In-utero exposure to mercury and other trace metals pose a significant threat to child health and development, but exposures and health impacts in artisanal and small-scale gold mining (ASGM) environments are poorly defined.We describe the CONAMAD study design, a prospective birth cohort consisting of multiparous women (18 and over) living in rural and peri-urban Peruvian Amazon communities exposed to ASGM.Pregnant women are enrolled from health posts across four zones of Madre de Dios, Peru. Data are collected at enrollment, childbirth, and (planned) 36-48 months. At enrollment, hair samples for mercury assessment, demographic and clinical data are obtained. At birth, we obtain venous and cord blood, placenta, hair, toenails, and saliva.Two hundred seventy mothers were enrolled at an average 20 weeks gestational age with no differences in maternal characteristics across zones. Two hundred fifteen mothers were successfully followed at birth. We obtained 214 maternal and cord blood samples, 211 maternal and 212 infant hair samples, 212 placenta samples, 210 infant saliva samples, and 214 infant dried blood spots. Data collected will allow for testing our primary hypotheses of maternal malnutrition modifying ratios of cord:maternal blood total mercury (tHg), cord blood:maternal hair tHg, and infant:maternal hair tHg, and whether chemical mixtures  have synergistic effects on infant neurodevelopment.CONAMAD is designed to collect and store samples for future processing and hypothesis testing associated with in-utero mercury exposure and child development. We have completed the exposure assessments and will conduct a follow-up of mothers to evaluate early child development outcomes, including developmental delay and growth. These data offer insights into disease mechanisms, exposure prevention, and policy guidance for countries where ASGM is prevalent. Methylmercury (MeHg) exposure from dietary intake is likely to lead to detrimental internal doses in susceptible groups, particularly in developing fetuses due to its ability to cross the blood brain barrier 2. When e579101114Neurobehavioral and immune function can also be impaired from perinatal exposure to MeHg with other potentially toxic trace elements , as compared to those exposed in adulthood , possibl182021Figure 1). The population of Madre de Dios has high exposure incidence to dietary mercury due to ASGM-related mercury pollution and consumption of high trophic level fish [24Cohorte de Nacimiento de Madre de Dios (Birth Cohort of Madre de Dios), which roughly translates as “with mothers” in Spanish. NIEHS (1R21ES026960) provided the primary funding for the design and data collection for this study. Funding for data collection was also provided by the Doris Duke Clinical International Research Fellowship and the Duke Center for Latin American and Caribbean Studies. Additional funding for data processing was provided by the NIEHS Superfund Research Program (P42ES010356) and the Josiah Charles Trent Memorial Foundation Endowment Fund (Grant #17–23). The 36–48-month follow-up is funded by the State University of New York-Albany. The study was motivated by the 2016 El Niño event for which we hypothesized that large-scale flooding from El Niño would increase mercury methylation and bioaccumulation, resulting in higher dietary mercury exposure. We specified three hypotheses:In November 2016, we initiated a birth cohort to enable assessment of peri-conceptional, late gestation and early life stages as potential critical windows of developmental exposure to MeHg. The study is in Madre de Dios, Peru, a southwestern region of the Amazon  will lead to greater than additive impacts on infant neurocognitive function via indirect  and direct effects (infant exposure ➔ cognitive deficits).Funding was used to maximize data collected from mothers and newborns and store the majority of data for future analysis. Data collected include: hair, toenails, blood and placenta for assessment of multi-panel trace elements exposure; clinical health outcomes ; saliva and dried blood spots (DBS) for epigenetic analysis from newborn and their siblings from the same biological parents (as a control); neurocognitive impairment ; and socio-demographic and nutritional measures from survey data. Here we describe the study design, enrollment and birth characteristics of mother-child dyads, and initial findings from enrollment and birth data.Figure 1), and has the lowest population density in Peru .Madre de Dios (MDD) borders Brazil and Bolivia (digenous ) and onedigenous . FamilieLike many regions around the world, MDD is undergoing rapid development. The construction of the Interoceanic Highway  32 and ex33343536243839404142Centros de Salud can attend “imminent” births. If a woman arrives at a birthing clinic but is not near initiating labor, she must return home, stay with nearby family, or pay for subsidized housing, which tend to be in disrepair. Thus, it is not uncommon for women, with a controlled, normal pregnancy, to wait until the last minute for an “imminent” delivery that can be attended in her local health facility.In Peru, the Ministry of Health recommends a minimum of 6 antenatal care visits. Clinical care during a prenatal visit usually includes administration (or prescription) of iron tablets or syrup, medicine for intestinal parasites, maternal weight, blood pressure, blood sample (for anemia assessment), belly width, and neonatal tetanus shot, among others. Women usually give births in designated birthing clinics . Other Investigacion de Migracion, Ambiente y Salud or IMAS) [We leveraged two two-stage cluster samples to design the birth cohort: the 2014 Encuesta Demografica y de Salud Familiar ; and ouor IMAS) 46. ENDESIMAS exhibited agreement for most indicators of interest. The total fertility rate was approximately 3 births per woman, 12% of WCBA were married , and WCBA received between 8 and 9 years of schooling. Anemia prevalence among WCBA was 29.1% in ENDES versus 43% in IMAS . Surveys reported WCBA with similar prevalence of short stature , an indicator of increased risk for adverse maternal and child health outcomes, and for prevalence of overweight or obese (body mass index or BMI > 25), which was 66% of WCBA. Both surveys indicated high prenatal and childbirth health care utilization, with 93% of pregnant women from both surveys reporting at least 4 prenatal care visits with the median gestational age of the first visit occurring at three months. Overall, 96% of women from ENDES and 87% from IMAS who gave birth in the five years prior to the survey did so in a health clinic or hospital, and 97% (ENDES) and 87% (IMAS) of births were attended by a health professional . Additional data in ENDES not reported in IMAS included maternal report of underweight at birth , very small size at birth (1.5%) and smaller size than average at birth (21%). Both surveys indicated an approximate stunting prevalence around 10% in children under five; however, ENDES reported 51% of children under five are anemic, while IMAS estimated 70%.ENDES and CONAMAD is a prospective birth cohort to measure in-utero  mercury exposure in pregnant mothers and the impact of exposure on newborn health and development. Human subjects research approval is through the Universidad Peruana Cayetano-Heredia (UPCH) Comité Institucional de Ética (CIE) para Humanos (SIDISI 66471), the DIRESA Madre de Dios, and the Duke University Office of Research Support via an Inter-Institutional Agreement with UPCH. Follow-up was further approved by the University at Albany Offices of Research Support via Inter-Institutional Agreements with UPCH.Figure 1): (1) Huepetuhe vicinity, representing older and larger gold mining areas; (2) La Pampa vicinity, representing more recent and emerging gold mining; (3) the Puerto Maldonado urban and sub-urban vicinity; and (4) Iberia vicinity, representing non-ASGM impacted communities. These four categories were defined from prior data [41a priori, that mercury exposure would be highest in zones 1 and 2, moderate in zone 3 and low in zone 4. We note that the highest concentration of indigenous communities is in zone 1; however, no indigenous mothers were enrolled.Enrollment occurred in four zones across MDD to capture pregnant women living in four general categories of environmental mercury exposure . Pregnant women were enrolled during any trimester prior to 30 weeks gestation. To be eligible, pregnant woman met the following criteria: (1) primary residence in MDD; (2) aged 18–30 years old; (3) reside with a domestic partner or spouse; and (4) have at least one additional child from the same biological parents as the unborn fetus . Exclusion criteria were: (1) clinical diagnosis of type II diabetes before pregnancy; (2) current smoker; (3) residence outside of MDD for more than two weeks during pregnancy (assessed at the time of enrollment); and (4) planning to give birth outside of MDD. Women were invited to participate by study nurses and health professionals at each of the regional hospitals/birth centers or by fieldworkers in close communication with medical personnel. If they agreed to participate, eligibility was ascertained and, if eligible, the interviewer would either review the consent form with the mother or she could review the form herself. If a woman could not read or could not speak Spanish, we provided local translation for her. Consent included approval for individual chemical exposure results to be shared with the local DIRESA and long-term storage of their biological samples for future testing. Once consent was obtained, clinical records of consenting women were stamped to increase the likelihood that a fieldworker would be notified and present at the time of birth. An enrollment survey was administered, a hair sample was obtained to measure prenatal mercury exposure, and a subset of women were given a silicone wristband to wear for seven days to measure exposure to volatile organic compounds (VOCs) and polycyclic aromatic hydrocarbons (PAHs). The enrollment survey includes transcription of prenatal care clinical records, including weight, blood pressure, and supplements administered, diet (frequency of fish consumption by species), and prenatal complications. Previous work in the region by our team on diet helped inform food consumption questions [The study design involves prenatal enrollment and data collection at birth and 36–48 months of age post-partum, with each contact point involving targeted data collection uestions 46..Figure 2At birth, data collected include: a postnatal survey; maternal and newborn hair samples, newborn toenails, and maternal and cord blood for trace elements exposure assessment (focusing primarily on Hg); placenta; and dried blood spots (DBS) and saliva samples for epigenetic testing . Fieldworkers were able to travel to most birthing centers within an hour, except for Iberia, which is four hours from Puerto Maldonado. Medical personnel at each hospital/birthing center were trained on how to take blood, placenta, hair, and nail samples, in case the fieldworker was not present. Three tufts of hair, from the mother and newborn (if possible) were obtained from the occipital region of the scalp. The hair specimens were cut with stainless steel scissors, secured on self-adhesive note paper with the proximal end fixed in the adhesive part of the paper, and placed in plastic zip lock bags with 2–3 silica gel desiccants. Newborn toenails, if possible, were collected using baby clippers and placed in paper envelope bags. Two 6 ml tubes of maternal blood were obtained using BD Vacutainer® Trace Element Plastic Blood Collection Tubes (with EDTA). Four 6 ml tubes of cord blood were similarly obtained by drawing blood from the already cut umbilical cord after birth. Four placenta punch biopsies were obtained (one punch from each of the four quadrants of the placenta) and stored in cryovials filled with AllProtect® tissue reagent as a stabilizer. Cord blood and placenta samples were obtained in a separate room from the mother to avoid contamination. Saliva was obtained from the newborn using a saliva sponge tip. Dried blood spots from the mother and infant (via heel stick) were obtained using Whatman cards and hemoglobin levels for anemia were evaluated.Biomarkers we plan to process include total hair Hg, total blood Hg, and other trace elements for maternal and cord blood samples. Banked (stored) samples include: placenta, saliva, DBS, bracelets for VOCs and PAHs. Duplicate and triplicate samples of hair, and blood will also be used for future stable isotope analysis, epigenetic testing, and cortisol.Figure 2). Anthropometrics, a DBS, and a hemoglobin test will be obtained from all individuals . Hair and toenail samples are obtained from the child and their parents and sibling. Saliva is obtained from the child (if not obtained at birth) and sibling. Blood pressure is obtained from adults (parents and grandparents). Finally, the child will be administered the Ages and Stages Questionnaire (ASQ-3) to screen for developmental delays or disorders  [49We initiated a 24-month follow-up in December 2019 with child ages expected to range from 20–33 months, with most visits occurring at approximately 24 months of age. This follow-up was halted due to COVID-19 with plans to re-start the study by January 2022. Data are planned to be collected from the child (newborn), the child’s parents, sibling, and, if present in the household or living nearby, maternal grandparents ( months) , which h months) 4950. ChiCONAMAD is designed to achieve 80% power with 250 mother-child dyads at the 0.05 significance level to detect a 20% increase in the Hgc:Hgm  in mothers pregnant during El Niño or whose infants were born having low birthweight compared to mothers pregnant after El Niño . Unfortunately, funding and IRB approval were not obtained until October 2016, after the El Niño event. Therefore, we re-computed power to detect an increase in the Hgc:Hgm ratio for malnourished mothers compared to non-malnourished mothers, as measured by short stature and/or anemia during pregnancy. Research indicates that iron deficiency, a cause of anemia, can increase maternal absorption of Hg . In addi52Table 1). Women were regularly enrolled over time . The average gestational age of enrollment was 20 weeks (Table 1), and there were no differences in enrollment timing by zone with 21% of women enrolled during their first trimester, 53% in their second, and 26% in their third (Table 1). Other characteristics of mothers at enrollment did not vary significantly by zone, including median maternal age (28 years) and prevalence of short stature (16.6%), with highest prevalence of short stature in mining areas (zones 1 and 2). Median age of male partners was 30 years and most women lived in a consensual union (94%). The most common forms of employment were driving a taxi (19%), farming (16%) and mining (14%). Other types of employment included construction (9%), machine operation and mechanic (8%), and administrative work (7%). We did not record the place of birth for the mother at enrollment; however, for their male partners, 39% were born in Madre de Dios and 35% in Cusco. Overall, 48% of male partners were born in an Amazon region of Peru and 48% in the highlands.Two hundred seventy pregnant women were enrolled between November 2016 and July 2018 in 12 health posts across the four zones. In 2017, MDD recorded a total of 4137 births ; thus, oInstituto de Investigaciones de la Amazonia Peruana (IIAP) near Puerto Maldonado prior to shipment to our laboratory at Duke University. For each woman, we created a unique ID consisting of the two-digit year of enrollment, two-digit initials of the enrolling health post, two-digit month of enrollment, and two-digit number representing the n-th mother enrolled during that month. This code will identify the maternal family, with the mother assigned the letter A, newborn B, spouse/partner C and so on. All biomarkers  are recorded with the name of the participant and their unique ID.Surveys and biomarkers were stored in a secure office (and freezer) in the A secure ACCESS database was created for enrollment, birth and follow-up survey data. 20% of surveys were screened for data entry errors and if error detection exceeded 10%, all surveys were screened. ACCESS data at enrollment and birth were exported and variables recoded for future analysis in SAS V9.4, R or STATA.–1 Hg2+ dissolved in 5% HNO3 (Brooks Rand). At the start of each batch run, the instrument calibration was confirmed with analysis of a standard reference material (SRM) hair samples (ERM DB001 Hg: 0.365 mg/kg) and verification of measured value within the certified concentration value. Analyses of analytical blanks and the SRM as repeated every 10 samples to verify instrument performance.Trace element analyses were performed on hair and blood samples in the Hsu-Kim laboratory at Duke University using similar protocols are previously described 3342. Bri33–1 gold solution preserved in 5% HCl. Prior to ICP-MS analysis, the samples were diluted with a 2% HNO3/0.5% HCl (v/v) diluent. The diluent included internal standards at a 20 µg L–1 concentration . All elements were analyzed in helium reaction gas mode, except for Se and Fe which were analyzed in hydrogen mode. An aliquot of caprine blood SRM  was analyzed in parallel with each sample batch of 24. Additionally, for each batch of 24 samples, two blood samples were selected and digested in triplicate. The ICP-MS instrument was calibrated and tuned daily at the start of each batch run. Calibrations were verified by an aqueous SRM (High Purity Standards: CRM-TMDW-A). Instrument blanks and calibration checks were performed at least once per 30 samples in the batch run.The concentrations of Hg and other trace elements  were quantified in whole blood samples by inductively coupled plasma mass spectrometry (ICP-MS) (Agilent Technologies 7900). First, a 0.5 mL aliquot of a thawed blood sample was transferred to pre-cleaned polypropylene test tubes and digested with 1 mL of ultra-trace clean nitric acid  and 0.05 ml of ultra-trace clean hydrochloric acid  for two hours at 65°C. After two hours, the samples were allowed to cool and 1 ml of ultra-trace clean hydrogen peroxide was added  to the mixture. The blood was digested for an additional hour. After the digestions, the samples were cooled and spiked with 10 microliters of a 4 mg LResults from metals analysis will be compared against measures of newborn development, including APGAR score, anemia , birth weight and size, two-year growth , and cognitive development (via the ASQ-3).Table 2). One of these 215 mothers had twins and an additional enrolled mother had a stillbirth. Among these women, we obtained 214 maternal and cord blood samples, 211 maternal and 212 infant hair samples, 212 placenta samples, 210 saliva samples, and 214 DBS. Lab processing of trace element contents for blood samples are complete; analysis of hair samples is ongoing. All other samples are stored for future use in a -80C freezer at Duke.Of the 270 pregnant mothers enrolled, we successfully followed up 215 (80%) mothers at childbirth  and 11% of short stature. Anemia was high across all zones, while short stature was more prevalent in zones 2 and 4. The total number of prenatal care visits was highest in zone 2 (7.9 visits) and lowest in zone 3 (6.9 visits). Hypertension prevalence during pregnancy was low overall (6% of pregnancies) but exceeded 8% in zones 1 and 2. Overall prevalence of cesarean births was 18%, but 11.7% were emergency cesareans and occurred primarily in zone 1.There were 55 women lost to birth follow-up. Of these, 28 (51%) were lost due to their failure to notify the study team when the birth was occurring. Another 14 (25%) gave birth in another location . The remaining lost to follow-up include two mothers who had an abortion and 11 with variable reasons such as lacking cell phones to call the study team, nervous about the study, or had a rapid labor such that they delivered at home or somewhere outside the health post. These women lost to follow-up did not significantly differ from those followed up by maternal age, zone, gestational age at enrollment, or stature. However, women not followed at birth had significantly fewer prior pregnancies compared to those who were followed . All women are being re-contacted for follow-up regardless of whether birth data were collected.Characteristics of the newborn are important indicators for impacts of in-utero exposure as well as indicators of early development during the first two years of life. We use several indicators to determine fetal growth outcomes. First, we use World Health Organization definitions for preterm births in three categories based on gestational age  . Second,Table 3). The average gestational age was 39 weeks, with only 5% of births occurring preterm. Similarly, 5% of children were classified as small-for-gestational age and 2% were born under 2500 g. APGAR scores were normal for almost all children. A surprising result was the large size of newborns: 35% were considered large gestational age and 56% were over 3500 g at birth. None of these birth outcomes varied by zone of enrollment (Table 3).Of the 216 children enrolled, 55.6% were male, which was relatively consistent across zones , clinical and health outcomes , and socio-demographic data that enable cross-cutting analyses of environmental factors impacting human health. This is of primary interest to local and regional health and environmental agencies, with whom we have fostered a strong partnership. DIRESA- Madre de Dios has provided access to clinical records as well as helped inform physicians and nurses of our study. The high rate of post-natal visitation has also enabled the use of in-depth clinical data on the children regarding vaccination dates, anthropometry, illnesses, medications, and nutritional supplements, among others.One potential limitation of this study is the relatively small sample size compared to birth cohorts in the Seychelles and Faroe Islands. However, as noted earlier, our enrollment sample of 270 mothers represents between 4.4% and 7% of all births in the region. In addition, our sample size of 215 mother-child dyads still allows us to test our major hypotheses and it is possible for us to augment the sample using the same protocols if additional data are needed for future tests. Another potential limitation in our study is the low prevalence of low birthweight (2%) and small for gestational age (5%) compared to 4.3% and 1.2% reported from ENDES, respectively. It is possible that, given our engagement with mothers, risk for low birthweight decreased, but we expect differences in our clinical definition of these outcomes vs. maternal recall in ENDES.Another limitation of our study is the delay in the original 24-month follow-up due to COVID-19. The study design will remain unchanged when we restart; however, we may suffer from loss of participants due to migration or death.https://sites.globalhealth.duke.edu/panlab/) and email the PI, William Pan (William.pan@duke.edu).We welcome collaborations with other researchers. We have a collaborative research team consisting of experts in environmental science, toxicology, epidemiology, biostatistics, and clinical research. Researchers interested in collaborating can visit our website for more information (The additional file for this article can be found as follows:10.5334/aogh.3152.s1Supplemental Figure 1.CONAMAD Mother’s Date of Enrollment and Expected Date of Birth at the time of enrollment.


In DTR/DTRCd11b mice, depletion of maternal CD11b+ myeloid cells caused preterm birth, neonatal death, and postnatal growth impairment, accompanied by uterine cytokine and leukocyte changes indicative of a proinflammatory response, while adoptive transfer of WT macrophages prevented preterm birth and partially rescued neonatal loss. In a model of intra-amniotic inflammation–induced preterm birth, macrophages polarized in vitro to an M2 phenotype showed superior capacity over nonpolarized macrophages to reduce uterine and fetal inflammation, prevent preterm birth, and improve neonatal survival. We conclude that macrophages exert a critical homeostatic regulatory role in late gestation and are implicated as a determinant of susceptibility to spontaneous preterm birth and fetal inflammatory injury.Macrophages are commonly thought to contribute to the pathophysiology of preterm labor by amplifying inflammation — but a protective role has not previously been considered to our knowledge. We hypothesized that given their antiinflammatory capability in early pregnancy, macrophages exert essential roles in maintenance of late gestation and that insufficient macrophages may predispose individuals to spontaneous preterm labor and adverse neonatal outcomes. Here, we showed that women with spontaneous preterm birth had reduced CD209  Preterm birth affects approximately 15 million infants annually and is the most common cause of death in children under 5 years of age . PretermPreterm labor is a syndrome of multiple etiologies , but a lIncreasingly, macrophages are understood to exert not only immune-regulatory roles but also trophic and remodeling roles that facilitate development and maintain homeostasis in a wide range of peripheral tissues , 30. Ear+ macrophages in DTR/DTRCd11b transgenic mice, and 3) adoptive transfer of in vitro M2-polarized macrophages into mice administered intra-amniotic LPS to induce preterm birth. The results demonstrated a hitherto unappreciated homeostatic role for macrophages in late gestation, exerted by suppression of proinflammatory cytokines to maintain uterine quiescence, sustain pregnancy, and protect the fetus from premature birth and inflammatory injury that compromise normal development and postnatal survival.We proposed that in late gestation, uterine macrophages may exert not only proinflammatory actions but also antiinflammatory homeostatic effects to sustain gestation prior to labor initiation at term. In the current study, we hypothesized that macrophages have essential roles in maintenance of late-gestation pregnancy, and that insufficient macrophages may predispose individuals to spontaneous preterm labor and adverse neonatal outcomes. To test this hypothesis, we undertook a series of experiments including 1) immunophenotyping of decidual macrophages from women with either term or spontaneous preterm birth, 2) targeted depletion of CD11bn = 69 women with a term birth  and n = 29 women with a spontaneous preterm birth . Women were matched for age, BMI, race, and delivery mode . Macrophages with a proinflammatory phenotype   on 16 dpc , and pregnancy and neonatal outcomes were assessed  , 43. Cd1assessed . Adminis/WT dams , consist at term .DTR/DTRCD11b mice given DT were severely affected, but pups of CD11bWT/WT mice given DT were healthy. DTR/DTRCD11b and CD11bWT/WT dams given DT and DTR/DTRCD11b dams given PBS all delivered a similar number of pups (data not shown), but in DTR/DTRCD11b mice given DT, 74% of pups were either born dead or failed to survive the first 24 hours compared with control groups, for which postnatal survival was 100%  also shortened gestational length  and decrDTR/DTRCd11b dams  were reduced by more than 90% in the maternal peripheral blood, myometrium, and decidua 24 hours after DT administration in DTR/DTRCd11b dams compared with control DTR/DTRCD11b dams given PBS or CD11bWT/WT dams given DT  were elevated to a variable degree in the maternal peripheral blood and myometrium of some DTR/DTRCd11b dams after DT administration, but neutrophil recruitment was not evident in the decidua  were generated in vitro and adoptively transferred to DT-injected DTR/DTRCd11b dams  arrays. The first array, targeting innate and adaptive immune response genes, revealed extensive (more than 8–10-fold) upregulation of Il6 and Irf1, as well as elevated Cd14, Tlr9, and Il1b upon CD11b+ myeloid cell depletion  were enriched in women who delivered a healthy term neonate , chemokines , inflammasome-related mediators , and cell adhesion molecules , while Ccl5 was upregulated   , 57, concretion) , 35. An regnancy , 60, whiDTR/DTRCd11b model but not the intra-amniotic LPS model most likely relates to the proinflammatory insult of LPS. Presumably, M2 macrophages were more potent suppressors of inflammation than BMDMs by virtue of their phenotype being further skewed and more stably embedded. In another study, macrophage depletion using anti-F4/80 antibody suppressed preterm birth caused by intravaginal LPS  (WT/WTCd11b (WT FVB/N) were mated with BALB/c males , and the presence of a vaginal plug indicated 0.5 dpc. DTR/DTRCd11b dams were administered 25 ng/g of DT on 16 dpc or 17 dpc to deplete CD11b+ cells. Control DTR/DTRCd11b dams were given 200 μL PBS, and WT/WTCd11b dams were given 25 ng/g DT. Some DTR/DTRCd11b and WT/WTCd11b dams also received progesterone or BMDMs. C57BL/6 dams received an ultrasound-guided intra-amniotic injection of 100 ng LPS. After treatments, dams were monitored to identify delivery time or euthanized for tissue recovery. Preterm birth was defined as delivery within 48 hours of intervention, and the rate was calculated as the proportion of the total number of pregnant mice. Pup survival and weight were evaluated at birth, at 12–24 hours after birth, and at 8 and 21 days of age. Further details are provided in Supplemental Methods.Female io, USA)  and Cd11WT/WTCd11b mice was incubated in media supplemented with 20% L929 conditioned media as described (+ M2-polarized BMDMs from C57BL/6-Tg(CAG-EGFP)131Osb/LeySop mice to allow tracking of passively transferred macrophages recruited into gestational tissues. Further details are provided in Supplemental Methods.Bone marrow from escribed . C57BL/6DTR/DTRCd11b dams 24 hours after administration of DT or PBS on 16 dpc, and macrophages were detected with rat anti–mouse F4/80 by immunohistochemistry. Single-cell suspensions were prepared from the uterine myometrium, uterine decidua, placenta, and fetal liver, as previously described  and placenta were collected from escribed , from Cd+ cells, were analyzed for IL-10, IL-12p70, TNF, and TGF-β concentrations by using Quantikine ELISA Kits . Maternal plasma and amniotic fluid cytokines were measured by using the ProcartaPlex Mouse Cytokine & Chemokine Panel 1A 36-plex  and Luminex 100 SystemFill. cDNA from decidual tissue collected 24 hours after intervention was analyzed by using QIAGEN RT² Profiler Mouse Innate & Adaptive Immune Response and RT² Profiler Mouse Inflammatory Response & Autoimmunity PCR arrays , following the manufacturer’s instructions. Supernatants from myometrial and decidual tissue macrophages, sorted by positive selection of F4/80R arrays . InteracR arrays . Fetal bR arrays . FurtherU test for continuous variables, Fisher’s exact test for binary nominal variables, and χ2 test for categorical nominal variables. For mouse data, Fisher’s exact test was performed to evaluate preterm birth rate and other categorical data, and Kaplan-Meier survival curves were used to plot and compare neonatal survival data (Mantel-Cox test). A Shapiro-Wilk test was performed to determine whether data were normally distributed. Comparisons among multiple groups were performed by using 1-way ANOVA and a post hoc 2-tailed t test, by using the 2-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli to correct for multiple comparisons in normally distributed data sets, or the Kruskal-Wallis test followed by Mann-Whitney U test for non-normally distributed data sets. For RT-qPCR arrays, the –ΔCt values were normalized by calculating the z score of each gene, and then heatmaps were created to represent the mean of the z score of –ΔCt and hierarchical clustering by using Euclidian distance. P values were considered significant when P was less than 0.05.Statistical analyses were performed by using SPSS v19.0 (IBM) and Prism v8.3.0 (GraphPad). For human demographic data, the group comparisons were performed by using the Mann-Whitney Eunice Kennedy Shriver National Institute of Child Health and Human Disease (WSU IRB 082403MP2F and WSU IRB 110605MP2F). All participating women provided written informed consent prior to sample collection.The collection and utilization of human biological materials for research purposes were approved by the IRBs of Wayne State University (WSU) and the DTR/DTRCd11b mice were cared for and used in experiments as detailed in the National Health and Medical Research Council of Australia (NHMRC) Australian Code of Practice for the Care and Use of Animals for Scientific Purposes with ethics approval from the Animal Ethics Committee, University of Adelaide (ethics IDs: M-2010-055 and M-2012-153). A Genetically Modified Organisms Dealing Authorization was obtained from the Institutional Biosafety Committee, University of Adelaide (ID: 10511). The use of C57BL/6, FVB/V, and C57BL/6-Tg(CAG-EGFP)131Osb/LeySop mice in experiments was approved by the IACUC at WSU . The authors adhered to the NIH Guide for the Care and Use of Laboratory Animals .SAR and NGL designed the study; NGL, VGF, PYC, HMG, KRD, and MTB performed research; NGL, SAR, VGF, PYC, and HMG analyzed data; RR provided clinical materials; NGL and SAR wrote the paper.


This study aimed to clarify predictors of preterm birth in pregnancy of women with systemic lupus erythematosus (SLE). We investigated the predictors of preterm birth before pregnancy from the perspective of the importance of preconception care.We analysed fetal outcomes of 108 pregnancies in 74 SLE patients in a retrospective study. We compared pre-pregnancy clinical characteristics and disease activity in these women between the preterm birth and full-term birth groups to select predictive factors for preterm birth before pregnancy.P = 0.029). Multivariate analysis identified history of lupus nephritis  and low C3 level (< 85 mg/dl) at pre-pregnancy  as risk factors for preterm birth. The greater the number of these risk factors, the higher was the preterm birth rate (P = 0.0007). In the case of SLEDAI score ≤ 4, the preterm birth rate was higher in the pre-pregnancy low C3 group (< 85 mg/dl) (42.1%) than in the high C3 group (C3 ≥ 85 mg/dl) (14.7%) (P = 0.018).Eighty-three of 108 pregnancies resulted in live births, of which 27 (25.0%) were preterm births. Pre-pregnancy serum complement 3 (C3) level was significantly lower in the preterm birth group (77.0 mg/dl) than the full-term birth group (87.5 mg/dl) (For patients with a history of LN, treatment management focusing on pre-pregnancy serum complement levels is very important. Systemic lupus erythematosus (SLE) occurs primarily in women of reproductive age . Recent Furthermore, pregnant patients with SLE have poor birth outcomes such as miscarriage, preterm birth, and fetal growth restriction, and there are many cases of preterm birth even if the pregnancy is successful –5. IncreWe investigated the factors that influence preterm birth in pregnancies complicated by SLE using a retrospective case-control study method. We report on the predictors of preterm birth before pregnancy, especially in terms of the importance of preconception care.The subjects were pregnant SLE patients who were admitted to Osaka Medical College Hospital between January 2000 and October 2020. All patients were managed in our hospital from pre-pregnancy to delivery, and all data were also collected in our hospital.All of these patients fulfilled the 1997 American College of Rheumatology revised criteria for SLE ) or number of subjects. Statistical analyses were performed with the chi-squared test or Fisher’s exact test for binary data and with the Mann–Whitney In total, 108 pregnancies in 74 patients were analysed, and the patients’ characteristics are shown in Table One patient was complicated with active lupus nephritis, but there were no active major organ lesions in the other patients. In the past, 29 patients (26.9%) were diagnosed as having lupus nephritis. The serum C3 level before pregnancy was 81 (74–92) mg/dl. Anti-dsDNA antibody and anti-RO/SS-A antibody were positive in 73 (67.6%) and 38 patients (35.2%), respectively. Forty-six patients (43%) were also positive for lupus anticoagulant activity, anti-cardiolipin IgG antibody, or anti-β-2GP1 antibody. Fifteen patients were complicated by APS and 38 by Sjögren’s syndrome. Ninety-one patients (84.3%) were treated with 10 (5–12) mg/dl of GC before pregnancy. GC pulse therapy was previously administered in 27 patients (25.0%). LDA was administered to 15 aPL-positive patients with a history of obstetrical abnormalities, and UFH was co-administered from the first trimester.Figure P < 0.001). There was no difference in the positive rate and antibody titer of anti-dsDNA antibody, or those in the other autoantibodies, between the two groups. Pre-pregnancy serum C3 levels were significantly lower in the preterm birth group (77.5 mg/dl) than in the full-term birth group (87.5 mg/dl) (P = 0.027). In terms of treatment, there was no significant difference in the dose of GC administered before pregnancy, but the past use of GC pulse therapy was higher in the preterm birth group (P = 0.009).Table P < 0.001; 88 vs 107 mg/dl, P < 0.001; and 100 vs 107 mg/dl, P = 0.024, respectively). However, there was no significant difference between the two groups in anti-dsDNA antibody titer and SLEDAI score during pregnancy. The dose of GC during pregnancy was significantly higher in the preterm birth group in the second and third trimesters .Figure p = 0.043). We then performed a multivariate analysis to select risk factors for preterm birth before pregnancy. Candidate risk factors obtained from the results of univariate analysis comparisons between the two groups included age at pregnancy, which is commonly cited as a risk of preterm birth, history of lupus nephritis, past use of GC pulse therapy, and serum C3 level < 85 mg/dl. The multivariate analysis using these variables identified a history of lupus nephritis  and pre-pregnancy serum C3 level < 85 mg/dl  as the risk factors for preterm birth (Table P = 0.0347).We focused on the serum C3 level as a predictor of preterm birth before pregnancy based on a comparison of patient backgrounds in the preterm and full-term birth groups. We performed a ROC analysis to determine a cut-off value for pre-pregnancy serum C3 level that could indicate risks for preterm birth. As a result, the cut-off value for the serum C3 level was set to 84 mg/dl .Figure n = 50) and caesarean section group (n = 33). There was no difference in serum C3 levels between the two groups. However, the frequency of lupus nephritis history was significantly higher in the caesarean section group than that in the spontaneous delivery group (p = 0.042).We compared the serum C3 levels and history of lupus nephritis between the spontaneous delivery group (P = 0.018). The number of weeks of delivery was 37 (35–38) in the low C3 group, which was significantly earlier than that in the high C3 group 38 (37–39) (P = 0.044).In pregnancies with a SLEDAI score > 4, 5 of 7 (71.4%) had preterm birth, and with a SLEDAI score ≤ 4, 21 of 72 (29.1%) had preterm birth. Figure Among the 83 women delivering live babies, 7 (8.4%) had SLE disease activity (medium disease activity or higher) before pregnancy. The pre-pregnancy serum C3 level was 81 (74–92) mg/dl of all. In addition, the presence of proteinuria or cellular casts of 0.5 g/day or more was observed before pregnancy in one woman.Twenty women (24.0%) had increased SLE disease activity (increase in the SLEDAI score of > 3) during pregnancy. Of the women who had undergone GC therapy before pregnancy, 43 (51.8%) had an increased level of GC (8.9 mg/day). GC was newly introduced in 4 women (4.8%) after their pregnancy was discovered. Due to SLE relapse during pregnancy, immunosuppressants were newly initiated during pregnancy in 5 women.We compared SLE disease activity between the preterm and full-term birth groups Fig. . There wWe investigated predictors of preterm birth in a retrospective case-control study from the perspective of preconception care for pregnancies in women with SLE. The preterm birth group had lower serum C3 levels than the full-term birth group before pregnancy, which indicated that a pre-pregnancy serum C3 level < 85 mg/dl and a history of lupus nephritis are potential risk factors for preterm birth in pregnancies with SLE. We also found that the higher the number of these risk factors in pregnant patients with SLE, the higher was the preterm birth rate. Furthermore, it was clarified that the risk of preterm birth increases in cases without SLE disease activity (SLEDAI score ≤ 4), but with low C3 level (< 85mg/dl) before pregnancy. To our knowledge, there have been no previous reports showing a relation between pre-pregnancy serum C3 levels and preterm birth in pregnancies with SLE. This is the first report to show that the combination of pre-pregnancy serum C3 level and a history of lupus nephritis may predict preterm birth in pregnancies with SLE.Pregnancy in women with SLE carries a higher risk of pregnancy complications than that in healthy individuals, and preterm births are reported to be 2.5 times higher . It is iCurrently, there are limited reports on the management and treatment of SLE before pregnancy, which is important from the perspective of preconception care . The preLupus nephritis and its history are associated with poor fetal outcomes and are also known as the risk of preterm birth in pregnancy with SLE . This stIn pregnancy with SLE, it is necessary to pay attention to exacerbation of SLE disease activity and, at the same time, management of the mother. Pregnancy is known to trigger exacerbations of SLE . So far,This study has some limitations. First, it was performed with a small number of patients at a single institution. However, this enabled us to perform the study with unified criteria for monitoring the data of each patient. Second, the type of lupus nephritis could not be identified by renal biopsy in some patients, and therefore, evaluation of the outcome of pregnancy based upon disease type was not possible. Third, changes in the types and usage of immunosuppressants in the pregnant patients with SLE over time may have affected study outcomes. Moreover, although the outcome of pregnancy and SLE disease activity in the mothers could be evaluated up to 1 year after delivery, subsequent growth of the children or long-term outcomes of the mothers could not be evaluated.In preconception care of pregnancies in women with SLE, it is important to reduce the risk of obstetric complications such as preterm birth and improve the prognosis of the mothers and children by fully controlling disease activity and strict pregnancy planning. The present study showed that management focusing on the history of lupus nephritis in addition to serum C3 levels in preconception care is important for predicting and preventing preterm birth in women with SLE. Prediction of preterm birth stratified by these risk factors will help rheumatologists and obstetricians who are struggling to manage pregnancies with SLE. In addition, this preconception care may increase the safety of the life events of pregnancy and childbirth, which have been a challenge for SLE patients. We hope that further research will enable SLE patients to become pregnant, give birth safely, and enrich their lives by spreading preconception care to all pregnant women with SLE.


Scientific Reports 10.1038/s41598-021-84500-6, published online 23 March 2021Correction to: The original version of this Article contained errors in References 27, 43 and 44, where hyperlinks to external data were omitted and as a result were incorrectly given as:27. Kramarenko, A. V. Demo of non-contac polatimetric cardiograph testing, 43. Kramarenko, A. V. Demo of an industrial application (pump work control) of a non-contact rf registration device .44. Kramarenko, A. V. Demo of a car driver monitoring system .The correct references are listed below:https://www.youtube.com/watch?v=gOGvGjJ2QnI (Accessed 23 Jul 2020a).27. Kramarenko, A. V. Demo of non-contac polatimetric cardiograph testing, https://www.youtube.com/watch?v=z-1pzf3iUyM (Accessed 3 Nov 2020b).43. Kramarenko, A. V. Demo of an industrial application (pump work control) of a non-contact rf registration device, https://www.youtube.com/watch?v=yB9uKd3MNxU (Accessed 13 Oct 2020c).44. Kramarenko, A. V. Demo of a car driver monitoring system, The original Article has been corrected.


Resuscitation at birth of infants with Congenital Diaphragmatic Hernia (CDH) remains highly challenging because of severe failure of cardiorespiratory adaptation at birth. Usually, the umbilical cord is clamped immediately after birth. Delaying cord clamping while the resuscitation maneuvers are started may: (1) facilitate blood transfer from placenta to baby to augment circulatory blood volume; (2) avoid loss of venous return and decrease in left ventricle filling caused by immediate cord clamping; (3) prevent initial hypoxemia because of sustained uteroplacental gas exchange after birth when the cord is intact. The aim of this trial is to evaluate the efficacy of intact cord resuscitation compared to immediate cord clamping on cardiorespiratory adaptation at birth in infants with isolated CDH. The Congenital Hernia Intact Cord (CHIC) trial is a prospective multicenter open-label randomized controlled trial in two balanced parallel groups. Participants are randomized either immediate cord clamping (the cord will be clamped within the first 15 s after birth) or to intact cord resuscitation group . The primary end-point is the number of infants with APGAR score <4 at 1 min or <7 at 5 min. One hundred eighty participants are expected for this trial. To our knowledge, CHIC is the first study randomized controlled trial evaluating intact cord resuscitation on newborn infant with congenital diaphragmatic hernia. Better cardiorespiratory adaptation is expected when the resuscitation maneuvers are started while the cord is still connected to the placenta.  Congenital Diaphragmatic Hernia (CDH) is a rare disease (1/3000 pregnancies) caused by diaphragmatic defect with ascension of the abdominal content into the thoracic cavity. Pulmonary consequences of CDH present a broad spectrum of severity . FailureThe CDH EURO Consortium proposes intubating newborn infants with CDH immediately after birth to limit PPHN . TreatmeTraditionally, the umbilical cord is clamped and cut immediately after birth. Clamping the umbilical cord immediately increases systemic peripheral resistance, resulting in an increase in arterial pressure (afterload). However, as the placental circulation receives 30–50% of fetal cardiac output, cord clamping transiently reduces venous return (by 30–50%), which, combined with the increase in afterload, decreases cardiac output . FollowiIn 2015, the International Liaison Committee on Resuscitation (ILCOR) recommended that the cord should not be cut for at least 1 to 3 min after birth in infants not requiring resuscitation . This reThe ex-utero intrapartum treatment (EXIT) procedure has been proposed during a variety of surgical procedures, mainly cervical mass resection, performed at birth to secure the fetal airway or ensure successful transition to postnatal environment . The aimWe have shown in three previous studies in normal newborn lambs and in newborn lambs with PPHN that blood gases did not change 30 min after birth despite lack of breathing compared to the blood gases obtained before birth ,14,15.We also evaluated the safety, feasibility and impacts of intact cord resuscitation (ICR) on cardiorespiratory adaptation at birth in newborn infants with CDH in a prospective pilot study . Resuscitation before cord clamping was possible for all infants in the ICR group. The cord was clamped at 7 ± 3 min after birth. No increase in maternal or neonatal adverse events was observed during the period of ICR. A multicenter randomized clinical study is required to confirm the benefit of intact cord resuscitation in CDH infants on cardiorespiratory adaptation at birth. To know whether intact cord resuscitation improves initial cardiorespiratory adaptation at birth is a major issue. The primary endpoint is the rate of infants with APGAR score < 4 at 1 min or <7 at 5 min.This primary objective is to show the efficacy of intact cord resuscitation compared to immediate cord clamping on cardiorespiratory adaptation at birth in full term newborn infants with isolated CDH. APGAR score is used to assess cardiorespiratory adaptation at birth. APGAR score is based on clinical assessment of color, heart rate, grimace, muscle tone, and respiratory effort. It is used worldwide by neonatal caregivers, both as a measure of the infant’s clinical status as well as a measure of the infant’s response to resuscitation. APGAR score can be assessed during resuscitation maneuvers, including intubation. Slight inter-observer variation may exist in intubated baby mainly due to lack of standardization of scoring respiratory effort states. In order to limit inter-observer variability, we will standardize respiratory effort scoring as previously reported . In ordeAPGAR is universally recognized as a major prognostic variable of neonatal outcome . More sp-Frequency of postpartum hemorrhage (PPH) defined by blood loss ≥ 500 mL;-Frequency of severe PPH, defined by measured blood loss ≥ 1000 mL;-Blood loss volume at 15 min after birth;-Total postpartum blood loss volume ;To ensure maternal safety of the procedure, blood loss will be carefully monitored after birth. A graduated collector bag for blood, placed under the woman’s buttocks just after delivery of the child, will be used systematically to measure the blood lost through the vagina in the immediate postpartum period. This bag will be left in place at least for 15 min. The following maternal safety endpoints will be assessed:-Frequency of infants with the need for epinephrine administration and/or fluid resuscitation;-Frequency of infants with the need for chest compressions;-Pre-ductal SpO2, and heart rate at 1, 5, and 10 min after birth: a pulse oxymeter sensor will be placed at the right hand as soon as possible (within the first minute after birth), which then will be connected to a pulse oxymeter; -Blood gases and plasma lactate concentration at one hour after birth (H1): these quantitative variables can be considered as objective markers of early cardiorespiratory adaptation at birth. -Blood gases, pre- and postductal SpO2, lactate, FiO2 set to obtain preductal SpO2 90–94%, ventilatory parameters , heart rate, blood pressure, and urine output at H1, H24, H48, H72, D7, D28;-Volume of fluid resuscitation during the first 24 h;-Frequency of infants with the need for vasoactive drugs during the first 24 h;-Frequency of infants with the need for pulmonary vasodilator during the first 24 h;-Hemoglobin concentration at H24;-Echocardiographic parameters  at H6, H24, H48, D7, D28;To assess the effect of intact cord resuscitation compared to immediate cord clamping on cardiorespiratory adaptation of infants after birth, the following secondary endpoints will be assessed:-Infant mortality rate at 90-day after birth;-○mechanical ventilation free-days , ○extracorporeal membranous oxygenation free-days ○pulmonary vasodilator treatment free-days ,○O2 supplementation free-days ,○parenteral nutrition free-days  ○Total duration of hospitalization,Infant morbidity outcomes assessed within the first 90 days after birth:To assess the effect of intact cord resuscitation compared to immediate cord clamping on infants’ mortality and morbidity, we choose to assess the number of free-days from medical support in order to address the effect of mortality on the assessment of morbidity. Because of the randomization process, we can assume that the two groups will be similar in term of severity. In the hypothesis that intact cord resuscitation decreases the mortality, it means that more severe CDH infant may survive, which may in turn increase the apparent morbidity. In case of death, the number of free-days from medical support is zero, which will not modify the number of free-days from medical support in the surviving population. Therefore, the following secondary end-points will be assessed:-Number of refusals to participating with the protocol: the reasons for refusal will be recorded ; -Anxiety and depression level assessed by Hospital Anxiety and Depression Scale, HADS. HADS is an auto-questionnaire translated in French, including seven questions to assess anxiety and seven questions to assess depression. HADS questionnaire lasts 3 to 6 min, and is mostly well accepted. Both parents will be requested to answer the questionnaire within the first 3 days after birth, in a calm and neutral room at the maternity ward. Psychologist in charge of the parents will be informed of the results of the test to adapt family support if required; -Semi-structured interviews will be proposed by a psychologist to the parents whatever the issue, at the end of the study period (90 ± 7 days after birth) to assess their personal experience of the resuscitating period at birth, including both early/delayed cord clamping and close/remote resuscitation maneuvers. The interviews tape recordings will be transcribed and then analyzed using thematic coding. The anonymized data will be independently coded by three researchers and compared for consistency of interpretation. The themes that emerged following the final coding will be used for a qualitative analysis of the parental verbatim;To assess parental acceptability and psychological impact of starting resuscitation while the cord is intact as compared to the immediate cord clamping group:The secondary objectives of the studies are as follows:This is a prospective multicenter open-label randomized controlled trial in two balanced parallel groups. Antenatal diagnosis of CDHNo severe additional malformation or chromosomal diseasesFull term No inclusion in another antenatal trialWritten informed consents from the parentsThe fetuses which required Fetoscopic Endoluminal Tracheal Occlusion (FETO procedure) will also be included in the study.Preterm birth before 37 weeks gestational ageOther severe malformation(s) or chromosomal diseasesNeed for emergency cesarean section (“red code”)Twin pregnancyPostnatal diagnosis of additional severe malformation(s) or chromosomal diseasesAfter assessing the inclusion and exclusion criteria, the parents will be informed of the study protocol and the intact cord resuscitation procedure . The written parental consents are requested before 37 weeks gestational age. A web-based randomization with electronic case-report form (eCRF) system is performed at admission for delivery at the maternity ward. Prenatal o/e LHR on ultrasounds and lung volumetry on fetal MRI allow to classify patients in according the severity of the CDH  ,21. RandThe newborn infant should be intubated routinely without bag and mask ventilation, with an endotracheal tube 3.5 mm;Ventilation in the delivery room should be done with a peak pressure below 25 cmH2O, and an initial FiO2 = 1;The goal of treatment in the delivery room is achieving heart rate > 120 beats/min and increasing preductal SpO2 or achieving acceptable preductal SpO2 targets between 80 and 95%;An oro- or nasogastric tube with continuous or intermittent suction should be placed;Group 1: Immediate cord clamping with transfer to the resuscitation roomThe infants with CDH will be included in the study and randomized in one of the two groups. Severity subgroups is considered as a stratified factor in randomization. A dynamic randomization procedure using the Pocock and Simon minimization method is used . WhateveIn the immediate cord clamping group, the cord will be clamped within the first 20 s after birth and the infant will be transferred to the resuscitation room. The newborn infant will be intubated and mechanically ventilated as quickly as possible on the resuscitation table as recommended in the Programme National de Soins. After cardiorespiratory stabilization , the infant will be transferred to the neonatal intensive care unit (NICU). Oxytocin is infused to the mother as recommended in the local protocol .Group 2: Intact cord resuscitation with resuscitation maneuvers performed on a dedicated trolley placed close to the motherIn the intact cord resuscitation group, the umbilical cord will be kept intact during the initial phase of the resuscitation. The infant will be placed on a specifically designed compact trolley with a warmed platform, suitable for commencing resuscitation between the mother’s legs in case of vaginal birth or near the operating table beside the mother in case of cesarean section. This trolley will be fully equipped for resuscitation, including a suction device, gas flowmeter/blender, ventilator, and monitoring system. Its height can be adjusted to position the infant close to the maternal perineum. The infant will be intubated and mechanically ventilated on this trolley; special care will be taken to prevent stretching, compression, or kinking of the cord. The cord will be clamped after cardiorespiratory stabilization will be obtained  or in case of spontaneous placental expulsion. Oxytocin is infused to the mother just after cord clamping. The infant will be then transferred to the NICU. See Except the timing of cord clamping, the national guidelines will be applied during the course of the prenatal and postnatal management.Here is a photograph of the trolley  :Some differences with usual care during implementation of the protocol are as follows:In both groups, although APGAR score is routinely assessed in every baby at 1 and 5 mn, special care will be taken to optimize scoring taken into account the fact that most infants with CDH are intubated at 5 min. APGAR score can be assessed during resuscitation maneuvers, including intubation. Slight inter-observer variation may exist in intubated baby mainly due to lack of standardization of scoring respiratory effort states . In ordean infant who is apneic and requires intubation and ventilation should receive the minimum value of 0 for respiratory effort;an infant who requires artificial ventilation at birth due to irregular or shallow ventilation should score 1;To assess whether an artificially ventilated infant is apnoeic or not, ventilation should be stopped briefly, when possible, to check for the presence of spontaneous respiratory movements .Preductal SpO2 and heart rate are routinely assessed in every baby with CDH. They will be recorded at 1, 5 and 10 min after birth in both groups.Blood gases and lactate concentration are routinely assessed in every baby with CDH. These parameters will be recorded at H1, H12, H24, H48, H72, D7, D28, in both groups. Compared with usual care, no additional blood samples are required for the protocol;In both groups, although noninvasive echocardiography is routinely assessed in every baby with CDH, left and right mean blood flow velocities, pulmonary artery pressure will be recorded at H6, H24, H48, D7, D28. Compared with usual care, no additional echocardiography is required for the protocol;Acceptability by the parents and psychological impact of intact cord resuscitation will be assessed using a questionnaire and a semi-directive interview.The healthcare providers  in charge of the baby or the mother may stop the procedure—i.e., clamp the cord—if they judge it is required for the safety of the patient. The reason for stopping the procedure and the timing at cord clamping will be recorded. In that case, the patient is not excluded from the study and the variable is recorded as planned in the protocol. The patient data are analyzed according to intention to treat principle.The follow-up of the infants is planned till the age of 3 months (90 days), corresponding to the last visit. If the patient is transferred in another hospital, the variable will be recorded till the age of 3 months.The primary objective is to show the superiority of intact cord resuscitation  compared to immediate cord clamping (control group) to improve cardiorespiratory adaptation at birth of CDH infant. The primary endpoint is defined as the rate of infants with low APGAR score, defined by a APGAR score < 4 at 1 min or <7 at 5 min of life. On the basis of literature, we expected that 37% of CDH infant in control arm will have a low APGAR score. To show a 50% relative reduction , with a type I error of 5% (two-sided test), and a power of 80%, we calculated that 90 subjects (mother/infant) per group are needed, which means a total of 180 subjects.Statistical analyses will be independently performed by the Biostatistics Department of University of Lille under the responsibility of Professor Alain Duhamel. For the data analysis, statisticians will be unaware of the treatment group allocation. Data will be analyzed using the SAS software  and all statistical tests will be performed with a two-tailed alpha risk of 0.05. All analyses will be performed in all randomized patients based on their original group of randomizations, according to the intention-to-treat principle. No interim analysis will be performed. A detailed statistical analysis plan will be written and finalized prior to the database lock. Baseline characteristics will be described for each group. Quantitative variables will be expressed as mean (standard deviation), or median (interquartile range) for non-Gaussian distribution. Categorical variables will be expressed as frequencies and percentages. Normality of distribution will be assessed graphically and using the Shapiro–Wilk test.The Data and Safety Monitoring Board (DSMB) is an independent consultative board asked to express an opinion to the sponsor of the study on the benefit/risk ratio and the management of the clinical trial.A DSMB will be set up, composed of the following members, at least: two pediatricians, two obstetricians, one anesthetist, and one methodologist.Its members will not participate in the study and will be independent from the investigation centers. They will be nominated by the sponsor of the study and will participate as volunteers in the respect of confidentiality of the data. -If the number of Serious Adverse Event (SAE) within the first 24 h after birth is twice higher in the “intact cord resuscitation” group after the inclusion of 30 patients,-If the DSMB judges that a reported SAE caused by the procedure requires to stop the study,-If inclusion rate is less than 25% of the total inclusion objectives, 24 months after starting the inclusion in the study. The board will meet after the inclusion of 30, 90 and 150 patients, to address those points, but it could be summoned on other circumstances:-If a mother’s death occurs within three months after she gave birth to her child,-At the request of any PI involved in the study.The DSMB will analyze the management of the study as well as its benefit/risk exposure and might recommend to stop the study in certain circumstances:Presently, 11 centers are participating to the study: two additional centers are expected to include CDH infants. The first center started enrolling patients for 6 months. At the present time, 10 patients have been recruited. For now, parental consents have been obtained in all cases. Recruitment is expected to be completed by December 2023.Resuscitation at birth of infants with congenital diaphragmatic hernia remains highly challenging because of severe failure of cardiorespiratory adaptation at birth.The traditional approach in the delivery room is immediate cord clamping followed by intubation. Initiating resuscitation prior to umbilical cord clamping (UCC) may support this transition, to avoid the loss of venous return and decrease in left ventricle filling caused by cord clamping, ideally increase in pulmonary blood flow should precede cord clamping. This would enable pulmonary venous return to replace umbilical venous return as the primary source for left ventricle preload and to minimize swings in left ventricular output caused by cord clamping. In infants with CDH, the cord is clamped while the pulmonary blood flow is still low. Evidence indicates that utero-placental gas exchange continues after birth when the cord is intact. A previous experimental study in newborn lambs showed that heart rates and right ventricle output were markedly decreased within 120 s of cord clamping. In contrast, if umbilical cord clamping was delayed until after ventilation had commenced, these large changes in heart rate, arterial pressures and flows were greatly reduced, resulting in a much more stable cardiovascular transition after birth ,26. We hPreliminary study has demonstrated the feasibility of intact cord resuscitation. Starting mechanical ventilation of the infant newborn with congenital diaphragmatic hernia before clamping the cord allowed a potential benefit to improved blood pressure at 1 h of life compared to those resuscitated traditionally .p < 0.05). Mean blood pressure was significantly higher in the intact cord resuscitation than in the immediate cord clamp group at H1 (52 ± 7 vs. 42 ± 7 mmHg), H6 (47 ± 3 vs. 40 ± 5 mmHg) and H12 (44 ± 2 vs. 39 ± 3 mmHg) (p < 0.05). Therefore, commencing resuscitation and initiating ventilation while the infant is still attached to the placenta is feasible in infants with CDH. The procedure is safe, and may support the cardiorespiratory transition at birth in infants with CDH [Lefebvre et al. demonstrated that Intact cord resuscitation was associated with higher APGAR scores at 1 and 5 min after birth. The pH was higher and the plasma lactate concentration was significantly lower at one hour after birth in the intact cord resuscitation than in the immediate cord clamp group (pH = 7.17 ± 0.1 vs. 7.08 ± 0.2; lactate = 3.6 ± 2 vs. 6.6 ± 4 mmol/L, with CDH .To our knowledge, CHIC trial is the first prospective multicenter randomized controlled trial to assess the impact of delayed cord clamping during resuscitation of newborn infant with congenital diaphragmatic hernia. Future publication on the primary and secondary results of the CHIC trial study will make a substantial contribution to this important orientation of research and the results will provide clear evidence on the impact of delayed cord clamping during resuscitation of the newborn infant with congenital diaphragmatic hernia.


Solanum melongena L.) peel, a metabolomic analysis was performed in six cultivars with different peel colors. A total of 167 flavonoids, including 16 anthocyanins, were identified based on a UPLC-MS/MS approach. Further analysis revealed that the delphinidins/flavonoids ratio was consistent with the purple coloration of eggplant peels, and SmF3′5′H expression level was consistent with the delphinidin 3-O-glucoside and delphinidin 3-O-rutinoside contents, the main anthocyanins in the purple-peels eggplant cultivars identified in this study. SmMYB113 overexpression promoted anthocyanins accumulation in eggplant peels and pulps. Metabolomic analysis revealed that delphinidins were still the main anthocyanins class in the peels and pulps of SmMYB113-OE4, but most anthocyanins were glycosylated at the 5-position of the B-ring. Our results provide new insights into the anthocyanin composition of eggplant peels and demonstrate the importance of SmMYB113 in stimulating anthocyanin biosynthesis in eggplant fruits.To understand the color formation mechanism in eggplant ( Anthocyanins are colored water-soluble pigments that give flowers, fruits, and tubers blue, red, or purple coloration. Studies have reported that anthocyanins can have anti-diabetic, anti-cancer, anti-inflammatory, anti-microbial, and anti-obesity effects and prevent cardiovascular diseases . TherefoSolanum melongena L.) has received much attention as a functional food. The eggplant extracts have potent antioxidative properties, suggesting their possible involvement in reducing the risk of various disorders  pigment, while delphinidin appears blue-red or purple. Pelargonidin gives an orange hue to flowers and red to some fruits and berries. Glycosylated cyanidin, delphinidin, and pelargonidin are the most common pigments in nature . PeonidiSmMYB113, orthologous to AtMYB75, promote anthocyanins biosynthesis in tobacco leaves in transient expression assays  have been identified to be one of the most important transcriptional regulators in anthocyanins biosynthesis, acting as activators or repressors . Anthocyn assays  and eggpn assays . HoweverSmMYB113 were also investigated using metabolome technology. This study aimed to improve our understanding of the mechanisms of eggplant coloration and to explore whether overexpression of SmMYB113 alters their anthocyanin profiles.In this study, we report the metabolic profiles of flavonoids that may contribute to different pigmentation in eggplants. Six eggplant cultivars with different peel colors representing all possible eggplant colors were examined . In addiS. melongena and one Solanum aethiopicum (mainly cultivated and popular in Africa), and SmMYB113-OE transgenic eggplants were planted in the solar greenhouse at Shandong Agricultural University. Peels of the six eggplant cultivars and peels and pulps of SmMYB113-OE transgenic eggplants were harvested from nine representative fruits of each type in triplicate. The samples were immediately frozen in liquid nitrogen after harvest, and the frozen samples were subjected to anthocyanin content measurement, gene expression analysis, and flavonoids metabolomic analysis.Six eggplant cultivars   includinThe anthocyanin contents were extracted using methods detailed in 1; MS, Tandem mass spectrometry, Applied Biosystems 6500 QTRAP2). The column contained water ACQUITY UPLC HSS T3 C18 . The solvent system contained water (0.04% acetic acid) and acetonitrile (0.04% acetic acid). The gradient program was 95:5 V/V at 0 min, 5:95 V/V at 11.0 min, 5:95 V/V at 12.0 min, 95:5 V/V at 12.1 min, and 95:5 V/V at 15 min. The flow rate was 0.4 ml min–1 at 40°C and the injection volume was 2 μl. Data were acquired using multiple reaction monitoring (MRM) with a triple quadrupole tandem mass spectrometer and processed using Analyst 1.6.1 software.Peels or pulps samples were prepared for flavonoid metabolomic analysis according to Actin gene (GU984779.1) was used as a standard. The relative expression was calculated using the 2–ΔΔCt method . Next, 1 μg RNA was reverse-transcribed into cDNA using a PrimeScript RT Reagent Kit with gDNA Eraser (TaKaRa). Quantitative real-time PCR (QRT-PCR) analysis was performed using SYBR Premix Ex Taq II Kit (TaKaRa) and LightCycler 96 system . The t method .SmMYB113 coding sequence was inserted into the pRI 101 vector containing the 35S-CaMV promoter. The fusion vector was transferred into Agrobacterium strain LBA4404 and introduced into cultivar No. 108. The cut-cotyledons were pre-cultured on MS medium in the dark for 2 days. Then the cotyledons were infected with Agrobacterium (OD600 about 0.6) for 15–20 min. The infected-cotyledons were placed on the differentiation MS medium without Kan and co-cultured in the dark for 2–3 days. The differentiation MS medium consisted of 200 mg L–1 carbenicillin (Cb), 3.0 mg L–1 6-benzylaminopurine (6-BA), 0.1 mg L–1 thidiazuron (TDZ), 7 g L–1 agar, 30 g L–1 sucrose, and 100 mg L–1 kanamycin. After the shoots were differentiated, the explants were placed on MS medium with 200 mg L–1 carbenicillin (Cb) for roots differentiation.The Eggplant is a widespread vegetable bearing different colored peels . AnthocyAnthocyanin biosynthesis is a branch of flavonoid biosynthesis, and colored flavonoids  and their glycosides contribute to the diversity of colors in leaves, fruits, and flowers . In eggpConsidering that No. 109 had the highest relative content of total flavonoids, but the lowest anthocyanin/flavonoid ratio, it was speculated that orange peel color resulted from flavones, flavanols, and isoflavonoids . As showSmF3′H and SmF3′5′H in the peels of eggplant fruits were analyzed using qRT-PCR. The expression ratio between SmF3′5′H and SmF3′H was also calculated. The SmF3′5′H/SmF3′H expression ratio was associated with the anthocyanin profiles variation among the six eggplant cultivars | ≥ 1 and VIP ≥ 1, 147 (in the peels) and 128 (in the pulps) flavonoids differed significantly between SmMYB113-OE4 and WT plants. Most flavonoids contents were primarily increased via SmMYB113 overexpression, except for 10 in peels and 11 in pulps which were downregulated galactoside was the main anthocyanin present in the black carrot cultivar ‘Deep Purple’. The study of R2R3MYBs promoting anthocyanin biosynthesis has mainly focused on regulating structural gene expression  assay were performed between SmMYB113 and Sm3GT and Sm5GT promoters. However, no transcriptional regulation was found  in taproots resulted in cyanidin 3-xylosyl(sinapoylglucosyl)galactoside accumulation and not cyanidin 3-xylosyl(feruloylglucosyl)galactoside accumulation. Although R2R3-MYB played a critical role in anthocyanin accumulation in plant tissues, other anthocyanin regulators also establish anthocyanin biosynthesis equilibrium. R2R3-MYB overexpression in plants cannot change the main anthocyanin class. However, it can disrupt their equilibrium by activating or enhancing the expression of downstream modifier genes, resulting in anthocyanin structure change.Further analysis showed that the delphinidins were the main anthocyanins in the peels of No. 64, No. 76, and B113-OE4 , indicatB113-OE4 , 4E. ComIn addition, previous studies have reported that delphinidin-3-(p-coumaroylrutinoside)-5-glucoside(nasunin) , “TulipaSmF3′5′H expression level in eggplant peels was sufficient to explain the purple color, so the SmF3′5′H/SmF3′H expression ratio was unnecessary. Simultaneously, targeted metabolic profiling of flavonoids was performed on the fruit peels and pulps of SmMYB113 overexpressing eggplant lines. SmMYB113 overexpression significantly increased the anthocyanins and flavonols contents in peels and pulps. Compared with WT, the delphinidin/flavonoid ratio was significantly increased, while no difference or small decreases in the cyanidin/flavonoid and pelargonin/flavonoid ratios were found. The delphinidins significantly accumulated as a result of SmMYB113 overexpression in this study differed slightly from the data reported in other studies. Altogether, our data provide a glimpse into the flavonoid metabolites in eggplant peels with different colors and SmMYB113-overexpressed eggplant peels.Anthocyanins are important chemical components leading to the purple coloration of eggplant peels. This study used targeted metabolic profiling of flavonoids to investigate the flavonoid and anthocyanin structures of six eggplant cultivars with different peel colors. This method detected 167 flavonoid metabolites, including 16 anthocyanins, with various modifications. The 16 anthocyanins could be classified in cyanidins, delphinidins, and pelargonins. According to our data, the purple color of eggplant peels was positively correlated with the delphinidin/flavonoid ratio, and “Mirtillin” and “Tulipanin” were the major anthocyanins in the purple eggplant peels. In addition, The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/GY: investigation and data curation. LL: investigation. MW: project administration. JL: project administration, data curation, software, and writing – original draft, review and editing. FY: project administration, and writing – review and editing. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


This cohort study investigates the association of antenatal fetal growth restriction with mode of delivery and outcomes among neonates who are small or appropriate for gestational age. Is antenatal fetal weight estimation associated with mode of delivery and neonatal outcomes of neonates who are small and appropriate for gestational age (SGA and AGA)?In this cohort study of 100 198 neonates, suspicion of fetal growth restriction (FGR) in neonates who were AGA was associated with cognitive bias in clinicians and with an increase in risk for cesarean delivery by more than 70%. Regardless of antenatal diagnosis of FGR, neonates who were SGA had increased rates of low Apgar scores and increased neonatal intensive care unit admission.These findings suggest that decision-making under risk circumstances among obstetricians may be associated with increased interventions but not improved neonatal outcomes and that such outcomes may be explained by application of prospect theory. Antenatal diagnosis of fetal weight is challenging, and the detection rate of fetal growth restriction (FGR) is low. Neonates with FGR are known to have an increased rate of obstetric intervention during labor, but the association of antenatal fetal weight estimation with mode of delivery and neonatal outcomes among neonates who are small and appropriate for gestational age (SGA and AGA) has not been reported.To evaluate the association of antenatal fetal weight estimation with mode of delivery and neonatal outcomes among neonates who are SGA and AGA, applying psychological concepts of cognitive bias and prospect theory to a model of clinical behavior.This cohort study was conducted between 2019 and 2020 using data from 2006 to 2018 at a tertiary care center in Jerusalem, Israel. Participants were 100 198 term singleton neonates without anomalies who were categorized into 4 groups according to the presence of an antenatal suspicion of FGR and final birth weight. Neonates with false positives  and neonates with true positives  were compared with neonates with AGA antenatal fetal weight estimation, including neonates with false negatives  and neonates with true negatives . Data were analyzed from July 2019 to July 2020.Fetal weight estimation was performed according to sonographic and clinical evaluation at admission to labor, with FGR defined as a birth weight less than the 10th percentile for gestational age. Sonographic fetal weight estimation was performed according to Hadlock formula. Clinical weight estimation was performed by trained obstetricians.The primary outcomes were obstetric intervention and mode of delivery; the secondary outcomes were neonatal Apgar score (with low Apgar score defined as <7) and neonatal intensive care unit (NICU) admission rates.P < .001) and a significantly increased rate of cesarean delivery . Increased NICU admission was found for neonates who were SGA compared with neonates who were AGA , as was increased rate of low Apgar score . In a multivariable model comparing group 1-FP, group 2-TP, and group 3-FN with group 4-TN, suspicion of FGR was independently associated with increased risk of caesarean delivery among neonates in group 1-FP .Among 100 198 neonates eligible for the study , there were 5671 neonates in group 1-FP, 3040 neonates in group 2-TP, 8508 neonates in group 3-FN, and 82 979 neonates in group 4-TN. Mean (SD) maternal age was 28.6 (5.7) years. Among 8711 neonates with suspected FGR, 34.9% were below the 10th percentile at birth, while 65.1% were AGA. Neonates with suspected FGR had a significantly increased rate of induction of labor (group 1-FP: 649 neonates [11.4%] and group 2-TP: 969 neonates [31.9%]) compared with neonates in group 3-FN (1055 neonates [12.4%]) and group 4-TN (7136 neonates [8.6%]) (This study found that antenatal diagnosis of FGR was independently associated with an increase in risk of caesarean delivery by 70% in neonates who were AGA without improvement in neonatal outcomes. These findings suggest that such outcomes may be explained by application of prospect theory and may be associated with cognitive bias in clinical decision-making. The term small for gestational age (SGA) represents newborns whose birth weight is less than the 10th percentile for gestational age.1 Birth weight from the 10th to 90th percentile is defined appropriate for gestational age (AGA).In accordance with the international definitions of fetal and neonatal weight, the term 3Neonates who are SGA have increased rates of perinatal and long-term complications. Timely recognition of fetal compromise and delivery may be warranted in order to prevent intrauterine death or long-term disability.5 The detection rate of FGR is reported to be below 40%.9The antenatal diagnosis of fetal weight is challenging and is based on the combination of sonographic and clinical estimations.11 with approximately 30% of fetuses with FGR estimated to be delivered via CD.13 In addition to the obvious detrimental associations of CD and operative vaginal deliveries with maternal morbidity, the neonatal benefit of CD in neonates with FGR remains unclear, and the optimal mode of delivery of neonates who are SGA remains to be determined.15Various international guidelines do not indicate cesarean delivery (CD) for fetuses with FGR; however, more obstetric interventions during labor and delivery are reported among these fetuses,Given that definite fetal weight is unknown prior to delivery, obstetric clinical decision-making is primarily based on antenatal assessment of fetal and maternal status. This situation can be viewed as an extension of prospect theory: making decisions under risk with uncertain outcomes while subjected to major biases, such as confirmation bias. We sought to investigate cognitive bias associated with fetal weight estimation and other maternal characteristics by evaluating mode of delivery and neonatal outcomes of neonates who were SGA and AGA.STROBE) reporting guideline for cohort studies.This retrospective cohort study was conducted at a single tertiary center, Shaare Zedek Medical Center, between January 1, 2006, and December 31, 2018. Ethical committee approval was obtained from the Shaare Zedek Medical Center institutional review board. Informed consent was waived by the ethical committee given that the study exclusively used deidentified retrospective data. This study follows the Strengthening the Reporting of Observational Studies in Epidemiology  without anomalies were categorized according to the presence of antenatal suspicion of FGR and actual birth weight. FGR was defined as fetal weight estimation less than the 10th percentile and was based on nationally accepted population growth curves.Neonates with FGR were categorized into 2 groups according to birth weight: neonates with false positives  and neonates with true positives . These neonates were compared with neonates with an AGA fetal weight estimation: neonates with false negatives  and neonates with true negatives  . The priFor categorical variables, statistics are provided as numbers and percentages. For continuous variables, statistics are described by mean values with SDs or median values with IQRs.2 test for categorical variables. The association of categorical variables with continuous measurements was tested by 1-way analysis of variance and Kruskal-Wallis tests.Comparisons between groups were performed using Pearson χMultivariable logistic regression models were fitted to estimate the independent association of antenatal weight estimation with rates of CDs and instrumental deliveries (IDs). Models were adjusted for the following maternal and neonatal covariates: maternal age, nulliparity, prior CD, prior abortion, maternal hypertensive disorder, use of assisted reproductive technology (ART), any background maternal clinical condition, and neonatal birth weight. Models were conducted in 2 blocks. Block 1 included group 1-FP, group 2-TP, and group 3-FN, with group 4-TN as the reference group. The 3 variables were forced into the model using the enter method. Block 2 included further adjustment for maternal and gestational characteristics and was performed using the backward stepwise (or logistic regression) method. Odds ratios (ORs) and 95% CIs were reported.P value < .05 was considered statistically significant. Analyses were carried out using SPSS Statistics for Windows version 25.0 (IBM). Data were analyzed from July 2019 to July 2020.All tests were 2-sided. A We identified 100 198 neonates  eligible for the study. This included 5671 neonates in group 1-FP, 3040 neonates in group 2-TP, 8508 neonates in group 3-FN, and 82 979 neonates in group 4-TN .P < .001), hypertensive disorder , ART , and prior CD .Mean (SD) maternal age was 28.6 (5.7) years. Study groups were different in several maternal parameters. Among mothers of neonates in group 1-FP and group 2-TP, compared with group 3-FN and group 4-TN, there were significantly increased rates of nulliparity , elective primary CD , and overall CD . Neonates who were SGA  had significantly increased rates of ID compared with neonates who were AGA  , third-stage placental complications , maternal hemoglobin drop greater than 3 g/dL , and maternal need for blood transfusions . There were no significant differences in rates of maternal readmission or puerperal fever  were different in several parameters from neonates without suspected FGR  regardless of birth weight. Those in group 1-FP and group 2-TP, compared with group 3-FN and group 4-TN, had a significantly increased rate of induction of labor .Multivariable logistic regression models comparing group 1-FP, group 2-TP, and group 3-FN with group 4-TN were fitted to estimate the independent association of antenatal fetal weight estimation with CD and ID rates . Models P = .49). A subanalysis of the data revealed that the increased CD rate in this group (18.3%) was associated with higher odds of nulliparity . When nulliparity was excluded from covariates, suspicion of FGR was associated with CD rate in group 2-TP . Lack of suspicion of FGR, despite SGA birth weight in group 3-FN, was associated with decreased CD risk .True-positive diagnosis of FGR  was not associated with risk of CD . For group 2-TP , the model showed an association with increased ID risk . When SGA was not suspected , an association with increased ID risk was found .For ID, FGR false-positive diagnosis  was associated with decreased risk of ID  birth weight was 3165 (359) g for group 1-FP, 2496.3 (277) g for group 2-TP, 2630.94 (234) group 3-FN, and 3323.77 (158) for group 4-TN. Among 8711 neonates with suspected FGR, 34.9% were below the 10th percentile of birth weight according to national population-based growth curves, while 65.1% were AGA.P < .001) and 5 minutes . Similarly, increased NICU admission rate was found for SGA groups  compared with AGA groups  , as were rates of prolonged NICU stay  and prolonged neonatal hospitalization   were found at 1 minute  .This cohort study found that neonates suspected of FGR had increased rates of obstetric intervention, including induction of labor and primary and overall CD, regardless of actual birth weight. In multivariate analysis, suspicion of FGR in neonates who were AGA was independently associated with an increase in risk for CD by more than 70%. However, in neonates who were SGA, there was no independent association of suspicion of FGR with increased CD, and the increased CD rate in this group was associated with maternal background characteristics, such as nulliparity. Regardless of suspicion of FGR, neonates who were SGA had decreased Apgar scores and increased NICU admission rates.6 that found poor antenatal detection of low birth weight and an increase in obstetric interventions. Antenatal detection of fetal weight is assessed via sonographic or clinical estimation, which depend on multiple factors that may be associated with the accuracy of the evaluation, including equipment, calculation formula, operator or clinician experience, and maternal body mass index.18 The reported percentage error is variable and considered to be approximately 10%. The agreement between sonographic and clinical assessment is high.19More than 65% of neonates antenatally suspected for FGR were AGA birth weight as recorded after delivery. However, these neonates had significantly increased rates of obstetric intervention, including overall and primary CD rate and induction of labor. Our findings are in accordance with a previous publication20 found that routine late-pregnancy ultrasound in low-risk or unselected populations was not associated with benefits for the mother or baby but was associated with an increased CD rate. It may be inferred that poor detection of fetal weight is associated with a large part of the increase in CD in neonates who are AGA.Bricker et alOur findings suggest the cardinal role of obstetrician cognitive bias in the process of obstetric decision-making. As found in the multivariate model in neonates who were AGA, suspicion of FGR was associated with an increase in risk of CD by more than 70%. This interesting finding may be explained by understanding social sciences concepts that interpret human behavior and decision-making under stress.Confirmation bias may be an appropriate explanation for the association of antenatal assessment of FGR with mode of delivery, regardless of actual birth weight. Confirmation bias refers to the human tendency to interpret data in favor of prior beliefs to support a predetermined choice. Applying this bias to the course of events in the delivery suite, we may assume that the clinician has a baseline belief that a small fetus is compromised and has a high probability of experiencing adverse outcomes under contractions. It is well established that fetal heart rate is subjectively interpreted by experts and depends on their experience. We suggest that when analyzing fetal heart rate during labor, the obstetrician may be more prone to interpret heart rate abnormalities as severe decelerations in a fetus with suspected FGR. This outcome associated with confirmation bias may lead the obstetrician to advise for a prompt intervention via an instrumental or cesarean delivery. Similarly, we suggest that suspicion of FGR may be associated with clinical judgment of proper labor progress, hastening the decision to end labor with emergency CD. In other cases, mothers are not granted a trial of labor and undergo an elective CD as reflected by the increased rate of primary-elective CD in group 1-FP compared with group 4-TN (12.0% vs 4.6%).21 and deals mainly with monetary outcomes and financial decision-making under uncertainty. The basic principles regarding decisions with uncertain outcomes may be readily applied to obstetric management at admission to labor. According to prospect theory, human nature is risk-averse, meaning that individuals will take actions to avoid potential losses even at the sacrifice of more likely potential gains. Regarding clinical decision-making for mode and timing of delivery in a parturient with suspicion of FGR, the possible outcomes may be considered losses or gains in the obstetrician’s eyes. When a CD is medically justified, the outcome of not performing an urgent CD is considered a major loss given that the neonate may die or experience a severe disability. Alternatively, the possible gain of not performing a CD consists of avoiding immediate maternal morbidity and financial burden, along with economic and health consequences of repeat CD in the future. A risk-averse obstetrician may choose CD even when the probability of adverse neonatal outcome is small, because the loss looms larger than any potential gain that is not as salient as the loss.Prospect theory was initially described by Kahneman and Tversky in 1979Additionally, medical liability and the obstetrician’s sense of personal responsibility may be greater in the former scenario than the latter. When a prompt intervention is chosen and a healthy baby is born who is SGA or AGA, it may be difficult to question the intervention given that it seems that the main goal was achieved. Exceedingly high CD rates are not a priority in individual patient treatment, but rather the immediate obstetric outcome is the priority. These arguments may explain the predominance of CD when FGR is suspected among neonates who are AGA. Interestingly, in the multivariate model, the lack of suspicion of FGR was associated with a protective outcome, with a decrease in the risk of CD by 30%. Possibly, an antenatal estimation of adequate fetal weight may allow the clinician to be tolerable toward fetal heart rate changes and labor progress, allowing successful vaginal delivery.22 with a shortened period of anticipated fetal compromise during delivery, which may encourage clinicians to favor a trial of labor.Maternal characteristics, such as nulliparity, presence of hypertensive disorder, ART conception, and prior CD, were independent risk factors associated with CD and were found to be more prevalent in the population of neonates suspected of FGR. When adjusted for these factors, increased risk of CD in neonates with suspected FGR who were SGA was not associated with the suspicion but rather with other background maternal characteristics, mainly the increased rate of nulliparity. This may be associated with the expectation that multiparous mothers will have a shorter overall and second stage of labor duration,The rate of IDs among the false-positive group 1-FP was not higher than that of the true-negative group 4-TN (4.4% vs 5.5%). A possible explanation may be that ID is most often performed for nonreassuring fetal heart rate at the second stage of labor. Neonates who are AGA have less fetal distress than those who are SGA, who have fewer reserves to withstand labor stress and the interference of blood flow during contractions. This finding is further supported by the result that group 2-TP was comparable to group 3-FN in having an increased rate of ID, .26 The adverse outcomes may be explained by the natural pathophysiologic events commonly termed placental syndrome.28 The primary events of trophoblastic invasion and formation of proper placental blood vessels are crucial to the establishment of successful placental function that will further promote adequate fetal growth.31 Nulliparous mothers and those with ART conceptions are at risk of poor placental formation and increased vascular-based morbidity.33In our cohort, neonates who were SGA  had increased rates of a low Apgar score, NICU admission, and prolonged hospitalization regardless of antenatal weight assessment. Concomitantly, mothers of neonates who were SGA had increased rates of postpartum placental complications , postpartum hemorrhage, hemoglobin drop of more than 3 g/dL in the first 48 hours postpartum, and need for blood products. These findings are supported by previous reports of poor perinatal outcomes among neonates who were SGA, including increased mortality and cerebral palsy rates.In this cohort of neonates who were SGA, obstetric intervention was not associated with improved neonatal outcomes. This finding suggests that fetal insult may be chronic during pregnancy rather than acute intrapartum.Constant vigilance and personal awareness of possible cognitive bias in obstetric decision-making is the first step toward limiting the association of these biases with maternal and neonatal outcomes among neonates who are AGA.This study raises issues and hypotheses to be further investigated in future studies. First, future studies may need to use novel artificial intelligence techniques to attempt to increase the accuracy of sonographic weight estimation. Second, our findings suggest that novel algorithms for obstetric clinical decision-making should incorporate outcomes associated with confirmation bias and other cognitive biases into calculations of delivery management. Third, this study’s findings suggest that cost calculations should be performed to estimate the financial cost associated with unnecessary CD vs the possible expenses of neonatal disability or death.This study has several limitations. First, the study’s retrospective nature prevents the determination of the causative effect of obstetrician’s cognitive bias and poor neonatal outcomes. Second, arterial pH values are not available for all neonates. Thus, pH data are not included in the study, although these data may contribute to the understanding of physiologic mechanisms underlying the poor neonatal outcomes. Third, umbilical cord Doppler values are not available for all neonates and thus not included in the study, although this information could improve the detection rate of fetuses at risk for poor neonatal outcomes. Fourth, placental pathology specimens are not available for all neonates. Thus, this information is not included in the study, although it may contribute to the understanding of the pathophysiology of increased hemorrhagic morbidity and third-stage placental complications among mothers of neonates who are SGA. At this point, we cannot provide long-term developmental outcomes of study group participants. Fifth, we did not discriminate clinical from sonographic weight estimation given that these data are recorded collectively in our patient records.We found that antenatal fetal weight estimation and diagnosis of FGR were independently associated with clinical decision-making, increased rate of induction of labor, and an increase in risk for CD by more than 70% in neonates who were AGA. Prospect theory and cognitive bias were applied to interpret our findings.