Implication of transcription factor FOXD2 dysfunction in syndromic congenital anomalies of the kidney and urinary tract (CAKUT).

Congenital anomalies of the kidney and urinary tract (CAKUT) are the predominant cause for chronic kidney disease below age 30 years. Many monogenic forms have been discovered due to comprehensive genetic testing like exome sequencing. However, disease-causing variants in known disease-associated genes only explain a proportion of cases. Here, we aim to unravel underlying molecular mechanisms of syndromic CAKUT in three unrelated multiplex families with presumed autosomal recessive inheritance. Exome sequencing in the index individuals revealed three different rare homozygous variants in FOXD2, encoding a transcription factor not previously implicated in CAKUT in humans: a frameshift in the Arabic and a missense variant each in the Turkish and the Israeli family with segregation patterns consistent with autosomal recessive inheritance. CRISPR/Cas9-derived Foxd2 knockout mice presented with a bilateral dilated kidney pelvis accompanied by atrophy of the kidney papilla and mandibular, ophthalmologic, and behavioral anomalies, recapitulating the human phenotype. In a complementary approach to study pathomechanisms of FOXD2-dysfunction-mediated developmental kidney defects, we generated CRISPR/Cas9-mediated knockout of Foxd2 in ureteric bud-induced mouse metanephric mesenchyme cells. Transcriptomic analyses revealed enrichment of numerous differentially expressed genes important for kidney/urogenital development, including Pax2 and Wnt4 as well as gene expression changes indicating a shift toward a stromal cell identity. Histology of Foxd2 knockout mouse kidneys confirmed increased fibrosis. Further, genome-wide association studies suggest that FOXD2 could play a role for maintenance of podocyte integrity during adulthood. Thus, our studies help in genetic diagnostics of monogenic CAKUT and in understanding of monogenic and multifactorial kidney diseases.

Fetal phenotype and diagnosis of autosomal dominant Robinow syndrome due to novel DVL1 variant.

Due to abnormal prenatal ultrasound findings of femoral shortening and flattened facial profile, a G2P0 pregnant patient underwent an amniocentesis at 15 weeks of gestation for proband-only exome sequencing. Bioinformatic filtering for genes included on the laboratory's extended skeletal dysplasia panel identified a heterozygous, likely pathogenic, frameshift variant in DVL1 NM_001330311.2:c.1575_1582dup; (p.Pro528ArgfsTer149). Pathogenic variants in DVL1 are associated with autosomal dominant Robinow syndrome (ADRS), a genetic disorder characterized by skeletal dysplasia with genital and craniofacial abnormalities. Prenatal ultrasound in the third trimester noted shortened long bones (first percentile for gestational age), macrocephaly with frontal bossing, short and upturned nose with a wide nasal root, triangular mouth, low pedal arches concerning for rocker-bottom feet, and ambiguous genitalia. A postnatal exam by Medical Genetics confirmed the prenatal findings in addition to hypertelorism, brachydactyly with broad thumbs and halluces, clinodactyly of second fingers, rigid gums with a frontal frenulum, and a sacral dimple. This case describes a novel variant in DVL1 identified in a fetus with prenatal and postnatal phenotypic features consistent with ADRS. To our knowledge, this is the first reported case of a prenatal molecular diagnosis of the dominant form of Robinow syndrome and the third case to describe prenatal ultrasound findings associated with this diagnosis.

Outcomes for expanded polytetrafluoroethylene strip in frontalis suspension surgery.

PURPOSE: This study evaluates surgical outcomes and complication rates of frontalis suspension with expanded polytetrafluoroethylene (ePTFE). METHODS: This retrospective cohort study reviewed all patients undergoing frontalis suspension surgery using ePTFE as the sling material from January 1 2012 to March 3 2020 by a single surgeon at a single academic center. Two different surgical techniques were evaluated in the placement of the sling material. Demographic, clinical, and operative data were extracted. Outcome data including postoperative lid height, reoperation, and complication rate were extracted for the cohort and compared between the two surgical techniques. Descriptive statistics were utilized. RESULTS: Sixty-four eyes from 49 unique patients were included in this study. Forty-three (67.2%) patients had isolated congenital blepharoptosis; 14 (21.9%) had blepharophimosis, ptosis, and epicanthus inversus syndrome (BPES); and 2 (3.1%) had cranial nerve III palsy. Fifty-one (79.7%) patients had no prior blepharoptosis surgery. Lid crease incision and stab incision techniques were utilized for 24 (37.5%) and 40 (62.5%) eyes, respectively. Overall, 21 (32.8%) eyes required reoperation with ePTFE to achieve appropriate eyelid height or contour. Only one patient experienced implant infection, requiring removal of ePTFE sling after a second reoperation. There were no cases of implant exposure or granuloma formation noted during the study period. CONCLUSION: An ePTFE strip soaked in cefazolin prior to utilization in surgery is a viable material for frontalis suspension surgery, with a lower infectious or inflammatory complication rate than previously reported. However, reoperation rate was still relatively high.

Bi-allelic Pathogenic Variants in HS2ST1 Cause a Syndrome Characterized by Developmental Delay and Corpus Callosum, Skeletal, and Renal Abnormalities.

Heparan sulfate belongs to the group of glycosaminoglycans (GAGs), highly sulfated linear polysaccharides. Heparan sulfate 2-O-sulfotransferase 1 (HS2ST1) is one of several specialized enzymes required for heparan sulfate synthesis and catalyzes the transfer of the sulfate groups to the sugar moiety of heparan sulfate. We report bi-allelic pathogenic variants in HS2ST1 in four individuals from three unrelated families. Affected individuals showed facial dysmorphism with coarse face, upslanted palpebral fissures, broad nasal tip, and wide mouth, developmental delay and/or intellectual disability, corpus callosum agenesis or hypoplasia, flexion contractures, brachydactyly of hands and feet with broad fingertips and toes, and uni- or bilateral renal agenesis in three individuals. HS2ST1 variants cause a reduction in HS2ST1 mRNA and decreased or absent heparan sulfate 2-O-sulfotransferase 1 in two of three fibroblast cell lines derived from affected individuals. The heparan sulfate synthesized by the individual 1 cell line lacks 2-O-sulfated domains but had an increase in N- and 6-O-sulfated domains demonstrating functional impairment of the HS2ST1. As heparan sulfate modulates FGF-mediated signaling, we found a significantly decreased activation of the MAP kinases ERK1/2 in FGF-2-stimulated cell lines of affected individuals that could be restored by addition of heparin, a GAG similar to heparan sulfate. Focal adhesions in FGF-2-stimulated fibroblasts of affected individuals concentrated at the cell periphery. Our data demonstrate that a heparan sulfate synthesis deficit causes a recognizable syndrome and emphasize a role for 2-O-sulfated heparan sulfate in human neuronal, skeletal, and renal development.

A homozygous frame-shift variant in PROSER1 is associated with developmental delay, hypotonia, genitourinary malformations, and distinctive facial features.

We report four children from three related families who presented with a similar phenotype characterized by developmental delay, hypotonia, seizures, failure-to-thrive, strabismus, drooling, recurrent otitis media, hearing impairment, and genitourinary malformations. They also shared common facial features including arched eyebrows, prominent eyes, broad nasal bridge, low-hanging columella, open mouth, thick lower lip, protruding tongue, large low-set ears, and parietal bossing. Exome sequencing for affected individuals revealed a homozygous frame-shift variant, c.1833del; p.(Thr612Glnfs*22), in PROSER1 which encodes the proline and serine rich protein 1 (PROSER1). PROSER1 has recently been found to be part of the histone methyltransferases KMT2C/KMT2D complexes. PROSER1 stabilizes TET2, a member of the TET family of DNA demethylases which is involved in recruiting the enhancer-associated KMT2C/KMT2D complexes and mediating DNA demethylation, activating gene expression. Therefore, PROSER1 may play vital and potentially general roles in gene regulation, consistent with the wide phenotypic spectrum observed in the individuals presented here. The consistent phenotype, the loss-of-function predicted from the frame-shift, the co-segregation of the phenotype in our large pedigree, the vital role of PROSER1 in gene regulation, and the association of related genes with neurodevelopmental disorders argue for the loss of PROSER1 to be the cause for a novel recognizable syndrome.

A homozygous ROR2 variant in a family with atypical Robinow syndrome and tetramelic transverse deficiency of autopods.

We present five members of a consanguineous Pakistani kinship with the most severe familial tetramelic transverse autopod deficiency reported to date and additionally having some of the common autosomal recessive Robinow syndrome-1 (RRS1) features including short stature, short neck, severe vertebral anomalies of kyphoscoliosis, hemivertebrae, fusion of thoracic vertebrae, broad forehead, and dental crowding. We mapped the locus of this atypical RRS and detected homozygous 8-nucleotide deletion c.1353_1360del (p.(Met452Alafs*4)) in ROR2, the gene responsible for RRS1. We did not find any other variant shared by all affected individuals that could possibly act as a modifier of limb defect. Autopods are affected in RRS1, but severe autopod deficiency is not a characteristic feature. Over 30 biallelic variants dispersed throughout the gene are known in ROR2-related RS, with no genotype-phenotype correlation for specific RRS1 features. Considering together with the sporadic case homozygous for variant p.(Arg442*) and the case homozygous for p.(Arg441Thrfs*16) in a family where heterozygous members have brachydactyly type B1, we propose that homozygous truncating variants that originate at residues 441-452 can cause severe autopod reduction anomalies, suggesting some genotype-phenotype correlation for this particular phenotype.

Clinical and molecular characterization of four patients with Robinow syndrome from different families.

Robinow syndrome (RS) is a rare heterogeneous disorder characterized by short stature, short-limbs, craniofacial, oro-dental abnormalities, vertebral segmentation defects, and frequently genital hypoplasia. Both autosomal dominant and recessive patterns of inheritance are observed with many causative genes. Here, we present the phenotypes and genotypes of four children with RS from different Indian families. Sequence variants were identified in genes ROR2, DVL1, and DVL3. Our results expand the mutational spectrum of RS and we also highlight the radiological changes in the radius and ulna in patients with ROR2 sequence variants which are primarily characteristic for ROR2 related RS but have been reported in WNT5A related RS.

Craniofacial phenotypes associated with Robinow syndrome.

Robinow syndrome is characterized by mesomelic limb shortening, hemivertebrae, and genital hypoplasia. Due to low prevalence and considerable phenotypic variability, it has been challenging to definitively characterize features of Robinow syndrome. While craniofacial abnormalities associated with Robinow syndrome have been broadly described, there is a lack of detailed descriptions of genotype-specific phenotypic craniofacial features. Patients with Robinow syndrome were invited for a multidisciplinary evaluation conducted by specialist physicians at our institution. A focused assessment of the craniofacial manifestations was performed by a single expert examiner using clinical examination and standard photographic images. A total of 13 patients with clinical and molecular diagnoses consistent with either dominant Robinow syndrome (DRS) or recessive Robinow syndrome (RRS) were evaluated. On craniofacial examination, gingival hyperplasia was nearly ubiquitous in all patients. Orbital hypertelorism, a short nose with anteverted and flared nares, a triangular mouth with a long philtrum, cleft palate, macrocephaly, and frontal bossing were not observed in all individuals but affected individuals with both DRS and RRS. Other anomalies were more selective in their distribution in this patient cohort. We present a comprehensive analysis of the craniofacial findings in patients with Robinow Syndrome, describing associated morphological features and correlating phenotypic manifestations to underlying genotype in a manner relevant for early recognition and focused evaluation of these patients.

Expanding the KIF4A-associated phenotype.

Kinesin super family (KIF) genes encode motor kinesins, a family of evolutionary conserved proteins, involved in intracellular trafficking of various cargoes. These proteins are critical for various physiological processes including neuron function and survival, ciliary function and ciliogenesis, and cell-cycle progression. Recent evidence suggests that alterations in motor kinesin genes can lead to a variety of human diseases, including monogenic disorders. Neuropathies, impaired higher brain functions, structural brain abnormalities and multiple congenital anomalies (i.e., renal, urogenital, and limb anomalies) can result from pathogenic variants in many KIF genes. We expand the phenotype associated with KIF4A variants from developmental delay and intellectual disability with or without epilepsy to a congenital anomaly phenotype with hydrocephalus and various brain anomalies at the more severe end of phenotypic manifestations. Additional anomalies of the kidneys and urinary tract, congenital lymphedema, eye, and dental anomalies seem to be variably associated and overlap with clinical signs observed in other kinesinopathies. Caution still applies to missense variants, but hopefully, future work will further establish genotype-phenotype correlations in a larger number of patients and functional studies may give further insights into the complex function of KIF4A.

P63-related disorders: Dermatological characteristics in 22 patients.

In P63-related ectodermal dysplasias (ED), the clinical characteristics focus on extra-cutaneous manifestations. The dermatological phenotype remains incompletely characterized. We report the dermatological features of 22 patients carrying a TP63 mutation. Erosions, erythroderma and pigmentary anomalies are characteristics of P63-related ED. Our data suggest that patients might be classified into two major P63-related disorders: AEC and EEC. RHS and ADULT represent mild AEC and EEC forms, respectively.

Popliteal Pterygium Syndrome With Syngnathia.

Popliteal pterygium syndrome is a condition characterized by skin webs on the popliteal fossa, which may impair mobility unless surgically repaired. Affected individuals may also have syndactyly on the fingers and/or toes. Most people with this disorder present cleft lip and cleft palate and they can have syngnathia, that is a congenital adhesion between maxilla and mandible by fibrous bands, which affects the opening of the mouth. The case that we report is about a 2-month-old male, who presented skin webs bilaterally on the popliteal fossa, syndactyly between the IV and the V toe of the right foot and between the III and the IV toe of the left foot, and genital malformations. He was born with complete bilateral cleft lip and complete cleft palate on the left side and incomplete cleft palate on the right side and syngnathia with 4 fibrous bands between the mandibular arch and the maxilla arch on the right side, which affected the opening of the mouth. The case of our patient is very interesting because there have been few reported patients affected by popliteal pterygium syndrome with syngnathia.

A piggyBac insertion disrupts Foxl2 expression that mimics BPES syndrome in mice.

Blepharophimosis, ptosis, epicanthus inversus syndrome (BPES) is an autosomal dominant genetic disorder characterized by small palpebral fissures and other craniofacial malformations, often with (type I) but could also without (type II) premature ovarian failure. While mutations of the forkhead transcription factor FOXL2 are associated with and likely be responsible for many BPES cases, how FOXL2 affects craniofacial development remain to be understood. Through a large-scale piggyBac (PB) insertion mutagenesis, we have identified a mouse mutant carrying a PB insertion ∼160 kb upstream of the transcription start site (TSS) of Foxl2. The insertion reduces, but not eliminates, the expression of Foxl2. This mutant, but not its revertant, displays BPES-like conditions such as midface hypoplasia, eyelid abnormalities and female subfertility. Further analysis indicates that the mutation does not affect mandible, but causes premature fusion of the premaxilla-maxilla suture, smaller premaxilla and malformed maxilla during midface development. We further identified an evolutionarily conserved fragment near the insertion site and observed enhancer activity of this element in tissue culture cells. Analyses using DNase I hypersensitivity assay and chromosome conformation capture assay in developing maxillary and periocular tissues suggest that the DNA region near the insertion site likely interacts with Foxl2 TSS. Therefore, this mutant presents an excellent animal model for mechanistic study of BPES and regulation of Foxl2.

A wet dressing for male genital surgery: A phase II clinical trial.

PURPOSE: This study was to confirm the safety and efficacy of BC dressing when used in surgical male wound healing at the urogenital area. METHODS: Open, non-controlled clinical study of phase II. A total of 141 patients, among those children, adolescents and adults with hypospadias (112), epispadias (04), phymosis (13) and Peyronie's disease (12) that had a BC dressing applied over the operated area after surgery. A written informed consent was obtained from all participants. Study exclusion criteria were patients with other alternative treatment indications due to the severity, extent of the injury or the underlying disease. The outcomes evaluated were efficacy, safe and complete healing. The costs were discussed. RESULTS: In 68% patients, the BC dressing fell off spontaneously. The BC was removed without complications in 13% of patients at the outpatient clinic during the follow-up visit and 17% not reported the time of removal. In 3% of the cases, the dressing fell off early. Complete healing was observed between 8th and 10th days after surgery. The BC dressings have shown a good tolerance by all the patients and there were no reports of serious adverse events. CONCLUSION: The bacterial cellulose dressings have shown efficacy, safety and that can be considered as a satisfactory alternative for postoperative wound healing in urogenital area and with low cost.

Urinary excretion of polyols and sugars in children with chronic kidney disease.

BACKGROUND: The urinary concentrations of monosaccharides and polyols are used for diagnosing inborn errors of metabolism and renal tubular disorders. Reference values are age-related and depend on the method of detection. However, the influence of the renal function is often still neglected. In this study we examined the urinary excretion of monosaccharides and polyols in children with various degrees of chronic kidney disease (CKD), but with no known metabolic or renal tubular disorders. CASE DIAGNOSIS/TREATMENT: In 25 patients with CKD stage 1-5, urinary concentrations of 18 monosaccharides and polyols were measured by gas chromatography-mass spectrometry (GC-MS) in random urinary samples and were compared with age-related reference values. Serum creatinine was measured at the time of the urine sample, and the height-independent estimated glomerular filtration rate (eGFR-Pottel) was calculated. Urinary excretions of monosaccharides and polyols were above the reference values in 8-88% of all patients. A significant difference between CKD stage 1-2 compared with CKD stage 3-5 was found for allose, arabitol and sorbitol (p < 0.05) and for arabinose, fucose, myoinositol, ribitol, xylitol, and xylose (p < 0.01). CONCLUSIONS: We show that the excretion of polyols and sugars depends on eGFR, which warrants a cautious interpretation of the results in patients with CKD.

Cervicovaginal anastomosis by Gore-Tex in Mullerian agenesis.

OBJECTIVE: To determine usage of polytetrafluoroethylene, Gore-Tex, as a stent in patients with congenital cervicovaginal agenesis. DESIGN: Follow-up study. SETTING: Tertiary center referral hospital. PATIENTS: Eight patients with Mullerian agenesis who had congenital cervical agenesis, associated with partial or complete vaginal aplasia. INTERVENTIONS: Laparoscopic uterovaginal anastomosis with placement of a polytetrafluoroethylene stent for reconstruction of cervical agenesis. In cases associated with vaginal agenesis, modified McIndoe vaginoplasty was done concomitantly. MEASUREMENTS AND MAIN RESULTS: Eight patients with cervical agenesis who had partial and complete vaginal aplasia underwent vaginoplasty. Mean (SD) follow-up after surgery (3 years old). All patients except three of them, (62.5 %) experienced relief of abdominal pain and regular menstruation. Due to ascending infection three of them (37.5 %) needed reoperation. So, they underwent laparotomy. Two of these three patients had total abdominal hysterectomy and one case tolerated resection of right obstructed side of didelphys uterus, then she became pregnant spontaneously. Four out of these eight cases were married and the others were single. One out of our married patients conceived three times. Her first pregnancy was aborted. She delivered two alive babies after inserting transabdominal Shirodkar suture in her other pregnancies. CONCLUSION: Uterovaginal anastomosis assisted laparoscopy with polytetrafluoroethylene (Gore-Tex) is effective and hopeful to relieve patients' symptoms, menstruation regularity, and saving fertility. If ascending infection threatening health happens, hysterectomy will be recommended.

VACTERL (vertebral defects, anal atresia, tracheoesophageal fistula with esophageal atresia, cardiac defects, renal and limb anomalies) association: disease spectrum in 25 patients ascertained for their upper limb involvement.

OBJECTIVE: To review the clinical characteristics in a series of 25 patients with VACTERL (vertebral defects, anal atresia, tracheoesophageal fistula with esophageal atresia, cardiac defects, renal and limb anomalies) association who were ascertained for upper limb involvement. STUDY DESIGN: The study involved a review of clinical and radiologic data from patients with VACTERL association collected by a hand surgery clinic between 2004 and 2013. RESULTS: Radial axis involvement was found in all 25 patients (100%), with severe thumb function impairment in 79% and complete absence of the radius in roughly 33%. Costovertebral anomalies were the most frequent feature, found in 23 patients (92%). All 3 core features (anal atresia, tracheoesophageal fistula with esophageal atresia, and costovertebral anomalies) were present in only 12% of the patients. Twelve patients (48%) had abnormalities not part of the VACTERL spectrum, showing a specific pattern of non-VACTERL-type malformations, including genitourinary abnormalities (12%), single umbilical artery (8%), and tethered cord (8%). Previously unreported clinical findings were concurrent hypoplasia of both the odontoid process and the coccyx in 2 patients and an isolated sacral dimple in 2 patients. CONCLUSION: Upper limb involvement in VACTERL association is a specific feature of the radial axis that occurs in monolateral form in approximately 75% of cases and, when bilateral, always occurs in a nonsymmetrical fashion. Odontoid and coccygeal hypoplasia and sacral dimple are newly reported malformations of the VACTERL phenotype.

An osteosclerotic form of Robinow syndrome.

Robinow syndrome (RS) is a clinically and genetically heterogenous condition primarily characterized by short stature, mesomelia, genital hypoplasia, oral abnormalities, and a facial gestalt that includes hypertelorism, a short nose, and a broad mouth. The disorder exists in both a dominant and a more severe recessive form. Here two unrelated cases of sporadic RS are described with the additional finding of axial and appendicular osteosclerosis. These two patients, coupled with three additional patients previously described in the literature, may represent a distinct sub-phenotype of this condition.

Syngnathia and obstructive apnea in a case of popliteal pterygium syndrome.

UNLABELLED: We describe an infant with popliteal pterygia, syngnathia, cleft lip and palate, and retrognathia diagnosed with popliteal pterygium syndrome (PPS). The neonatal course was complicated by severe obstructive apnea necessitating tracheostomy. CONCLUSION: This report illustrates the potential for airway compromise in PPS patients and the need for thorough neonatal airway assessment.

Anesthetic and dental management of a child with IMAGe syndrome.

IMAGe syndrome (OMIM 300290) is a rare multisystem disorder that has a broad phenotypic presentation. Though variable, this disorder mainly consists of Intrauterine growth retardation, Metaphyseal dysplasia, Adrenal hypoplasia congenita, and Genital abnormalities. Patients with IMAGe syndrome present as an uncommon yet important challenge for dentists and anesthesiologists due to their wide range of dysmorphic facial features, adrenal insufficiency, electrolyte imbalances, and need for steroid replacement. The purpose of this case report is to describe the successful anesthetic management of a pediatric patient diagnosed with IMAGe syndrome who presented for full mouth dental rehabilitation.

Craniofacial studies in chicken embryos confirm the pathogenicity of human FZD2 variants associated with Robinow syndrome.

Robinow syndrome is a rare disease caused by variants of seven WNT pathway genes. Craniofacial features include widening of the nasal bridge and jaw hypoplasia. We used the chicken embryo to test whether two missense human FZD2 variants (1301G>T, p.Gly434Val; 425C>T, p.Pro142Lys) were sufficient to change frontonasal mass development. In vivo, the overexpression of retroviruses with wild-type or variant human FZD2 inhibited upper beak ossification. In primary cultures, wild-type and variant human FZD2 significantly inhibited chondrogenesis, with the 425C>T variant significantly decreasing activity of a SOX9 luciferase reporter compared to that for the wild type or 1301G>T. Both variants also increased nuclear shuttling of ß-catenin (CTNNB1) and increased the expression of TWIST1, which are inhibitory to chondrogenesis. In canonical WNT luciferase assays using frontonasal mass cells, the variants had dominant-negative effects on wild-type FZD2. In non-canonical assays, the 425C>T variant failed to activate the reporter above control levels and was unresponsive to exogenous WNT5A. This is the first single amino acid change to selectively alter ligand binding in a FZD receptor. Therefore, FZD2 missense variants are pathogenic and could lead to the altered craniofacial morphogenesis seen in Robinow syndrome.