Single-cell RNA-sequencing analysis of early sea star development.

Echinoderms represent a broad phylum with many tractable features to test evolutionary changes and constraints. Here, we present a single-cell RNA-sequencing analysis of early development in the sea star Patiria miniata, to complement the recent analysis of two sea urchin species. We identified 20 cell states across six developmental stages from 8 hpf to mid-gastrula stage, using the analysis of 25,703 cells. The clusters were assigned cell states based on known marker gene expression and by in situ RNA hybridization. We found that early (morula, 8-14 hpf) and late (blastula-to-mid-gastrula) cell states are transcriptionally distinct. Cells surrounding the blastopore undergo rapid cell state changes that include endomesoderm diversification. Of particular import to understanding germ cell specification is that we never see Nodal pathway members within Nanos/Vasa-positive cells in the region known to give rise to the primordial germ cells (PGCs). The results from this work contrast the results of PGC specification in the sea urchin, and the dataset presented here enables deeper comparative studies in tractable developmental models for testing a variety of developmental mechanisms.

Vasa, a regulator of localized mRNA translation on the spindle.

Localized mRNA translation is a biological process that allows mRNA to be translated on-site, which is proposed to provide fine control in protein regulation, both spatially and temporally within a cell. We recently reported that Vasa, an RNA-helicase, is a promising factor that appears to regulate this process on the spindle during the embryonic development of the sea urchin, yet the detailed roles and functional mechanisms of Vasa in this process are still largely unknown. In this review article, to elucidate these remaining questions, we first summarize the prior knowledge and our recent findings in the area of Vasa research and further discuss how Vasa may function in localized mRNA translation, contributing to efficient protein regulation during rapid embryogenesis and cancer cell regulation.

The LOTUS domain is a conserved DEAD-box RNA helicase regulator essential for the recruitment of Vasa to the germ plasm and nuage.

DEAD-box RNA helicases play important roles in a wide range of metabolic processes. Regulatory proteins can stimulate or block the activity of DEAD-box helicases. Here, we show that LOTUS (Limkain, Oskar, and Tudor containing proteins 5 and 7) domains present in the germline proteins Oskar, TDRD5 (Tudor domain-containing 5), and TDRD7 bind and stimulate the germline-specific DEAD-box RNA helicase Vasa. Our crystal structure of the LOTUS domain of Oskar in complex with the C-terminal RecA-like domain of Vasa reveals that the LOTUS domain occupies a surface on a DEAD-box helicase not implicated previously in the regulation of the enzyme's activity. We show that, in vivo, the localization of Drosophila Vasa to the nuage and germ plasm depends on its interaction with LOTUS domain proteins. The binding and stimulation of Vasa DEAD-box helicases by LOTUS domains are widely conserved.

The embryonic origin of primordial germ cells in the tardigrade Hypsibius exemplaris.

Primordial germ cells (PGCs) give rise to gametes - cells necessary for the propagation and fertility of diverse organisms. Current understanding of PGC development is limited to the small number of organisms whose PGCs have been identified and studied. Expanding the field to include little-studied taxa and emerging model organisms is important to understand the full breadth of the evolution of PGC development. In the phylum Tardigrada, no early cell lineages have been identified to date using molecular markers. This includes the PGC lineage. Here, we describe PGC development in the model tardigrade Hypsibius exemplaris. The four earliest-internalizing cells (EICs) exhibit PGC-like behavior and nuclear morphology. The location of the EICs is enriched for mRNAs of conserved PGC markers wiwi1 (water bear piwi 1) and vasa. At early stages, both wiwi1 and vasa mRNAs are detectable uniformly in embryos, which suggests that these mRNAs do not serve as localized determinants for PGC specification. Only later are wiwi1 and vasa enriched in the EICs. Finally, we traced the cells that give rise to the four PGCs. Our results reveal the embryonic origin of the PGCs of H. exemplaris and provide the first molecular characterization of an early cell lineage in the tardigrade phylum. We anticipate that these observations will serve as a basis for characterizing the mechanisms of PGC development in this animal.

Hypoxic perfusion of pulmonary arterial vasa vasorum increases pulmonary arterial pressure.

The pathophysiology of pulmonary hypertension (PH) is not fully understood. Here, we tested the hypothesis that hypoxic perfusion of the vasa vasorum of the pulmonary arterial (PA) wall causes PH. Young adult pig lungs were explanted and placed into a modified ex vivo lung perfusion unit (organ care system, OCS) allowing the separate adjustment of parameters for mechanical ventilation, as well as PA perfusion and bronchial arterial (BA) perfusion. The PA vasa vasorum are branches of the BA. The lungs were used either as the control group (n = 3) or the intervention group (n = 8). The protocol for the intervention group was as follows: normoxic ventilation and perfusion (steady state), hypoxic BA perfusion, steady state, and hypoxic BA perfusion. During hypoxic BA perfusion, ventilation and PA perfusion maintained normal. Control lungs were kept under steady-state conditions for 105 min. During the experiments, PA pressure (PAP) and blood gas analysis were frequently monitored. Hypoxic perfusion of the BA resulted in an increase in systolic and mean PAP, a reaction that was reversible upon normoxic BA perfusion. The PAP increase was reproducible during the second hypoxic BA perfusion. Under control conditions, the PAP stayed constant until about 80 min of the experiment. In conclusion, the results of the current study prove that hypoxic perfusion of the vasa vasorum of the PA directly increases PAP in an ex situ lung perfusion setup, suggesting that PA vasa vasorum function and wall ischemia may contribute to the development of PH.NEW & NOTEWORTHY Hypoxic perfusion of the vasa vasorum of the pulmonary artery directly increased pulmonary arterial pressure in an ex vivo lung perfusion setup. This suggests that the function of pulmonary arterial vasa vasorum and wall ischemia may contribute to the development of pulmonary hypertension.

Incidence and causes of perinatal death in prenatally diagnosed vasa previa: a systematic review and meta-analysis.

OBJECTIVE: This study aimed to estimate the perinatal mortality associated with prenatally diagnosed vasa previa and to determine what proportion of those perinatal deaths are directly attributable to vasa previa. DATA SOURCES: The following databases have been searched from January 1, 1987, to January 1, 2023: PubMed, Scopus, Web of Science, and Embase. STUDY ELIGIBILITY CRITERIA: Our study included all studies (cohort studies and case series or reports) that had patients in which a prenatal diagnosis of vasa previa was made. Case series or reports were excluded from the meta-analysis. All cases in which prenatal diagnosis was not made were excluded from the study. METHODS: The programming language software R (version 4.2.2) was used to conduct the meta-analysis. The data were logit transformed and pooled using the fixed effects model. The between-study heterogeneity was reported by I2. The publication bias was evaluated using a funnel plot and the Peters regression test. The Newcastle-Ottawa scale was used to assess the risk of bias. RESULTS: Overall, 113 studies with a cumulative sample size of 1297 pregnant individuals were included. This study included 25 cohort studies with 1167 pregnancies and 88 case series or reports with 130 pregnancies. Moreover, 13 perinatal deaths occurred among these pregnancies, consisting of 2 stillbirths and 11 neonatal deaths. Among the cohort studies, the overall perinatal mortality was 0.94% (95% confidence interval, 0.52-1.70; I2=0.0%). The pooled perinatal mortality attributed to vasa previa was 0.51% (95% confidence interval, 0.23-1.14; I2=0.0%). Stillbirth and neonatal death were reported in 0.20% (95% confidence interval, 0.05-0.80; I2=0.0%) and 0.77% (95% confidence interval, 0.40-1.48; I2=0.0%) of pregnancies, respectively. CONCLUSION: Perinatal death is uncommon after a prenatal diagnosis of vasa previa. Approximately half of the cases of perinatal mortality are not directly attributable to vasa previa. This information will help in guiding physicians in counseling and will provide reassurance to pregnant individuals with a prenatal diagnosis of vasa previa.

Vasa previa in singleton pregnancies: diagnosis and clinical management based on an international expert consensus.

BACKGROUND: There are limited data to guide the diagnosis and management of vasa previa. Currently, what is known is largely based on case reports or series and cohort studies. OBJECTIVE: This study aimed to systematically collect and classify expert opinions and achieve consensus on the diagnosis and clinical management of vasa previa using focus group discussions and a Delphi technique. STUDY DESIGN: A 4-round focus group discussion and a 3-round Delphi survey of an international panel of experts on vasa previa were conducted. Experts were selected on the basis of their publication record on vasa previa. First, we convened a focus group discussion panel of 20 experts and agreed on which issues were unresolved in the diagnosis and management of vasa previa. A 3-round anonymous electronic survey was then sent to the full expert panel. Survey questions were presented on the diagnosis and management of vasa previa, which the experts were asked to rate on a 5-point Likert scale (from "strongly disagree"=1 to "strongly agree"=5). Consensus was defined as a median score of 5. Following responses to each round, any statements that had median scores of ≤3 were deemed to have had no consensus and were excluded. Statements with a median score of 4 were revised and re-presented to the experts in the next round. Consensus and nonconsensus statements were then aggregated. RESULTS: A total of 68 international experts were invited to participate in the study, of which 57 participated. Experts were from 13 countries on 5 continents and have contributed to >80% of published cohort studies on vasa previa, as well as national and international society guidelines. Completion rates were 84%, 93%, and 91% for the first, second, and third rounds, respectively, and 71% completed all 3 rounds. The panel reached a consensus on 26 statements regarding the diagnosis and key points of management of vasa previa, including the following: (1) although there is no agreement on the distance between the fetal vessels and the cervical internal os to define vasa previa, the definition should not be limited to a 2-cm distance; (2) all pregnancies should be screened for vasa previa with routine examination for placental cord insertion and a color Doppler sweep of the region over the cervix at the second-trimester anatomy scan; (3) when a low-lying placenta or placenta previa is found in the second trimester, a transvaginal ultrasound with Doppler should be performed at approximately 32 weeks to rule out vasa previa; (4) outpatient management of asymptomatic patients without risk factors for preterm birth is reasonable; (5) asymptomatic patients with vasa previa should be delivered by scheduled cesarean delivery between 35 and 37 weeks of gestation; and (6) there was no agreement on routine hospitalization, avoidance of intercourse, or use of 3-dimensional ultrasound for diagnosis of vasa previa. CONCLUSION: Through focus group discussion and a Delphi process, an international expert panel reached consensus on the definition, screening, clinical management, and timing of delivery in vasa previa, which could inform the development of new clinical guidelines.

Third-trimester fetoscopic ablation therapy for types II and III vasa previa.

BACKGROUND: Vasa previa is an obstetrical condition in which fetal vessels located near the cervix traverse the fetal membranes unprotected by underlying placenta. Type I vasa previa arises directly from a velamentous cord root, whereas types II and III arise from an accessory lobe or a distal lobe of the same placenta, respectively. Fetoscopic laser ablation for types II and III vasa previa is a novel therapeutic option with benefits that include surgical resolution of the vasa previa, avoidance of prolonged hospitalization, and opportunity for a term vaginal delivery. The potential risks of fetoscopy can be mitigated by delaying laser surgery until a gestational age of 31 to 33 weeks, immediately before anticipated hospitalized surveillance. OBJECTIVE: This study aimed to assess feasibility and outcomes of types II and III vasa previa patients treated via fetoscopic laser ablation in the third trimester. STUDY DESIGN: This is a retrospective study of singleton pregnancies with types II and III vasa previa treated with fetoscopic laser ablation at a gestational age ≥31 weeks at a single center between 2006 and 2022. Pregnancy and newborn outcomes were assessed. Continuous variables are expressed as mean±standard deviation. RESULTS: Of 84 patients referred for vasa previa, 57 did not undergo laser ablation: 19 either had no or resolved vasa previa, 25 had type I vasa previa (laser-contraindicated), and 13 had type II or III vasa previa but declined laser treatment. Of the remaining 27 patients who underwent laser ablation, 7 were excluded (laser performed at <31 weeks and/or twins), leaving 20 study patients. The mean gestational age at fetoscopic laser ablation was 32.0±0.6 weeks, and total operative time was 62.1±19.6 minutes. There were no perioperative complications. All patients had successful occlusion of the vasa previa vessels (1 required a second procedure). All patients were subsequently managed as outpatients. The mean gestational age at delivery was 37.2±1.8 weeks, the mean birthweight was 2795±465 g, and 70% delivered vaginally. Neonatal intensive care unit admission occurred in 3 cases: 1 for respiratory distress syndrome and 2 for hyperbilirubinemia requiring phototherapy. There were no cases of neonatal transfusion, intraventricular hemorrhage, sepsis, patent ductus arteriosus, or death. CONCLUSION: Laser ablation for types II and III vasa previa at 31 to 33 gestational weeks was technically achievable and resulted in favorable outcomes.

Vasa vasorum interna in the carotid wall of active forms of Takayasu arteritis evidenced by ultrasound localization microscopy.

Introduction: Takayasu arteritis (TA) is associated with microvascularization of the wall of large arteries and is related to inflammation. Ultrasound localization microscopy (ULM), combining ultrafast ultrasound imaging with microbubble (MB) injection, can track the path of MBs within the arterial wall and thus provide imaging of the vasa vasorum. From the analysis of MB tracks in the common carotid arteries of patients with active TA, we report the presence of microvessels in connection with the carotid lumen (i.e., vasa vasorum interna [VVI]). Methods: ULM maps were obtained on five patients with active disease in the observational single-center series of the TAK-UF study. MB tracks connected to the carotid lumen were automatically identified, allowing the reconstruction of VVI. Results: MB tracking allows us to observe a microvascular network on the inner part of the wall, with some vessels in communication with the carotid lumen. This type of vessel was identified in all patients with active TA (n = 5) with a median of 2.2 [1.1-3.0] vessels per acquisition (2D longitudinal view of 3 cm of the common carotid artery). The blood flow within these vessels is mainly centrifugal; that is, toward the adventitia (88% [54-100] of MB tracks with flow directed to the outer part of the wall). Conclusion: VVI are present in humans in the case of active TA and emphasize the involvement of the intima in the pathological process. ClinicalTrials.gov Identifier: NCT03956394.

Prenatal diagnosis and postnatal outcome of Type-III vasa previa: systematic review of literature.

OBJECTIVE: Type-III vasa previa (VP) is a rare form of VP, not necessarily associated with other placental or vascular anomalies, in which aberrant vessels run from the placenta to the amniotic membranes, near the internal cervical os, before returning to the placenta. Early diagnosis of Type-III VP is important but technically challenging. The objective of this study was to gather the current available evidence on the perinatal diagnosis and outcome of Type-III VP. METHODS: A systematic review of the literature on the perinatal diagnosis of atypical Type-III VP was carried out in PubMed, MEDLINE and EMBASE accordingto PRISMA guidelines from inception to March 2023. Data extraction and tabulation were performed by two operators and checked by a third senior author. The quality of the included studies was evaluated using the National Institutes of Health tool for the quality assessment of case-series studies. Our local ultrasound database was searched for previously unreported recent cases. Characteristics of prenatally and postnatally diagnosed Type-III VP, including clinical features and perinatal outcomes, were summarized using descriptive statistics. RESULTS: Eighteen cases of Type-III VP were included, of which 16 were diagnosed prenatally (14 cases were retrieved from 10 publications and two were unpublished cases from our center) and two were diagnosed postnatally (retrieved from two publications). All prenatal cases were diagnosed on transvaginal ultrasound at a mean gestational age of 29 weeks (median, 31 weeks; range, 19-38 weeks). Conception was achieved with in-vitro fertilization in 4/16 (25.0%) cases. There were no prenatal symptoms in 15/18 (83.3%) cases, while in two (11.1%) cases there was vaginal bleeding and in one (5.6%) preterm labor occurred. In 15/18 (83.3%) cases, at least one placental abnormality was observed, including low-lying insertion (9/17), succenturiate or accessory lobe (1/17), velamentous cord insertion (3/18) and marginal insertion (9/18). All prenatally diagnosed cases were liveborn and were delivered by Cesarean section before rupture of membranes at a median gestational age of 35 weeks (range, 32-38 weeks) without neonatal complications. Emergency Cesarean section was performed in 2/16 (12.5%) cases with a prenatal diagnosis and 1/2 (50.0%) cases with a postnatal diagnosis (P = 0.179). Among those with data available, an Apgar score of ≤ 7 was observed in the prenatally vs postnatally diagnosed group in 5/13 vs 1/1 cases, respectively, at the 1-min evaluation and 3/13 vs 1/1 cases, respectively, at the 5-min evaluation. CONCLUSIONS: The prenatal diagnosis of Type-III VP is challenging, with few cases reported in the literature; however, it is crucial for minimizing the risk of adverse outcome by enabling early-term elective Cesarean delivery prior to rupture of membranes. Given that clinical manifestations and risk factors are non-specific, and that Type-III VP cannot be excluded when there is a normal cord insertion or a singular placental mass, systematic screening by transvaginal ultrasound in the general pregnant population is recommended, particularly in those with a low-lying or morphologically abnormal placenta and those who conceived using assisted reproductive technology. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Incorporation of vasa previa screening into a routine anomaly scan: A single center cohort study.

INTRODUCTION: Vasa previa (VP), defined as unprotected fetal vessels traversing the membranes over the cervix, is associated with a high perinatal mortality when undiagnosed prenatally. Conversely, prenatal diagnosis with ultrasound and cesarean delivery before the membranes rupture is associated with excellent outcomes. However, controversy exists regarding screening for VP. In the UK, routine screening for VP is not recommended. The objective of this study was to report the incidence of VP and our experience in the detection of VP with a universal screening protocol at the time of the second-trimester fetal anomaly scan with third-trimester confirmation in an unselected population of pregnancies. MATERIAL AND METHODS: We performed a single-center historical cohort study of all pregnant women who underwent routine second-trimester anomaly screening scans at West Middlesex University Hospital, London, UK, between 2012 and 2016. Over 5 years, every patient undergoing routine anomaly screening was evaluated for VP using a systematic protocol during their 20-week anomaly scan. Suspected cases of VP were rescanned in the third trimester by specialist sonographers with an interest in VP. The primary outcomes were the incidence and detection of VP. RESULTS: During the study period, 24 690 anatomy scans were performed. A total of 64 patients were identified as having potential VP at the second-trimester anomaly screening scan, of which 19 were confirmed by the specialist sonographer in the third trimester and at delivery. The screen positive rate was 0.26% (95% confidence interval [CI] 0.20%-0.32%). VP at birth was found in 19/24690 births (1:1299 [95% CI: 1:832-1:2030] births). Universal screening for VP using our protocol had a sensitivity of 100% and a specificity of 99.78% (95% CI: 99.72%-99.84%). The false-positive rate of the second-trimester screen was 0.18% (95% CI: 0.13-0.24). There were no false positives or false negatives at delivery. Of the 19 patients with confirmed VP, 17 had scheduled cesarean deliveries, and two required emergency deliveries due to antepartum hemorrhage. One baby died, giving a perinatal mortality of 5%. CONCLUSIONS: VP complicates approximately 1:1300 pregnancies. Routine screening for VP yielded a 100% detection rate. We suggest the inclusion of structured VP assessment in standard fetal anomaly screening programs.

Impact of Universal Vasa Previa Screening With Color Doppler During the OB Ultrasound Anatomy Scan.

OBJECTIVES: Our institution introduced universal vasa previa (VP) screening utilizing transabdominal ultrasound with color Doppler for all pregnancies at the second trimester anatomy scan. Our study sought to describe the clinical impact of this intervention. METHODS: Radiology records from the 12 months pre- and post-intervention were queried for "vasa previa." Records included for analysis were those with a first-time diagnosis or discussion of VP at the anatomy scan. Cases were categorized by outcome: (Group 1) True VP, with subgroups A, unresolved by time of delivery and B, resolved by delivery; (Group 2) False positives; (Group 3) Possible VP without definitive diagnosis; and (Group 4) VP ruled out, for example, "no features of VP." Group size was expressed as a percentage of total anatomy scans during pre- or post-intervention periods respectively. Absolute and relative percent change were calculated for each group. RESULTS: In the pre-intervention period, 1 case (0.36% of total scans) was categorized in Group 1A, 1 case (0.36%) in Group 3, and 7 cases (2.53%) in Group 4. In the post-intervention period, 2 cases (0.30%) were in Group 1A, 4 cases (0.61%) in Group 1B, 2 cases (0.30%) in Group 2, 1 case (0.15%) in Group 3, and 7 cases (1.06%) in Group 4. There was a +153% relative change in true positives, from 0.36 to 0.91%. CONCLUSIONS: Universal color Doppler screening may have increased detection (sensitivity) while simultaneously increasing false positives (decreased specificity). While decreasing sensitivity is not ideal, this is acceptable given the potential catastrophic outcome of a missed VP.

Diagnosis and Analysis of Vasa Previa Types With Flow HD Glass Body.

OBJECTIVES: To explore the value of applying flow high definition (HD) glass body in prenatal diagnosis of vasa previa and to preliminarily discuss the types of vasa previa. METHODS: Two-dimensional ultrasound, flow HD, and flow HD glass body were used to image the umbilical cord insertion site and placenta, observe the cervical internal os and surrounding areas, and retrospectively analyze cases of vasa previa. RESULTS: There were 15 cases of vasa previa, including 14 cases of singleton pregnancies and 1 case of twin pregnancy, with a total of 22 vasa previa, including 10 veins and 12 arteries. There was 1 case with 3 vessels, 5 cases with 2 vessels, and 9 cases with a single vessel. Among them, in 3 cases of vasa previa detected at 12, 14, and 24 weeks, respectively, the vasa previa were relocated to a normal position at 24, 29, and 35 weeks of gestation when re-examined. Routine 2-dimensional ultrasound examination in this group showed tubular or circular hypoechoic areas near the cervical internal os, but vasa previa could not be confirmed. Flow HD could display color blood flow at and near the cervical internal os in 15 cases, but it was difficult to continuously show the course and source of the blood vessels under the chorion. Flow HD glass body from multiple angles could display the relationship between 15 cases of 22 vasa previa and the placenta and cervix. Combined with color Doppler blood flow spectra, flow HD glass body could determine the types of vasa previa. CONCLUSIONS: Flow HD glass body imaging can clearly display vasa previa, showing their origin and the spatial relationship with the cervix and placenta in a 3-dimensional manner, displaying the course and attachment points of umbilical vessels under the chorion. It can observe the area of interest at any angle, and combined with color Doppler blood flow spectra, it can judge the vasa previa of the umbilical vein, providing a more definite imaging basis for clinical management.

The potential impact of universal screening for vasa previa in the prevention of stillbirths.

OBJECTIVES: To estimate the number of pregnancies complicated by vasa previa annually in nine developed countries, and the potential preventable stillbirths associated with undiagnosed cases. We also assessed the potential impact of universal screening for vasa previa on reducing stillbirth rates. METHODS: We utilized nationally-reported birth and stillbirth data from public databases in the United States, United Kingdom, Canada, Germany, Ireland, Greece, Sweden, Portugal, and Australia. Using the annual number of births and the number and rate of stillbirths in each country, and the published incidence of vasa previa and stillbirth rates associated with the condition, we estimated the expected annual number of cases of vasa previa, those that would result in a livebirth, and the potential preventable stillbirths with and without prenatal diagnosis. RESULTS: There were 6,099,118 total annual births with 32,550 stillbirths, corresponding to a summary stillbirth rate of 5.34 per 1,000 pregnancies. The total expected vasa previa cases was estimated to be 5,007 (95 % CI: 3,208-7,201). The estimated number of livebirths would be 4,937 (95 % CI: 3,163-7,100) and 3,610 (95 % CI: 2,313-5,192) in pregnancies with and without a prenatal diagnosis of VP. This implies that prenatal diagnosis would potentially prevent 1,327 (95 % CI: 850-1,908) stillbirths in these countries, corresponding to a potential reduction in stillbirth rate by 4.72 % (95 % CI: 3.80-5.74) if routine screening for vasa previa was performed. CONCLUSIONS: Our study highlights the importance of universal screening for vasa previa and suggests that prenatal diagnosis of prevention could potentially reduce 4-5 % of stillbirths.

Facial nerve palsy after middle meningeal artery embolization for chronic subdural hematoma: a case report.

Middle meningeal artery embolization (MMAE) has emerged as a safe and efficacious alternative to surgery for the treatment of new or recurrent chronic subdural hematoma (CSDH). Several complications such as facial palsy may suddenly occur even in the absence of evident dangerous anastomoses in the angiogram. We herein present a case-report of left facial nerve palsy after MMAE.

Fetoscopic laser photocoagulation: a medically reasonable treatment option in the management of types II and III vasa previa.

Vasa previa is a condition where unprotected fetal vessels cross the cervix within the membranes, posing a considerable risk of fetal death or severe morbidity if the membranes rupture before or during delivery. There has not been a definitive in utero treatment for this condition. Patients are typically closely monitored and hospitalized in the early third trimester and scheduled for cesarean delivery before term. This approach poses considerable physical, social, psychological, and financial challenges for pregnant patients and their families. Furthermore, fetal vessel rupture may lead to severe hypoxic-ischemic injury and consequent neurodevelopmental impairment. Finally, babies delivered early due to vasa previa may face both the short- and long-term consequences of prematurity. Recently, fetoscopic laser photocoagulation using a single-port fetoscope has emerged as a potential therapeutic option for patients with types II and III vasa previa. This innovative approach aims to reduce hospital stays, increases the chance of successful vaginal delivery, and potentially allows pregnancies to reach full term, providing lifelong benefits for the infant. Preliminary clinical studies on human subjects have demonstrated promising results concerning the feasibility, safety, and efficacy of this intervention for a subset of patients with types II and III vasa previa. After reviewing the current state of the art, we argued that offering fetoscopic laser photocoagulation in specialized centers under IRB supervision meets the ethical obligations of beneficence and non-maleficence for both pregnant and fetal patients, as well as the autonomy-based obligations for pregnant patients.

Characterization of potential spermatogonia biomarker genes in the European eel (Anguilla anguilla).

Identification of specific molecular markers for spermatogonial stem cells in teleost is crucial for enhancing the efficacy of reproductive biotechnologies in aquaculture, such as transplantation and surrogate production in fishes. Since it is not yet possible to distinguish spermatogonial stem cells of European eel (Anguilla anguilla) using specific molecular markers, we isolated spermatogonial cells from immature European eels to find these potential markers. We attempted this by studying three candidate genes: vasa, nanos2, and dnd1. Two vasa (vasa1 and vasa2) genes, nanos2, and dnd1 were identified, characterized, and studied in the muscle, testis, and isolated spermatogonia. Our results showed that vasa1 and vasa2 had the highest levels of expression when measured by qPCR. In situ hybridization and immunochemistry assays showed that the four genes were localized explicitly in type A spermatogonia. However, vasa1 and vasa2 exhibited stronger signals in the immature testicular tissue than the other two potential markers. According to this, vasa1 and vasa2 were found to be the most effective markers for spermatogonial cells in the European eel.

A conserved node in the regulation of Vasa between an induced and an inherited program of primordial germ cell specification.

Primordial germ cells (PGCs) are specified by diverse mechanisms in early development. In some animals, PGCs are specified via inheritance of maternal determinants, while in others, in a process thought to represent the ancestral mode, PGC fate is induced by cell interactions. Although the terminal factors expressed in specified germ cells are widely conserved, the mechanisms by which these factors are regulated can be widely diverse. Here we show that a post-translational mechanism of germ cell specification is conserved between two echinoderm species thought to employ divergent germ line segregation strategies. Sea urchins segregate their germ line early by an inherited mechanism. The DEAD-box RNA - helicase Vasa, a conserved germline factor, becomes enriched in the PGCs by degradation in future somatic cells by the E3-ubiquitin-ligase Gustavus (Gustafson et al., 2011). This post-translational activity occurs early in development, substantially prior to gastrulation. Here we test this process in germ cell specification of sea star embryos, which use inductive signaling mechanisms after gastrulation for PGC fate determination. We find that Vasa-GFP protein becomes restricted to the PGCs in the sea star even though the injected mRNA is present throughout the embryo. Gustavus depletion, however, results in uniform accumulation of the protein. These data demonstrate that Gustavus-mediated Vasa turnover in somatic cells is conserved between species with otherwise divergent PGC specification mechanisms. Since Gustavus was originally identified in Drosophila melanogaster to have similar functions in Vasa regulation (Kugler et al., 2010), we conclude that this node of Vasa regulation in PGC formation is ancestral and evolutionarily transposable from the ancestral, induced PGC specification program to an inherited PGC specification mechanism.

GLH-1/Vasa represses neuropeptide expression and drives spermiogenesis in the C. elegans germline.

Germ granules harbor processes that maintain germline integrity and germline stem cell capacity. Depleting core germ granule components in C. elegans leads to the reprogramming of germ cells, causing them to express markers of somatic differentiation in day-two adults. Somatic reprogramming is associated with complete sterility at this stage. The resulting germ cell atrophy and other pleiotropic defects complicate our understanding of the initiation of reprogramming and how processes within germ granules safeguard the totipotency and immortal potential of germline stem cells. To better understand the initial events of somatic reprogramming, we examined total mRNA (transcriptome) and polysome-associated mRNA (translatome) changes in a precision full-length deletion of glh-1, which encodes a homolog of the germline-specific Vasa/DDX4 DEAD-box RNA helicase. Fertile animals at a permissive temperature were analyzed as young adults, a stage that precedes by 24 â€‹h the previously determined onset of somatic reporter-gene expression in the germline. Two significant changes are observed at this early stage. First, the majority of neuropeptide-encoding transcripts increase in both the total and polysomal mRNA fractions, suggesting that GLH-1 or its effectors suppress this expression. Second, there is a significant decrease in Major Sperm Protein (MSP)-domain mRNAs when glh-1 is deleted. We find that the presence of GLH-1 helps repress spermatogenic expression during oogenesis, but boosts MSP expression to drive spermiogenesis and sperm motility. These insights define an early role for GLH-1 in repressing somatic reprogramming to maintain germline integrity.

Isolation of vasa vasorum endothelial cells from pulmonary artery adventitia: Implementation to vascular biology research.

Isolated endothelial cells are valuable in vitro model for vascular research. At present, investigation of disease-relevant changes in vascular endothelium at the molecular level requires established endothelial cell cultures, preserving vascular bed-specific phenotypic characteristics. Vasa vasorum (VV) form a microvascular network around large blood vessels, in both the pulmonary and systemic circulations, that are critically important for maintaining the integrity and oxygen supply of the vascular wall. However, despite the pathophysiological significance of the VV, methods for the isolation and culture of vasa vasorum endothelial cells (VVEC) have not yet been reported. In our prior studies, we demonstrated the presence of hypoxia-induced angiogenic expansion of the VV in the pulmonary artery (PA) of neonatal calves; an observation which has been followed by a series of in vitro studies on isolated PA VVEC. Here we present a detailed protocol for reproducible isolation, purification, and culture of PA VVEC. We show these cells to express generic endothelial markers, (vWF, eNOS, VEGFR2, Tie1, and CD31), as well as progenitor markers (CD34 and CD133), bind lectin Lycopersicon Esculentum, and incorporate acetylated low-density lipoproteins labeled with acetylated LDL (DiI-Ac-LDL). qPCR analysis additionally revealed the expression of CD105, VCAM-1, ICAM-1, MCAM, and NCAM. Ultrastructural electron microscopy and immunofluorescence staining demonstrated that VVEC are morphologically characterized by a developed actin and microtubular cytoskeleton, mitochondrial network, abundant intracellular vacuolar/secretory system, and cell-surface filopodia. VVEC exhibit exponential growth in culture and can be mitogenically activated by multiple growth factors. Thus, our protocol provides the opportunity for VVEC isolation from the PA, and potentially from other large vessels, enabling advances in VV research.