an important pathway in the regulation of bone formation. To further investigate the role of LRP4 in the bone formation process, we generated an Lrp4 mutated sclerosteosis mouse model by introducing the p.Arg1170Gln mutation in the mouse genome. Extensive analysis of the bone phenotype of the Lrp4R1170Q/R1170Q knock-in (KI) mouse showed the presence of increased trabecular and cortical bone mass as a consequence of increased bone formation by the osteoblasts. In addition, three-point bending analysis also showed that the increased bone mass results in increased bone strength. In contrast to the human sclerosteosis phenotype, we could not observe syndactyly in the forelimbs or hindlimbs of the Lrp4 KI animals. Finally, we could not detect any significant changes in the bone formation and resorption markers in the serum of the mutant mice. However, the serum sclerostin levels were strongly increased and the level of sclerostin in the tibia was decreased in Lrp4R1170Q/R1170Q mice, confirming the role of LRP4 as an anchor for sclerostin in bone. In conclusion, the Lrp4R1170Q/R1170Q mouse is a good model for the human sclerosteosis phenotype caused by mutations in LRP4 and can be used in the future for further investigation of the mechanism whereby LRP4 regulates bone formation. © 2017 American Society for Bone and Mineral Research.
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Glicoproteínas/metabolismo , Homocigoto , Hiperostosis , Mutación Missense , Receptores de LDL , Sindactilia , Tibia/metabolismo , Vía de Señalización Wnt , Proteínas Adaptadoras Transductoras de Señales , Sustitución de Aminoácidos , Animales , Modelos Animales de Enfermedad , Técnicas de Sustitución del Gen , Glicoproteínas/genética , Humanos , Hiperostosis/genética , Hiperostosis/metabolismo , Hiperostosis/patología , Péptidos y Proteínas de Señalización Intercelular , Proteínas Relacionadas con Receptor de LDL , Ratones , Ratones Noqueados , Receptores de LDL/genética , Receptores de LDL/metabolismo , Sindactilia/genética , Sindactilia/metabolismo , Sindactilia/patología , Tibia/patologíaRESUMEN
We report recurrence of osteopathia striata with cranial sclerosis (OSCS) in two full siblings conceived by unaffected parents. Molecular confirmation of OSCS in both siblings was achieved by identification of a novel heterozygous mutation in the WTX gene. Neither parent had clinical features of OSCS nor was the pathogenic mutation demonstrable in DNA extracted from both peripheral blood leucocytes and buccal cells. This case demonstrates germline mosaicism in OSCS and represents the third report of mosaicism affecting the germline in families with OSCS. Previous reports were of parental gonadosomal mosaicism, with one showing recurrence in multiple children. Our observation adds to a body of evidence that suggests that germline mosaicism in OSCS may occur more frequently than believed previously and may have implications for counselling families with OSCS.
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Mutación de Línea Germinal , Mosaicismo , Osteosclerosis/diagnóstico , Osteosclerosis/genética , Hermanos , Proteínas Adaptadoras Transductoras de Señales/genética , Huesos/diagnóstico por imagen , Huesos/patología , Análisis Mutacional de ADN , Femenino , Humanos , Lactante , Imagen por Resonancia Magnética , Mutación , Linaje , Radiografía , Proteínas Supresoras de Tumor/genéticaRESUMEN
Mutations in the LRP4 gene, coding for a Wnt signaling coreceptor, have been found to cause several allelic conditions. Among these, two are characterized by a strong skeletal involvement, namely sclerosteosis and Cenani-Lenz syndrome. In this work, we evaluated the role of LRP4 in the pathophysiology of these diseases. First, we report a novel LRP4 mutation, leading to the substitution of arginine at position 1170 in glutamine, identified in a patient with sclerosteosis. This mutation is located in the central cavity of the third ß-propeller domain, which is in line with two other sclerosteosis mutations we previously described. Reporter assays demonstrate that this mutation leads to impaired sclerostin inhibition of Wnt signaling. Moreover, we compared the effect of this novel variant to mutations causing Cenani-Lenz syndrome and show that impaired membrane trafficking of the LRP4 protein is the likely mechanism underlying Cenani-Lenz syndrome. This is in contrast to sclerosteosis mutations, previously shown to impair the binding between LRP4 and sclerostin. In addition, to better understand the biology of LRP4, we investigated the circulating sclerostin levels in the serum of a patient suffering from sclerosteosis owing to a LRP4 mutation. We demonstrate that impaired sclerostin binding to the mutated LRP4 protein leads to dramatic increase in circulating sclerostin in this patient. With this study, we provide the first evidence suggesting that LRP4 is responsible for the retention of sclerostin in the bone environment in humans. These findings raise potential concerns about the utility of determining circulating sclerostin levels as a marker for other bone-related parameters. Although more studies are needed to fully understand the mechanism whereby LRP4 facilitates sclerostin action, it is clear that this protein represents a potent target for future osteoporosis therapies and an interesting alternative for the antisclerostin treatment currently under study.
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Proteínas Morfogenéticas Óseas , Marcadores Genéticos , Hiperostosis , Proteínas Relacionadas con Receptor de LDL , Mutación Missense , Sindactilia , Proteínas Adaptadoras Transductoras de Señales , Sustitución de Aminoácidos , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Marcadores Genéticos/genética , Células HEK293 , Humanos , Hiperostosis/genética , Hiperostosis/metabolismo , Proteínas Relacionadas con Receptor de LDL/genética , Proteínas Relacionadas con Receptor de LDL/metabolismo , Unión Proteica , Dominios Proteicos , Sindactilia/genética , Sindactilia/metabolismoRESUMEN
Sclerosing bone dysplasias are a heterogeneous group of rare diseases marked by increased BMD caused by either increased bone formation or by decreased bone resorption. In this study we have focused on craniotubular hyperostoses mainly affecting the long bones and the skull. Currently, there are three causative genes identified namely LRP5, SOST and LRP4. All three genes are involved in the canonical Wnt signalling pathway. These findings support the role of this pathway in regulating bone formation. The secreted Frizzled related proteins (sFRPs) can modulate the Wnt signalling pathway by binding to Wnt ligands or Frizzled receptors. Studies using mice showed that two members of this family, sFRP1 and sFRP4, have an important effect on bone formation. Sfrp1-/- mice have increased BMD values especially after peak BMD was reached. On the contrary, sfrp4 overexpression mice exhibit reduced BMD. Therefore, we selected sFRP1 and sFRP4, two members of the secreted Frizzled related protein (sFRP) family, as candidate genes for mutation analysis in patients with craniotubular hyperostosis. Using Sanger sequencing we screened the exons and intron/exon boundaries of sFRP1 and sFRP4 in 53 patients. In all patients mutations in LRP5, SOST and LRP4 were excluded. We identified two unknown heterozygous variants both in sFRP1. The first variant in sFRP1 is an intronic variant which, according to prediction programs, does not affect the splicing of the gene. The second variant (p.Trp131Arg/-) was identified in a young boy whose healthy mother does not carry the variant. In conclusion, our studies indicate that mutations neither in sFRP1 nor in sFRP4 are a common cause of craniotubular hyperostoses. As a consequence, further research will be necessary to identify the disease causing gene(s) in this group of patients.
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Hiperostosis/genética , Péptidos y Proteínas de Señalización Intercelular/genética , Proteínas de la Membrana/genética , Mutación , Proteínas Proto-Oncogénicas/genética , Cráneo/patología , Animales , Densidad Ósea , Estudios de Cohortes , Femenino , Masculino , Ratones , LinajeRESUMEN
Osteoporosis is a common disease characterized by an increased susceptibility to fracture. It is a complex disorder resulting from the interaction of several polymorphisms in different genes and environmental factors. Since we recently reported a role for low density lipoprotein-related protein (LRP)-4 in monogenic disorders with bone overgrowth, we now wanted to evaluate whether genetic variation in the LRP4 gene has an effect on the susceptibility to osteoporosis in a population based cohort from the Odense Androgen Study. We chose to genotype four common (minor allele frequency (MAF)≥0.05) and non-synonymous coding polymorphisms located in the extracellular region of the LRP4 protein: rs3816614 (A/g), rs2306029 (G/a), rs2306033 (C/t) and rs6485702 (G/a) (large and small characters indicate major and minor alleles, respectively). Bone mineral density (BMD) measurements of the hip, the spine and whole body as well as different hip geometry parameters were available for a total of 1404 Danish men from two age groups ([20-29 years]: n=804; [60-74 years]: n=600). Using linear regression analysis adjusted for age, height and weight, we found significant associations between both rs2306029 and rs6485702 and BMD at all sites except the lumbar spine. The most significant association was found with whole body BMD (p=4.7×10(-5)). In addition, we found these two polymorphisms to be associated with different geometry parameters especially of the femoral shaft. Analysis of the two associated SNPs in the separate age groups demonstrated that most associations are only present in the youngest group of Danish men. In the group of elderly men, one Bonferroni corrected association between whole body BMD and rs6485702 was found to be significant. Subsequently, all polymorphisms were included in haplotype analyses using the PLINK software (v1.07). After adjusting for age, height and weight, two out of five common haplotypes (MAF≥0.01) were found to be of particular interest in the regulation of hip and whole body BMD (AGCG, AACA). Additional analysis suggested that these latter associations are driven by the association of rs6485702. We suggest, based on these results and the localisation of the variant in the third ß-propeller domain of LRP4, that the variant has possibly a functional effect. Hereby, we conclude that common variation in the LRP4 gene determines hip and whole body BMD and thus confirm previous results from different GWAs. In addition, our data proves an additional role for LRP4 in regulating hip structure. Finally, interaction analysis for LRP4 with SOST and LRP5 showed interaction with LRP5 for femoral shaft geometry.
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Densidad Ósea/fisiología , Haplotipos/genética , Cadera/anatomía & histología , Proteínas Relacionadas con Receptor de LDL/genética , Anciano , Densidad Ósea/genética , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Vía de Señalización Wnt/genética , Vía de Señalización Wnt/fisiologíaRESUMEN
INTRODUCTION: Sclerosteosis (OMIM 269500) is a rare autosomal recessive condition characterized by increased bone density associated with syndactyly. It is linked to a genetic defect in the SOST gene coding for sclerostin. So far, six different loss-of-function mutations in SOST have been reported in patients with sclerosteosis. Our objective was to sequence and identify mutation in the SOST and LRP5 genes which are known to be causal for craniotubular hyperostosis in a patient from India. PATIENT AND METHODS: A 22year old woman presented with typical features of sclerosteosis in form of progressive visual and hearing loss, syndactyly and radiographs revealing increased density of bone. Genomic sequencing of the SOST gene as well as exons 2, 3 and 4 of the LRP5 gene was performed. RESULTS: We identified a novel homozygous mutation in the (SOST) gene, characterized as one nucleotide insertion resulting in a frame shift mutation and loss of functional sclerostin. Her parents were also found to have a similar but heterozygous mutation in the (SOST) gene. CONCLUSION: A novel frame shift mutation in the (SOST) gene causing loss of functional sclerostin was identified in a patient with sclerosteosis and her parents.