Placental Membranes Research Articles

Visualization of microbes by 16S in situ hybridization in term and preterm placentas without intraamniotic infection

Numerous reports have documented bacteria in the placental membranes and basal plate decidua in the absence of immunopathology using histologic techniques. Similarly, independent metagenomic characterizations have identified an altered taxonomic makeup in association with spontaneous preterm birth. Here we sought to corroborate these findings by localizing presumptive intactbacteria using molecular histology within the placental microanatomy. Here we examined for microbes in term and preterm gestations using a signal-amplified 16S universal in situ hybridization probe set for bacterial rRNA, alongside traditional histologic methods of Warthin-Starry and Gram stains, as well as clinical culture methodologies. We further sought to differentiate accompanying 16S gene sequencing taxonomic profiles from germ-free (gnotobiotic) mouse and extraction and amplicon contamination controls. Placentas were collected from a total of 53 subjects, composed of term labored (n= 4) and unlabored cesarean deliveries (n= 22) and preterm vaginal (n= 18) and cesarean deliveries (n= 8); a placenta from a single subject with clinical and histologic evident choriomanionitis was employed as a positive control (n= 1). The preterm cohort included spontaneous preterm birth with (n= 6) and without (n= 10) preterm premature rupture of membranes, as well as medically indicated preterm births (n= 10). Placental microbes were visualized using an in situ hybridization probe set designed against highly conserved regions of the bacterial 16S ribosome, which produces an amplified stable signal using branched DNA probes. Extracted bacterial nucleic acids from these same samples were subjected to 16S rRNA metagenomic sequencing (Illumina, V4) for course taxonomic analysis, alongside environmental and kit contaminant controls. A subset of unlabored, cesarean-delivered term pregnancies were also assessed with clinical culture for readily cultivatable pathogenic microbes. Molecular in situ hybridization of bacterial rRNA enabled visualization and localization of low-abundance microbes after systematic high-power scanning. Despite the absence of clinical or histologic chorioamnionitis in 52 of 53 subjects, instances of 16S rRNA signal were confidently observed in 13 of 16 spontaneous preterm birth placentas, which was not significantly different from term unlabored cesarean specimens (18 of 22; P > .05). 16S rRNA signal was largely localized to the villous parenchyma and/or syncytiotrophoblast, and less commonly the chorion and the maternal intervillous space. In all term and unlabored cesarean deliveries, visualization of evident placental microbes by in situ hybridization occurred in the absence of clinical or histologic detection using conventional clinical cultivation, hematoxylin-eosin, and Gram staining. In 1 subject, appreciable villous bacteria localized to an infarction, where 16S microbial detection was confirmed by Warthin-Starry stain. In all instances, parallel sample principle coordinate analysis using Bray-Cutis distances of 16S rRNA gene sequencing data demonstrated consistent taxonomic distinction from all negative or potential contamination controls (P= .024, PERMANOVA). Classification from contaminant filtered data identified a distinct taxonomic makeup among term and preterm cohorts when compared with contaminant controls (false discovery rate <0.05). Presumptively intact placental microbes are visualized as low-abundance, low-biomass and sparse populations within the placenta regardless of gestational age and mode of delivery. Their taxonomic makeup is distinct from contamination controls. These findings further support several previously published findings, including our own, which have used metagenomics to characterize low-abundance and low-biomass microbial communities in the placenta.

The telomere gestational clock: increasing short telomeres at term in the mouse

The biologic mechanism(s) regulating the length of gestation are currently poorly understood. After peaking at the blastocyst stage, the average telomere lengths have been reported to shorten during the remainder of gestation in the placenta and fetal membranes in both human and mouse pregnancies, thereby providing a potential countdown biologic clock. These previous studies have reported changes in the average telomere lengths, whereas it has now been shown that the shortest telomeres, not the average telomere lengths, are the mediators of telomere dysfunction which limits cellular survival and results in aging. These studies sought to assess for the first time a significant increase in short telomeres in the fetal membrane and placental tissue near the end of pregnancy in the mouse. Placental and fetal membrane tissues were harvested from timed-pregnant CD-1 mice on gestational days 14-18 prior to the onset of parturition. Telomere lengths were determined for 30 DNA samples (5 each forgestational days 14, 16, and 18 from placentas and fetal membranes) using acommercial high-throughput quantitative fluorescence in situ hybridization technique. Quantitative measurements of representative short telomeres (ie, 3kband 5 kb telomere fragments) were performed for 29-30 DNA samples (4-7 each for gestational days 14, 15, 16, 17, and 18 from placentas, fetal membranes, and maternal liver) using a real-time quantitative polymerase chain reaction modification of the classic telomere restriction fragment technique. The median telomere lengths of fetal membrane tissue decreased from gestational days 14-18 (18,705-16,364 kb) and were significantly shorter than telomeres in placental tissue (P < .05). Representative histograms for the distribution of telomere lengths in mouse fetal membranes (as shown in the Figure) confirm a curve skewed to the left (toward shorter telomere lengths).The relative quantity of the representative short telomeres (ie, 3 kb and 5 kb fragments) increased significantly as gestation progressed in both placenta and fetal membrane tissue. In gestational day 18 fetal membranes, the relative quantity of 3 kb and 5 kb telomeres increased 5.5-fold and 9.3-fold compared with gestational day 14 tissues (P < .05). In placental tissue the relative quantity of 3 kb and 5 kb telomeres increased 9.3-fold and 7.8-fold compared with gestational day 14 tissues (P < .05). Studies performed using adult liver tissue demonstrated little variation of the representative short telomeres and no significant difference between the nonpregnant and pregnant samples. These mouse studies have demonstrated that the distribution of telomere lengths in fetal membrane and placental tissues are skewed toward shorter lengths and that the quantity of representative short telomeres increase significantly prior to parturition. The telomere gestational clock is a novel hypothesis supported by several preliminary mouse studies and interesting associations in human pregnancies between maternal conditions and telomere lengths. (eg, stress, education, pollution, neighborhood quality, and race). As such, the current hypothesis generating study provides a foundation for future research regarding the potential role for a telomere-based biologic clock that determines gestational length in human and other mammalian pregnancies.

Oral and uro-vaginal intra-amniotic infection in women with preterm delivery: A case-control study.

The aim of the present study was to establish the association between the presence of oral and uro-vaginal microorganisms in the placental membrane and preterm delivery (PTD), the premature rupture of membranes (PRM), and the clinical signs of intra-amniotic infection. Eighty-four women with PTD and 127 women with delivery at term were assessed for the PRM, clinical signs of intra-amniotic infection, and the presence of periodontitis. Twenty-seven microorganisms were identified in the placental tissue using nested polymerase chain reaction (PCR). Porphyromonas gingivalis (P.gingivalis) was quantified by droplet digital PCR. The prevalence of microorganisms was 9.47% (20/211). P.gingivalis was the most prevalent (12/211, 5.68%). Mycoplasma hominis, Ureaplasma urealyticum, Staphylococcus spp, and Fusobacterium nucleatum were isolated at a very low frequency in the placenta. Candida albicans was associated with PTD (P=0.027). Periodontitis was associated with clinical signs of infection (odds ratio [OR]=3.8, 95% confidence interval [CI]: 1.28-13.5) and with PTD (OR=1.99; 95% CI: 1.07-3.72). The presence of P.gingivalis in the placenta was not associated with perinatal complications. Detecting microorganisms in the placenta by nested PCR is not relevant, as it has a poor association with clinical variables that establish the diagnosis of chorioamnionitis. However, periodontitis was associated with the clinical signs of intra-amniotic infection and PTD.

Allogeneic Skin Substitutes Versus Human Placental Membrane Products in the Management of Diabetic Foot Ulcers: A Narrative Comparative Evaluation of the Literature.

Outcomes following standard wound care (SWC) for diabetic foot ulcers (DFUs) remain suboptimal. Supplementing SWC with tissue engineered allogeneic cellular wound therapies represents an emerging treatment strategy. This review aimed to evaluate the efficacy and safety of allogeneic skin substitutes and human placental membrane allografts in the management of DFUs. Ovid MEDLINE and Embase databases were searched from inception to October 2017. Any randomized controlled trial (RCT) with an allogeneic skin substitute or placental membrane allograft intervention group was included. Our primary outcome measure was the proportion of completely healed ulcers. Secondary outcome measures included time to complete wound healing and local adverse event rates. Each study was assessed for risk of bias and the quality of evidence was appraised using the GRADE (Grading of Recommendations, Assessment, Development, and Evaluation) approach. Moderate quality evidence from 11 included RCTs demonstrated that both allogeneic cellular approaches improve the proportion of completely healed ulcers at 6 and 12 weeks. One RCT showed that a placental membrane allograft was superior to an allogeneic skin substitute, although this has yet to be repeated in other studies. The addition of allogeneic cellular wound products to SWC improves DFU outcomes. Further studies are required to conclusively establish if placental membrane allografts are superior to allogeneic skin substitutes.

Comparative evaluation of clinical efficacy of coronally advanced flap alone and in combination with placental membrane and demineralized freeze-dried bone allograft in the treatment of gingival recession.

Background:Guided tissue regeneration is a very commonly employed technique for treating recession defects. However, achieving the required space beneath the membrane is often difficult.Aim and Objectives:The purpose of the present study was to compare and evaluate the clinical efficacy of coronally advanced flap (CAF) alone or in combination with amnion membrane (AM) or chorion membrane (CM) and demineralized freeze-dried bone allografts (DFDBAs) in the management of gingival recession (GR) defects.Materials and Methods:The sample size (Miller's Class I and Class II GR-type defects) consisted of thirty patients which were stratified into three groups randomly (ten for each group). Group A patients were treated with only CAF and Group B patients were treated by CAF in combination with CM and DFDBA. Similarly, Group C patients were treated by CAF in combination with AM and DFDBA. Clinical parameters such as gingival index, GR depth, GR width, relative attachment level, and width of keratinized tissue were assessed at baseline and 3 months postoperatively.Results:Inter- and intragroup data were analyzed by paired t-test. The percentage of root coverage obtained in the study groups was 60%, 78%, and 63%, respectively, for Groups A, B, and C. Statistically significant difference was obtained in the clinical parameters of Group B which was treated with CM and DFDBA.Conclusions:Combination of DFDBA and placental membrane (chorion/amnion) in CAF procedure provided an additional benefit over CAF alone in the treatment of Class I and II GR defects.

Contributions of Drug Transporters to Blood-Placental Barrier.

The placenta is the only organ linking two different individuals, mother and fetus, termed as blood-placental barrier. The functions of the blood-placental barrier are to regulate material transfer between the maternal and fetal circulation. The main functional units are the chorionic villi within which fetal blood is separated by only three or four cell layers (placental membrane) from maternal blood in the surrounding intervillous space. A series of drug transporters such as P-glycoprotein (P-GP), breast cancer resistance protein (BCRP), multidrug resistance-associated proteins (MRP1, MRP2,MRP3,MRP4,andMRP5), organic anion-transporting polypeptides (OATP4A1, OATP1A2, OATP1B3, and OATP3A1), organic anion transporter 4 (OAT4), organic cation transporter 3 (OCT3), organic cation/carnitine transporters (OCTN1andOCTN2), multidrug and toxin extrusion 1 (MATE1), and equilibrative nucleoside transporters (ENT1 and ENT2) have been demonstrated on the apical membrane of syncytiotrophoblast, some of which also expressed on the basolateral membrane of syncytiotrophoblast or fetal capillary endothelium. These transporters are involved in transport of most drugs in the placenta, in turn, affecting drug distribution in fetus. Moreover, expressions of these transporters in the placenta often vary along with the gestational ages and are also affected by pathophysiological factor. This chapter will mainly illustrate function and expression of these transporters in placentas, their contribution to drug distribution in fetus, and their clinical significance.

Impairment of production and posttranslational changes of placental nuclear and membrane proteins with complicated pregnancy

The content of nuclear and membrane proteins of the placenta, as well as posttranslational modification of these proteins in physiological pregnancy and placental insufficiency (PI) were studied. Differential centrifugation, electrophoresis in polyacrylamide gel, spectrophotometric methods were used. It was found that with PN there is a decrease in the degree of production of the studied proteins of varying degrees relative to control parameters. For chromatin proteins, a more pronounced decrease in the content of non-histone proteins was found in comparison with histones. Among histone fractions, the maximum decrease was detected in the H2A fraction. The degree of change in the amount of membrane proteins depends on the detergent used. Changes in posttranslational protein modifications disorders are characterized by a decrease in the content of amine and amide (especially difficult to hydrolyze) groups and an increase in carbonyl derivatives of proteins. The revealed changes in the composition and structure of the nuclear and membrane proteins of the placenta, performing numerous regulatory functions, can be triggering links in the chain of molecular damage in the placenta at PI.

521: Intra-amniotic injection alters the intrauterine microbiome in a primate model of inflammatory preterm birth

Identification of causative bacteria in chorioamnionitis has been unyielding, and our prior work examining the intrauterine microbiome in association with histological chorioamnionitis demonstrates the presence of bacteria in the placental membranes that is normally found within oral and vaginal cavities. Here, we aimed to examine changes in the intrauterine microbiome that occur in a primate model of inflammatory preterm birth. Sterile intra-amniotic injections (IA) of saline, lipopolysaccharide (LPS), IL-1B, or Ureaplasma parvum were given to preterm (126-132 days) Rhesus macaques (n=23) with ultrasound guidance. Delivery of offspring occurred via cesarean within one week post-injection. Sterile techniques was utilized in the collection and DNA extraction of oral swabs, stool, placental tissue, vaginal swabs, and meconium. Next-generation sequencing (16S and whole genome shotgun (WGS)) of DNA was performed with negative controls, and resultant sequences were quality filtered prior to analysis. Analysis of 16S sequencing was performed with QIIME, LEfSe, GENE-E, and PICRUSt. Analysis of WGS sequencing was performed using HUMAnN and MG-RAST. Integrative analysis was used to determine the relationships of the microbiome alterations in each cohort with relation to bacterial metabolic pathways. As expected, microbiota of the maternal oral and vaginal cavities was not significantly altered by intra- amniotic injection of LPS, IL-1B, nor U. parvum when examining alpha (Fig. A) and beta diversity. However, intrauterine microbiota were significantly altered by both alpha (p<0.03, Fig. A) and beta (p=0.02, Fig. B) diversity with intra-amniotic injection of inflammatory mediators in comparison to control animals. Furthermore, we found clustering of samples by control or inflammatory injection when examining hierarchal clustering by Spearman’s correlation (Fig. C). This study demonstrates the presence of bacteria in the intrauterine environment due to alterations occurring in placenta microbiota with exposure to inflammatory mediators. Additionally, we find that inflammatory mediators significantly alter the intrauterine microbiome in a clinically relevant primate model.

1010: Polybrominated diphenyl ethers (PBDE) exposed placental and fetal membrane show differential protein expression in exosomes

Flame retardant chemical polybrominated diphenyl ethers (PBDE) exposure is associated with spontaneous preterm birth (PTB). We have shown that PBDE causes fetal membrane senescence and injury to placental cells. To determine if PBDE exposure and cellular level changes are reflected in exososomes released from these tissues, fetal membrane and placental explants were exposed to PBDE congener 99 and profiled their exosomal protein cargo Human fetal membrane and placenta explants from term, not in labor cesarean sections were exposed to PBDE-99, one of the harmful congeners, 25 μM for 48 hours. Exosomes were isolated from media by differential centrifugation followed by size exclusion chromatography. Exososome characterization was done by cryoelectron microscopy (morphology), nanoparticle tracking analysis (size and quantity), western blot (markers) and mass spectroscopy (cargo proteins). Differentially expressed proteins were analyzed by Ingenuity Pathway Analysis (IPA) to identify specific pathways. Membrane and placental explant exosomes shared similar characteristics and markers (Fig 1C, Table I). Compared to respective controls PBDE-99 exposed FM derived exosomes showed acute phase inflammatory response whereas inflammatory pathways were not dominant in placental exosomes (Fig 1D). PBDE exposure indicated FM cell necrosis that was not evident in placenta. Placenta had more immune cell activities than FM (Fig 1 D). Environmental pollutant PBDE-99 did not produce any characteristic changes in exosomes derived from FM or placenta. However, exosome cargo reflected varying pathologic effects by PBDE on FM and placenta. FM appears to be more susceptible to the deleterious effects of PBDE-99 compared to placenta.

Prevalence, Morphology, Embryogenesis, and Diagnostic Utility of Umbilical Cord Vitelline Vascular Remnants.

The objective of this study is to better understand embryonic vitelline vascular remnants in the umbilical cord, to assess their prevalence, to categorize their morphology, and then finally to describe and assess inflammation arising from these structures. During routine placental sign out, the author noted the presence or absence of vitelline vessel remnants for 1 year; when present, he assessed their histologic patterns and noted whether there were neutrophils marginating from the remnants and into the adjacent Wharton's jelly and whether there was any other evidence of amniotic fluid infection in sections of placental disc, membranes, or cord. All cord sections with vitelline vessel remnants were immunostained for CD15 to document any infiltrates, to highlight patterns of infiltration, and to evaluate whether mild cases of umbilical phlebitis were associated with these lesions and were at risk of being missed. Vitelline vessel remnants were present in 4.2% of placentas examined. There were 5 vitelline vessel remnant histologic patterns identified providing insight into the vitelline vessel circulation. Funisitis, primarily neutrophilic, arising from vitelline vessel remnants was present in 70.3% of the 37 cords with vitelline vessel remnants. The presence of vitelline vessel remnant funisitis documents continued active circulation in these vascular structures, and vitelline vessel remnant funisitis was associated with the presence of other placental histological evidence of amniotic fluid infection in 53.8% of cases. The author also reviews normal embryology and the pathology of vitelline vessel remnants.

A quality improvement pilot assessment of the safety and associated outcomes of a viable cryopreserved umbilical tissue allograft as an adjunct surgical wrap in peroneus brevis tendon repair.

Surgical tendon repairs of the lower extremity are frequently associated with post-operative (post-op) risks that result in poor patient outcomes. Initially, increased pain levels may contribute to extended post-op rehabilitation while the development of tissue adhesions and fibrosis limit long-term functionality through reduced range of motion. Several surgical methods describing incorporation of various augmentative graft materials in tendon repair exist. However, reports demonstrating technique and both short- and long-term patient outcomes are lacking. Recently, advances in tissue preservation technology have led to the commercialization of human placental allografts. Of these available allogeneic biomaterials, the components found in human placental membranes may provide anti-inflammatory, antimicrobial, anti-adhesive, and antifibrotic properties to benefit surgical outcomes.Here, the authors introduce and technically describe the use of a viable cryopreserved umbilical tissue (vCUT) (Stravix, Osiris Therapeutics, Inc., Columbia, MD) as a complementary surgical wrap in primary tendon repair, with particular focus on the peroneus brevis. A pilot study was undertaken to assess the safety and potential for secondary rehabilitative outcomes associated with the use of vCUT in 5 tendon repair cases. The use of vCUT as a surgical tendon wrap was evaluated via the following primary endpoints at post-op day 7:1. presence of erythema, tenderness, heat and/or swelling;2. pain score;3. patient use of narcotic medication; and4. the development of adverse events during any point during the post-op course, defined in this study as dehiscence, confirmed infection, fluid collection or drainage.Secondary investigative endpoints included clinical and rehabilitative outcome measures for comparative pain reduction and transition times to both controlled ankle movement (CAM) boot and normal shoe ambulation.All patients were followed for an average of 24.15 months (range 16.75–26.5 months) after surgery. For primary safety measures, erythema, tenderness, drainage, heat, and swelling was absent in all 5 surgical sites. None of the patients required post-op use of narcotics past day 7. The potential for long-term rehabilitative improvement with adjunct use of vCUT was also demonstrated through reduced pain and reduced transition times to functional and non-assisted ambulation in normal shoewear as compared to historical controls managed without vCUT.This surgical technique is simple and safe for patients and preliminary findings have demonstrated favorable clinical and rehabilitative outcomes over historically observed controls.

Coxiella burnetii and Q fever in Brazil: a public health issue

Q fever is a zoonosis that is broadly distributed worldwide, despite few reports associated with its occurrence in Brazil. Coxiella burnetii, namely the causative agent of Q fever is a gram-negative coccobacillus and an obligate intracellular bacterial parasite of the order of Legionellales. The microorganism is generally present in the urine and feces of infected animals and can be found in large quantities in placental membranes of at-term or aborted animals. The inhalation of particles suspended in the air or contaminated aerosols is the most common form of bacterial contact. Q Fever is a self-limited disease, and often progresses in a benign way. In cases where the disease progresses to the chronic form, endocarditis is the most prevalent manifestation. Clinical diagnosis is difficult since the symptoms are similar to many other diseases. In confirmed cases, antibiotic therapy is the treatment indicated. Given the lack of knowledge about the disease and the difficulty of diagnosis, it is believed that Q fever is more common than generally believed in Brazil.

MiR-21 Contributes to Human Amniotic Membrane-Derived Mesenchymal Stem Cell Growth and Human Amniotic Membrane-Derived Mesenchymal Stem Cell-Induced Immunoregulation.

Human amniotic membrane-derived mesenchymal stem cells (hAM-MSCs) are considered a new and favorable source of stem cells for cell replacement-based therapy. Some microRNAs (miRNAs) have been reported to participate in the regulation of immune responses. Our aim was to investigate the effects of miR-21 on the biological characteristics, immunoregulatory properties, and potential mechanisms of hAM-MSCs. hAM-MSCs were isolated from the placental amnion membrane of a newborn. Cell proliferation, cell cycle, apoptosis, and expressions of cell surface markers were measured by CCK-8 and flow cytometric assays in hAM-MSCs. The expression of mesenchymal-specific antigens vimentin and stage-specific embryonic antigen-4 (SSEA-4) were identified by immunofluorescence staining. Tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein-1 (MCP-1), and interleukin-10 (IL-10) expressions in the cocultured supernatant of hAM-MSCs and peripheral blood mononuclear cells (PBMC) were detected via enzyme-linked immunosorbent assays (ELISA). Flow cytometric analyses revealed that the positive expression rates of the cell surface markers CD29, CD44, CD73, and CD90 in hAM-MSCs were 97.3%, 96.3%, 97.8%, and 98.2%, respectively, while the rates of CD34 and CD45 expression were only 0.6% and 0.84%, respectively. The immunofluorescent staining results showed that vimentin and SSEA-4 were positive in hAM-MSCs. CCK-8 assays revealed that miR-21 overexpression significantly promoted hAM-MSC proliferation. Cell cycle analyses revealed that the number of hAM-MSCs-miR-21+ cells during the synthesis phase (S phase) was significantly increased. miR-21 overexpression also significantly inhibited apoptosis in hAM-MSCs. The ELISA analyses revealed that miR-21 overexpression enhanced the inhibitory effect of hAM-MSCs on the secretion of TNF-α and MCP-1 as well as the promotive effect on the secretion of IL-10 in PBMC cocultured with miR-21-hAM-MSCs. In addition, miR-21 downregulation reduced the inhibitory effect of hAM-MSCs on the secretion of TNF-α, MCP-1, and the promotive effect on the secretion of IL-10 in PBMC cocultured with anti-miR-21-hAM-MSCs. Our data showed that miR-21 promoted hAM-MSCs proliferation, inhibited apoptosis, and was involved in controlling the immunoregulatory capacity of hAM-MSCs.

Comparative expression patterns of selected membrane transporters in differentiated BeWo and human primary trophoblast cells

Although placental membrane transporters have an important impact on materno-fetal nutrient transfer, placental cell models are poorly characterized regarding transporter expression. We assessed the mRNA expression of 26 physiologically important solute carriers and ABC transporters in BeWo (b30 clone) and primary human trophoblast cells (PHT) before and after syncytialization. 77% of the transporters showed similar mRNA expression changes between BeWo and PHT after syncytialization. Selected transporters, however, were either lacking in BeWo or showed different trends after syncytialization. In conclusion, BeWo cells generally represent an apt model for transporter studies, but their suitability should be confirmed for each transporter by comparison with PHT.

The Human Microbiome and Child Growth - First 1000 Days and Beyond.

The assembly of microbial communities within the gastrointestinal tract during early life plays a critical role in immune, endocrine, metabolic, and other host developmental pathways. Environmental insults during this period, such as food insecurity and infections, can disrupt this optimal microbial succession, which may contribute to lifelong and intergenerational deficits in growth and development. Here, we review the human microbiome in the first 1000 days - referring to the period from conception to 2 years of age - and using a developmental model, we examine the role of early microbial succession in growth and development. We propose that an 'undernourished' microbiome is intergenerational, thereby perpetuating growth impairments into successive generations. We also identify and discuss the intertwining host-microbe-environment interactions occurring prenatally and during early infancy, which may impair the trajectories of healthy growth and development, and explore their potential as novel microbial targets for intervention.

A short periconceptional exposure to maternal type-1 diabetes is sufficient to disrupt the feto-placental phenotype in a rabbit model

Tight metabolic control of type-1 diabetes is essential during gestation, but it could be crucial during the periconception period. Feto-placental consequences of maternal type-1 diabetes around the time of conception need to be explored.Using a rabbit model, type-1 diabetes was induced by alloxan 7 days before mating. Glycemia was maintained at 15–20 mmol/L with exogenous insulin injections to prevent ketoacidosis. At 4 days post-conception (dpc), embryos were collected from diabetic (D) or normoglycemic control (C) dams, respectively, and transferred into non-diabetic recipients. At 28dpc, D- and C-feto-placental units were collected for biometry, placental analyses and lipid profiles.D-fetuses were growth-retarded, hyperglycemic and dyslipidemic compared to C-fetuses. The efficiency of D-placentas was associated with an increased gene expression related to nutrient supply and lipid metabolism whereas volume density of fetal vessels decreased.Fetal plasma, placental and fetal liver membranes had specific fatty acid signatures depending on embryonic origin. Tissues from D-fetuses contained more omega-6 polyunsaturated fatty acids. The concentrations of docosahexaenoic acid decreased while linoleic acid increased in the heart of D-fetuses.This study demonstrates that a short exposure to maternal type-1 diabetes in the periconception window, until the blastocyst stage, is able to irreversibly malprogram the feto-placental phenotype, through precocious and persistent structural and molecular adaptations of placenta.

OP11.06: Do placental and amniotic membrane pathological changes occur following open and fetoscopic neural tube defect (NTD) repair?

OP11.06: Do placental and amniotic membrane pathological changes occur following open and fetoscopic neural tube defect (NTD) repair?

Immunohistochemical demonstration of RECK protein and interleukin-6 in fetal membranes from singleton pregnancies with late preterm delivery, intact membranes and histological chorioamnionitis

ABSTRACT We investigated whether chorioamnionitis affects immunohistochemical demonstration of RECK protein and interleukin-6 (IL-6) expression in fetal placental membranes following late preterm delivery with intact membranes. Fetal membranes of 28 women with single pregnancy, preterm delivery and histologically documented chorioamnionitis at gestational age 34−366/7 weeks constituted the chorioamnionitis study group. The control group consisted of 28 fetal membranes from women with preterm deliveries at the same gestational age without histological chorioamnionitis. Immunohistochemistry was performed using monoclonal antibodies against RECK protein and IL-6. We found a statistically significant difference in RECK expression between the chorioamnionitis and control groups; however, we found no difference in IL-6 expression between the groups. We demonstrated that RECK expression is down-regulated in fetal membranes from pregnancies with spontaneous late preterm birth and intact membranes, which suggests its role in preterm parturition. Equal expression of IL-6 in fetal membranes of pregnancies with and without histological chorioamnionitis is an intriguing and unexpected observation that requires further investigation.

Proliferative, Migratory, and Transition Properties Reveal Metastate of Human Amnion Cells

Amnion epithelial cell (AEC) shedding causes microfractures in human placental membranes during gestation. However, microfractures are healed to maintain membrane integrity. To better understand the cellular mechanisms of healing and tissue remodeling, scratch assays were performed using primary AECs derived from normal term not in labor membranes. AECs were grown under different conditions: i) normal cultures (control), ii) oxidative stress (OS) induction by cigarette smoke extract (CSE), iii) co-treatment of CSE and antioxidant N-acetyl-l-cysteine, and iv) treatment with amniotic fluid (AF). Cell migration time and distance, changes in intermediate filament (cytokeratin-18 and vimentin) expressions, and cellular senescence were determined. Control AECs in culture exhibited a metastate with the expression of both cytokeratin-18 and vimentin. During healing, AECs proliferated, migrated, and transitioned from epithelial to mesenchymal phenotype with increased vimentin. Wound healing was associated with mesenchymal to epithelial transition (MET). CSE-induced OS and senescence prevented wound healing in which cells sustained mesenchymal state. N-acetyl-l-cysteine reversed CSE's effect to aid wound closure through MET. AF accelerated cellular transitions and healing. Our data suggest that AECs undergo epithelial to mesenchymal transition during proliferation and migration and MET at the injury site to promote healing. AF accelerated whereas OS diminished cellular transitions and healing. OS-inducing pregnancy risk factors may diminish remodeling capacity contributing to membrane dysfunction, leading to preterm birth.

Transport mechanism of ursodeoxycholic acid in human placental BeWo cells.

Ursodeoxycholic acid (UDCA) is a first-line drug to treat intrahepatic cholestasis of pregnancy (ICP). However, its effects on the fetus are not clearly known. To better guide its clinical use, we aimed to study the mechanism underlying the placental transport of UDCA. The uptake and efflux of UDCA across placental apical membranes were studied using BeWo cells; effects of different exposure durations, UDCA concentrations, temperatures, and inhibitors of transporters were studied. A transwell assay was performed, and UDCA concentration in both fetal and maternal sides was measured using LC-MS/MS. Higher unidirectional transport of UDCA was observed in the basolateral-to-apical direction than in the apical-to-basolateral direction. Ko143 and verapamil, which are typical inhibitors of efflux transporters, significantly increased UDCA transport from different directions. UDCA uptake from the apical membrane of BeWo cells was time-dependent, but sodium-independent. It was inhibited by inhibitors of energy metabolism and of organic anion transporters, indicating an active transport mechanism. UDCA uptake from the apical membranes of BeWo cells could be mediated by organic anion-transporting polypeptides, whereas its efflux could be mediated by breast cancer resistance protein and multidrug resistant protein 3. The results of the present study may provide a basis for UDCA use in pregnancy.