Tridimensional Visualization and Analysis of Early Human Development.

Generating a precise cellular and molecular cartography of the human embryo is essential to our understanding of the mechanisms of organogenesis in normal and pathological conditions. Here, we have combined whole-mount immunostaining, 3DISCO clearing, and light-sheet imaging to start building a 3D cellular map of the human development during the first trimester of gestation. We provide high-resolution 3D images of the developing peripheral nervous, muscular, vascular, cardiopulmonary, and urogenital systems. We found that the adult-like pattern of skin innervation is established before the end of the first trimester, showing important intra- and inter-individual variations in nerve branches. We also present evidence for a differential vascularization of the male and female genital tracts concomitant with sex determination. This work paves the way for a cellular and molecular reference atlas of human cells, which will be of paramount importance to understanding human development in health and disease. PAPERCLIP.

Spectrum of Diabetic Neuropathy: New Insights in Diagnosis and Treatment.

Diabetic neuropathy is a highly prevalent complication of diabetes. It consists of a broad range of neuropathic conditions, such as distal symmetric polyneuropathy and various forms of autonomic neuropathies involving the cardiovascular, gastrointestinal, and urogenital systems. Prevention or diagnosis in early stages of disease is crucial to prevent symptomatic onset and progression, particularly in the absence of current disease-modifying therapies. In this review, we describe the four main types of diabetic neuropathy. We review current understanding with respect to diagnosis and treatment while highlighting knowledge gaps and future directions.

Biased precursor ingression underlies the center-to-pole pattern of male sex determination in mouse.

During mammalian development, gonadal sex determination results from the commitment of bipotential supporting cells to Sertoli or granulosa cell fates. Typically, this decision is coordinated across the gonad to ensure commitment to a single organ fate. When unified commitment fails in an XY mouse, an ovotestis forms in which supporting cells in the center of the gonad typically develop as Sertoli cells, while supporting cells in the poles develop as granulosa cells. This central bias for Sertoli cell fate was thought to result from the initial expression of the drivers of Sertoli cell fate, SRY and/or SOX9, in the central domain, followed by paracrine expansion to the poles. However, we show here that the earliest cells expressing SRY and SOX9 are widely distributed across the gonad. In addition, Sertoli cell fate does not spread among supporting cells through paracrine relay. Instead, we uncover a center-biased pattern of supporting cell precursor ingression that occurs in both sexes and results in increased supporting cell density in the central domain. Our findings prompt a new model of gonad patterning in which a density-dependent organizing principle dominates Sertoli cell fate stabilization.

Human prostate organoid generation and the identification of prostate development drivers using inductive rodent tissues.

The reactivation of developmental genes and pathways during adulthood may contribute to pathogenesis of diseases such as prostate cancer. Analysis of the mechanistic links between development and disease could be exploited to identify signalling pathways leading to disease in the prostate. However, the mechanisms underpinning prostate development require further characterisation to interrogate fully the link between development and disease. Previously, our group developed methods to produce prostate organoids using induced pluripotent stem cells (iPSCs). Here, we show that human iPSCs can be differentiated into prostate organoids using neonatal rat seminal vesicle mesenchyme in vitro. The organoids can be used to study prostate development or modified to study prostate cancer. We also elucidated molecular drivers of prostate induction through RNA-sequencing analyses of the rat urogenital sinus and neonatal seminal vesicles. We identified candidate drivers of prostate development evident in the inductive mesenchyme and epithelium involved with prostate specification. Our top candidates included Spx, Trib3, Snai1, Snai2, Nrg2 and Lrp4. This work lays the foundations for further interrogation of the reactivation of developmental genes in adulthood, leading to prostate disease.

In vitro induction of prostate buds from murine urogenital epithelium in the absence of mesenchymal cells.

The prostate is a male reproductive gland which secretes prostatic fluid that enhances male fertility. During development and instigated by fetal testosterone, prostate cells arise caudal to the bladder at the urogenital sinus (UGS), when the urogenital mesenchyme (UGM) secretes signals to the urogenital epithelium (UGE). These initial mesenchymal signals induce prostate-specific gene expression in the UGE, after which epithelial progenitor cells form prostatic buds. Although many important factors for prostate development have been described using UGS organ cultures, those necessary and sufficient for prostate budding have not been clearly identified. This has been in part due to the difficulty to dissect the intricate signaling and feedback between epithelial and mesenchymal UGS cells. In this study, we separated the UGM from the UGE and tested candidate growth factors to show that when FGF10 is present, testosterone is not required for initiating prostate budding from the UGE. Moreover, in the presence of low levels of FGF10, canonical WNT signaling enhances the expression of several prostate progenitor markers in the UGE before budding of the prostate occurs. At the later budding stage, higher levels of FGF10 are required to increase budding and retinoic acid is indispensable for the upregulation of prostate-specific genes. Lastly, we show that under optimized conditions, female UGE can be instructed towards a prostatic fate, and in vitro generated prostate buds from male UGE can differentiate into a mature prostate epithelium after in vivo transplantation. Taken together, our results clarify the signals that can induce fetal prostate buds in the urogenital epithelium in the absence of the surrounding, instructive mesenchyme.

Archaea in the Human Microbiome and Potential Effects on Human Infectious Disease.

Archaea represent a separate domain of life, next to bacteria and eukarya. As components of the human microbiome, archaea have been associated with various diseases, including periodontitis, endodontic infections, small intestinal bacterial overgrowth, and urogenital tract infections. Archaea are generally considered nonpathogenic; the reasons are speculative because of limited knowledge and gene annotation challenges. Nevertheless, archaeal syntrophic principles that shape global microbial networks aid both archaea and potentially pathogenic bacteria. Evaluating archaea interactions remains challenging, requiring clinical studies on inflammatory potential and the effects of archaeal metabolism. Establishing a culture collection is crucial for investigating archaea functions within the human microbiome, which could improve health outcomes in infectious diseases. We summarize potential reasons for archaeal nonpathogenicity, assess the association with infectious diseases in humans, and discuss the necessary experimental steps to enable mechanistic studies involving archaea.

Concerted morphogenesis of genital ridges and nephric ducts in the mouse captured through whole-embryo imaging.

During development of the mouse urogenital complex, the gonads undergo changes in three-dimensional structure, body position and spatial relationship with the mesonephric ducts, kidneys and adrenals. The complexity of genital ridge development obscures potential connections between morphogenesis and gonadal sex determination. To characterize the morphogenic processes implicated in regulating gonad shape and fate, we used whole-embryo tissue clearing and light sheet microscopy to assemble a time course of gonad development in native form and context. Analysis revealed that gonad morphology is determined through anterior-to-posterior patterns as well as increased rates of growth, rotation and separation in the central domain that may contribute to regionalization of the gonad. We report a close alignment of gonad and mesonephric duct movements as well as delayed duct development in a gonad dysgenesis mutant, which together support a mechanical dependency linking gonad and mesonephric duct morphogenesis.

Changes in microbial composition and interaction patterns of female urogenital tract and rectum in response to HPV infection.

BACKGROUND: Previous studies have shown that changes in the microbial community of the female urogenital tract are associated with Human papillomavirus (HPV) infection. However, research on this association was mostly focused on a single site, and there are currently few joint studies on HPV infection and multiple sites in the female urogenital tract. METHODS: We selected 102 healthy women from Yunnan Province as the research object, collected cervical exfoliation fluid, vaginal, urethral, and rectal swabs for microbial community analysis, and measured bacterial load, and related cytokine content. The link between HPV, microbiota, and inflammation was comprehensively evaluated using bioinformatics methods. FINDINGS: The impact of HPV infection on the microbial composition of different parts varies. We have identified several signature bacterial genera that respond to HPV infection in several detection sites, such as Corynebacterium, Lactobacillus, Campylobacter, and Cutibacterium have been detected in multiple sites, reflecting their potential significance in cross body sites HPV infection responses. There was a solid microbial interaction network between the cervix, vagina, and urethra. The interrelationships between inflammatory factors and different bacterial genera might also affect the immune system's response to HPV infection. INTERPRETATION: It might be an effective strategy to prevent and treat HPV infection by simultaneously understanding the correlation between the microbial changes in multiple parts of the female urogenital tract and rectum and HPV infection, and controlling the microbial network related to HPV infection in different parts.

Delphi Consensus on Diagnostic Criteria for LUMBAR Syndrome.

OBJECTIVE: To develop consensus on diagnostic criteria for LUMBAR syndrome, the association of segmental infantile hemangiomas that affect the Lower body with Urogenital anomalies, Ulceration, spinal cord Malformations, Bony defects, Anorectal malformations, Arterial anomalies and/or Renal anomalies. STUDY DESIGN: These diagnostic criteria were developed by an expert multidisciplinary and multi-institutional team based on analysis of peer-reviewed data, followed by electronic-Delphi consensus of a panel of 61 international pediatric specialists. RESULTS: After 2 Delphi rounds, a 92% or higher level of agreement was reached for each Delphi statement. 98% of panelists agreed with the diagnostic criteria, and 100% agreed the criteria would be useful in clinical practice. The diagnosis of LUMBAR requires the presence of a segmental, or patterned, infantile hemangioma of the lumbosacral, sacrococcygeal, or pelvic cutaneous regions plus one additional criterion of the urogenital, spinal, bony, anorectal, arterial, or renal organ systems. CONCLUSIONS: These diagnostic criteria will enhance clinical care by improving screening, detection, and overall awareness of this poorly understood neurocutaneous disorder. The criteria can be utilized by a wide variety of pediatric subspecialists. In addition, formal criteria will improve phenotypic uniformity among LUMBAR syndrome cohorts and a patient registry, allowing investigators to assess clinical features, long-term outcomes, and results of genetic sequencing in a standardized manner. Finally, these criteria will serve as a starting point for prospective studies to establish formal screening and management guidelines.

Transrectal ultrasound for intraoperative interstitial needle guidance in cervical cancer brachytherapy.

OBJECTIVE: This study aimed to prospectively assess the visibility of interstitial needles on transrectal ultrasound (TRUS) in cervical cancer brachytherapy patients and evaluate its impact on implant and treatment plan quality. MATERIAL AND METHODS: TRUS was utilized during and after applicator insertion, with each needle's visibility documented through axial images at the high-risk clinical target volume's largest diameter. Needle visibility on TRUS was scored from 0 (no visibility) to 3 (excellent discrimination, margins distinct). Quantitative assessment involved measuring the distance between tandem and each needle on TRUS and comparing it to respective magnetic resonance imaging (MRI) measurements. Expected treatment plan quality based on TRUS images was rated from 1 (meeting all planning objectives) to 4 (violation of High-risk clinical target volume (CTVHR) and/or organ at risk (OAR) hard constraints) and compared to the final MRI-based plan. RESULTS: Analysis included 23 patients with local FIGO stage IB2-IVA, comprising 41 applications with a total of 230 needles. A high visibility rate of 99.1% (228/230 needles) was observed, with a mean visibility score of 2.5 ±â€¯0.7 for visible needles. The maximum and mean difference between MRI and TRUS measurements were 8 mm and -0.1 ±â€¯1.6 mm, respectively, with > 3 mm discrepancies in 3.5% of needles. Expected treatment plan quality after TRUS assessment exactly aligned with the final MRI plan in 28 out of 41 applications with only minor deviations in all other cases. CONCLUSION: Real-time TRUS-guided interstitial needle placement yielded high-quality implants, thanks to excellent needle visibility during insertion. This supports the potential of TRUS-guided brachytherapy as a promising modality for gynecological indications.

Female non-obstetric urogenital fistula repair: long-term patient-reported outcomes and a scoping literature review.

OBJECTIVE: To investigate long-term and patient-reported outcomes, including sexual function, in women undergoing urogenital fistula (UGF) repair, addressing the lack of such data in Western countries, where fistulas often result from iatrogenic causes. PATIENTS AND METHODS: We conducted a retrospective analysis at a tertiary referral centre (2010-2023), classifying fistulas based on World Health Organisation criteria and evaluating surgical approaches, aetiology, and characteristics. Both objective (fistula closure, reintervention rates) and subjective outcomes (validated questionnaires) were assessed. A scoping review of patient-reported outcome measures in UGF repair was also performed. RESULTS: The study included 50 patients: 17 (34%) underwent transvaginal and 33 (66%) transabdominal surgery. History of hysterectomy was present in 36 patients (72%). The median (interquartile range [IQR]) operating time was 130 (88-148) min. Fistula closure was achieved in 94% of cases at a median (IQR) follow-up of 50 (16-91) months and reached 100% after three redo fistula repairs. Seven patients (14%) underwent reinterventions for stress urinary incontinence after transvaginal repair (autologous fascial slings). Patient-reported outcomes showed median (IQR) scores on the International Consultation on Incontinence Questionnaire Female Lower Urinary Tract Symptoms Modules (ICIQ-FLUTS) of 5 (3-7) for filling symptoms, 1 (0-2) for voiding symptoms and 4.5 (1-9) for incontinence symptoms. The median (IQR) score on the ICIQ Female Sexual Matters Associated with Lower Urinary Tract Symptoms Module (ICIQ-FLUTSsex) was 3 (1-5). The median (IQR) ICIQ Satisfaction (ICIQ-S) outcome score and overall satisfaction with surgery item score was 22 (18.5-23.5) and 10 (8.5-10), respectively. Higher scores indicate higher symptom burden and treatment satisfaction, respectively. Our scoping review included 1784 women, revealing mixed aetiology and methodological and aetiological heterogeneity, thus complicating cross-study comparisons. CONCLUSIONS: Urogenital fistula repair at a specialised centre leads to excellent outcomes and high satisfaction. Patients with urethrovaginal fistulas are at increased risk of stress urinary incontinence, possibly due to the original trauma site of the fistula.

The utility of Martius fat pad in the repair of urogenital fistulae: review of current evidence.

OBJECTIVE: To present the contemporary evidence on transvaginal urogenital fistulae (UGF) repair with Martius fat pad (MFP), compared to direct graftless fistula repair. METHODS: We reviewed all available studies reporting lower UGF repair via the transvaginal approach in MEDLINE, Embase, and Cochrane Central Register of Controlled Trials (CENTRAL). The primary outcome of interest was the fistula closure rates. When available, patients' baseline characteristics, indications for surgery, and early and late postoperative complications with focus on MFP-related complications are reported. RESULTS AND DISCUSSION: In obstetric fistulae, tissue interposition has been almost completely abandoned, with contemporary large series reporting closure rates of >90% with graftless repair, even for complex fistulae. Similarly, most simple, non-irradiated iatrogenic fistulae can be closed safely without or with tissue interposition with success rates ranging between 86% and 100%. However, MFP is valuable in fistulae with difficulty achieving tension-free and layered closure, with significant tissue loss, urethral involvement and with poorly vascularised tissues after radiotherapy, with reported success rates between 80% and 97% in those challenging situations. CONCLUSION: A UGF repair should be individualised after considering the specific characteristics and complexity of the procedure. MFP interposition is probably unnecessary for the majority of low (obstetric) fistulae within otherwise healthy tissues. However, MFP may still have a place to maximise outcomes in low-income settings, in select cases with higher (iatrogenic) fistulae, and in most cases with radiotherapy.

Prophylactic application of vaginal lactic acid bacteria against urogenital pathogens and its prospective use in sanitary suppositories.

Beneficial and pathogenic microbes coexist in the vaginal canal, where a diminishing population of lactic acid bacteria may cause recurring urogenital infections. Probiotic bacteria Lactobacillus crispatus, Lactobacillus gasseri, Lactobacillus vaginalis, and pathogenic microbes Enterococcus faecalis, Enterobacter cloacae, Shigella sp., Staphylococcus epidermidis, and Escherichia fergusonii were isolated from vaginal swabs. Lactobacillus sp. and their probiotic culture free supernatant (PCFS) inhibited the growth of the above-mentioned urogenital pathogens. L. crispatus produced both lactic acid and hydrogen peroxide, exhibiting the best antimicrobial potential against the studied pathogens. Lyophilized L. crispatus had a shelf life of 12 months and the lyophilized PCFS also retained its antibacterial property with a minimum inhibition concentration of 1 µg/µL. Carboxy-methyl cellulose-alginate, a green alternative to super-absorbent polymers, was encapsulated with L. crispatus cells. The probiotic in its encapsulated state retained its viability for 21 days, and the bead showed 30% solvent absorptive capacity. PCFS-laced non-woven fabric displayed antibacterial property with no change in its physicochemical properties. These probiotic and postbiotic formulations have excellent prophylactic potential for urogenital infections. Such formulations can be exploited as additives in sanitary suppositories to enhance vaginal health.

Pediatric urogenital schistosomiasis diagnosed in France.

BACKGROUND: Schistosomiasis affects approximately 230 million people worldwide. There is an increased incidence of schistosomiasis cases in France acquired from outside the country. This increases the risk of schistosomiasis outbreaks as observed in Corsica. Clinicians from non-endemic regions are not accustomed to diagnosing and managing this pathology. The objective of this study is to provide a better description of the clinical and paraclinical characteristics and disease evolution of affected children. METHODS: Through the French Pediatric Nephrology Society and the Pediatric Infectious Pathology Group, we contacted all French pediatric centers that may have treated children with urinary schistosomiasis between 2013 and 2019. Age, sex, comorbidities, and clinical, biological, and radiological data (at discovery and follow-up) were collected retrospectively. RESULTS: A total of 122 patients from 10 different centers were included. The median age was 14 years and the sex ratio M/F was 4:1. Hematuria was present in 82% of the patients while urinary tract abnormality was found in 36% of them. Fourteen patients (11%) displayed complicated forms of urinary schistosomiasis including 10 patients with chronic kidney disease. A total of 110 patients received treatment with praziquantel, which was well-tolerated and led to clinical resolution of symptoms in 98% of cases. CONCLUSION: Patients with schistosomiasis present frequent kidney, urinary, or genital involvement. Systematic screening of patients returning from endemic areas is therefore recommended, especially since treatment with antiparasitic drugs is effective and well-tolerated. Enhancing medical knowledge of this pathology among all practitioners is essential to improve care and outcomes.

The incidence of major bleeding in adult patients with urogenital and gynecological cancer being treated with direct oral anticoagulants (DOACs): a systematic review.

BACKGROUND: Direct oral anticoagulants (DOACs) are the mainstay of treatment for venous thromboembolism (VTE) and non-valvular atrial fibrillation (AF), with or without an underlying cancer. Patients with cancer have a 2-3-fold increase in risk for bleeding complications compared to non-cancer patients taking anticoagulant therapy, however the incidence of bleeding for urogenital and gynecological cancers on DOACs are uncertain. AIMS: To assess the bleeding risk associated with the use of DOACs in patients with urogenital and/or gynecological cancers. METHOD: We conducted a systematic review of randomized controlled trials (RCTs) and prospective cohort studies to address the safety of DOACs for VTE and AF when used in patients with urogenital and/or gynecological malignancy. The primary outcomes assessed were major and clinically relevant non-major (CRNMB) bleeding, with minor bleeding considered as a secondary outcome. MEDLINE, EMBASE and COCHRANE Central Registry of Controlled Trials were searched up to and including Oct 28, 2022. The study protocol was registered in PROSPERO (CRD42022370981). Studies were independently assessed for inclusion and data extracted in duplicate. RESULT: Seven studies met our inclusion criteria (Fig. 1): 2 RCTs and 5 prospective cohort studies. A total of 676 patients treated with DOACs were included, 628 (92.8%) had VTE and 48 (7.1%) had AF. In patients with VTE treated with DOACs, the pooled major bleeding rate was 2.1%, 95% confidence intervals (CI) 0.9-3.3% (Fig. 2). Pooled estimates could not be determined for AF patients given small event and patient numbers. CONCLUSION: Major bleeding rates in urogenital and/or gynecological cancer patients treated with DOACs are similar to that of the general cancer population.

Female Urethroplasty with a Buccal Mucosa Graft using a Supraurethral Approach.

INTRODUCTION AND HYPOTHESIS: In the setting of recurrent female urethral stricture, urethroplasty offer the best chance of cure. However, which approach (dorsal or ventral) and which tissue (buccal mucosa, vaginal graft, vaginal flap) remain areas of controversy. In this article and accompanying video, we describe female urethroplasty with a supraurethral approach using a buccal mucosa graft. METHODS: A stricture of 3 cm in length was observed in the mid urethra. A supraurethral semi-lunar incision was made and dissection was performed up to the stricture. A dorsal urethrotomy was performed and a 3 × 2 cm oral mucosal graft was harvested from the left cheek. The mucosal graft was anastomosed to both urethral edges with running sutures. The graft was fixed to the supraurethral tissue with quilting sutures. A urethral catheter and a suprapubic catheter were left in place for 3 weeks. RESULTS: Following removal of the catheters, the patient was able to void satisfactorily with no incontinence. No complications were observed in the urethral area or at the graft harvest site. CONCLUSIONS: Buccal mucosa graft urethroplasty with a supraurethral approach is a reliable method in the treatment of female urethral stricture.

Incidence of Urinary Incontinence During the Postpartum Period in Rajavithi Hospital: A Prospective Cohort Study.

INTRODUCTION AND HYPOTHESIS: The objective was to study the incidence of urinary incontinence (UI), associated risk factors and quality of life (QOL) in postpartum women. METHODS: A prospective study was conducted with 406 postpartum women at Rajavithi Hospital and followed up over the phone between June 2020 and September 2021. Inclusion criteria were singleton pregnant women aged 18-45 years, and gestational age ≥ 37 weeks. Baseline characteristics (age, body mass index, birthweight, gestational age, parity, delivery type, smoking, and alcohol and caffeine intake) were recorded. UI was defined as a score ≥ 16.7% using the Urogenital Distress Inventory. Incontinence-related QOL was evaluated using the Incontinence Impact Questionnaire: a score of ≥ 70 indicated poor QOL. Outcomes were assessed during the postpartum period at 2 days, 6 weeks, 3 months, and 6 months. Multivariate logistic regression was used to analyze risk factors for UI. RESULTS: The incidence of self-reported UI at 2 days, 6 weeks, 3 months, and 6 months postpartum were 39%, 3%, 1%, and 0% respectively. Caffeine consumption during pregnancy was only a risk factor for UI (adjusted RR 1.61, 95%CI 1.27-2.05, p < 0.001) after adjusting for age, BMI, birthweight, parity, delivery type, alcohol, smoking, and pelvic floor exercise. Three women with UI had poor QOL, whereas all women without UI reported a good QOL. CONCLUSION: In our study sample, urinary incontinence was found in one-third of women during the early postpartum period, but for most women symptoms improved with the first 6 weeks and all resolved at 6 months. In this study, caffeine consumption during pregnancy was the only risk factor for UI.

Long-term Sexual Function and Quality of Life in Women Following Substitution Urethroplasty.

PURPOSE: To assess the long-term quality of life (QOL) and sexual function (SF) in women who underwent either dorsal on-lay (DO) or ventral inlay (VI) urethroplasty for urethral stricture disease. METHODOLOGY: Between January 2016 and September 2022, women who underwent either dorsal on-lay (DO) or ventral inlay (VI) urethroplasties and had at least a six-month follow-up been included. Using the Female Sexual Function Index (FSFI) and WHO-QOL bref questionnaires, the QOL and SF were evaluated. Scores were compared between the two groups after being examined for internal validity. A sub-group analysis was carried out based on the procedure's success. RESULTS: With follow-up periods ranging from 6 to 86 months, 25 patients who received VI urethroplasty and 10 patients who underwent DO urethroplasty were included. Both scores demonstrated strong internal consistency. The cumulative QOL and FSFI scores were comparable in both groups (p = 0.53 and p = 0.83, respectively). Significantly high scores were noted in the physical health domain (76.5 ± 9.9 vs 62.33 ± 10.97; p = 0.03; (95% CI = 0.72-24.4)) and the environmental domain (75.75 ± 3.84 vs 66.00 ± 4.24; p = 0.01 (95% CI = 2.64-16.85) in patients with successful VI and DO urethroplasties respectively. Addictions, low socioeconomic status and protracted symptom duration were associated with low QOL scores. Old age was related to low FSFI scores. CONCLUSION: Substitution urethroplasty, despite the approach, showed good QOL and SF scores. Long symptom duration, addictions, and poor socioeconomic status were associated with low QOL whereas old age independently influenced low FSFI scores.

Functional Anatomy of Urogenital Hiatus Closure: the Perineal Complex Triad Hypothesis.

INTRODUCTION: Urogenital hiatus enlargement is a critical factor associated with prolapse and operative failure. This study of the perineal complex was performed to understand how interactions among its three structures: the levator ani, perineal membrane, and perineal body-united by the vaginal fascia-work to maintain urogenital hiatus closure. METHODS: Magnetic resonance images from 30 healthy nulliparous women with 3D reconstruction of selected subjects were used to establish overall geometry. Connection points and lines of action were based on perineal dissection in 10 female cadavers (aged 22-86 years), cross sections of 4 female cadavers (aged 14-35 years), and histological sections (cadavers aged 16 and 21 years). RESULTS: The perineal membrane originates laterally from the ventral two thirds of the ischiopubic rami and attaches medially to the perineal body and vaginal wall. The levator ani attaches to the perineal membrane's cranial surface, vaginal fascia, and the perineal body. The levator line of action in 3D reconstruction is oriented so that the levator pulls the medial perineal membrane cranio-ventrally. In cadavers, simulated levator contraction and relaxation along this vector changes the length of the membrane and the antero-posterior diameter of the urogenital hiatus. Loss of the connection of the left and right perineal membranes through the perineal body results in diastasis of the levator and a widened hiatus, as well as a downward rotation of the perineal membrane. CONCLUSION: Interconnections involving the levator ani muscles, perineal membrane, perineal body, and vaginal fascia form the perineal complex surrounding the urogenital hiatus in an arrangement that maintains hiatal closure.

Biallelic ANGPT2 loss-of-function causes severe early-onset non-immune hydrops fetalis.

BACKGROUND: Hydrops fetalis, a pathological fluid accumulation in two or more body compartments, is aetiologically heterogeneous. We investigated a consanguineous family with recurrent pregnancy loss due to severe early-onset non-immune hydrops fetalis. METHODS AND RESULTS: Whole exome sequencing in four fetuses with hydrops fetalis revealed that they were homozygous for the angiopoietin-2 (ANGPT2) variant Chr8 (GRCh37/Hg19): 6385085T>C, NM_001147.2:c.557A>G. The substitution introduces a cryptic, exonic splice site predicted to result in loss of 10 nucleotides with subsequent shift in reading frame, leading to a premature stop codon. RNA analysis in the heterozygous parents demonstrated loss of detectable mutant allele, indicative of loss-of-function via nonsense-mediated mRNA decay. Serum ANGPT2 levels were reduced in the parents. In a pregnancy with a healthy, heterozygous child, transiently increased fetal nuchal translucency was noted. CONCLUSION: Pathogenic heterozygous ANGPT2 missense variants were recently shown to cause autosomal dominant primary lymphoedema. ANGPT2 is a ligand of the TIE1-TIE2 (tyrosine kinase with immunoglobulin-like and epidermal growth factor-like domains 1 and 2) pathway. It is critical to the formation and remodelling of blood and lymphatic vessels and is involved in vessel maintenance. ANGPT2 knockout mice die from generalised lymphatic dysfunction. We show here that a homozygous pathogenic variant causes loss-of-function and results in severe early-onset hydrops fetalis. This is the first report of an autosomal recessive ANGPT2-related disorder in humans.