GWAS of thyroid dysgenesis identifies a risk locus at 2q33.3 linked to regulation of Wnt signaling.

Congenital hypothyroidism due to thyroid dysgenesis (TD), presented as thyroid aplasia, hypoplasia or ectopia, is one of the most prevalent rare diseases with an isolated organ malformation. The pathogenesis of TD is largely unknown, although a genetic predisposition has been suggested. We performed a genome-wide association study (GWAS) with 142 Japanese TD cases and 8380 controls and found a significant locus at 2q33.3 (top single nucleotide polymorphism, rs9789446: P = 4.4 × 10-12), which was replicated in a German patient cohort (P = 0.0056). A subgroup analysis showed that rs9789446 confers a risk for thyroid aplasia (per allele odds ratio = 3.17) and ectopia (3.12) but not for hypoplasia. Comprehensive epigenomic characterization of the 72-kb disease-associated region revealed that it was enriched for active enhancer signatures in human thyroid. Analysis of chromosome conformation capture data showed long-range chromatin interactions of this region with promoters of two genes, FZD5 and CCNYL1, mediating Wnt signaling. Moreover, rs9789446 was found to be a thyroid-specific quantitative trait locus, adding further evidence for a cis-regulatory function of this region in thyroid tissue. Specifically, because the risk rs9789446 allele is associated with increased thyroidal expression of FDZ5 and CCNYL1 and given the recent demonstration of perturbed early thyroid development following overactivation of Wnt signaling in zebrafish embryos, an enhanced Wnt signaling in risk allele carriers provides a biologically plausible TD mechanism. In conclusion, our work found the first risk locus for TD, exemplifying that in rare diseases with relatively low biological complexity, GWAS may provide mechanistic insights even with a small sample size.

Rap1 small GTPase is essential for maintaining pulmonary endothelial barrier function in mice.

Vascular permeability is dynamically but tightly controlled by vascular endothelial (VE)-cadherin-mediated endothelial cell-cell junctions to maintain homeostasis. Thus, impairments of VE-cadherin-mediated cell adhesions lead to hyperpermeability, promoting the development and progression of various disease processes. Notably, the lungs are a highly vulnerable organ wherein pulmonary inflammation and infection result in vascular leakage. Herein, we showed that Rap1, a small GTPase, plays an essential role for maintaining pulmonary endothelial barrier function in mice. Endothelial cell-specific Rap1a/Rap1b double knockout mice exhibited severe pulmonary edema. They also showed vascular leakage in the hearts, but not in the brains. En face analyses of the pulmonary arteries and 3D-immunofluorescence analyses of the lungs revealed that Rap1 potentiates VE-cadherin-mediated endothelial cell-cell junctions through dynamic actin cytoskeleton reorganization. Rap1 inhibits formation of cytoplasmic actin bundles perpendicularly binding VE-cadherin adhesions through inhibition of a Rho-ROCK pathway-induced activation of cytoplasmic nonmuscle myosin II (NM-II). Simultaneously, Rap1 induces junctional NM-II activation to create circumferential actin bundles, which anchor and stabilize VE-cadherin at cell-cell junctions. We also showed that the mice carrying only one allele of either Rap1a or Rap1b out of the two Rap1 genes are more vulnerable to lipopolysaccharide (LPS)-induced pulmonary vascular leakage than wild-type mice, while activation of Rap1 by administration of 007, an activator for Epac, attenuates LPS-induced increase in pulmonary endothelial permeability in wild-type mice. Thus, we demonstrate that Rap1 plays an essential role for maintaining pulmonary endothelial barrier functions under physiological conditions and provides protection against inflammation-induced pulmonary vascular leakage.

RMRP-related short stature: A report of six additional Japanese individuals with cartilage hair hypoplasia and literature review.

Biallelic pathogenic variants in RMRP, the gene encoding the RNA component of RNase mitochondrial RNA processing enzyme complex, have been reported in individuals with cartilage hair hypoplasia (CHH). CHH is prevalent in Finnish and Amish populations due to a founder pathogenic variant, n.71A > G. Based on the manifestations in the Finnish and Amish individuals, the hallmarks of CHH are prenatal-onset growth failure, metaphyseal dysplasia, hair hypoplasia, immunodeficiency, and other extraskeletal manifestations. Herein, we report six Japanese individuals with CHH from four families. All probands presented with moderate short stature with mild metaphyseal dysplasia or brachydactyly. One of them had hair hypoplasia and the other immunodeficiency. By contrast, the affected siblings of two families showed only mild short stature. We also reviewed all previously reported 13 Japanese individuals. No n.71A > G allele was detected. The proportions of Japanese versus Finnish individuals were 0% versus 70% for birth length < -2.0 SD, 84% versus 100% for metaphyseal dysplasia and 26% versus 88% for hair hypoplasia. Milder manifestations in the Japanese individuals may be related to the difference of genotypes. The mildest form of CHH phenotypes is mild short stature without overt skeletal alteration or extraskeletal manifestation and can be termed "RMRP-related short stature".

Biochemical monitoring of 21-hydroxylase deficiency: a clinical utility of overnight fasting urine pregnanetriol.

PURPOSE OF REVIEW: 21-Hydroxylase deficiency (21-OHD), the most common form of congenital adrenal hyperplasia, is an autosomal recessive disorder caused by pathogenic variants in CYP21A2. Although this disorder has been known for several decades, many challenges related to its monitoring and treatment remain to be addressed. The present review is written to describe an overview of biochemical monitoring of this entity, with particular focus on overnight fasting urine pregnanetriol. RECENT FINDINGS: We have conducted a decade-long research project to investigate methods of monitoring 21-OHD in children. Our latest studies on this topic have recently been published. One is a review of methods for monitoring 21-OHD. The other was to demonstrate that measuring the first morning PT level may be more practical and useful for biochemical monitoring of 21-OHD. The first morning pregnanetriol (PT), which was previously reported to reflect a long-term auxological data during the prepubertal period, correlated more significantly than the other timing PT in this study, with 17-OHP, before the morning medication. SUMMARY: In conclusion, although the optimal method of monitoring this disease is still uncertain, the use of overnight fasting urine pregnanetriol (P3) as a marker of 21-OHD is scientifically sound and may be clinically practical.

Intracranial germinoma in the lateral ventricle with polydipsia and polyuria: a case report and literature review.

Central nervous system germ cell tumors (CNSGCTs) are rare neoplasms which usually develop in the midline structures. They are occasionally involved in off-midline structures of the brain. Here, we report an extremely rare case of an intracranial germinoma in the lateral ventricle. The patient was a 10-year-old boy with a 1-year history of polydipsia and polyuria. Brain magnetic resonance imaging (MRI) showed a relatively homogeneously enhancing lesion in the lateral ventricle, and the posterior pituitary gland was not hyperintense on T1-weighted imaging. Subependymoma was suspected, and tumor removal operation was performed; however, because the intraoperative pathological investigation revealed germinoma, we could only perform partial removal of the tumor. Postoperative histology also confirmed germinoma. Then, the patient received chemotherapy, followed by radiation therapy. MRI showed no recurrence for 6 years after treatment. Intracranial germinoma in the lateral ventricle is extremely rare. The diagnosis is occasionally challenging, especially when the tumors are located in atypical locations. This paper presents a literature review of previously described CNSGCTs of the lateral ventricle to improve awareness of CNSGCTs in atypical locations. We also consider the relationship between imaging findings and clinical manifestations.

An elderly patient with 17α-hydroxylase deficiency misdiagnosed as primary aldosteronism: a case report.

BACKGROUND: 17α-hydroxylase deficiency (17OHD) is a rare autosomal recessive disorder. Aldosterone levels are usually low in patients with 17OHD. However, among the approximately 150 cases of 17OHD reported to date, aldosterone levels were not low in all cases. Therefore, some 17OHD cases may have been misdiagnosed as primary aldosteronism (PA) cases. Often before puberty, 17OHD is diagnosed because of abnormal genital morphology and menstrual irregularities. However, we report a very rare case of 17OHD in an elderly patient with a high aldosterone/renin ratio (ARR) similar to that in PA. CASE PRESENTATION: A 63-year-old Japanese woman was transferred to our medical facility for the evaluation of bilateral adrenal hypertrophy, which was incidentally discovered during an abdominal examination after cholecystectomy. The patient had hypokalemia and a high aldosterone/renin ratio. Her medical history included hypertension and right intracerebral capsular hemorrhage at the age of 30 years. Additional testing revealed low cortisol, high adrenocorticotropic hormone, and low testosterone and dehydroepiandrosterone sulfate, indicating congenital adrenal hyperplasia. Genetic analysis revealed a mutation in the CYP17A1 gene and a karyotype of 46, XY; hence, she was diagnosed with 17OHD. CONCLUSION: 17OHD can resemble PA. The combination of a high ARR and low cortisol level should trigger the consideration of 17OHD.

Current status of transition medicine for 21-hydroxylase deficiency in Japan: from the perspective of pediatric endocrinologists.

To manage of 21-hydroxylase deficiency (21-OHD), transition medicine from pediatric to adult health care is an important process and requires individually optimized approaches. We sent cross-sectional questionnaire surveys on the current status of transition from pediatric to adult health care in 21-OHD patients to all councillors of the Japanese Society for Pediatric Endocrinology. Many pediatric departments (42.2%) experienced adult 21-OHD patients, and 115 patients (53 males, mean age of 26) in 46 institutions were identified. Whereas almost two-thirds of pediatric endocrinologists regarded the problems of counterparts and cooperation as hindrance of transition medicine, the major reason for continuing to be treated in pediatrics was the patient's own request. The prevalence of long-term complications including obesity, osteoporosis, infertility, menstrual disorder, gender dysphoria, and testicular adrenal rest tumor were 27.5%, 8.8%, 11.1%, 26.3%, 7.1%, 12.5%, respectively, which is comparable to those of other cohorts previously reported. However, several items, especially infertility and osteoporosis were not checked well enough in adult 21-OHD patients treated in pediatrics. Though 44 of 62 female patients had genital reconstructive surgery, more than half of them were not followed up by gynecologists or pediatric urologists. Quite a few adult 21-OHD patients had been followed up in pediatrics even after coming of age; however, surveillance by pediatric endocrinologists of gynecological, reproductive, and mental problems may be insufficient. Therefore, multidisciplinary approaches should be required in transition medicine for 21-OHD and prerequisite for graduation of pediatrics. Pediatric endocrinologists will need to play a leading role in the development of transition systems.

Iatrogenic Cushing syndrome due to topical corticosteroids after skin graft surgery for giant nevus: A case report.

Iatrogenic Cushing syndrome (ICS) may be caused by exposure to corticosteroid therapy. We report a case of ICS caused by topical corticosteroids applied to the skin after skin graft surgery. We presume that the development of ICS, in this case, was attributed to a combination of skin barrier disruption, usage of high-potency corticosteroids, broad application area/ large quantity of the corticosteroid, and the extended treatment period. Patients may be at risk for ICS after extensive skin graft surgery if large amounts of very potent topical corticosteroids are applied for an extended period to prevent scar formation or relieve itching.

Establishing intellectual disability as the key feature of patients with biallelic RNPC3 variants.

Some mammalian genes contain both major and minor introns, the splicing of which require distinctive major and minor spliceosomes, respectively; these genes are referred to as minor intron containing-genes. RNPC3 (RNA-binding domain-containing protein 3) is one of the proteins that are unique to the minor spliceosome U11/U12 di-snRNP. Only two families with biallelic pathogenic variants in the RNPC3 gene encoding the protein have been reported so far, and the affected members in both families had proportional short stature. While the affected members of the originally identified family did not have intellectual disability, the patients from the other family exhibited intellectual disability. Here, we report on a patient with severe primordial microcephalic dwarfism and intellectual disability who carried compound heterozygous variants in RNPC3 (NM_017619.3): c.261dup, p.Leu88Thrfs*11 and c.1228T>G, p.Phe410Val. The single nucleotide substitution c.1228T>G had a very high predictive score for pathogenicity: the p.Phe410 residue is highly conserved down to fish. Based on ACMG (American College of Medical Genetics and Genomics) guideline, this non-synonymous variant was scored as likely pathogenic. This documentation of yet another patient with biallelic RNPC3 variants exhibiting intellectual disability lends further support to the notion that intellectual disability is a key feature of the spectrum of RNPC3-related disorders.

Identification of the first promoter-specific gain-of-function SOX9 missense variant (p.E50K) in a patient with 46,XX ovotesticular disorder of sex development.

SOX9, a transcription factor, is expressed in the undifferentiated XX and XY gonads. SRY induces significant upregulation of SOX9 expression in XY gonads. Loss-of-function SOX9 variants cause testicular dysgenesis in 46,XY patients, while duplication of the total gene or the upstream regulatory region results in testicular development in 46,XX patients. However, gain-of-function (GoF) SOX9 variants have not been reported previously. We report the case of a 16-year-old female patient with a 46,XX karyotype who had masculinized external genitalia and unilateral ovotestis. Next-generation sequencing-based genetic screening for disorders of sex development led to the identification of a novel SOX9 variant (p.Glu50Lys), transmitted from the phenotypically normal father. Expression analysis showed that E50K-SOX9 enhanced transactivation of the luciferase reporter containing the testis enhancer sequence core element compared with that containing the wildtype-SOX9. This GoF activity was not observed in the luciferase reporter containing Amh, the gene for anti-Müllerian hormone. We genetically engineered female mice (Sox9E50K/E50K ), and they showed no abnormalities in the external genitalia or ovaries. In conclusion, a novel SOX9 variant with a promoter-specific GoF activity was identified in vitro; however, the disease phenotype was not recapitulated by the mouse model. At present, the association between the GoF SOX9 variant and the ovotestis phenotype remains unclear. Future studies are needed to verify the possible association.

High-power 813-nm Tm3+-doped ZBLAN fiber MOPA with signal-light photodarkening suppression.

The photodarkening suppression effect of an 813-nm Tm3+-doped ZBLAN fiber amplifier has been investigated. We have experimentally observed that a photodarkened fiber can be bleached by an 813-nm light and that the photoinduced loss during the amplifier operation is effectively suppressed with the help of the high-power signal. To the best of our knowledge, this is the first investigation of the photodarkening suppression by using a higher power signal in Tm3+-doped fiber amplifiers. Based on these signal photodarkening suppression effects, we have designed a multistage fiber amplifier, and demonstrated the stable operation of the 1.15-W fiber master oscillator power amplifier at 813 nm without additional photobleaching light.

Quantitative monitoring of circulating tumor DNA in patients with advanced pancreatic cancer undergoing chemotherapy.

According to cancer genome sequences, more than 90% of cases of pancreatic ductal adenocarcinoma (PDAC) harbor active KRAS mutations. Digital PCR (dPCR) enables accurate detection and quantification of rare mutations. We assessed the dynamics of circulating tumor DNA (ct-DNA) in patients with advanced PDAC undergoing chemotherapy using dPCR. KRAS G12/13 mutation was assayed by dPCR in 47 paired tissue- and ct-DNA samples. The 21 patients were subjected to quantitative ct-DNA monitoring at 4 to 8-week intervals during chemotherapy. KRAS mutation was detected in 45 of those 47 patients using tissue DNA. In the KRAS mutation-negative cases, next-generation sequencing revealed KRAS Q61K and NRAS Q61R mutations. KRAS mutation was detected in 23/45 cases using ct-DNA (liver or lung metastasis, 18/19; mutation allele frequency [MAF], 0.1%-31.7%; peritoneal metastasis, 3/9 [0.1%], locally advanced, 2/17 [0.1%-0.2%]). In the ct-DNA monitoring, the MAF value changed in concordance with the disease state. In the 6 locally advanced cases, KRAS mutation appeared concurrently with liver metastasis. Among the 6 cases with liver metastasis, KRAS mutation disappeared during the duration of stable disease or a partial response, and reappeared at the time of progressive disease. The median progression-free survival was longer in cases in which KRAS mutation disappeared after an initial course of chemotherapy than in those in which it was continuously detected (248.5 vs 50 days, P < .001). Therefore, ct-DNA monitoring enables continuous assessment of disease state and could have prognostic utility during chemotherapy.

DNA Methylation Status of SHOX-Flanking CpG Islands in Healthy Individuals and Short Stature Patients with Pseudoautosomal Copy Number Variations.

SHOX resides in the short arm pseudoautosomal region (PAR1) of the sex chromosomes and escapes X inactivation. SHOX haploinsufficiency underlies idiopathic short stature (ISS) and Leri-Weill dyschondrosteosis (LWD). A substantial percentage of cases with SHOX haploinsufficiency arise from pseudoautosomal copy number variations (CNVs) involving putative enhancer regions of SHOX. Our previous study using peripheral blood samples showed that some CpG dinucleotides adjacent to SHOX exon 1 were hypomethylated in a healthy woman and methylated in a woman with gross X chromosomal rearrangements. However, it remains unknown whether submicroscopic pseudoautosomal CNVs cause aberrant DNA methylation of SHOX-flanking CpG islands. In this study, we examined the DNA methylation status of SHOX-flanking CpG islands in 50 healthy individuals and 10 ISS/LWD patients with pseudoautosomal CNVs. In silico analysis detected 3 CpG islands within the 20-kb region from the translation start site of SHOX. Pyrosequencing and bisulfite sequencing of genomic DNA samples revealed that these CpG islands were barely methylated in peripheral blood cells and cultured chondrocytes of healthy individuals, as well as in peripheral blood cells of ISS/LWD patients with pseudoautosomal CNVs. These results, in conjunction with our previous findings, indicate that the DNA methylation status of SHOX-flanking CpG islands can be affected by gross X-chromosomal abnormalities, but not by submicroscopic CNVs in PAR1. Such CNVs likely disturb SHOX expression through DNA methylation-independent mechanisms, which need to be determined in future studies.

Effects of pre- and post-pubertal dihydrotestosterone treatment on penile length in 5α-reductase type 2 deficiency.

Steroid 5α-reductase type 2 deficiency (5αRD2) is a congenital disorder of sex development caused by impairment of conversion from testosterone (T) to 5α-dihydrotestosterone (DHT). DHT deficiency leads to various degrees of undervirilized external genitalia including micropenis, primarily correlated with mutations of the SRD5A2 gene that encodes 5α-reductase type 2. Four Japanese boys with isolated micropenis were diagnosed as 5αRD2 by elevated ratios of serum T/DHT, and decreased ratios of urinary 5α/5ß-reduced steroid metabolites. Genetic analyses for SRD5A2 identified that the four patients shared a hypomorphic mutation R227Q that has a residual activity related to the mild-form of 5αRD2. For prepubertal micropenis, DHT was transdermally applied to the four patients at the ages of 4-11 year, increasing a median of stretched penile lengths (SPLs) from 2.6 cm (-2.5 SD) to 4.4 cm (-0.2 SD). Nevertheless, the post-pubertal penile growth was apparently retarded, despite normal levels of T secreted from well-developed testes. The second course of DHT treatment underwent at ages of 12-18 year, but unable to normalize SPLs at a range of 6.0 to 7.0 cm (-3.4 to -2.4 SD). The prostate volumes of two patients were variable at 8.1 and 21 cm3, and a sperm cell count of one patient was normal as young adult. DHT treatment contributes to development of the penis and prostate, which are favorable for the potential fertility of 5αRD2 adults. Meanwhile, the retarded penile growth and a risk of prostate overgrowth may complicate the post-pubertal management with DHT for 5αRD2 males.

Prefrontal modulation during chewing performance in occlusal dysesthesia patients: a functional near-infrared spectroscopy study.

OBJECTIVES: Neuropsychological associations can be considerable in occlusal dysesthesia (OD) patients who routinely complain of persistent occlusal discomfort, and somatization effects in the superior medial prefrontal cortex and the temporal and parietal regions are also present. However, the relationship between physical activity, i.e., chewing, prefrontal cognitive demand, and psychiatric states in OD patients remains unclear. We investigated this relationship in this study. MATERIALS AND METHODS: OD patients (n = 15) and healthy control (n = 15; HC) subjects were enrolled in this study. Occlusal contact, chewing activities of the masticatory muscles, prefrontal activities, and psychiatric states such as depression and somatization, of the participants were evaluated. Functional near-infrared spectroscopy was used to determine prefrontal hemodynamics and the Symptom Checklist-90-R was used to score the psychiatric states. RESULTS: We observed a significant association between prefrontal deactivation during chewing and somatization subscales in OD patients. Further, there were no significant differences with regard to the occlusal state and chewing physical activities between the OD patients and HC subjects. CONCLUSIONS: Chewing-related prefrontal deactivation may be associated with somatization severity in OD patients. CLINICAL RELEVANCE: fNIRS is a functional imaging method that uses the principal of neuro-vascular couplings. It is applicable for evaluation of psychiatric state based on prefrontal cortex blood flow in patients with psychiatric disorders.

Subpopulations of vomeronasal sensory neurons with coordinated coexpression of type 2 vomeronasal receptor genes are differentially dependent on Vmn2r1.

The mouse vomeronasal organ is specialized in the detection of pheromones. Vomeronasal sensory neurons (VSNs) express chemosensory receptors of two large gene repertoires, V1R and V2R, which encode G-protein-coupled receptors. Phylogenetically, four families of V2R genes can be discerned as follows: A, B, C, and D. VSNs located in the basal layer of the vomeronasal epithelium coordinately coexpress V2R genes from two families: Approximately half of basal VSNs coexpress Vmn2r1 of family C with a single V2R gene of family A8-10, B, or D ('C1 type of V2Rs'), and the other half coexpress Vmn2r2 through Vmn2r7 of family C with a single V2R gene of family A1-6 ('C2 type V2Rs'). The regulatory mechanisms of the coordinated coexpression of V2Rs from two families remain poorly understood. Here, we have generated two mouse strains carrying a knockout mutation in Vmn2r1 by gene targeting in embryonic stem cells. These mutations cause a differential decrease in the numbers of VSNs expressing a given C1 type of V2R. There is no compensatory expression of Vmn2r2 through Vmn2r7. VSN axons coalesce into glomeruli in the appropriate region of the accessory olfactory bulb in the absence of Vmn2r1. Gene expression profiling by NanoString reveals a differential and graded decrease in the expression levels across C1 type of V2Rs. There is no change in the expression levels of C2 type of V2Rs, with two exceptions that we reclassified as C1 type. Thus, there appears to be a fixed probability of gene choice for a given C2 type of V2R.

Discordant fetal phenotype of hypophosphatasia in two siblings.

Hypophosphatasia (HPP) is an autosomal recessive metabolic disorder with impaired bone mineralization due to mutations in the ALPL gene. The genotype-phenotype correlation of this disorder has been widely described. Here, we present two affected siblings, whose fetal phenotypes were discordant. A 31-year-old Japanese woman, G0P0, was referred to our institution because of fetal micromelia. After obstetric counseling, the pregnancy was terminated at 21 weeks' gestation. Post-mortem radiographs demonstrated severely defective mineralization of the skeleton. The calvarial, spinal, and tubular bones were mostly missing. Only the occipital bones, mandible, clavicles, ribs, one thoracic vertebra, ilia, and tibia were relatively well ossified. The radiological findings suggested lethal HPP. Genetic testing for genomic DNA extracted from the umbilical cord identified compound heterozygous mutations in the ALPL gene (c.532T>C, p.Y178H; c.1559delT, p.Leu520Argfs*86). c.532T>C was a novel variant showing no residual activity of the protein by the functional analysis. The parents were heterozygous carriers. In the next pregnancy, biometric values on fetal ultrasonography at 20 and 26 weeks' gestation were normal. At 34 weeks, however, a small chest and shortening of distal long bones came to attention. The neonate delivered at 41 weeks showed serum ALP of <5U/L. Radiological examination showed only mild thoracic hypoplasia and metaphyseal mineralization defects of the long bones. ALP replacement therapy was introduced shortly after birth, and the neonate was discharged at day 22 without respiratory distress. Awareness of discordant fetal phenotypes in siblings with HPP precludes a diagnostic error, and enables early medical intervention to mildly affected neonates.

Cover Image, Volume 176A, Number 1, January 2018.

The cover image, by Satoru Ikenoue et al., is based on the Clinical Report Discordant fetal phenotype of hypophosphatasia in two siblings, DOI: 10.1002/ajmg.a.38531.