Fetal autopsy for the diagnosis of skeletal dysplasia and comparison with prenatal ultrasound findings over a 16-year period.

OBJECTIVES: To evaluate the relationship between prenatal ultrasonography (USG) and fetal autopsy findings. METHODS: Among 453 pregnancy terminations performed because of fetal anomalies on prenatal USG, 54 with skeletal dysplasia on fetal autopsy were included in this retrospective study. RESULTS: The most common diagnoses among the 54 fetal autopsies were osteogenesis imperfecta (n=12), dysostosis (n=10), achondroplasia (n=9), arthrogryposis (n=6), and thanatophoric dysplasia (n=6). The prenatal USG and fetal autopsy findings showed complete agreement in 35 cases (64.8%), partial agreement in nine cases (16.6%), and disagreement in 10 cases (18.5%). CONCLUSIONS: Fetal autopsy via perinatal pathology is essential for precise identification of the type of skeletal dysplasia; it should be routinely performed to confirm the diagnosis of prenatally detected fetal anomalies. Autopsy is vital for accurate prenatal diagnosis and the 'gold standard' technique for the identification of clinically important abnormalities.

Thanatophoric dysplasia type 1 with temporal lobe dysplasia: Report of a case along with differential diagnosis.

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.

Early prenatal presentation of the cartilage-hair hypoplasia / anauxetic dysplasia spectrum of disorders mimicking recurrent thanatophoric dysplasia.

Three sibling fetuses identified with limb shortening and thoracic narrowing at twelve weeks' gestation on first trimester ultrasound examination are presented. The parents were non-consanguineous, Caucasian, healthy, of normal stature and had a healthy normal daughter. The radiographic abnormalities were highly suggestive of thanatophoric dysplasia, but molecular analysis failed to identify a pathogenic variant in FGFR3. The three fetuses were found to have identical compound heterozygous mutations in RMRP in trans, one inherited from the mother and one from the father. This represents the early prenatal presentation and fetal findings of metaphyseal dysplasia type McKusick (Cartilage-hair hypoplasia; CHH)/anauxetic dysplasia spectrum of disorders.

Clinical features and molecular genetic analysis of thanatophoric dysplasia type I in a neonate with a de novo c.2419 T > C (p. Ter807Arg) (X807R) mutation in FGFR3.

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 39 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 (38 cm, <3rd percentile), redundant skin folds, a narrow thorax with a bust of 29.5 cm (3-5th percentile), and macrocephaly with a head circumference of 36 cm (>97th percentile). The proposita had postnatal dyspnea and unfortunately died of respiratory failure at the age of 13 days. Molecular genetic analysis revealed a mutation of c.2419 T > C (p. Ter807Arg) (X807R) in FGFR3. Live-born infants with TDI are exceedingly rare, and we hereby report a newborn with a c.2419 T > C mutation in FGFR3, emphasizing phenotype with clinical characteristics and ultrasonographic and X-ray findings, to raise awareness about the heterogeneous patterns of TD.

Multiplex PCR in noninvasive prenatal diagnosis for FGFR3-related disorders.

Thanatophoric dysplasia and achondroplasia are allelic disorders caused by a constitutively active mutation in the FGFR3 gene. Because thanatophoric dysplasia is a lethal disorder and achondroplasia is non-lethal, they need to be distinguished after ultrasound identification of fetal growth retardation with short limbs. Accordingly, we have developed a noninvasive prenatal test using cell-free fetal DNA in the maternal circulation to distinguish thanatophoric dysplasia and achondroplasia. A multiplex PCR system encompassing five mutation hotspots in the FGFR3 gene allowed us to efficiently identify the responsible mutation in cell-free DNA in all examined pregnancies with a suspected thanatophoric dysplasia or achondroplasia fetus. This system will be helpful in the differential diagnosis of thanatophoric dysplasia and achondroplasia in early gestation and in couples concerned about the recurrence of thanatophoric dysplasia due to germinal mosaicism.

Mutant FGFR3 associated with SADDAN disease causes cytoskeleton disorganization through PLCγ1/Src-mediated paxillin hyperphosphorylation.

K650M/E substitutions in the Fibroblast growth factor receptor 3 (FGFR3) are associated with Severe Achondroplasia with Developmental Delay and Acanthosis Nigricans (SADDAN) and Thanatophoric Dysplasia type II (TDII), respectively. Both SADDAN and TDII present with affected endochondral ossification marked by impaired chondrocyte functions and growth plate disorganization. In vitro, K650M/E substitutions confer FGFR3 constitutive kinase activity leading to impaired biosynthesis and accumulation of immature receptors in endoplasmic reticulum (ER)/Golgi. From those compartments, both SADDAN-FGFR3 and TDII-FGFR3 receptors engender uncontrolled signalling, activating PLCγ1, signal transducer and activator of transcription 1, 3 and 5 (STAT1/3/5) and ERK1/2 effectors. Here, we investigated the impact of SADDAN-FGFR3 and TDII-FGFR3 signalling on cytoskeletal organization. We report that SADDAN-FGFR3, but not TDII-FGFR3, affects F-actin organization by inducing tyrosine hyperphosphorylation of paxillin, a key regulator of focal adhesions and actin dynamics. Paxillin phosphorylation was upregulated at tyrosine 118, a functional target of Src and FAK kinases. By using Src-deficient cells and a Src kinase inhibitor, we established a role played by Src activation in paxillin hyperphosphorylation. Moreover, we found that SADDAN-FGFR3 induced FAK phosphorylation at tyrosines 576/577, suggesting its involvement as a Src co-activator in paxillin phosphorylation. Interestingly, paxillin hyperphosphorylation by SADDAN-FGFR3 caused paxillin mislocalization and partial co-localization with the mutant receptor. Finally, the SADDAN-FGFR3 double mutant unable to bind PLCγ1 failed to promote paxillin hyperphosphorylation, pointing to PLCγ1 as an early player in mediating paxillin alterations. Overall, our findings contribute to elucidate the molecular mechanism leading to cell dysfunctions caused by SADDAN-FGFR3 signalling.

HDAC6 deficiency or inhibition blocks FGFR3 accumulation and improves bone growth in a model of achondroplasia.

Mutations that cause increased and/or inappropriate activation of FGFR3 are responsible for a collection of short-limbed chondrodysplasias. These mutations can alter receptor trafficking and enhance receptor stability, leading to increased receptor accumulation and activity. Here, we show that wildtype and mutant activated forms of FGFR3 increase expression of the cytoplasmic deacetylase HDAC6 (Histone Deacetylase 6) and that FGFR3 accumulation is compromised in cells lacking HDAC6 or following treatment of fibroblasts or chondrocytes with small molecule inhibitors of HDAC6. The reduced accumulation of FGFR3 was linked to increased FGFR3 degradation that occurred through a lysosome-dependent mechanism. Using a mouse model of Thanatophoric Dysplasia Type II (TDII) we show that both HDAC6 deletion and treatment with the small molecule HDAC6 inhibitor tubacin reduced FGFR3 accumulation in the growth plate and improved endochondral bone growth. Defective endochondral growth in TDII is associated with reduced proliferation and poor hypertrophic differentiation and the improved bone growth was associated with increased chondrocyte proliferation and expansion of the differentiation compartment within the growth plate. These findings further define the mechanisms that control FGFR3 accumulation and contribute to skeletal pathology caused by mutations in FGFR3.

Endoplasmic reticulum stress-mediated apoptosis contributes to a skeletal dysplasia resembling platyspondylic lethal skeletal dysplasia, Torrance type, in a novel Col2a1 mutant mouse line.

In humans, mutations in the COL2A1 gene encoding the α1(II) chain of type II collagen, create many clinical phenotypes collectively termed type II collagenopathies. However, the mechanisms generating this diversity remain to be determined. Here we identified a novel Col2a1 mutant mouse line by screening a large-scale N-ethyl-N-nitrosourea mutant mouse library. This mutant possessed a p.Tyr1391Ser missense mutation in the C-propeptide coding region, and this mutation was located in positions corresponding to the human COL2A1 mutation responsible for platyspondylic lethal skeletal dysplasia, Torrance type (PLSD-T). As expected, p.Tyr1391Ser homozygotes exhibited lethal skeletal dysplasias resembling PLSD-T, including extremely short limbs and severe dysplasia of the spine and pelvis. The secretion of the mutant proteins into the extracellular space was disrupted, accompanied by an abnormally expanded endoplasmic reticulum (ER) and the up-regulation of ER stress-related genes in chondrocytes. Chondrocyte apoptosis was severely induced in the growth plate of the homozygotes. These findings strongly suggest that ER stress-mediated apoptosis caused by the accumulated mutant proteins in ER contributes to skeletal dysplasia in Co12a1 mutant mice and PLSD-T patients.

Pathophysiological analyses of cortical malformation using gyrencephalic mammals.

One of the most prominent features of the cerebral cortex of higher mammals is the presence of gyri. Because malformations of the cortical gyri are associated with severe disability in brain function, the mechanisms underlying malformations of the cortical gyri have been of great interest. Combining gyrencephalic carnivore ferrets and genetic manipulations using in utero electroporation, here we successfully recapitulated the cortical phenotypes of thanatophoric dysplasia (TD) by expressing fibroblast growth factor 8 in the ferret cerebral cortex. Strikingly, in contrast to TD mice, our TD ferret model showed not only megalencephaly but also polymicrogyria. We further uncovered that outer radial glial cells (oRGs) and intermediate progenitor cells (IPs) were markedly increased. Because it has been proposed that increased oRGs and/or IPs resulted in the appearance of cortical gyri during evolution, it seemed possible that increased oRGs and IPs underlie the pathogenesis of polymicrogyria. Our findings should help shed light on the molecular mechanisms underlying the formation and malformation of cortical gyri in higher mammals.

Thanatophoric dysplasia type 1 with cloverleaf skull in a dichorionic twin.

Here is reported for the first time, a case of thanatophoric dysplasia type 1 with cloverleaf skull in a (Mexican) dichorionic female twin. The patient's main clinical and radiographic findings included severe limb shortening, narrow thorax shape; short ribs, marked platyspondyly, curved short femurs, and a cloverleaf skull. The female twin sib had normal growth parameters and phenotypic appearance. According to the literature, cloverleaf skull in thanatophoric dysplasia type 1 is rare, even more so in dichorionic twins. Moreover, the present observation confirms that thanatophoric dysplasia type 1 patients may show phenotypic heterogeneity related to cloverleaf skull and other congenital anomalies. Therefore, a careful family history along with clinical, radiological, and molecular investigations is suggested, in order to achieve an accurate parental counseling for thanatophoric dysplasia.

A case of thanatophoric dysplasia type 2: a novel mutation.

Thanatophoric dysplasia (TD) is a lethal form of skeletal dysplasia with short-limb dwarfism. Two types distinguished with their radiological characteristics have been defined clinically. The femur is curved in type 1, while it is straight in type 2. TD is known to be due to a mutation in the fibroblast growth factor receptor 3 (FGFR3) gene. We report a male patient who showed clinical findings congruent with TD type 2 and a new mutation in the FGFR3 gene, a finding which has not been reported previously.

Statin treatment rescues FGFR3 skeletal dysplasia phenotypes.

Gain-of-function mutations in the fibroblast growth factor receptor 3 gene (FGFR3) result in skeletal dysplasias, such as thanatophoric dysplasia and achondroplasia (ACH). The lack of disease models using human cells has hampered the identification of a clinically effective treatment for these diseases. Here we show that statin treatment can rescue patient-specific induced pluripotent stem cell (iPSC) models and a mouse model of FGFR3 skeletal dysplasia. We converted fibroblasts from thanatophoric dysplasia type I (TD1) and ACH patients into iPSCs. The chondrogenic differentiation of TD1 iPSCs and ACH iPSCs resulted in the formation of degraded cartilage. We found that statins could correct the degraded cartilage in both chondrogenically differentiated TD1 and ACH iPSCs. Treatment of ACH model mice with statin led to a significant recovery of bone growth. These results suggest that statins could represent a medical treatment for infants and children with TD1 and ACH.

Dysmorphic choroid plexuses and hydrocephalus associated with increased nuchal translucency: early ultrasound markers of de novo thanatophoric dysplasia type II with cloverleaf skull (Kleeblattschaedel).

Prenatal diagnosis of thanatophoric dysplasia (TD) type II presenting in the first trimester with increased nuchal translucency (NT) and cloverleaf skull (Kleeblattschaedel) have been scantly reported in the medical record. Abnormal choroid plexus has been seen in association with fetal anomalies. Here we described a case of increased NT associated with indented choroid plexuses, early onset hydrocephalus and cloverleaf skull in a fetus subsequently diagnosed at early second trimester to carry a de novo mutation encoding for TD type II. The findings of dysmorphic choroid plexus, early onset hydrocephalus and cloverleaf skull at first trimester scan may be early, useful ultrasound markers of TD type II. Molecular analysis to control for possible overlapping syndromes were performed and resulted negative. Postmortem X-ray and 3D-CT scan confirmed the cloverleaf skull, narrow thorax, straight femur with rhizomelic shortening of the limbs and the presence of a communicating hydrocephalus.

Brain malformation with loss of normal FGFR3 expression in thanatophoric dysplasia type I.

Thanatophoric dysplasia is a lethal form of chondrodysplastic dwarfism in which the cerebral cortex displays a unique and complex malformation. We report a female case of thanatophoric dysplasia type I (TD1) with FGFR3 mutation. In this case, fetal ultrasonography at the 18th week of gestation led to a prenatal diagnosis of TD1 with characteristic bone features. The subject was stillborn at the 21st week of gestation, showing marked shortening of the long bones, small thorax and curved short femurs, but without a cloverleaf skull. The temporal lobe was enlarged and hyperconvoluted, appearing as broad gyri and deep sulci, which were composed of focal polymicrogyria-like shallow sulci and heterotopic neuroblastic nests in the intermediate zone and marginal zone. Abundant precursor cells, immunoreactive for nestin and Ki-67 were observed with scattered mitoses in the thickened inner intermediate and subventricular zones of the temporal and occipital lobes. The cytoarchitecture from the entorhinal cortex to Ammon's horn was disorganized with leptomeningeal glioneuronal heterotopia, immunoreactive for doublecortin and nestin. The expression of FGFR3 was virtually not discernible in the temporal and occipital lobes or in the hippocampus. Genetic analysis revealed a point mutation at C8526T (R248C) in the exon 7 of FGFR3. This is the first report that demonstrates that overproduction of intermediate progenitor cells might be induced by FGFR3 mutation in a human TD1 case.

Thanatophoric dysplasia: autopsy findings over a 25-year period.

The aim of our study was to retrospectively assess morphological findings in thanatophoric dysplasia, particularly, in how many cases were cerebral manifestations with temporal lobe dysplasia identified. We also wanted to register and analyze the proportions between lung, brain, and body weight. Criteria for inclusion were an autopsy performed during the period ranging from 1985 to 2009 with a diagnosis of thanatophoric dysplasia. During a 25-year period 25 cases of thanatophoric dysplasia were registered. Temporal lobe dysplasia was recognized in 52% of the cases, and after 1998 temporal lobe dysplasia was described in all cases. In 19 cases the brain/body weight ratio was increased, and in all cases the lung/body weight ratio was below the corresponding ratio calculated according to standard measurements. In all but one case the ratio of brain to lung weight was increased. This study focuses on morphological findings, stressing the importance of temporal lobe dysplasia in confirming a diagnosis of thanatophoric dysplasia. Lung/body, brain/body, and brain/lung weight ratios confirm macrocephaly and lung hypoplasia, which are constant findings in cases involving thanatophoric dysplasia. Femur and brain morphology inclusive histology remains the ultimate tool for confirmation of this lethal condition, although it has to be seen in a context inclusive of radiological examination.