Juvenile Paget’s disease (JPD) is a rare heritable osteopathy characterized biochemically by markedly increased serum alkaline phosphatase (ALP) activity emanating from generalized acceleration of skeletal turnover. Affected infants and children typically suffer bone pain and fractures and deformities, become deaf, and have macrocranium. Some who survive to young adult life develop blindness from retinopathy engendered by vascular microcalcification. Most cases of JPD are caused by osteoprotegerin (OPG) deficiency due to homozygous loss-of-function mutations within the TNFRSF11B gene that encodes OPG. We report a 3-year-old Iranian girl with JPD and craniosynostosis who had vitamin D deficiency in infancy. She presented with fractures during the first year-of-life followed by bone deformities, delayed development, failure-to-thrive, and pneumonias. At 1 year-of-age, biochemical studies of serum revealed marked hyperphosphatasemia together with low-normal calcium and low inorganic phosphate and 25-hydroxyvitamin D levels. Several family members in previous generations of this consanguineous kindred may also have had JPD and vitamin D deficiency. Mutation analysis showed homozygosity for a unique missense change (c.130T>C, p.Cys44Arg) in TNFRSF11B that would compromise the cysteine-rich domain of OPG that binds receptor activator of NF-κB ligand (RANKL). Both parents were heterozygous for this mutation. The patient’s serum OPG level was extremely low and RANKL level markedly elevated. She responded well to rapid oral vitamin D repletion followed by pamidronate treatment given intravenously. Our patient is the first Iranian reported with JPD. Her novel mutation in TNFRSF11B plus vitamin D deficiency in infancy was associated with severe JPD uniquely complicated by craniosynostosis. Pamidronate treatment with vitamin D sufficiency can be effective treatment for the skeletal disease caused by the OPG deficiency form of JPD.
alkaline phosphatase; bone remodeling; craniosynostosis; deafness; hyperphosphatasemia; hyperphosphatasia; osteoclast; osteolysis; osteoprotegerin; pamidronate; retinopathy; rickets; tumor necrosis factor; vascular calcification; vitamin D deficiency
We report a 32-year-old man and his 59-year-old mother with a unique and extensive variant of Camurati-Engelmann disease (CED) featuring histopathological changes of osteomalacia and alterations within TGFβ1 and TNFSF11 encoding TGFβ1 and RANKL, respectively. He suffered leg pain and weakness since childhood and reportedly grew until his late 20s, reaching 7 feet in height. He had deafness, perforated nasal septum, torus palatinus, disproportionately long limbs with knock-knees, low muscle mass, and pseudoclubbing. Radiographs revealed generalized skeletal abnormalities, including wide bones and cortical and trabecular bone thickening in keeping with CED, except that long bone ends were also affected. Lumbar spine and hip BMD Z-scores were + 7.7 and + 4.4, respectively. Biochemical markers of bone turnover were elevated. Hypocalciuria accompanied low serum 25-hydroxyvitamin D (25[OH]D) levels. Pituitary hypogonadism and low serum insulin-like growth factor (IGF)-1 were present. Karyotype was normal. Despite vitamin D repletion, iliac crest histology revealed severe osteomalacia. Exon 1 of TNFRSF11A (RANK), exons 2, 3, and 4 of LRP5, and all coding exons and adjacent mRNA splice junctions of TNFRSF11B (OPG), SQSTM1 (sequestosome 1), and TNSALP (tissue nonspecific alkaline phosphatase) were intact. His asymptomatic and less dysmorphic 5′11″ mother, also with low serum 25(OH)D, had milder clinical, radiological, biochemical, and histopathological findings. Both individuals were heterozygous for a novel 12-bp duplication (c.27_38dup, p.L10_L13dup) in exon 1 of TGFβ1, predicting four additional leucine residues in the latency-associated-peptide segment of TGFβ1, consistent with CED. The son was also homozygous for a single base transversion in TNFSF11, predicting a nonconservative amino acid change (c.107C > G, p.Pro36Arg) in the intracellular domain of RANKL that was heterozygous in his nonconsanguineous parents. This TNFSF11 variant was not found in the SNP Database, nor in published TNFSF11 association studies, but it occurred in four of the 134 TNFSF11 alleles (3.0%) we tested randomly among individuals without CED. Perhaps the unique phenotype of this CED family is conditioned by altered RANKL activity. © 2011 American Society for Bone and Mineral Research.
DEAFNESS; DIAPHYSEAL DYSPLASIA; OSTEOMALACIA; OSTEOSCLEROSIS; PERIOSTITIS; PREDNISONE; TORUS PALATINUS
Dysosteosclerosis (DSS) is the form of osteopetrosis distinguished by the presence of skin findings such as red-violet macular atrophy, platyspondyly and metaphyseal osteosclerosis with relative radiolucency of widened diaphyses. At the histopathological level, there is a paucity of osteoclasts when the disease presents. In two patients with DSS, we identified homozygous or compound heterozygous missense mutations in SLC29A3 by whole-exome sequencing. This gene encodes a nucleoside transporter, mutations in which cause histiocytosis–lymphadenopathy plus syndrome, a group of conditions with little or no skeletal involvement. This transporter is essential for lysosomal function in mice. We demonstrate the expression of Slc29a3 in mouse osteoclasts in vivo. In monocytes from patients with DSS, we observed reduced osteoclast differentiation and function (demineralization of calcium surface). Our report highlights the pleomorphic consequences of dysfunction of this nucleoside transporter, and importantly suggests a new mechanism for the control of osteoclast differentiation and function.
Dysosteosclerosis (DSS), an extremely rare dense bone disease, features short stature and fractures and sometimes optic atrophy, cranial nerve palsy, developmental delay, and failure of tooth eruption in infancy or early childhood consistent with osteopetrosis (OPT). Bone histology during childhood shows unresorbed primary spongiosa from deficient osteoclast action. Additionally, there is remarkable progressive flattening of all vertebrae and, by adolescence, paradoxical metaphyseal osteopenia with thin cortical bone. Reports of consanguinity indicate autosomal recessive inheritance, yet more affected males than females suggest X-linked recessive inheritance. We investigated a nonconsanguineous girl with DSS. Osteosclerosis was discovered at age 7 months. Our studies, spanning ages 11 to 44 months, showed weight at approximately 50th percentile, and length diminishing from approximately 30th percentile to –2.3 SD. Head circumference was +4 SD. The patient had frontal bossing, blue sclera, normal teeth, genu valgum, and unremarkable joints. Radiographs showed orbital and facial sclerosis, basilar thickening, bone-in-bone appearance of the pelvis, sclerotic long bone ends, and fractures of ribs and extremities. Progressive metaphyseal widening occurred as vertebrae changed from ovoid to flattened and became beaked anteriorly. A hemogram was normal. Consistent with OPT, serum parathyroid hormone (PTH) concentrations reflected dietary calcium levels. Serum bone alkaline phosphatase, osteocalcin, and TRACP-5b were subnormal. The iliac crest contained excessive primary spongiosa and no osteoclasts. No mutations were identified in the splice sites or exons for the genes encoding chloride channel 7, T-cell immune regulator 1, OPT-associated transmembrane protein 1, and monocyte colony-stimulating factor (M-CSF) and its receptor C-FMS, ANKH, OPG, RANK, and RANKL. Genomic copy-number microarray was unrevealing. Hence, DSS is a distinctive OPT of unknown etiology featuring osteoclast deficiency during early childhood. How osteopenia follows is an enigma of human skeletal pathobiology. © 2010 American Society for Bone and Mineral Research.
bone remodeling; erlenmeyer flask deformity; osteoclast; osteosclerosis; RANKL; skeletal dysplasia
Mucopolysaccharidosis type VII (MPS VII) is characterized by deficient β-glucuronidase (GUSB) activity, which leads to accumulation of chondroitin, heparan and dermatan sulfate glycosaminoglycans (GAGs), and multisystemic disease. MPS VII patients can develop kypho-scoliotic deformity and spinal cord compression due to disease of intervertebral discs, vertebral bodies, and associated tissues. We have previously demonstrated in MPS VII dogs that intervertebral discs degenerate, vertebral bodies have irregular surfaces, and vertebral body epiphyses have reduced calcification, but the pathophysiological mechanisms underlying these changes are unclear. We hypothesized that some of these manifestations could be due to upregulation of destructive proteases, possibly via the binding of GAGs to Toll-like receptor 4 (TLR4), as has been proposed for other tissues in MPS models. In this study, the annulus fibrosus of the intervertebral disc of 6 month-old MPS VII dogs had cathepsin B and K activities that were 117- and 2-fold normal, respectively, which were associated with elevations in mRNA levels for cathepsins as well as TLR4. The epiphyses of MPS VII dogs had a marked elevation in mRNA for the cartilage-associated gene collagen II, consistent with a developmental delay in the conversion of the cartilage to bone in this region. A spine from a human patient with MPS VII exhibited similar increased cartilage in the vertebral bodies adjacent to the end plates, disorganization of the intervertebral discs, and irregular vertebral end plate morphology. These data suggest that the pathogenesis of destructive changes in the spine in MPS VII may involve upregulation of cathepsins. Inhibition of destructive proteases, such as cathepsins, might reduce spine disease in patients with MPS VII or related disorders.
Lumbar spine; mucopolysaccharidosis VII; bone; intervertebral disc; cathepsins; inflammation; dog
Hypophosphatasia (HPP) is the inborn error of metabolism characterized by deficiency of alkaline phosphatase activity leading to rickets or osteomalacia and to dental defects. HPP occurs from loss-of-function mutations within the gene that encodes the tissue-nonspecific isozyme of alkaline phosphatase (TNAP). TNAP knockout (Alpl−/−, a.k.a. Akp2−/−) mice closely phenocopy infantile HPP, including the rickets, vitamin B6-responsive seizures, improper dentin mineralization, and lack of acellular cementum. Here, we report that lack of TNAP in Alpl−/− mice also causes severe enamel defects, which are preventable by enzyme replacement with mineral-targeted TNAP (ENB-0040). Immunohistochemistry was used to map the spatiotemporal expression of TNAP in the tissues of the developing enamel organ of healthy mouse molars and incisors. We found strong, stage-specific expression of TNAP in ameloblasts. In the Alpl−/− mice, histological, μCT, and scanning electron microscopy analysis showed reduced mineralization and disrupted organization of the rods and inter-rod structures in enamel of both the molars and incisors. All of these abnormalities were prevented in mice receiving from birth daily subcutaneous injections of mineral-targeting, human TNAP (sALP-FcD10, a.k.a. ENB-0040) at 8.2 mg/kg/day for up to 44 days. These data reveal an important role for TNAP in enamel mineralization, and demonstrate the efficacy of mineral-targeted TNAP to prevent enamel defects in HPP.
Hypophosphatasia is an inborn error of metabolism characterized by deficient activity of the tissue-nonspecific isoenzyme of alkaline phosphatase (TNSALP) and skeletal disease due to impaired mineralization of cartilage and bone matrix. We investigated two independently generated TNSALP gene knock-out mouse strains as potential models for hypophosphatasia. Homozygous mice (−/−) had < 1% of wild-type plasma TNSALP activity; heterozygotes had the predicted mean of ~50%. Phosphoethanolamine, inorganic pyrophosphate, and pyridoxal 5′-phosphate are putative natural substrates for TNSALP and all were increased endogenously in the knock-out mice. Skeletal disease first appeared radiographically at ~10 days of age and featured worsening rachitic changes, osteopenia, and fracture. Histologic studies revealed developmental arrest of chondrocyte differentiation in epiphyses and in growth plates with diminished or absent hypertrophic zones. Progressive osteoidosis from defective skeletal matrix mineralization was noted but not associated with features of secondary hyperparathyroidism. Plasma and urine calcium and phosphate levels were unremarkable. Our findings demonstrate that TNSALP knock-out mice are a good model for the infantile form of hypophosphatasia and provide compelling evidence for an important role for TNSALP in postnatal development and mineralization of the murine skeleton.
Adaptor protein-2 (AP2), a central component of clathrin-coated vesicles (CCVs), is pivotal in clathrin-mediated endocytosis which internalises plasma membrane constituents such as G protein-coupled receptors (GPCRs)1-3 . AP2, a heterotetramer of alpha, beta, mu and sigma subunits, links clathrin to vesicle membranes and binds to tyrosine-based and dileucine-based motifs of membrane-associated cargo proteins1,4. Here, we show that AP2 sigma subunit (AP2S1) missense mutations, which all involved the Arg15 residue (Arg15Cys, Arg15His and Arg15Leu) that forms key contacts with dileucine-based motifs of CCV cargo proteins4, result in familial hypocalciuric hypercalcemia type 3 (FHH3), an extracellular-calcium homeostasis disorder affecting parathyroids, kidneys and bone5-7 These AP2S1 mutations occurred in >20% of FHH patients without calcium-sensing GPCR (CaSR) mutations which cause FHH18-12. AP2S1 mutations decreased the sensitivity of CaSR-expressing cells to extracellular-calcium and reduced CaSR endocytosis, likely through a loss of interaction with a C-terminus CaSR dileucine-based motif whose disruption also decreased intracellular signalling. Thus, our results reveal a new role for AP2 in extracellular-calcium homeostasis.
Hypophosphatasia (HPP) is the metabolic bone disease caused by loss-of-function mutation within the gene that encodes the “tissue nonspecific” isoenzyme of alkaline phosphatase (TNSALP). Perinatal HPP is usually fatal due to respiratory insufficiency, and infantile HPP often has a similar outcome although no formal study into the natural history of these severe forms of HPP has been undertaken. We reviewed our 80-year (1927–2007) cohort of 15 Canadian patients with perinatal HPP. All had Mennonite heritage. Family linkage studies indicated that nine were homozygous for a TNSALP disease allele, likely Gly334Asp. Three patients had parents who were carriers for the Gly334Asp allele by mutation analysis. One patient was confirmed by mutation analysis to be homozygous for the TNSALP Gly334Asp mutation. One patient who had only one Mennonite parent was a genetic compound for the Gly334Asp mutation and the Val382Ile mutation. This patient’s sibling was also affected. All 15 patients had profound skeletal hypomineralization, severe rickets, and respiratory insufficiency. All died by 9 months of age, usually soon after birth, from pulmonary failure.
Hypophosphatasia (HPP) features rickets or osteomalacia from tissue-nonspecific alkaline phosphatase (TNSALP) deficiency due to deactivating mutations within the ALPL gene. Enzyme replacement therapy with a bone-targeted, recombinant TNSALP (sALP-FcD10, renamed ENB-0040) prevents manifestations of HPP when initiated at birth in TNSALP knockout (Akp2−/−) mice. Here, we evaluated the dose-response relationship of ENB-0040 to various phenotypic traits of Akp2−/− mice receiving daily subcutaneous (SC) injections of ENB-0040 from birth at 0.5, 2.0, or 8.2 mg/kg for 43 days. Radiographs, μCT, and histomorphometric analyses documented better bone mineralization with increasing doses of ENB-0040. We found a clear, positive correlation between ENB-0040 dose and prevention of mineralization defects of the feet, rib cage, lower limbs, and jaw bones. According to a dose-response model, the ED80 (the dose prevents the bone defects in 80% of mice) was 3.2, 2.8 and 2.9 mg/kg/day for these sites, respectively. Long bones seemed to respond to lower daily doses of ENB-0040. There was also a positive relationship between ENB-0040 dose and survival. Median survival, body weight, and bone length all improved with increasing doses of ENB-0040. Urinary PPi concentrations remained elevated in all treatment groups, indicating that while this parameter is a good biochemical marker for diagnosing HPP, it may not be a good follow up marker for evaluating response to treatment when administering bone-targeted TNSALP. These dose-response relationships strongly support the pharmacological efficacy of ENB-0040 for HPP, and provide the experimental basis for the therapeutic range of ENB-0040 chosen for clinical trials.
alkaline phosphatase; calcification; ENB-0040; rickets; osteomalacia
Osteogenesis imperfecta (OI) is most often caused by mutations in the type I procollagen genes (COL1A1/COL1A2). We identified two children with substitutions in the type I procollagen C-propeptide cleavage site, which disrupt a unique processing step in collagen maturation and define a novel phenotype within OI. The patients have mild OI caused by mutations in COL1A1 (Patient 1: p.Asp1219Asn) or COL1A2 (Patient 2: p.Ala1119Thr), respectively. Patient 1 L1-L4 DXA z-score was +3.9 and pQCT vBMD was +3.1; Patient 2 had L1-L4 DXA z-score of 0.0 and pQCT vBMD of −1.8. Patient BMD contrasts with radiographic osteopenia and histomorphometry without osteosclerosis. Mutant procollagen processing is impaired in pericellular and in vitro assays. Patient dermal collagen fibrils have irregular borders. Incorporation of pC-collagen into matrix leads to increased bone mineralization. FT-IR imaging confirms elevated mineral/matrix ratios in both patients, along with increased collagen maturation in trabecular bone, compared to normal or OI controls. Bone mineralization density distribution revealed a marked shift toward increased mineralization density for both patients. Patient 1 has areas of higher and lower bone mineralization than controls; Patient 2’s bone matrix has a mineral content exceeding even classical OI bone. These patients define a new phenotype of high BMD OI and demonstrate that procollagen C-propeptide cleavage is crucial to normal bone mineralization.
Osteogenesis imperfecta; C-propeptide; collagen; C-proteinase; mineralization; high bone mass
Mucopolysaccharidosis I (MPS I) and MPS VII are due to loss-of-function mutations within the genes that encode the lysosomal enzymes α-L-iduronidase and β-glucuronidase, respectively, and result in accumulation of glycosaminoglycans and multisystemic disease. Both disorders are associated with elastin fragmentation and dilatation of the aorta. Here, the pathogenesis and effect of gene therapy on aortic disease in canine models of MPS was evaluated. We found that cathepsin S is upregulated at the mRNA and enzyme activity level, while matrix metalloproteinase 12 (MMP-12) is upregulated at the mRNA level, in aortas from untreated MPS I and MPS VII dogs. Both of these proteases can degrade elastin. In addition, mRNA levels for the interleukin 6-like cytokine oncostatin M were increased in MPS I and MPS VII dog aortas, while mRNA for tumor necrosis factor α and toll-like receptor 4 were increased in MPS VII dog aortas. These cytokines could contribute to upregulation of the elastases. Neonatal intravenous injection of a retroviral vector expressing β-glucuronidase to MPS VII dogs reduced RNA levels of cathepsin S and MMP-12 and aortic dilatation was delayed, albeit dilatation developed at late times after gene therapy. A post-mortem aorta from a patient with MPS VII also exhibited elastin fragmentation. We conclude that aortic dilatation in MPS I and MPS VII dogs is likely due to degradation of elastin by cathepsin S and/or MMP-12. Inhibitors of these enzymes or these cytokine-induced signal transduction pathways might reduce aortic disease in patients with MPS.
Mucopolysaccharidosis; canine; lysosomal storage disease; cathepsin S; elastin; aorta; gene therapy
Familial benign hypocalciuric hypercalcaemia (FBHH) is a genetically heterogeneous disorder that consists of three designated types, FBHH1, FBHH2 and FBHH3, whose chromosomal locations are 3q21.1, 19p and 19q13, respectively. FBHH1 is caused by mutations of a calcium-sensing receptor (CaSR), but the abnormalities underlying FBHH2 and FBHH3 are unknown. FBHH3, also referred to as the Oklahoma variant (FBHHOk), has been mapped to a 12cM interval, flanked by D19S908 and D19S866. To refine the location of FBHH3, we pursued linkage studies using 24 polymorphic loci. Our results establish a linkage between FBHH3 and 17 of these loci, and indicate that FBHH3 is located in a 4.1 Mb region flanked centromerically by D19S112 and telomerically by rs245111, which in the syntenic region on mouse chromosome 7 contains four Casr-related sequences (Gprc2a-rss). However, human homologues of these Gprc2a-rss were not found and a comparative analysis of the 22.0 Mb human and 39.3 Mb mouse syntenic regions showed evolutionary conservation of two segments that were inverted with loss from the human genome of 11.6 Mb that contained the four Gprc2a-rss. Thus, FBHH3 cannot be attributed to Gprc2a-rss abnormalities. DNA sequence analysis of 12 other genes from the interval that were expressed in the parathyroids and/or kidneys did not detect any abnormalities, thereby indicating that these genes are unlikely to be the cause of FBHH3. The results of this study have refined the map location of FBHH3, which will facilitate the identification of another CaSR or a mediator of calcium homeostasis.
calcium; linkage; rearrangement
Familial benign hypocalciuric hypercalcaemia (FBHH) is a genetically heterogenous disorder that consists of three types designated, FBHH1, FBHH2 and FBHH3 whose chromosomal locations are 3q21.1, 19p and 19q13, respectively. FBHH1 is caused by mutations of a calcium sensing receptor (CaSR), but the abnormalities underlying FBHH2 and FBHH3 are unknown. FBHH3, also referred to as the Oklahoma variant (FBHHOk), has been mapped to a 12cM interval, flanked by D19S908 and D19S866. To refine the location of FBHH3, we pursued linkage studies using 24 polymorphic loci. Our results establish linkage between FBHH3 and 17 of these loci, and indicate that FBHH3 is located in a 4.1Mb region flanked centromerically by D19S112 and telomerically by rs245111, which in the syntenic region on mouse chromosome 7 contains 4 Casr related sequences (Gprc2a-rss). However, human homologues of these Gprc2a-rss were not found and a comparative analysis of the 22.0Mb human and 39.3Mb mouse syntenic regions showed evolutionary conservation of 2 segments that were inverted with loss from the human genome of 11.6Mb that contained the 4 Gprc2a-rss. Thus, FBHH3 cannot be due to Gprc2a-rss abnormalities. DNA sequence analysis of 12 other genes from the interval that were expressed in the parathyroids and/or kidneys did not detect any abnormalities, thereby indicating that these genes are unlikely to be the cause of FBHH3. The results of this study have refined the map location of FBHH3, which will facilitate the identification of another CaSR or a mediator of calcium homeostasis.
calcium; linkage; rearrangement
Hypophosphatasia (HPP) is the inborn error of metabolism that features rickets or osteomalacia caused by loss-of-function mutation(s) within the gene that encodes the tissue-nonspecific isozyme of alkaline phosphatase (TNALP). Consequently, natural substrates for this ectoenzyme accumulate extracellulary including inorganic pyrophosphate (PPi), an inhibitor of mineralization, and pyridoxal 5′-phosphate (PLP), a co-factor form of vitamin B6. Babies with the infantile form of HPP often die with severe rickets and sometimes hypercalcemia and vitamin B6-dependent seizures. There is no established medical treatment.
Materials and Methods
Human TNALP was bioengineered with the C terminus extended by the Fc region of human IgG for one-step purification and a deca-aspartate sequence (D10) for targeting to mineralizing tissue (sALP-FcD10). TNALP-null mice (Akp2−/−), an excellent model for infantile HPP, were treated from birth using sALP-FcD10. Short-term and long-term efficacy studies consisted of once daily subcutaneous injections of 1, 2, or 8.2 mg/kg sALP-FcD10 for 15, 19, and 15 or 52 days, respectively. We assessed survival and growth rates, circulating levels of sALP-FcD10 activity, calcium, PPi, and pyridoxal, as well as skeletal and dental manifestations using radiography, μCT, and histomorphometry.
Akp2−/− mice receiving high-dose sALP-FcD10 grew normally and appeared well without skeletal or dental disease or epilepsy. Plasma calcium, PPi, and pyridoxal concentrations remained in their normal ranges. We found no evidence of significant skeletal or dental disease.
Enzyme replacement using a bone-targeted, recombinant form of human TNALP prevents infantile HPP in Akp2−/− mice.
alkaline phosphatase; calcification; epilepsy; osteomalacia; rickets
Inclusion body myopathy with Paget disease of the bone (PDB) and/or frontotemporal dementia (IBMPFD, OMIM 167320), is a progressive autosomal dominant disorder caused by mutations in the Valousin-containing protein (VCP, p97 or CDC48) gene. IBMPFD can be difficult to diagnose. We assembled data on a large set of families to illustrate the number and type of misdiagnoses that occurred. Clinical analysis of 49 affected individuals in nine families indicated that 42 (87%) of individuals had muscle disease. The majority were erroneously diagnosed with limb girdle muscular dystrophy (LGMD), facioscapular muscular dystrophy, peroneal muscular dystrophy, late adult onset distal myopathy, spinal muscular atrophy, scapuloperoneal muscular dystrophy, or amyotrophic lateral sclerosis (ALS) among others. Muscle biopsies showed rimmed vacuoles characteristic of an inclusion body myopathy in 7 of 18 patients (39%), however, inclusion body myopathy was correctly diagnosed among individuals in only families 5 and 15. Frontotemporal dementia (FTD) was diagnosed in 13 individuals (27%) at a mean age of 57 years (range 48.9–60.2 years); however, several individuals had been diagnosed with Alzheimer disease. Histopathological examination of brains of three affected individuals revealed a pattern of ubiquitin positive neuronal intranuclear inclusions and dystrophic neurites. These families expand the clinical phenotype in IBMPFD, a complex disorder caused by mutations in VCP. The presence of PDB in 28 (57%) individuals suggests that measuring serum alkaline phosphatase (ALP) activity may be a useful screen for IBMPFD in patients with myopathy.
autosomal dominant; hereditary inclusion body myopathy; limb-girdle muscular dystrophy; Paget disease of bone; frontotemporal dementia; chromosome 9p13.3-12; VCP (valosin-containing protein)
MMPs, which degrade components of the ECM, have roles in embryonic development, tissue repair, cancer, arthritis, and cardiovascular disease. We show that a missense mutation of MMP13 causes the Missouri type of human spondyloepimetaphyseal dysplasia (SEMDMO), an autosomal dominant disorder characterized by defective growth and modeling of vertebrae and long bones. Genome-wide linkage analysis mapped SEMDMO to a 17-cM region on chromosome 11q14.3–23.2 that contains a cluster of 9 MMP genes. Among these, MMP13 represented the best candidate for SEMDMO, since it preferentially degrades collagen type II, abnormalities of which cause skeletal dysplasias that include Strudwick type SEMD. DNA sequence analysis revealed a missense mutation, F56S, that substituted an evolutionarily conserved phenylalanine residue for a serine in the proregion domain of MMP13. We predicted, by modeling MMP13 structure, that this F56S mutation would result in a hydrophobic cavity with misfolding, autoactivation, and degradation of mutant protein intracellularly. Expression of wild-type and mutant MMP13s in human embryonic kidney cells confirmed abnormal intracellular autoactivation and autodegradation of F56S MMP13 such that only enzymatically inactive, small fragments were secreted. Thus, the F56S mutation results in deficiency of MMP13, which leads to the human skeletal developmental anomaly of SEMDMO.
X-linked recessive hypoparathyroidism, due to parathyroid agenesis, has been mapped to a 906-kb region on Xq27 that contains 3 genes (ATP11C, U7snRNA, and SOX3), and analyses have not revealed mutations. We therefore characterized this region by combined analysis of single nucleotide polymorphisms and sequence-tagged sites. This identified a 23- to 25-kb deletion, which did not contain genes. However, DNA fiber–FISH and pulsed-field gel electrophoresis revealed an approximately 340-kb insertion that replaced the deleted fragment. Use of flow-sorted X chromosome–specific libraries and DNA sequence analyses revealed that the telomeric and centromeric breakpoints on X were, respectively, approximately 67 kb downstream of SOX3 and within a repetitive sequence. Use of a monochromosomal somatic cell hybrid panel and metaphase-FISH mapping demonstrated that the insertion originated from 2p25 and contained a segment of the SNTG2 gene that lacked an open reading frame. However, the deletion-insertion [del(X)(q27.1) inv ins (X;2)(q27.1;p25.3)], which represents a novel abnormality causing hypoparathyroidism, could result in a position effect on SOX3 expression. Indeed, SOX3 expression was demonstrated, by in situ hybridization, in the developing parathyroid tissue of mouse embryos between 10.5 and 15.5 days post coitum. Thus, our results indicate a likely new role for SOX3 in the embryonic development of the parathyroid glands.