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Introduction: Opioid use disorder (OUD) is implicated in major obstetrical diseases such as fetal growth restriction. Whether or not opioids directly impact placental trophoblast development and function remains unclear. We sought to examine the expression of opioid receptors (OPRs) in villous trophoblasts and the effect of opioids on placental transcriptomics., Methods: Trophoblast stem (TS) cells and primary human trophoblast (PHT) cells from healthy term placentas were used to assess OPR expression in conditions that enhance trophoblast stemness vs differentiation. Placental RNAseq was conducted using our retrospective cohorts of pregnant people with OUD vs controls, both without major obstetrical complications. RT-qPCR was used to determine the effect of fentanyl on the expression of putative opioid targets and stemness or differentiation-associated genes in TS and PHT cells., Results: Three main OPRs, including OPRM1, OPRD1, and OPRK1 were expressed in term PHT cells cultured in the stemness medium, whereas only OPRD1 and OPRK1 were expressed in TS cells. Interestingly, upon induction of differentiation, the expressed OPR mRNAs in TS or in PHT cells were downregulated. We found 286 differentially expressed long RNAs in placentas from the OUD participants vs controls. While three putative opioid targets differed their expression in stemness vs differentiation states of trophoblasts, fentanyl had no effect on their expression or the expression of major stemness or differentiation-relevant genes in TS and PHT cells., Discussion: Trophoblastic expression of OPRs and opioid RNA targets is impacted by cell differentiation, suggesting differential susceptibility of villous trophoblasts to the effect of opioids., Competing Interests: Declaration of competing interest The authors certify that there are no financial interests that influence the results and findings obtained through this study. Yoel Sadovsky is a consultant to Bio-Rad Laboratories, Inc., (Copyright © 2023. Published by Elsevier Ltd.)

Trophoblast injury is central to clinically relevant placenta dysfunction. We hypothesized that the mRNA of primary human trophoblasts, exposed to distinct injuries in vitro, capture transcriptome patterns of placental biopsies obtained from common obstetrical syndromes. We deployed a CIBERSORTx deconvolution method to correlate trophoblastic RNAseq-based expression matrices with the transcriptome of omics-defined placental dysfunction patterns in vivo. We found distinct trophoblast injury patterns in placental biopsies from women with fetal growth restriction and a hypertensive disorder, or in biopsies clustered by their omics analysis. Our RNAseq data are useful for defining the contribution of trophoblast injuries to placental dysfunction syndromes., Competing Interests: Declaration of competing interest The authors certify that there are no financial interests that influence the results and findings obtained through this study. As stated on the title page, Yoel Sadovsky is a consultant to Bio-Rad Laboratories, Inc., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Introduction: The chromosome 19 miRNA cluster (C19MC) encodes a large family of microRNAs (miRNAs) that are abundantly expressed in the placenta of higher primates and also in certain cancers. In the placenta, miRNAs from this cluster account for nearly 40% of all miRNAs present in trophoblasts. However, the function of these miRNAs in the placenta remains poorly understood. Recent observations reveal a role for these miRNAs in cell migration, and suggest that they are involved in the development and function of the human placenta. Here, we examine the placenta in transgenic mice expressing the human C19MC miRNAs., Methods: We produced transgenic mice using pronuclear microinjection of a bacterial artificial chromosome plasmid carrying the entire human C19MC locus and derived a homozygous line using crossbreeding. We performed morphological characterization and profiled gene expression changes in the placentas of the transgenic mice., Results: C19MC transgenic mice delivered on time with no gross malformations. The placentas of transgenic mice expressed C19MC miRNAs and were larger than wild type placentas. Histologically, we found that the transgenic placenta exhibited projections of spongiotrophoblasts that penetrated deep into the labyrinth. Gene expression analysis revealed alterations in the expression of several genes involved in cell migration, with evidence of enhanced cell proliferation., Discussion: Mice that were humanized for transgenically overexpressed C19MC miRNAs exhibit enlarged placentas with aberrant delineation of cell layers. The observed phenotype and the related gene expression changes suggest disrupted migration of placental cell subpopulations., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Introduction: Imprinted genes are preferentially expressed from one parentally inherited allele, and many are crucial to the regulation of placental function and fetal growth. Murine Krüppel-like factor 14 (Klf14) is a maternally expressed imprinted transcription factor that is a component of the Mest imprinted gene cluster on mouse chromosome 6. We sought to determine if loss of Klf14 expression alters the course of normal mouse extraembryonic development. We also used high-throughput RNA sequencing (RNAseq) to identify a set of differentially expressed genes (DEGs) in placentas with loss of Klf14., Methods: We generated a Klf14 knockout (Klf14 null ) mouse using recombineering and transgenic approaches. To identify DEGs in the mouse placenta we compared mRNA transcriptomes derived from 17.5dpc Klf14 matKO and wild-type littermate placentas by RNAseq. Candidate DEGs were confirmed with quantitative reverse transcription PCR (qPCR) on an independent cohort of male and female gestational age matched Klf14 matKO placentas., Results: We found that 17.5dpc placentas inheriting a maternal null allele (Klf14 matKO ) had a modest overgrowth phenotype and a near complete ablation of Klf14 expression. However, there was no effect on fetal growth. We identified 20 DEGs differentially expressed in Klf14 matKO placentas by RNAseq, and subsequently validated five that are highly upregulated (Begain, Col26a1, Fbln5, Gdf10, and Nell1) by qPCR. The most enriched functional gene-networks included those classified as regulating cellular development and metabolism., Conclusion: These results suggest that loss of the maternal Klf14 locus in the mouse placenta acts results in changes in gene expression patterns that modulate placental growth., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Introduction: Cultured primary human trophoblasts (PHT), derived from term placentas, are relatively resistant to infection by diverse viruses. The resistance can be conferred to non-trophoblastic cells by pre-exposing them to medium that was conditioned by PHT cells. This antiviral effect is mediated, at least in part, by microRNAs (miRNA) expressed from the chromosome 19 microRNA cluster (C19MC). Recently we showed that PHT cells and cells pre-exposed to PHT medium are also resistant to infection by Zika virus (ZIKV), an effect mediated by the constitutive release of the type III interferons (IFN) IFN lambda-1 and IFN lambda-2 in trophoblastic medium. We hypothesized that trophoblastic C19MC miRNA are active against ZIKV, and assessed the interaction of this pathway with IFN lambda-1 - mediated resistance., Methods: Term PHT cells were cultured using standard techniques. An osteosarcoma cell line (U2OS) was used as non-trophoblastic cells, which were infected with either ZIKV or vesicular stomatitis virus (VSV). Trophoblastic extracellular vesicles (EVs) were produced by gradient ultracentrifugation. RT-qPCR was used to determine viral infection, cellular or medium miRNA levels and the expression of interferon-stimulated genes., Results: We showed that C19MC miRNA attenuate infection of U2OS cells by ZIKV, and that C19MC miRNA or exosomes that contain C19MC miRNA did not influence the type III IFN pathway. Similarly, cell exposure to recombinant IFN lambda-1 had no effect on miRNA expression, and these pathways did not exhibit synergistic interaction., Discussion: PHT cells exert antiviral activity by at least two independent mechanisms, mediated by C19MC miRNA and by type III IFNs., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Fetal activity in multiple pregnancies, as expressed by maternal daily fetal movement recordings, was compared with that in singleton pregnancies. It was found that the mean fetal activity in 33 cases of twins, six cases of triplets, one case of quadruplets, and one case of quintuplets was generally higher than that obtained during singleton pregnancies. In most cases, fetal movements of triplets, quadruplets, and quintuplets were even higher than the movement of twins.

Traumatic intrauterine adhesions are known to be reversible. In some cases, following lysis of adhesions we find residual adhesions on hysterography. We have studied 11 cases in whom new adhesion formation, of different type and location have recurred following curettage for lysis of adhesions or in subsequent pregnancy. In each case there were between two and five different forms of intrauterine adhesions during the course of study. In all 11 cases hypomenorrhea was a prominent symptom. On premestrual curettage a scanty amount of endometrium was removed. Their obstetric history was characterized by sterility (four cases) and repeated early abortions, spontaneous or missed. Treatment consisted of repeated curettage and placement of an IUD. Four patients who subsequently become pregnant carried pregnancy to term or near term. Intrauterine adhesions may be considered as a localized manifestation of more generalized process of endometrial fibrosis. This may explain this syndrome of recurrent intrauterine adhesions and the paradoxical phenomenon of limited adhesions being associated with amenorrhea and hypomenorrhea.

The association between fetal heart rate (FHR) accelerations and fetal movements during uterine contractions was studied in 52 pregnant women near term or at the beginning of labor. FHR and uterine contractions were recorded by tococardiograph. At the same time, fetal movements, whether associated or not with contractions, were viewed by real-time ultrasound. During uterine contractions, 95.5% of the FHR accelerations were associated with fetal movements. Also, 90.9% of the accelerations which appeared when the uterus was not contracting were associated with fetal movements. Fetal movements were not seen in 91% of uterine contractions which were not associated with FHR accelerations. The suggestion is made that uterine contractions stimulate both fetal movements and FHR accelerations.

A case is presented in which external vibratory acoustic stimulation performed on a term fetus produced severe prolonged bradycardia, necessitating emergency cesarean section delivery. The possible pathogenesis is discussed.

The rate of fetal heart rate accelerations associated with fetal movements to total fetal movements of twin pregnancies was found to be significantly lower than that of pregnancies with singleton infants. The number of fetal heart rate accelerations was similar. As fetal heart rate accelerations are reflective of fetal movements, the results indicate increased fetal activity in twins that is related to an additive effect of two normally active fetuses.

The fetal response to vibratory acoustic stimulation during periods of low fetal activity and low fetal heart rate reactivity was studied in 10 healthy pregnant women at term. In each case, two periods of low reactivity were studied. Consecutive cases alternated: The vibratory acoustic stimulus was applied 10 minutes after the first nonreactive period in half of the cases; the remainder were stimulated during the second nonreactive period. The unstimulated period served as a control. After vibratory acoustic stimulation the baseline fetal heart rate, the mean number of fetal heart rate accelerations, and, the number of fetal movements were significantly increased with values in the control nonstimulated periods (p less than 0.0001). This consistent response to vibratory acoustic stimulation may prove to be clinically useful in altering periods of low reactivity observed during nonstress testing of normal fetuses.