Intraluminal chloride regulates lung branching morphogenesis: involvement of PIEZO1/PIEZO2.

BACKGROUND: Clinical and experimental evidence shows lung fluid volume as a modulator of fetal lung growth with important value in treating fetal lung hypoplasia. Thus, understanding the mechanisms underlying these morphological dynamics has been the topic of multiple investigations with, however, limited results, partially due to the difficulty of capturing or recapitulating these movements in the lab. In this sense, this study aims to establish an ex vivo model allowing the study of lung fluid function in branching morphogenesis and identify the subsequent molecular/ cellular mechanisms. METHODS: Ex vivo lung explant culture was selected as a model to study branching morphogenesis, and intraluminal injections were performed to change the composition of lung fluid. Distinct chloride (Cl-) concentrations (5.8, 29, 143, and 715 mM) or Cl- channels inhibitors [antracene-9-carboxylic acid (A9C), cystic fibrosis transmembrane conductance regulator inhibitor172 (CFTRinh), and calcium-dependent Cl- channel inhibitorA01 (CaCCinh)] were injected into lung lumen at two timepoints, day0 (D0) and D2. At D4, morphological and molecular analyses were performed in terms of branching morphogenesis, spatial distribution (immunofluorescence), and protein quantification (western blot) of mechanoreceptors (PIEZO1 and PIEZO2), neuroendocrine (bombesin, ghrelin, and PGP9.5) and smooth muscle [alpha-smooth muscle actin (α-SMA) and myosin light chain 2 (MLC2)] markers. RESULTS: For the first time, we described effective intraluminal injections at D0 and D2 and demonstrated intraluminal movements at D4 in ex vivo lung explant cultures. Through immunofluorescence assay in in vivo and ex vivo branching morphogenesis, we show that PGP9.5 colocalizes with PIEZO1 and PIEZO2 receptors. Fetal lung growth is increased at higher [Cl-], 715 mM Cl-, through the overexpression of PIEZO1, PIEZO2, ghrelin, bombesin, MLC2, and α-SMA. In contrast, intraluminal injection of CFTRinh or CaCCinh decreases fetal lung growth and the expression of PIEZO1, PIEZO2, ghrelin, bombesin, MLC2, and α-SMA. Finally, the inhibition of PIEZO1/PIEZO2 by GsMTx4 decreases branching morphogenesis and ghrelin, bombesin, MLC2, and α-SMA expression in an intraluminal injection-independent manner. CONCLUSIONS: Our results identify PIEZO1/PIEZO2 expressed in neuroendocrine cells as a regulator of fetal lung growth induced by lung fluid.

University students' opinion on gamete donor identification regimes.

PURPOSE: This study aims to evaluate the opinion of university students about the identification or nonidentification of gamete donation and the probability of donation according to the different regimes. METHODS: This was a cross-sectional observational study based on an online anonymous survey including questions about sociodemographic data, reasons for considering donations, information about the donation process and legislation, and their opinions about the different regimes and how they would influence donations. RESULTS: In total, 1393 valid responses were obtained, with a mean age of 24.0 years (SD = 4.8), most of the respondents being female (68.5%), living in a relationship (56.7%), and without children (88.4%). The main reasons for considering donation would be altruism and monetary compensation. Overall, it was found that participants were poorly informed about the donation procedure and legislation. Students revealed preference for nonidentified donation, and they were less likely to donate in an open identity regime. CONCLUSION: Most university students consider themselves poorly informed about gamete donation, express a preference for nonidentified gamete donation, and would less likely donate on an open identity basis. Thus, an identified regime may be less attractive to potential donors and lead to a decrease in the availability of gamete donors.

Diaphragmatic hernia repair porcine model to compare the performance of biodegradable membranes against Gore-Tex®.

BACKGROUND: Patch repair of congenital diaphragmatic hernia (CDH) using Gore-Tex® is associated with infection, adhesions, hernia recurrence, long-term musculoskeletal sequels and poor tissue regeneration. To overcome these limitations, the performance of two novel biodegradable membranes was tested to repair CDH in a growing pig model. METHODS: Twelve male pigs were randomly assigned to 3 different groups of 4 animals each, determined by the type of patch used during thoracoscopic diaphragmatic hernia repair (Gore-Tex®, polycaprolactone electrospun membrane-PCLem, and decellularized human chorion membrane-dHCM). After 7 weeks, all animals were euthanized, followed by necropsy for diaphragmatic evaluation and histological analysis. RESULTS: Thoracoscopic defect creation and diaphragmatic repair were performed without any technical difficulty in all groups. However, hernia recurrence rate was 0% in Gore-Tex®, 50% in PCLem and 100% in dHCM groups. At euthanasia, Gore-Tex® patches appeared virtually unchanged and covered with a fibrotic capsule, while PCLem and dHCM patches were replaced by either floppy connective tissue or vascularized and floppy regenerated membranous tissue, respectively. CONCLUSION: Gore-Tex® was associated with a higher survival rate and lower recurrence. Nevertheless, the proposed biodegradable membranes were associated with better tissue integration when compared with Gore-Tex®.

User satisfaction with an intrauterine system containing 52 mg levonorgestrel: a Portuguese study.

OBJECTIVE: To characterise patient satisfaction 6 and 12 months after insertion of a 52 mg levonorgestrel-releasing intrauterine system (LNG-IUS) in Portuguese women. STUDY DESIGN: A non-interventional and prospective study was conducted in Portuguese women of reproductive age who had Levosert® inserted. Two questionnaires (administered 6 and 12 months after 52 mg LNG-IUS insertion) were used to collect information on the patients' menstrual pattern, discontinuation rate, and satisfaction rate with Levosert®. RESULTS: A total of 102 women were enrolled, of which only 94 (92.2%) completed the study. Seven participants discontinued the use of the 52 mg LNG-IUS. At 6 and 12 months, 90.7% and 90.4% of participants, respectively, were either satisfied or very satisfied with the 52 mg LNG-IUS. At 6 and 12 months, 73.2% and 72.3% of participants, respectively, indicated that they were very likely to recommend the 52 mg LNG-IUS to a friend or family member. Most women (92.2%) continued to use the 52 mg LNG-IUS for the first year. The percentage of women who were 'much more satisfied' with Levosert® than with their previous contraceptive methods was 55.9% and 57.8% at 6 and 12 months, respectively, per questionnaire assessment. Satisfaction was associated with age (p = 0.004), amenorrhoea (p < 0.003) and absence of dysmenorrhoea (p = 0.003), but not with parity (p = 0.922). CONCLUSIONS: These data suggest that the continuation and satisfaction rates with Levosert® were very high, and that this system is well accepted among Portuguese women. Patient satisfaction was driven by a favourable bleeding pattern and absence of dysmenorrhoea.

Satisfaction with the 52 mg levonorgestrel­releasing intrauterine system was high among Portuguese women. Patient satisfaction was mainly driven by a favourable bleeding pattern and absence of dysmenorrhoea.

Fetal heart rate variability and infant self-regulation: the impact of mother's prenatal depressive symptoms.

BACKGROUND: Foetal heart rate (FHR) variability is considered a marker of foetal neurobehavioral development associated with infant self-regulation and thus may be an early precursor of the adverse impact of mother's prenatal depressive symptoms on infant self-regulation. OBJECTIVE: This study analysed the mediator role of FHR variability in the association between mother's prenatal depressive symptoms and infant self-regulation at three months. METHODS: The sample comprised 86 first-born infants and their mothers. Mothers reported on depressive symptoms at the first trimester of pregnancy and on depressive symptoms and infant self-regulation at three months postpartum. FHR variability was recorded during routine cardiotocography at the third trimester of pregnancy. A mediation model was tested, adjusting for mother's postnatal depressive symptoms. RESULTS: Higher levels of mother's prenatal depressive symptoms were associated with both lower FHR variability and lower infant self-regulation at three months. FHR variability was associated with infant self-regulation and mediated the association between mother's prenatal depressive symptoms and infant self-regulation at three months. CONCLUSION: Findings suggested FHR variability as an early precursor of infant self-regulation that underlies the association between mother's prenatal depressive symptoms and infant self-regulation. Infants of mothers with higher levels of prenatal depressive symptoms could be at risk of self-regulation problems, partially due to their lower FHR variability.

Vulvovaginal candidiasis and asymptomatic vaginal colonization in Portugal: Epidemiology, risk factors and antifungal pattern.

Vulvovaginal candidiasis (VVC) has been identified as a global issue of concern due to its clinical, social and economic implications. The emerging relevance of VVC makes it crucial to increase the knowledge on its epidemiological and etiological features in order to improve its prevention and treatment. Thus, this study aimed to reveal the incidence, microbiology, antifungal pattern and risk factors of VVC in Portugal. For that, high vaginal samples were collected from 470 symptomatic and asymptomatic participants; Candida spp. were identified with molecular techniques and their antifungal susceptibility was analyzed with E-tests. The results revealed an incidence of VVC among women with vulvovaginitis of 74.4%. Furthermore, 63.7% of asymptomatic women were colonized with Candida spp. Importantly, women with history of recurrent vaginal infections, those who use over-the-counter antifungals, oral contraceptive pills and non-cotton underwear were found to be at significantly higher risk of developing VVC. Candida albicans was the most common species (59%), followed by Candida glabrata (27%), in a total of eight distinct species, with similar distribution among colonized and infected participants. Of note, various isolates, especially of the most common species, showed low susceptibility towards fluconazole. In contrast, only few isolates showed low susceptibility towards caspofungin. Overall, this study suggests that the identification of species causing VVC and their antifungal susceptibility are urgently needed in clinical practice in order to improve the decision for the most adequate treatment. It also suggests that avoiding certain risk behaviors may prevent the development of VVC. LAY SUMMARY: Vaginal candidiasis (VVC) is a relevant infection worldwide. In this study, we identified several risk behaviors that may promote VVC and concluded that vaginal microbiologic analyses are urgently required in clinical practice in order to improve the prevention and treatment of this disease.

Pelvic floor dysfunction after vaginal delivery: MOODS-a prospective study.

INTRODUCTION AND HYPOTHESIS: Labor is a known risk factor for pelvic floor dysfunction (PFD); however, the impact of operative vaginal delivery (OVD), particularly spatulas, remains unclear. The aim of this study was to compare postpartum PFD symptoms in women undergoing spontaneous vaginal delivery (SVD) and those undergoing OVD. METHODS: An observational prospective study (MOODS: Maternal-neonatal Outcomes in Operative Vaginal Delivery) was enrolled at Hospital de Braga from February to October 2018. All singleton term OVD (Thierry spatulas and vacuum extractor) and a convenience SVD sample were recruited, in a 2:1 ratio. To assess PFD symptoms Pelvic Floor Distress Inventory-20 (PFDI-20) was applied at 3, 6, and 12 months postpartum. The questionnaire is divided into three subscales: Urinary (UDI), Colorectal-Anal (CRADI), and Pelvic Organ Prolapse Distress Inventory (POPDI). RESULTS: Of the 304 women recruited, 207 were included, 34.3% with SVD and 65.7% with OVD. Thierry spatulas were used in 53.7% of women undergoing OVD. Frequency of nulliparous (p < 0.001), episiotomy (p < 0.001), neuraxial anesthesia (p < 0.001), postpartum pain (p = 0.001) and occiput-posterior fetal position (p < 0.001) were significantly higher in OVD. Second phase of labor duration was longer in OVD (p = 0.001). At 3 months postpartum, women undergoing OVD and spatula-assisted delivery had higher UDI score, POPDI score, and global score, with no differences at 6 months and 1 year. After controlling for confounding variables, OVD and spatulas were still associated with greater POPDI scores at 3 months postpartum. CONCLUSIONS: Operative vaginal delivery, particularly with spatulas, seems to be associated with a higher prevalence of early PFD symptoms, mainly regarding pelvic organ prolapse.

The Role of Operative Vaginal Delivery in Postpartum Sexual Dysfunction: MOODS - A Prospective Study.

BACKGROUND: Childbirth brings many changes to women's life and sexual health. The influence of operative vaginal delivery on sexual function has produced inconsistent results. AIM: To evaluate the effects of mode of vaginal delivery (spontaneous or operative) in postpartum sexual function. METHODS: Descriptive prospective study (MOODS- Maternal-neonatal Outcomes in Operative Vaginal Delivery) including 304 women who had a singleton term vaginal delivery (operative or spontaneous in a relation 2:1). Women were invited to answer a questionnaire at 3, 6 months and 1 year postpartum. OUTCOMES MEASUREMENT: A validated questionnaire was applied, the Female Sexual Function Index (FSFI) Score, to evaluate effects of operative delivery on sexual health. Sexual dysfunction was defined by FSFI score <26.55. RESULTS: 211 women answered at least one questionnaire. Overall rate of sexual dysfunction was 62%, 43% and 48% at 3, 6 and 12 months respectively. At 3 months, total FSFI score was significantly lower in operative vaginal delivery group (mean±SD, 21.3±8.6 vs 24.9±7.9, P = 0.015). Arousal (P = 0.028), orgasm (P = 0.029), satisfaction (P = 0.015) and pain (P = 0.007) FSFI domains were also significantly inferior. At this time, 44% women in spontaneous delivery group and 70% in operative delivery group had sexual dysfunction (P = 0.0002). At 6 months, there were no differences in FSFI scores according the type of delivery. At 12 months, total FSFI score was similar in both groups, but pain domain was significantly lower in operative delivery (P = 0.004). Considering type of instrument (Thierry's Spatulas or Kiwi Vacuum), no differences were found regarding episiotomy, perineal trauma, obstetric anal sphincter injury or postpartum complications. FSFI scores did not differ between the two instruments at any time point. A logistic regression showed that, when controlled for perineal trauma, mode of delivery was independently associated with sexual dysfunction at 3 months (P = 0.02). CLINICAL IMPLICATIONS: Clinicians should assess women's sexual health during pregnancy and postpartum period in order to enhance their wellbeing. STRENGTHS/LIMITATIONS: Strengths include its prospective design, standardized questionnaire and the new perspectives about a different obstetrical instrument (Thierry's spatulas). Limitations include the absence of pre-pregnancy sexual function data and considerable drop-out rate. CONCLUSION: Sexual dysfunction affects a great proportion of newly mothers and in postpartum period mode of delivery and perineal trauma seem to play an important role. Although there was a progressive reduction over time, prevalence of sexual dysfunction at 6 months and 1 year postpartum was still considerable. The type of obstetrical instrument does not seem to influence short or long-term sexual function. de Sousa NQ, Borges AC, Sarabando R, et al. The Role of Operative Vaginal Delivery in Postpartum Sexual Dysfunction: MOODS - A Prospective Study. J Sex Med 2021;18:1075-1082.

Breast cancer post-surgical impact on women´s quality of life during chemotherapy treatment: A structural equation modelling approach.

OBJECTIVE: Breast cancer is an important public health problem that is increasing in incidence, being a stressor with a negative impact on women's quality of life. This study is focused on the evaluation of temporal precursors (one month before) of women's quality of life undergoing chemotherapy, considering post-surgical personal, clinical, cognitive and neuropsychophysiological factors, according to the Transactional Stress and Coping Model. METHODS: This longitudinal study included 112 patients with breast cancer. Data were collected in two different moments: before and during the adjuvant chemotherapy. Structural equation modelling was used to support a theoretically based model in which some antecedent factors impact patients' long-term quality of life through a set of mediators. RESULTS: The associations of breast symptoms, body image and sexual functioning with psychological distress and quality of life were totally mediated by illness perceptions, while the associations of working memory with psychological distress and quality of life were totally mediated by self-efficacy for coping. Patients with greater psychological distress showed higher levels of nadir cortisol. CONCLUSIONS: Results showed the importance of assessing patients' perceptions of their illness, prior to chemotherapy, as well as promoting more self-efficacy for coping, in order to improve women's emotional state and quality of life.

ROBO2 signaling in lung development regulates SOX2/SOX9 balance, branching morphogenesis and is dysregulated in nitrofen-induced congenital diaphragmatic hernia.

BACKGROUND: Characterized by abnormal lung growth or maturation, congenital diaphragmatic hernia (CDH) affects 1:3000 live births. Cellular studies report proximal (SOX2+) and distal (SOX9+) progenitor cells as key modulators of branching morphogenesis and epithelial differentiation, whereas transcriptome studies demonstrate ROBO/SLIT as potential therapeutic targets for diaphragm defect repair in CDH. In this study, we tested the hypothesis that (a) experimental-CDH could changes the expression profile of ROBO1, ROBO2, SOX2 and SOX9; and (b) ROBO1 or ROBO2 receptors are regulators of branching morphogenesis and SOX2/SOX9 balance. METHODS: The expression profile for receptors and epithelial progenitor markers were assessed by Western blot and immunohistochemistry in a nitrofen-induced CDH rat model. Immunohistochemistry signals by pulmonary structure were also quantified from embryonic-to-saccular stages in normal and hypoplastic lungs. Ex vivo lung explant cultures were harvested at E13.5, cultures during 4 days and treated with increasing doses of recombinant rat ROBO1 or human ROBO2 Fc Chimera proteins for ROBO1 and ROBO2 inhibition, respectively. The lung explants were analyzed morphometrically and ROBO1, ROBO2, SOX2, SOX9, BMP4, and ß-Catenin were quantified by Western blot. RESULTS: Experimental-CDH induces distinct expression profiles by pulmonary structure and developmental stage for both receptors (ROBO1 and ROBO2) and epithelial progenitor markers (SOX2 and SOX9) that provide evidence of the impairment of proximodistal patterning in experimental-CDH. Ex vivo functional studies showed unchanged branching morphogenesis after ROBO1 inhibition; increased fetal lung growth after ROBO2 inhibition in a mechanism-dependent on SOX2 depletion and overexpression of SOX9, non-phospho ß-Catenin, and BMP4. CONCLUSIONS: These studies provided evidence of receptors and epithelial progenitor cells which are severely affected by CDH-induction from embryonic-to-saccular stages and established the ROBO2 inhibition as promoter of branching morphogenesis through SOX2/SOX9 balance.

STATs in Lung Development: Distinct Early and Late Expression, Growth Modulation and Signaling Dysregulation in Congenital Diaphragmatic Hernia.

BACKGROUND: Congenital diaphragmatic hernia (CDH) is a life-threatening developmental anomaly, intrinsically combining severe pulmonary hypoplasia and hypertension. During development, signal transducers and activators of transcription (STAT) are utilized to elicit cell growth, differentiation, and survival. METHODS: We used the nitrofen-induced CDH rat model. At selected gestational time points, lungs were divided into two experimental groups, i.e., control or CDH. We performed immunohistochemistry and western blotting analysis to investigate the developmental expression profile of the complete family of STATs (STAT1-6), plus specific STATs activation (p-STAT3, p-STAT6) and regulation by SOCS (SOCS3) in normal lungs against those of diseased lungs. The normal fetal lung explants were treated with piceatannol (STAT3 inhibitor) in vitro followed by morphometrical analysis. RESULTS: Molecular profiling of STATs during the lung development revealed distinct early and late expression signatures. Experimental CDH altered the STATs expression, activation, and regulation in the fetal lungs. In particular, STAT3 and STAT6 were persistently over-expressed and early over-activated. Piceatannol treatment dose-dependently stimulated the fetal lung growth. CONCLUSION: These findings suggest that STATs play an important role during normal fetal lung development and CDH pathogenesis. Moreover, functionally targeting STAT signaling modulates fetal lung growth, which highlights that STAT3 and STAT6 signaling might be promising therapeutic targets in reducing or preventing pulmonary hypoplasia in CDH.

Use of etonogestrel implant beyond approved duration: prolonged contraceptive effectiveness.

BACKGROUND: The subdermal etonogestrel contraceptive implant has a current approved duration of 3 years. There is, however, recent and strong evidence of its effectiveness for 2 additional years during which no pregnancies were documented. CASE: We report a case of a 28-year-old woman who had an etonogestrel implant inserted 4 years earlier and wished to get pregnant. The implant was, however, deep in the triceps muscle and could not easily be removed. Her serum levels of etonogestrel remained positive even 6 years after insertion. As infertility investigation revealed endometriosis and bilateral tubal obstruction she was referred for assisted reproduction. Despite obtaining oocytes during an ovarian stimulation cycle, the endometrium was never above 4 mm, precluding embryo transfer. It was decided to remove the implant in order to improve endometrial conditions, after which the patient achieved pregnancy. CONCLUSION: This case report suggests that the etonogestrel implant is effective for even longer than the most recent studies have shown.

Resection of Sentinel Lymph Nodes by an Extraperitoneal Minilaparoscopic Approach Using Indocyanine Green for Uterine Malignancies: A Preclinical Comparative Study.

Background The sentinel lymph node (SLN) concept might minimize surgical aggressiveness in cervical and endometrial malignancies. The aim of the study was to test the feasibility and reliability of minilaparoscopic extraperitoneal SLN excision after indocyanine green (ICG) cervical injection using a high-definition near infrared (NIR) imaging system in an in vivo porcine model. The same procedure was performed using conventional laparoscopic instruments and both outcomes were compared. Methods Twenty-four animals were equally and randomly divided into a minilaparoscopic group (group A) and a 5-mm conventional laparoscopic group (group B). A high-definition NIR imaging system and a 30° ICG endoscope were used. First, ICG (0.5 mL) was injected in the paracervical region. The SLN coloring time was recorded. An extraperitoneal approach to the SLN was executed with the same CO2 retropneumoperitoneum pressures (10 mm Hg). In both groups, the times for SLN localization and excision, as well as complications, were registered. Finally, a laparotomy was then done to evaluate whether any stained SLN still remained. The same surgical team performed all experiments. Results SLNs were identified and extraperitoneally excised in all animals without major complications. The SLN localization varied between animals from external iliac to preaortic regions. The surgical times were shorter with minilaparoscopy (39.3 ± 13 minutes) than with conventional 5-mm instruments (51.3 ± 14.17 minutes; P = .042). In group B, one stained SLN remained and was only detected by laparotomy. Conclusions We confirmed the feasibility and reliability of extraperitoneal minilaparoscopic approach for identification, dissection, and excision of SLN using an NIR imaging system and ICG.

Neuroendocrine factors regulate retinoic acid receptors in normal and hypoplastic lung development.

KEY POINTS: Retinoic acid (RA) and ghrelin levels are altered in human hypoplastic lungs when compared to healthy lungs. Although considerable data have been obtained about RA, ghrelin and bombesin in the congenital diaphragmatic hernia (CDH) rat model, neuroendocrine factors have never been associated with the RA signalling pathway in this animal model. In this study, the interaction between neuroendocrine factors and RA was explored in the CDH rat model. The authors found that normal fetal lung explants treated with RA, bombesin and ghrelin showed an increase in lung growth. Hypoplastic lungs presented higher expression levels of the RA receptors α and γ. Moreover bombesin and ghrelin supplementation, in vitro, to normal lungs increased RA receptor α/γ expression whereas administration of bombesin and ghrelin antagonists to normal and hypoplastic lungs decreased it. These data reveal for the first time that there is a link between neuroendocrine factors and RA, and that neuroendocrine factors sensitise the lung to the RA action through RA receptor modulation. ABSTRACT: Congenital diaphragmatic hernia (CDH) is characterised by a spectrum of lung hypoplasia and consequent pulmonary hypertension, leading to high morbidity and mortality rates. Moreover, CDH has been associated with an increase in the levels of pulmonary neuroendocrine factors, such as bombesin and ghrelin, and a decrease in the action of retinoic acid (RA). The present study aimed to elucidate the interaction between neuroendocrine factors and RA. In vitro analyses were performed on Sprague-Dawley rat embryos. Normal lung explants were treated with bombesin, ghrelin, a bombesin antagonist, a ghrelin antagonist, dimethylsulfoxide (DMSO), RA dissolved in DMSO, bombesin plus RA and ghrelin plus RA. Hypoplastic lung explants (nitrofen model) were cultured with bombesin, ghrelin, bombesin antagonist or ghrelin antagonist. The lung explants were analysed morphometrically, and retinoic acid receptor (RAR) α, ß and γ expression levels were assessed via Western blotting. Immunohistochemistry analysis of RAR was performed in normal and hypoplastic lungs 17.5 days post-conception (dpc). Compared with the controls, hypoplastic lungs exhibited significantly higher RARα/γ expression levels. Furthermore considering hypoplastic lungs, bombesin and ghrelin antagonists decreased RARα/γ expression. Normal lung explants (13.5 dpc) treated with RA, bombesin plus RA, ghrelin plus RA, bombesin or ghrelin exhibited increased lung growth. Moreover, bombesin and ghrelin increased RARα/γ expression levels, whereas the bombesin and ghrelin antagonists decreased RARα/γ expression. This study demonstrates for the first time that neuroendocrine factors function as lung growth regulators, sensitising the lung to the action of RA through up-regulation of RARα and RARγ.

The role of ephrins-B1 and -B2 during fetal rat lung development.

UNLABELLED: BACKGROUND/ AIMS: The knowledge of the molecular network that governs fetal lung branching is an essential step towards the discovery of novel therapeutic targets against pulmonary pathologies. Lung consists of two highly branched systems: airways and vasculature. Ephrins and its receptors, Eph, have been implicated in cardiovascular development, angiogenesis and vascular remodeling. This study aims to clarify the role of these factors during lung morphogenesis. METHODS: Ephrins-B1, -B2 and receptor EphB4 expression pattern was assessed in fetal rat lungs between 15.5 and 21.5 days post-conception, by immunohistochemistry. Fetal rat lungs were harvested at 13.5 dpc, cultured during 4 days and treated with increasing doses of ephrins-B1 and -B2 and the activity of key signaling pathways was assessed. RESULTS: Ephrin-B1 presents mesenchymal expression, whereas ephrin-B2 and its receptor EphB4 were expressed by the epithelium. Both ephrins stimulated pulmonary branching. Moreover, while ephrin-B1 did not affect the pathways studied, ephrin-B2 supplementation decreased activity of JNK, ERK and STAT. This study characterizes the expression pattern of ephrins-B1, -B2 and EphB4 receptor throughout rat lung development. CONCLUSION: Our data highlight a possible role of ephrins as molecular stimulators of lung morphogenesis. Moreover, it supports the idea that classical vascular factors might play a role as airway growth promoters.

The monocarboxylate transporter inhibitor α-cyano-4-hydroxycinnamic acid disrupts rat lung branching.

BACKGROUND/AIMS: The human embryo develops in a hypoxic environment. In this way, cells have to rely on the glycolytic pathway for energy supply, leading to an intracellular accumulation of monocarboxylates such as lactate and pyruvate. These acids have an important role in cell metabolism and their rapid transport across the plasma membrane is crucial for the maintenance of intracellular pH homeostasis. This transport is mediated by a family of transporters, designated by monocarboxylate transporters (MCTs), namely isoforms 1 and 4. MCT1/4 expression is regulated by the ancillary protein CD147.The general aim of this study was to characterize the expression pattern of MCT1/4, CD147 and the glucose transporter GLUT1 during human fetal lung development and elucidate the role of MCTs in lung development. METHODS: The expression pattern of MCT1/4 and GLUT1 was characterized by immunohistochemistry and fetal lung viability and branching were evaluated by exposing rat fetal lung explants to CHC, an inhibitor of MCT activity. RESULTS: Our findings show that all the biomarkers are differently expressed during fetal lung development and that CHC appears to have an inhibitory effect on lung branching and viability, in a dose dependent way. CONCLUSION: We provide evidence for the role of MCTs in embryo lung development, however to prove the dependence of MCT activity further studies are waranted.

Associated factors to human papillomavirus vaccine adhesion in adult women: a cross-sectional study.

Background: The goal of human papillomavirus (HPV) vaccination is the decline of HPV related premalignant lesions, leading to prevention of cervical cancer. Vaccination against HPV is recommended until the age of 45 to prevent viral reinfections and reactivations. The aim of this study was to evaluate adhesion to HPV vaccination and their associated factors in adult women. Methods: Cross-sectional study in two tertiary hospitals, with a questionnaire distributed to women born between 1974 and 1992, from September till November 2019. Data collected included sociodemographic information, clinical information, knowledge about HPV, and the HPV vaccine and data regarding vaccine recommendation. Factors associated with vaccination were searched by bivariate and multivariate statistical analysis. Results: In 469 questionnaires, 25.4% (n = 119) women were vaccinated. The main reason for not vaccinating was the non-recommendation (n = 276; 70.2%). In bivariate analyses, vaccinated women were younger, predominantly not married, had higher educational level, and higher careers (P ≤ .001); an abnormal cytology, HPV infection or previous excision of the transformation zone were associated with a 3 to 4-fold increase in the odds of vaccination. Age, high-risk HPV infection, and knowing someone vaccinated remained factors independently associated with HPV vaccination in the multivariate analyses (P< .05). The recommendation of "vaccinate immediately" was independently associated with effectively doing it (P< .001). Conclusions: HPV vaccination is associated with vaccine recommendation, especially if it is recommended to do immediately. These results reinforce the need of health professionals to be aware of the impact that their recommendation has on adhesion to HPV vaccination.

A novel label-free electrochemical immunosensor for detection of surfactant protein B in amniotic fluid.

The deficiency of Surfactant Protein B (SPB) in Amniotic Fluid (AF) has been associated with severe respiratory pathologies in newborns such as respiratory distress syndrome (RDS). Early-stage diagnosis of these respiratory complications by monitoring the SPB in AF could be a significant clinical tool and the first step toward adopting efficient therapy. In this work, we report the development of a novel sensitive electrochemical immunosensor for the detection of SPB in human AF samples. The surface biofunctionalization steps were characterized by using Electrochemical Impedance Spectroscopy (EIS), cyclic voltammetry, and quartz crystal microbalance techniques. The performance of the immunosensor was investigated by EIS, showing a linear dynamic range between 2 ng/mL and 2000 ng/mL and a limit of detection of 0.1 ng/mL. The immunosensor was also selective and specific to other surfactant proteins presented in AF. To the best of our knowledge, this is the first developed electrochemical biosensor for SPB detection. More significant, the immunosensor developed is simpler than the traditional enzyme-linked immunosorbent assay, and present a higher sensitivity for SPB, indicating that it could be a promising alternative approach for SPB detection in the clinical diagnosis.

Local fetal lung renin-angiotensin system as a target to treat congenital diaphragmatic hernia.

Antenatal stimulation of lung growth is a reasonable approach to treat congenital diaphragmatic hernia (CDH), a disease characterized by pulmonary hypoplasia and hypertension. Several evidences from the literature demonstrated a possible involvement of renin-angiotensin system (RAS) during fetal lung development. Thus, the expression pattern of renin, angiotensin-converting enzyme, angiotensinogen, type 1 (AT1) and type 2 (AT2) receptors of angiotensin II (ANGII) was assessed by immunohisto-chemistry throughout gestation, whereas the function of RAS in the fetal lung was evaluated using fetal rat lung explants. These were morphometrically analyzed and intracellular pathway alterations assessed by Western blot. In nitrofen-induced CDH model, pregnant rats were treated with saline or PD-123319. In pups, lung growth, protein/DNA ratio, radial saccular count, epithelial differentiation and lung maturation, vascular morphometry, right ventricular hypertrophy and overload molecular markers, gasometry and survival time were evaluated. Results demonstrated that all RAS components were constitutively expressed in the lung during gestation and that ANGII had a stimulatory effect on lung branching, mediated by AT1 receptor, through p44/42 and Akt phosphorylation. This stimulatory effect on lung growth was mimicked by AT2-antagonist (PD-123319) treatment. In vivo antenatal PD-123319 treatment increased lung growth, ameliorated indirect parameters of pulmonary hypertension, improved lung function and survival time in nonventilated CDH pups, without maternal or fetal deleterious effects. Therefore, this study demonstrated a local and physiologically active RAS during lung morphogenesis. Moreover, selective inhibition of AT2 receptor is presented as a putative antenatal therapy for CDH.