CRL4-Cereblon complex in Thalidomide Embryopathy: a translational investigation.

The Cereblon-CRL4 complex has been studied predominantly with regards to thalidomide treatment of multiple myeloma. Nevertheless, the role of Cereblon-CRL4 in Thalidomide Embryopathy (TE) is still not understood. Not all embryos exposed to thalidomide develop TE, hence here we evaluate the role of the CRL4-Cereblon complex in TE variability and susceptibility. We sequenced CRBN, DDB1, CUL4A, IKZF1, and IKZF3 in individuals with TE. To better interpret the variants, we suggested a score and a heatmap comprising their regulatory effect. Differential gene expression after thalidomide exposure and conservation of the CRL4-Cereblon protein complex were accessed from public repositories. Results suggest a summation effect of Cereblon variants on pre-axial longitudinal limb anomalies, and heatmap scores identify the CUL4A variant rs138961957 as potentially having an effect on TE susceptibility. CRL4-Cereblon gene expression after thalidomide exposure and CLR4-Cereblon protein conservation does not explain the difference in Thalidomide sensitivity between species. In conclusion, we suggest that CRL4-Cereblon variants act through several regulatory mechanisms, which may influence CRL4-Cereblon complex assembly and its ability to bind thalidomide. Human genetic variability must be addressed not only to further understand the susceptibility to TE, but as a crucial element in therapeutics, including in the development of pharmacogenomics strategies.

Extremity Findings of Methotrexate Embryopathy.

Background: Methotrexate (MTX) is widely used as an immunosuppressant, chemotherapeutic, and abortifacient agent. It is also a potent teratogen, and intentional or unintentional exposure during pregnancy is associated with heterogeneous birth anomalies. Methods: We retrospectively reviewed a cohort of patients who presented to our clinic with limb anomalies in the setting of MTX embryopathy. Results: In our case series, we describe 7 cases of patients who had limb anomalies with heterogeneous functionality, from severely debilitating to completely asymptomatic. Most of the upper extremity anomalies in our group were managed conservatively. Conclusions: Methotrexate embryopathy is a rare but clinically important entity with phenotypic and functional variability. This series underscores the need for proper counseling of patients and raises concern regarding using this medication for the purpose of abortion.

The role of ESCO2, SALL4 and TBX5 genes in the susceptibility to thalidomide teratogenesis.

Thalidomide is widely used for several diseases; however, it causes malformations in embryos exposed during pregnancy. The complete understanding of the mechanisms by which thalidomide affects the embryo development has not yet been obtained. The phenotypic similarity makes TE a phenocopy of syndromes caused by mutations in ESCO2, SALL4 and TBX5 genes. Recently, SALL4 and TBX5 were demonstrated to be thalidomide targets. To understand if these genes act in the TE development, we sequenced them in 27 individuals with TE; we verified how thalidomide affect them in human pluripotent stem cells (hPSCs) through a differential gene expression (DGE) analysis from GSE63935; and we evaluated how these genes are functionally related through an interaction network analysis. We identified 8 variants in ESCO2, 15 in SALL4 and 15 in TBX5. We compared allelic frequencies with data from ExAC, 1000 Genomes and ABraOM databases; eight variants were significantly different (p < 0.05). Eleven variants in SALL4 and TBX5 were previously associated with cardiac diseases or malformations; however, in TE sample there was no association. Variant effect prediction tools showed 97% of the variants with potential to influence in these genes regulation. DGE analysis showed a significant reduction of ESCO2 in hPSCs after thalidomide exposure.

Limb deformity in a newborn. Is rifampicin just an innocent bystander?

OBJECTIVE: The first-line antituberculous agents for use during pregnancy have minimal teratogenic effects. The possibility of limb deformity during rifampin use, however, was reported by some researchers. CASE REPORT: A male newborn was born with a hypoplastic right forearm to a mother with tuberculosis who used isoniazid and rifampicin in the first two months of her pregnancy. CONCLUSIONS: The limb anomaly in our case might be attributed to rifampicin usage during the first 2 months of pregnancy. Caution should be given with regard to possible congenital malformations which could be associated with the treatment of pregnant women with antituberculous drugs.

Defectos transversos terminales de miembros por exposición prenatal al misoprostol / Terminal Transverse Defects Members by Prenatal Exposure to Misoprostol

La exposición prenatal al misoprostol se ha asociado con la ocurrencia de defectos transversos terminales en miembros superiores e inferiores: gastroquisis, hipodactilia e hipoglosia entre otros defectos congénitos. El mecanismo de acción del misoprostol como agente teratogénico ha sido atribuido a un proceso de disrupción vascular con alteración del flujo sanguíneo al feto, lo cual deriva en perfusión inadecuada, hipoxia y hemorragia. Se presenta este caso de niño de dos años de edad con defectos transversos terminales de miembros superiores e inferiores cuya madre utilizó misoprostol en el primer trimestre del embarazo con fines abortivos. Se realiza esta investigación con el objetivo de brindar información sobre los efectos teratogénicos del Misoprostol y alertar sobre las posibles consecuencias de la continuación del embarazo después de su uso con fines abortivos para prevenir el nacimiento de niños con defectos congénitos incompatibles para la vida o con discapacidad permanente

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Sensitivity to cadmium-chloride-induced forelimb ectrodactyly is independent of the p53 gene-dosage in the C57BL/6J mouse.

BACKGROUND: The p53 pathway plays an important role in the regulation of apoptosis, osteoblast differentiation, skeletal development, and teratogenic sensitivity. The administration of cadmium chloride (CdCl(2)) on gestational day 9 in susceptible mouse strains causes postaxial forelimb ectrodactyly in a percentage of fetuses through unknown mechanisms. In this study, the hypothesis that the p53 gene dosage might affect the incidence or severity of CdCl(2)-induced forelimb ectrodactyly was examined. METHODS: Heterozygous p53-null female mice, on the C57BL/6J background known to be sensitive to CdCl(2)-induced forelimb ectrodactyly, were mated with heterozygous males and then treated with a single intraperitoneal (ip) dose of CdCl(2) (4 mg x kg(-1)) at embryonic day (ED) 9. Embryos and fetuses, genotyped using DNA isolated from the yolk sacs, were collected at ED10 and examined for the pattern of cell death in the limb buds or collected at ED18 and examined for limb malformations. RESULTS: In the wild type and heterozygous p53 embryonic limb buds, CdCl(2)-induced apoptosis involved mesenchymal cells as well as the apical ectodermal ridge (AER), whereas CdCl(2)-induced apoptosis was restricted mainly to the AER in the homozygous p53-null limb buds. No difference in the incidence or severity of forelimb ectrodactyly in the embryos of different p53 genotypes was observed. CONCLUSION: Despite the fact that CdCl(2) induced both p53-dependent (in the mesenchyme) and p53-independent (in the AER) cell death in the developing limb bud, CdCl(2)-induced ectrodactyly was independent of the p53 gene dosage at the studied time point.

Effects of limited doses of retinyl palmitate at the critical time of limb morphogenesis in mouse embryos.

With a view to examine the effects of defined doses of retinyl palmitate (Vit. A) on limb morphogenesis and their effects at the critical time in mouse embryos, pregnant Swiss Webster albino mice were administered retinyl palmitate (10000 or 15000 IU/kg, i.p.) on different days of pregnancy. Vitamin A in 15000 IU/kg, i.p. dose was most effective as produced malformations in the forelimbs by day 10 in 28.6% mice and in the hindlimbs by day 11 in 20.6% mice. Further, two injections in a day with the lower dose (10000 IU/kg, i.p.) had more teratogenic effects than single 15000 IU/kg, i.p. injection. Two injections of either dose on day 10 resulted in higher embryo absorption.

Fetal sodium valproate exposure causes Baller-Gerold syndrome phenotype: both phenotypes in the same family.

Baller-Gerold syndrome (BGS) is characterized by craniosynostosis and preaxial upper-limb malformations, and it has an autosomal recessive inheritance. Valproate syndrome occurs after exposure to valproic acid in utero, and is characterized by trigonocephaly. Both syndromes can also present with other malformations. Herein, we report a female newborn and her brother who both had a history of fetal exposure to maternal anti-epileptic drugs, especially sodium valproate. On physical examination of the female patient, craniosynostosis, trigonocephaly, right radius aplasia and hypoplastic thumb, and cardiac and renal malformations were determined, and she was diagnosed with BGS phenotype. The brother's examination revealed trigonocephaly, polymastia and hypospadias, and he was diagnosed with valproate syndrome. Based on these patients, we aimed to add further evidence in the literature indicating that the use of sodium valproate alone and in combination with other anti-epileptic drugs throughout pregnancy can increase the risk of serious fetal congenital malformations depending on the doses.

Unilateral radius aplasia due to lamotrigine and oxcarbazepine use in pregnancy.

Lamotrigine (LTG) has been used in epilepsia patients for treatment of partial seizures. It can cross the placenta and there are limited data about its use in pregnancy and foetal adverse effects. Extremity and cardiac malformations, dysmorphic facial appearance, coanal atresia and upper respiratory and gastrointestinal anomalies have been reported because of LTG use in pregnancy. Oxcarbazepine (OXC) is one of the new antiepileptic agents. Although the drug and its metabolites can easily pass from placenta to the foetus, available data suggest that it can be safely used during pregnancy because no teratogenicity has been reported. One infant with atrial septal defect and patent ductus arteriosus due to use of LTG and OXC in pregnancy has been reported in literature. Here, we report a female infant with micrognatia, low-set ears, facial dysmorphism and unilateral radius aplasia born to a mother who used LTG 100 mg/day and OXC 1200 mg/day during pregnancy for seizures. To our knowledge, this is the first major anomaly case associated with the combined use of these drugs. This case can provide useful data about the teratogenicity of LTG and OXC combination therapy.

Fetal exposure to sodium valproate associated with Baller-Gerold syndrome: case report and review of the literature.

CASE REPORT: We report three patients with a history of maternal valproate use during pregnancy who presented with a combination of metopic suture synostosis and upper limb malformations, which could be diagnosed as Baller-Gerold syndrome (BGS). The patients underwent surgical treatment for the craniofacial deformity, during which standard frontocranial reconstruction was performed. REVIEW OF THE LITERATURE: Only 32 patients have been reported in the world literature and these cases support the emerging view that BGS is not a distinct syndrome, but should instead be considered to be an heterogeneous phenotype with variable etiology. CONCLUSIONS: Our case series suggests for the first time that fetal sodium valproate exposure may also cause this phenotype.

Cadmium-induced postaxial forelimb ectrodactyly: association with altered sonic hedgehog signaling.

Administration of CdSO(4) to C57BL/6 mice at day 9.5 of gestation induces a high incidence of postaxial forelimb ectrodactyly in the offspring. We propose that Cd(2+) exposure impairs the process of anterior/posterior formation in the limb bud, a process that is directed by Sonic hedgehog (Shh) signaling. We show that exposure of the mouse embryo to Cd(2+) disrupts Shh signaling as measured by polarizing activity of mouse limb bud ZPA grafted to a host chick wing, and activity of a Gli:luciferase reporter exposed to limb bud lysates. Yet the expression of Shh and its translation are not affected by Cd(2+) exposure. We propose that teratogen exposure affects the processing of Shh in the cells in which it is made.

Reduced somatosensory hand representation in thalidomide-induced dysmelia as revealed by fMRI.

The concept of cerebral plasticity suggests that the hand representation in somatosensory cortex is abnormal in congenital malformation disorders. To investigate this issue we studied 11 subjects with different degrees of upper extremity dysmelia due to thalidomide embryopathy in comparison to 10 control subjects. In the affected subjects fingers are typically missing in radio-ulnar order beginning with the thumb. Haemodynamic responses to electrical stimulation of the radial-most and ulnar-most fingers were measured in each subject using functional magnetic resonance tomography. The size of the hand area in the primary somatosensory cortex was estimated by calculating the Euclidian distance between corresponding activation peaks on the lateral postcentral gyrus. The cortical somatosensory hand representation was found to be significantly smaller in dysmelic subjects as compared with the control subjects (P <0.001). The shrinkage of the hand area was not proportional to the number of missing fingers. Furthermore, the cortical representation of the ulnar fingers in the dysmelic subjects was shifted towards the cortical thumb representation of the control group. We suggest that the unproportional reduction of the hand area together with the observed shift may reflect use-dependent rather than malformation-induced reorganization of the somatosensory hand area.

Use-dependent cortical plasticity in thalidomide-induced upper extremity dysplasia: evidence from somaesthesia and neuroimaging.

In this study cerebral reorganization was investigated in thalidomide-damaged subjects who use their feet to compensate for their malformed upper extremities. Tactile localization across toes was combined with fMRI to study use-dependent plasticity of the human somatosensory cortex. The manner of compensatory foot use was assessed by a questionnaire. In the behavioural experiment toes were stimulated with above threshold monofilaments and subjects had to report which toe was stimulated. When feet were employed for all everyday actions subjects made significantly fewer errors in the localization task. In subjects who use their feet only for specific actions such as grasping objects there were as many localization errors as in the control group of thalidomide-affected subjects with normal extremities. However, the patterns of mislocalizations were different with less errors occurring for the toe of the dominant foot involved in these actions. Functional MRI showed stronger haemodynamic responses to electrical stimulation of the toes in subjects using their feet for everyday actions as compared to controls. Our data show that long-term use of the feet for fine sensorimotor skills leads to better performance in tactile localization and changes in cerebral SI representation supporting the notion of use-dependent plasticity in the somatosensory cortex.