ABSTRACT
Thanatophoric dysplasia type 1 (TD1) is a lethal form of osteochondral dysplasia due to mutation of FGFR3 gene. In addition to severe shortening of the limbs there is temporo-occipital lobe dysplasia along with a range of other CNS anomalies. In this report we describe the radiological and anatomical features at autopsy in neonate with TD1 along with the CNS anomalies. We have also summarized the key distinguishing features of TD1 from other common types of osteochondral dysplasia. An accurate diagnosis is important for genetic counseling and impact on future pregnancies.
Subject(s)
Limb Deformities, Congenital/pathology , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/pathology , Autopsy , Extremities/pathology , Humans , Infant, Newborn , Male , Receptor, Fibroblast Growth Factor, Type 3/genetics , Temporal Lobe/pathology , Thanatophoric Dysplasia/diagnosis , Thanatophoric Dysplasia/mortalityABSTRACT
OBJECTIVE: The aim of this study is to explore the utility of rapid medical trio exome sequencing (ES) for prenatal diagnosis using the skeletal dysplasia as an exemplar. METHOD: Pregnant women who were referred for genetic testing because of ultrasound detection of fetal abnormalities suggestive of a skeletal dysplasia were identified prospectively. Fetal samples (amniocytes or cord blood), along with parental blood, were send for rapid copy number variations testing and medical trio ES in parallel. RESULTS: Definitive molecular diagnosis was made in 24/27 (88.9%) cases. Chromosomal abnormality (partial trisomy 18) was detected in one case. Sequencing results had explained the prenatal phenotype enabling definitive diagnoses to be made in 23 cases. There were 16 de novo dominant pathogenic variants, four dominant pathogenic variants inherited maternally or paternally, two recessive conditions with pathogenic variants inherited from unaffected parents, and one X-linked condition. The turnaround time from receipt of samples in the laboratory to reporting sequencing results was within 2 weeks. CONCLUSION: Medical trio ES can yield very timely and high diagnostic rates in fetuses presenting with suspected skeletal dysplasia. These definite diagnoses aided parental counseling and decision making in most of cases.
Subject(s)
Exome Sequencing/methods , Osteochondrodysplasias/diagnosis , Parents , Prenatal Care/methods , Abnormalities, Multiple/diagnosis , Abnormalities, Multiple/genetics , Achondroplasia/diagnosis , Achondroplasia/genetics , Adult , Brain Diseases/diagnosis , Brain Diseases/genetics , Campomelic Dysplasia/diagnosis , Campomelic Dysplasia/genetics , Carbohydrate Metabolism, Inborn Errors/diagnosis , Carbohydrate Metabolism, Inborn Errors/genetics , Congenital Disorders of Glycosylation/diagnosis , Congenital Disorders of Glycosylation/genetics , Female , Fetal Growth Retardation/diagnosis , Fetal Growth Retardation/genetics , Genetic Counseling/methods , Genetic Testing/methods , Humans , Ichthyosis/diagnosis , Ichthyosis/genetics , Limb Deformities, Congenital/diagnosis , Limb Deformities, Congenital/genetics , Male , Microcephaly/diagnosis , Microcephaly/genetics , Osteochondrodysplasias/genetics , Osteogenesis Imperfecta/diagnosis , Osteogenesis Imperfecta/genetics , Pathology, Molecular , Phosphoglycerate Dehydrogenase/deficiency , Phosphoglycerate Dehydrogenase/genetics , Pregnancy , Prenatal Diagnosis , Psychomotor Disorders/diagnosis , Psychomotor Disorders/genetics , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Seizures/diagnosis , Seizures/genetics , Thanatophoric Dysplasia/diagnosis , Thanatophoric Dysplasia/genetics , Time Factors , Trisomy 18 Syndrome/diagnosis , Ultrasonography, Prenatal , Young AdultABSTRACT
We present a case report that entails prenatal ultrasonography, postnatal characteristics, and molecular genetic analysis of a newborn who presented with thanatophoric dysplasia type I (TDI) with a mutation in the fibroblast growth factor receptor 3 gene (FGFR3). A malformed newborn with tachypnea, delivered by caesarean at the gestational age of 39â¯weeks, was the first child of nonconsanguineous parents by a spontaneous pregnancy. Features in prenatal ultrasonography and postnatal radiography were consistent with the diagnosis of TDI, presenting with short body length (38â¯cm, <3rd percentile), redundant skin folds, a narrow thorax with a bust of 29.5â¯cm (3-5th percentile), and macrocephaly with a head circumference of 36â¯cm (>97th percentile). The proposita had postnatal dyspnea and unfortunately died of respiratory failure at the age of 13â¯days. Molecular genetic analysis revealed a mutation of c.2419â¯Tâ¯>â¯C (p. Ter807Arg) (X807R) in FGFR3. Live-born infants with TDI are exceedingly rare, and we hereby report a newborn with a c.2419â¯Tâ¯>â¯C mutation in FGFR3, emphasizing phenotype with clinical characteristics and ultrasonographic and X-ray findings, to raise awareness about the heterogeneous patterns of TD.
Subject(s)
Mutation , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/genetics , Thanatophoric Dysplasia/pathology , Adult , Amino Acid Sequence , Base Sequence , Female , Gestational Age , Humans , Infant, Newborn , Phenotype , Pregnancy , Prognosis , Receptor, Fibroblast Growth Factor, Type 3/geneticsABSTRACT
INTRODUCTION: Skeletal dysplasias are a heterogeneous group of disorders comprising of more than 300 entities, many of which manifest in the prenatal period, emphasizing the importance of accurate prenatal diagnosis. Detection of a lethal skeletal dysplasia via prenatal ultrasound is often straightforward. However, establishing the specific diagnosis and detailed evaluation of intracranial anomalies are often challenging. Fetal magnetic resonance imaging (MRI) is superior to ultrasound in the detection of abnormal sulcation pattern, corpus callosal agenesis, and posterior fossa anomalies. Hence, it has the potential of delineating neuroimaging features that may not be fully elucidated by ultrasound. The objective of this article is to describe an unusual case of thanatophoric dysplasia (TD) with dysplastic tectal plate and resultant aqueductal stenosis diagnosed on fetal MRI. To the best of our knowledge, this has never been reported before in the literature. A comprehensive review of literature pertaining to TD-associated CNS abnormalities will also be included. CONCLUSIONS: Our reported case adds to the current limited knowledge of this rare entity and emphasizes the crucial role of fetal MRI in expanding the neuroimaging phenotypes of TD.
Subject(s)
Fetal Diseases/diagnostic imaging , Hydrocephalus/diagnostic imaging , Magnetic Resonance Imaging/methods , Prenatal Diagnosis/methods , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/diagnostic imaging , Female , Humans , Neuroimaging/methods , PregnancyABSTRACT
BACKGROUND: Thanatophoric dysplasia (TD) results from sporadic de novo mutations in the FGFR3 gene. Upon confirming intrauterine diagnosis of this perinatal disease, pregnancy termination is recommended. There is limited information on the natural history of longer-term survivors with type 1 TD. CASE REPORT: A full-term neonate was confirmed via postnatal genetic testing to have type 1 TD. At 28 days, chylous ascites developed. Medium-chain triglyceride use improved the ascites. Cerebral ventriculomegaly worsened throughout life. Death due to respiratory failure occurred at age 5 months. CONCLUSION: The chylous ascites in this child with type 1 TD and survival past the neonatal stage suggests that type 1 TD may be accompanied by abnormalities of the lymphatic channels. Moreover, ventriculomegaly can be progressive.
Subject(s)
Chylous Ascites/genetics , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/complications , Humans , Infant , Infant, Newborn , Male , Mutation , Receptor, Fibroblast Growth Factor, Type 3/genetics , Thanatophoric Dysplasia/geneticsABSTRACT
Leptomeningeal glioneuronal heterotopia (LGH) is a focal malformation of the cerebral cortex and frequently found in patients with thanatophoric dysplasia (TD). The pathophysiological mechanisms underlying LGH formation are still largely unclear because of difficulties in obtaining brain samples from human TD patients. Recently, we established a new animal model for analysing cortical malformations of human TD by utilizing our genetic manipulation technique for gyrencephalic carnivore ferrets. Here we investigated the pathophysiological mechanisms underlying the formation of LGH using our TD ferrets. We found that LGH was formed during corticogenesis in TD ferrets. Interestingly, we rarely found Ki-67-positive and phospho-histone H3-positive cells in LGH, suggesting that LGH formation does not involve cell proliferation. We uncovered that vimentin-positive radial glial fibers and doublecortin-positive migrating neurons were accumulated in LGH. This result may indicate that preferential cell migration into LGH underlies LGH formation. Our findings provide novel mechanistic insights into the pathogenesis of LGH in TD.
Subject(s)
Meningeal Neoplasms/physiopathology , Thanatophoric Dysplasia/physiopathology , Animals , Cell Movement/physiology , Cerebral Cortex/physiopathology , Disease Models, Animal , Ependyma/metabolism , Ependyma/physiopathology , Ependymoglial Cells/metabolism , Ferrets , Neuroglia/metabolism , Neurons/metabolism , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Thanatophoric Dysplasia/metabolism , Vimentin/metabolismABSTRACT
Chemokines are small secreted signaling proteins produced by a broad range of cells, including immune cells. Several studies have recently suggested potential roles of chemokines and their receptors in the pathophysiology of autism spectrum disorders (ASDs). SAM3 is a novel brain-specific chemokine-like molecule with an unknown physiological function. We explored the relevance of chemokines in the development of ASD in mice, with a focus on SAM3. We generated Sam3 gene knockout (KO) mice and characterized their behavioral phenotypes, with a focus on those relevant to ASD. Sam3-deficient mice displayed all three core phenotypes of ASD: impaired responses to social novelty, defects in social communication, and increased repetitive behavior. In addition, they showed increased anxiety. Interestingly, gender differences were identified for several behaviors: only male Sam3 KO mice exhibited increased anxiety and increased repetitive behaviors. Sam3 KO mice did not exhibit changes in other behaviors, including locomotor activities, fear learning and memory, and object recognition memory. These findings indicate that a deficiency of SAM3, a novel brain-specific chemokine-like molecule, may lead to the pathogenesis of ASDs and suggest the possibility that SAM3, a soluble factor, could be a novel therapeutic target for ASD treatment.
Subject(s)
Autism Spectrum Disorder/etiology , Brain/metabolism , Brain/physiopathology , Phenotype , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Animals , Anxiety/genetics , Anxiety/psychology , Autism Spectrum Disorder/psychology , Behavior, Animal , Disease Models, Animal , Female , Genetic Loci , Genotype , Maze Learning , Memory , Mice , Mice, Knockout , Organ Specificity/genetics , Social BehaviorABSTRACT
Bone fracture healing is processed through multiple biological stages that partly recapitulates the skeletal development process. FGFR3 is a negative regulator of chondrogenesis during embryonic stage and plays an important role in both chondrogenesis and osteogenesis. We have investigated the role of FGFR3 in fracture healing using unstabilized fracture model and found that gain-of-function mutation of FGFR3 inhibits the initiation of chondrogenesis during cartilage callus formation. Here, we created closed, stabilized proximal tibia fractures with an intramedullary pin in Fgfr3-/-mice and their littermate wild-type mice. Fracture healing was evaluated by radiography, micro-CT, histology, and real-time polymerase chain reaction (RT-PCR) analysis. The fractured Fgfr3-/- mice had increased formation of cartilaginous callus, more fracture callus, and more rapid endochondral ossification in fracture sites with up-regulated expressions of chondrogenesis related gene. The fractures of Fgfr3-/- mice healed faster with accelerated fracture callus mineralization and up-regulated expression of osteoblastogenic genes. The healing of fractures in Fgfr3-/- mice was accelerated in the stage of formation of cartilage and endochondral ossification. Downregulation of FGFR3 activity can be considered as a potential bio-therapeutic strategy for fracture treatment.
Subject(s)
Fracture Healing/physiology , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Animals , Cells, Cultured , Chondrogenesis/genetics , Chondrogenesis/physiology , Fracture Healing/genetics , Male , Mice , Osteogenesis/genetics , Osteogenesis/physiology , Real-Time Polymerase Chain Reaction , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Reverse Transcriptase Polymerase Chain Reaction , Tibial Fractures/genetics , Tibial Fractures/metabolism , X-Ray MicrotomographyABSTRACT
The papillomavirus (PV) E2 protein is a DNA binding, protein interaction platform that recruits viral and host factors necessary for transcription and replication. We recently discovered phosphorylation of a tyrosine (Y102) in bovine PV (BPV) E2. To identify the responsible factor, we tested several candidate tyrosine kinases that are highly expressed in keratinocytes for binding to BPV-1 E2. Fibroblast growth factor receptor 3 (FGFR3) coimmunoprecipitated with the BPV-1 E2 protein, as did human papillomavirus 31 (HPV-31) E2, which also colocalized with FGFR3 within the nucleus. A constitutively active mutant form of FGFR3 decreased BPV-1 and HPV-31 transient replication although this result also occurred in a BPV-1 E2 mutant lacking a previously identified phosphorylation site of interest (Y102). Furthermore, FGFR3 depletion in cell lines that maintain HPV-31 episomes increased viral copy number. These results suggest that FGFR3 kinase activity may regulate the PV reproductive program through phosphorylation of the E2 protein although this is unlikely to occur through the Y102 residue of HPV E2.IMPORTANCE The papillomavirus (PV) is a double-stranded DNA tumor virus infecting cervix, mouth, and throat tissues. The viral protein E2 is responsible for the replication of the virus. Understanding the mechanisms of the replicative life cycle of the virus may bring to light direct targets and treatments against viral infection. We recently found that the fibroblast growth factor receptor 3 (FGFR3) interacts with and mediates PV E2 function through phosphorylation of the E2 protein. Our study suggests that the function of the E2 protein may be regulated through a direct FGFR3 target during the maintenance stage of the PV life cycle.
Subject(s)
DNA-Binding Proteins/metabolism , Human papillomavirus 31/physiology , Oncogene Proteins, Viral/metabolism , Phosphotransferases/metabolism , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Viral Proteins/metabolism , Virus Replication/physiology , Animals , Cattle , DNA Replication , Human papillomavirus 31/enzymology , Humans , Phosphorylation , Plasmids , Receptor, Fibroblast Growth Factor, Type 3/chemistry , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Tyrosine/chemistryABSTRACT
BACKGROUND: The dimerization efficiency of FGFR3 transmembrane domain plays a critical role in the formation of a normal skeleton through the negative regulation of bone development. Recently, gain-of-function mutations in the transmembrane domain of FGFR3 has been described associated with an aberrant negative regulation, leading to the development of achondroplasia-group disorders, including achondroplasia (ACH), hypochondroplasia (HCH) and thanatophoric dysplasia (TD). Here, we describe a non-consanguineous Pakistani family with achondroplasia to explain hereditary basis of the disease. METHODS: PCR-based linkage analysis using microsatellite markers was employed to localize the disease gene. Gene specific intronic primers were used to amplify the genomic DNA from all affected as well as phenotypically healthy individuals. Amplified PCR products were then subjected to Sanger sequencing and RFLP analysis to identify a potentially pathogenic mutation. The impact of identified mutation on FGFR3 protein's structure and stability was highlighted through different bioinformatics tools. RESULTS: Genetic screening of the family revealed a previously reported heterozygous c.1138 G > A (p.G380R) mutation in the coding exon 8 of FGFR3 gene. Identified genetic variation was confirmed in all affected individuals while healthy individuals and controls were found genotypically normal. The results were further validated by RFLP analysis as c.1138 G > A substitution generates a unique recognition site for SfcI endonuclease. Following SfcI digestion, the electrophoretic pattern of three bands/DNA fragments for each patient is indicative of heterozygous status of the disease allele. In silico studies of the mutant FGFR3 protein predicted to adversely affect the stability of FGFR3 protein. CONCLUSIONS: Mutation in the transmembrane domain may adversely affect the dimerization efficiency and overall stability of the FGFR3, leading to a constitutively active protein. As a result, an uncontrolled intracellular signaling or negative bone growth regulation leads to achondroplasia. Our findings support the fact that p.G380R is a common mutation among diverse population of the world and like other countries, can be used as a molecular diagnosis marker for achondroplasia in Pakistan.
Subject(s)
Achondroplasia/genetics , Genetic Predisposition to Disease , Mutation/genetics , Receptor, Fibroblast Growth Factor, Type 3/genetics , Achondroplasia/diagnosis , Amino Acid Sequence/genetics , Genetic Testing/methods , Humans , Pakistan , Pedigree , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Signal Transduction/genetics , Thanatophoric Dysplasia/geneticsABSTRACT
Blood and lymphatic vasculatures are intimately involved in tissue oxygenation and fluid homeostasis maintenance. Assembly of these vascular networks involves sprouting, migration and proliferation of endothelial cells. Recent studies have suggested that changes in cellular metabolism are important to these processes. Although much is known about vascular endothelial growth factor (VEGF)-dependent regulation of vascular development and metabolism, little is understood about the role of fibroblast growth factors (FGFs) in this context. Here we identify FGF receptor (FGFR) signalling as a critical regulator of vascular development. This is achieved by FGF-dependent control of c-MYC (MYC) expression that, in turn, regulates expression of the glycolytic enzyme hexokinase 2 (HK2). A decrease in HK2 levels in the absence of FGF signalling inputs results in decreased glycolysis, leading to impaired endothelial cell proliferation and migration. Pan-endothelial- and lymphatic-specific Hk2 knockouts phenocopy blood and/or lymphatic vascular defects seen in Fgfr1/Fgfr3 double mutant mice, while HK2 overexpression partly rescues the defects caused by suppression of FGF signalling. Thus, FGF-dependent regulation of endothelial glycolysis is a pivotal process in developmental and adult vascular growth and development.
Subject(s)
Endothelial Cells/cytology , Endothelial Cells/metabolism , Fibroblast Growth Factors/metabolism , Glycolysis , Neovascularization, Physiologic , Signal Transduction , Animals , Cell Movement , Cell Proliferation , Female , Hexokinase/metabolism , Lymphangiogenesis , Lymphatic Vessels/cytology , Lymphatic Vessels/metabolism , Mice , Mice, Inbred C57BL , Proto-Oncogene Proteins c-myc/metabolism , Receptor, Fibroblast Growth Factor, Type 1/deficiency , Receptor, Fibroblast Growth Factor, Type 1/genetics , Receptor, Fibroblast Growth Factor, Type 1/metabolism , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Receptor, Fibroblast Growth Factor, Type 3/metabolismABSTRACT
Thanatophoric dysplasia (TD), the most common lethal skeletal dysplasia, is a de novo genetic disease caused by a mutation in the fibroblast growth factor receptor 3 (FGFR3) gene. "Thanatophoric" means "dead bearing" in Greek. Because FGFR3 is the main modulator of bone maturation, typical features of TD include short extremities, curved femur, clover-leaf skull, small narrow chest, and platyspondyly. TD can be classified into two subgroups according to the morphologic findings, with prominent curved femur suggesting type I TD (TD 1) and with marked clover-leaf skull and relatively straight long bones, favoring type II TD (TD 2). However, considering the genetic profiles, TD 1 and TD 2 could be confidently delineated. Here, we report five genotype-phenotype correlated autopsy cases of TD. Five cases had stigmata of TD on antenatal ultrasonography. Terminations were done at gestational age 16 to 28weeks, after family consultation. In autopsy, all fetuses showed short limbs and clover-leaf skull. The microscopic examination of the bones showed disorganized growth plate, consistent with TD. However, some differences existed in gross and microscopic findings between cases. In genetic analyses, three cases revealed missense mutation of Y373C, while the remaining two cases had missense mutation of S371C and S249C each. They were hot spot mutations of TD 1. A correlation between genotype and phenotype was not apparent due to the limited number of the cases. Therefore, a molecular work up to identify the mutation of FGFR3 is indispensable for TD diagnosis in the era of precision medicine for genetic consultation and future targeted therapy.
Subject(s)
Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Skull/abnormalities , Thanatophoric Dysplasia/genetics , Adult , Autopsy , Exons , Female , Fetus/abnormalities , Genotype , Gestational Age , Humans , Mutation, Missense , Phenotype , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Thanatophoric Dysplasia/diagnosisABSTRACT
OBJECTIVE: We present perinatal imaging findings and molecular genetic analysis of thanatophoric dysplasia type I (TD1) in a fetus. CASE REPORT: A 28-year-old woman was referred for genetic counseling at 22 weeks of gestation because of abnormal prenatal ultrasound findings. Level II ultrasound examination revealed a narrow chest, shortened and curved long limbs, protrusion of the abdomen, and macrocephaly. A tentative diagnosis of TD1 was made. After genetic counseling, the pregnancy was terminated and a malformed fetus was delivered. Postnatal radiography findings were consistent with the diagnosis of TD1, with additional findings of short ribs, platyspondyly, and horizontal acetabular roofs. Molecular genetic analysis using umbilical cord tissue revealed a heterozygous mutation of c.2419T>G (p.Ter807Gly) (X807G) in the fibroblast growth factor receptor 3 gene (FGFR3). CONCLUSION: A second-trimester fetus with a heterozygous c.2419T>G mutation in FGFR3 may present characteristic ultrasound and X-ray findings of TD1.
Subject(s)
Mutation , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Thanatophoric Dysplasia/genetics , Adult , Female , Humans , Pregnancy , Pregnancy Trimester, Second , Sequence Analysis, DNA , Thanatophoric Dysplasia/diagnostic imaging , Ultrasonography, PrenatalABSTRACT
UNLABELLED: PTH stimulates bone formation in Fgfr3 knockout mice through promotion of proliferation and differentiation in osteoblasts. INTRODUCTION: Previous studies showed that endogenous fibroblast growth factor 2 (FGF-2) is required for parathyroid hormone (PTH)-stimulated bone anabolic effects, however, the exact mechanisms by which PTH stimulate bone formation and the function of FGF receptors in mediating these actions are not fully defined. FGF receptor 3 (FGFR3) has been characterized as an important regulator of bone metabolism and is confirmed to cross-talk with PTH/PTHrP signal in cartilage and bone development. METHODS: Fgfr3 knockout and wild-type mice at 2-month-old and 4-month-old were intraperitoneally injected with PTH intermittently for 4 weeks and then the skeletal responses to PTH were assessed by dual energy X-ray absorptiometry (DEXA), micro-computed tomography (µCT) and bone histomorphometry. RESULTS: Intermittent PTH treatment improved bone mineral density (BMD) and femoral mechanical properties in both Fgfr3 (-/-) and wild-type mice. Histomorphometric analysis showed that bone formation and bone resorption were increased in both genotypes following PTH treatment. PTH treatment increased trabecular bone volume (BV/TV) in WT and Fgfr3-deficient mice. The anabolic response in Fgfr3-deficient and wild-type bone is characterized by an increase of both bone formation and resorption-related genes following PTH treatment. In addition, we found that Fgfr3 null osteoblasts (compared to wild-type controls) maintained normal abilities to response to PTH-stimulated increase of proliferation, differentiation, expression of osteoblastic marker genes (Cbfa1, Osteopontin and Osteocalcin), and phosphorylation of Erk1/2. CONCLUSIONS: Bone anabolic effects of PTH were not impaired by the absence of FGFR3, suggesting that the FGFR3 signaling may not be required for osteoanabolic effects of PTH activities.
Subject(s)
Parathyroid Hormone/pharmacology , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Animals , Blotting, Western , Bone Density/drug effects , Bone Remodeling/drug effects , Cells, Cultured , Male , Mice , Mice, Knockout , Osteoblasts/drug effects , Osteoblasts/metabolism , Real-Time Polymerase Chain Reaction , Receptor, Fibroblast Growth Factor, Type 3/geneticsABSTRACT
Protein-losing enteropathy is a primary or secondary manifestation of a group of conditions, and etiologies which are broadly divisible into those with mucosal injury on the basis of inflammatory and ulcerative conditions, mucosal injury without erosions or ulcerations, and lymphatic abnormalities. We describe the first case of protein-losing enteropathy in a pediatric patient, with severe skeletal dysplasia consistent with thanatophoric dysplasia type I and DNA analysis that revealed a c.1949A>T (p.Lys650Met) in exon 15 of the FGFR3 gene. She presented with protein-losing enteropathy in her 6th month. Post-mortem examination revealed lymphangiectasia in the small intestine. To our knowledge, this is the first report of intestinal lymphangiectasia as a complication of skeletal dysplasia resulting in severe protein-losing enteropathy. © 2016 Wiley Periodicals, Inc.
Subject(s)
Lymphangiectasis, Intestinal/diagnosis , Lymphangiectasis, Intestinal/genetics , Mutation , Protein-Losing Enteropathies/diagnosis , Protein-Losing Enteropathies/genetics , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Thanatophoric Dysplasia/diagnosis , Thanatophoric Dysplasia/genetics , Alleles , Amino Acid Substitution , Autopsy , Brain/pathology , Codon , Fatal Outcome , Female , Genetic Association Studies , Humans , Infant, Newborn , Intestinal Mucosa/pathology , Lung/pathology , Magnetic Resonance Imaging , Phenotype , RadiographyABSTRACT
Thanatophoric dysplasia is a type of short-limbed neonatal dwarfism that is usually lethal in the perinatal period. It is characterized by short limbs, a narrow, bell-shaped thorax, macrocephaly with a prominent forehead, and flattened vertebral bodies. These malformations result from autosomal dominant mutations in the fibroblast growth factor receptor 3 (FGFR3) gene. In this report, we describe a novel FGFR3 insertion mutation in a fetus with shortened limbs, curved femurs, and a narrow thorax. The diagnosis of thanatophoric dysplasia type 1 was suspected clinically, and FGFR3 sequencing showed a c.742_743insTGT variant, which predicts p.R248delinsLC. In vivo studies in zebrafish demonstrated that this mutation resulted in the overexpression of zebrafish Fgfr3, leading to the over-activation of downstream signaling and dorsalized embryos. To date, no insertions or deletions in FGFR3 have been reported to cause thanatophoric dysplasia types 1 or 2; therefore, this represents the first report to describe such a mutation. © 2016 Wiley Periodicals, Inc.
Subject(s)
Genetic Association Studies , Mutagenesis, Insertional , Phenotype , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Receptor, Fibroblast Growth Factor, Type 3/genetics , Thanatophoric Dysplasia/diagnosis , Thanatophoric Dysplasia/genetics , Abortion, Induced , Alleles , Animals , Autopsy , Exons , Female , Fetus , Gene Expression , Genotype , Humans , Mutation , Pregnancy , ZebrafishABSTRACT
Fibroblast growth factor receptor 3 (FGFR3) participates in bone remodeling. Both Fgfr3 global knockout and activated mice showed decreased bone mass with increased osteoclast formation or bone resorption activity. To clarify the direct effect of FGFR3 on osteoclasts, we specifically deleted Fgfr3 in osteoclast lineage cells. Adult mice with Fgfr3 deficiency in osteoclast lineage cells (mutant [MUT]) showed increased bone mass. In a drilled-hole defect model, the bone remodeling of the holed area in cortical bone was also impaired with delayed resorption of residual woven bone in MUT mice. In vitro assay demonstrated that there was no significant difference between the number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts derived from wild-type and Fgfr3-deficient bone marrow monocytes, suggesting that FGFR3 had no remarkable effect on osteoclast formation. The bone resorption activity of Fgfr3-deficient osteoclasts was markedly decreased accompanying with downregulated expressions of Trap, Ctsk, and Mmp 9. The upregulated activity of osteoclastic bone resorption by FGF2 in vitro was also impaired in Fgfr3-deficient osteoclasts, indicating that FGFR3 may participate in the regulation of bone resorption activity of osteoclasts by FGF2. Reduced adhesion but not migration in osteoclasts with Fgfr3 deficiency may be responsible for the impaired bone resorption activity. Our study for the first time genetically shows the direct positive regulation of FGFR3 on osteoclastic bone resorption. © 2016 American Society for Bone and Mineral Research.
Subject(s)
Bone Resorption/pathology , Bone and Bones/pathology , Cell Lineage , Gene Deletion , Osteoclasts/metabolism , Osteoclasts/pathology , Receptor, Fibroblast Growth Factor, Type 3/genetics , Animals , Bone Remodeling , Bony Callus/pathology , Cell Adhesion , Cell Movement , Gene Targeting , Mice , Organ Size , Osteogenesis , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Up-Regulation , Wound HealingABSTRACT
A Thanatophoric dysplasia, is a severe congenital anomaly which mostly causes stillbirth or death of the affected baby within hours due to respiratory insufficiency. The diagnosis of TD is typically suspected on ultrasound during the second trimester of pregnancy, when severe shortening of the long bones, frontal bossing, flattened vertebrae, and short ribs that result in a narrow thorax and bell-shaped abdomen, can be seen. Here, we present a case with prenatal ultrasonographic findings suggestive of TD, and highlight the patient's postnatal dysmorphic features and typical radiographic findings. The definitive diagnosis of TD type I (TDI) was made postnatally, when molecular genetic analysis revealed the previously described p.R248C mutation in FGFR3. This case is reported due to its relative long life span and the definitive molecular diagnosis that could be made during hospitalization.
Subject(s)
DNA Mutational Analysis , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/genetics , Fatal Outcome , Genetic Carrier Screening , Humans , Infant , Infant, Newborn , Karyotyping , Pathology, Molecular , Receptor, Fibroblast Growth Factor, Type 3/genetics , Survival , Thanatophoric Dysplasia/diagnosis , Thanatophoric Dysplasia/mortality , Ultrasonography, PrenatalABSTRACT
We present herein our first experience with a case of thanatophoric dysplasia (type I) that was diagnosed during the second trimester using three- and four-dimensional HDlive ultrasonography. The HDlive rendering mode clearly showed the anatomical features of thanatophoric dysplasia: external malformations and skeletal abnormalities, including extremely short limbs, flattened vertebral bodies, and short horizontal ribs, among others. HDlive can provide valuable, highly realistic images for the differential diagnosis of skeletal dysplasia. It may also play an important complementary role when conventional two- and three-dimensional ultrasonography does not provide sufficient definition.
Subject(s)
Imaging, Three-Dimensional/methods , Receptor, Fibroblast Growth Factor, Type 3/deficiency , Thanatophoric Dysplasia/diagnostic imaging , Ultrasonography, Prenatal/methods , Abortion, Induced , Adult , Female , Humans , Male , PregnancyABSTRACT
Here we describe an experimental tool, termed quantitative imaging Förster resonance energy transfer (QI-FRET), that enables the quantitative characterization of membrane protein interactions. The QI-FRET methodology allows us to acquire binding curves and calculate association constants for complex membrane proteins in the native plasma membrane environment. The method utilizes FRET detection, and thus requires that the proteins of interest are labeled with florescent proteins, either FRET donors or FRET acceptors. Since plasma membranes of cells have complex topologies precluding the acquisition of two-dimensional binding curves, the FRET measurements are performed in plasma membrane derived vesicles that bud off cells as a result of chemical or osmotic stress. The results overviewed here are acquired in vesicles produced with an osmotic vesiculation buffer developed in our laboratory, which does not utilize harsh chemicals. The concentrations of the donor-labeled and the acceptor-labeled proteins are determined, along with the FRET efficiencies, in each vesicle. The experiments utilize transient transfection, such that a wide variety of concentrations is sampled. Then, data from hundreds of vesicles are combined to yield dimerization curves. Here we discuss recent findings about the dimerization of receptor tyrosine kinases (RTKs), membrane proteins that control cell growth and differentiation via lateral dimerization in the plasma membrane. We focus on the dimerization of fibroblast growth factor receptor 3 (FGFR3), a RTK that plays a critically important role in skeletal development. We study the role of different FGFR3 domains in FGFR3 dimerization in the absence of ligand, and we show that FGFR3 extracellular domains inhibit unliganded dimerization, while contacts between the juxtamembrane domains, which connect the transmembrane domains to the kinase domains, stabilize the unliganded FGFR3 dimers. Since FGFR3 has been documented to harbor many pathogenic single amino acid mutations that cause skeletal and cranial dysplasias, as well as cancer, we also study the effects of these mutations on dimerization. First, we show that the A391E mutation, linked to Crouzon syndrome with acanthosis nigricans and to bladder cancer, significantly enhances FGFR3 dimerization in the absence of ligand and thus induces aberrant receptor interactions. Second, we present results about the effect of three cysteine mutations that cause thanatophoric dysplasia, a lethal phenotype. Such cysteine mutations have been hypothesized previously to cause constitutive dimerization, but we find instead that they have a surprisingly modest effect on dimerization. Most of the studied pathogenic mutations also altered FGFR3 dimer structure, suggesting that both increases in dimerization propensities and changes in dimer structure contribute to the pathological phenotypes. The results acquired with the QI-FRET method further our understanding of the interactions between FGFR3 molecules and RTK molecules in general. Since RTK dimerization regulates RTK signaling, our findings advance our knowledge of RTK activity in health and disease. The utility of the QI-FRET method is not restricted to RTKs, and we thus hope that in the future the QI-FRET method will be applied to other classes of membrane proteins, such as channels and G protein-coupled receptors.