Placentae of small appropriately-grown-for-gestational-age neonates exhibit sexually dimorphic transcriptomic changes representative of placental insufficiency.

INTRODUCTION: Previous studies have reported that neonates less than the 25th BWC especially if they were male, were more likely to be associated with birth complications suggesting small neonates often identified as appropriately grown are at risk of adverse outcomes. We have questioned whether smaller neonates not typically categorized as "small for gestational age" may not reach their genetically determined growth due to placental insufficiency. METHODS: RNA-Seq was performed on the Illumina NovaSeq 600 using term placentae from neonates that were less than the 10th birthweight centile (BWC) (n = 39), between the 10th and the 30th BWC (n = 15) or greater than the 30th BWC (n = 23). Bioinformatic analyses were conducted and statistical significance was assessed at a level of P < 0.05 for single comparisons or FDR <0.05 unless otherwise noted. RESULTS: Gene set enrichment analysis revealed differences between BWC groups and in relation to the sex of the placenta. Genes associated with hypoxia, inflammatory responses, estrogen responsive genes, and androgen responsive genes were enriched (FDR <0.1) for in placentae of neonates <10th BWC regardless of sex and also in male placentae of neonates between the 10th-30th BWC. Female placenta of neonates between the 10th-30th BWC were comparable to placentae of neonates >30th BWC. DISCUSSION: These findings provide evidence that small male neonates may be at a greater risk of an adverse outcome than females due to changes in gene expression that are associated with placental dysfunction. The current data raises questions of whether placental pathology for smaller appropriately grown neonates should be scientifically and clinically examined in more depth.

Asymmetry-Enhanced Motion of Urease-Powered Micromotors from Double Emulsion-Templated Microcapsules.

Autonomous motion of enzyme-powered motors has important implications for drug delivery, cell-cell communication, and protocell engineering. Although many of these systems are inspired by the motion of biological cells, most of them lack key structural features, like micrometer-sized boundaries and aqueous compartments, and rely on bubble propulsion to generation motion. In this study, we use droplet microfluidics to generate large populations of cell-sized microcapsules with poly(lactic-co-glycolic acid) shells and functionalize their surfaces with the enzyme urease to drive their motion. We adjust the number of surface functional groups for urease conjugation by preparing microcapsules with two different surfactants, poly(vinyl alcohol) (PVA) and poly(ethylene-alt-maleic anhydride) (PEMA). We also tune the surface roughness of the microcapsules by varying the concentration of silica nanoparticles in the droplet middle phase. We find that PEMA plays a crucial role in increasing the grafting density of urease on the surface of smooth microcapsules, leading to active motion in the presence of urea. In addition, rough microcapsules prepared with PEMA and loaded with comparable amounts of urease move up to three times faster than their smooth counterparts, which we believe is due to an asymmetric distribution of urease on the surface, giving rise to a preferred direction of motion. Taken together, these results provide new insights into the role that various stabilizing agents play in the induction of motion by enzymatic motors prepared from microfluidics, which is a potentially powerful tool for future preparation of motile protocells in biomedicine.

Sex-dependent differential transcript expression in the placenta of growth restricted infants.

INTRODUCTION: The pathological decrease of fetal growth during gestation can lead to subsequent poor health outcomes for the fetus. This process is commonly controlled by the placenta, the interface between mother and baby during gestation. Sex-specific gene expression has been implicated in placental function, therefore, there is a need to determine if it is important during reduced fetal growth. We therefore aimed to characterise placental gene expression at term to evaluate sex-specific genetic changes that occur in small for gestational age (SGA) infants. METHODS: RNA-sequencing of twelve human placental tissue samples collected from pregnancies yielding either term appropriate for gestational age (AGA) or SGA infants identified at delivery. Candidate genes associated with fetal size and fetal sex were identified using differential gene expression and weighted gene co-expression network analyses. Single-cell sequencing data was used for candidate validation and to estimate candidate transcript expression in specific placental cell populations. RESULTS: Differential gene expression and weighted gene co-expression network analyses identified 403 candidate transcripts associated with SGA infants. One hundred and three of these transcripts showed sex-specific expression. . Published placental sequencing datasets were used to validate the key expression results from the twelve placental samples initially studied; the sex-independent transcript expression for genes involved in cell cycle processes in males (7 transcripts) and endoplasmic reticulum stress in females (17 transcripts). DISCUSSION: This study identified the activation of multiple molecular mechanisms involved in the placental response to an adverse environmental stressor. Mechanisms such as disrupted protein synthesis were shared between infant biological sex when comparing AGA to SGA, whilst other pathways such as cell cycle and endoplasmic reticulum stress appear as independent/specific to either males or females when investigating reduced fetal growth. This data suggests that sexual dimorphism is an important consideration when examining placental dysfunction and poor fetal growth.

Low pKa of Lys promotes glycation at one complementarity-determining region of a bispecific antibody.

Protein glycation is a common, normally innocuous, post-translational modification in therapeutic monoclonal antibodies. However, when glycation occurs on complementarity-determining regions (CDRs) of a therapeutic monoclonal antibody, its biological activities (e.g., potency) may be impacted. Here, we present a comprehensive approach to understanding the mechanism of protein glycation using a bispecific antibody. Cation exchange chromatography and liquid chromatography-mass spectrometry were used to characterize glycation at a lysine residue within a heavy chain (HC) CDR (HC-CDR3-Lys98) of a bispecific antibody. Thermodynamic analysis revealed that this reaction is reversible and can occur under physiological conditions with an apparent affinity of 8-10 mM for a glucose binding to HC-CDR3-Lys98. Results from kinetic analysis demonstrated that this reaction follows Arrhenius behavior in the temperature range of 5°C-45°C and can be well predicted in vitro and in a non-human primate. In addition, this glycation reaction was found to be driven by an unusually low pKa on the ε-amino group of HC-CDR3-Lys98. Van't Hoff analysis and homology modeling suggested that this reaction is enthalpically driven, with this lysine residue surrounded by a microenvironment with low polarity. This study provides, to our knowledge, new insights toward a mechanistic understanding of protein glycation and strategies to mitigate the impact of protein glycation during pharmaceutical development.

Implementation of a structured practical activity to analyse student healthcare worker perceptions and compliance with prescribed infection control procedures.

BACKGROUND: Non-compliance with infection control guidelines has been reported within healthcare settings. Infection control education in undergraduate healthcare education programs forms a critical component in preparing student healthcare workers for vocational roles. METHODS: Clinical sciences students (nutrition science, paramedicine, pharmacy, podiatry, optometry studying for qualifications recognised by the Australian Health Practitioner Regulation Agency) self-reported hygiene perceptions and practices and collected microbiological swabs from personal or medical equipment items before and after recommended disinfection procedures. RESULTS: Cultivable microorganisms were isolated from 95% of student medical equipment items. Disinfection significantly reduced microbial growth on student medical equipment items (P < 0.05). CONCLUSIONS: Student perceptions of infection control procedures do not always correlate with infection control practice. Infection control education of undergraduate healthcare students requires ongoing assessment to ensure successful translation into clinical practice.

Limitations of 16S rRNA Gene Sequencing to Characterize Lactobacillus Species in the Upper Genital Tract.

The endometrial cavity is an upper genital tract site previously thought as sterile, however, advances in culture-independent, next-generation sequencing technology have revealed that this low-biomass site harbors a rich microbial community which includes multiple Lactobacillus species. These bacteria are considered to be the most abundant non-pathogenic genital tract commensals. Next-generation sequencing of the female lower genital tract has revealed significant variation amongst microbial community composition with respect to Lactobacillus sp. in samples collected from healthy women and women with urogenital conditions. The aim of this study was to evaluate our ability to characterize members of the genital tract microbial community to species-level taxonomy using variable regions of the 16S rRNA gene. Samples were interrogated for the presence of microbial DNA using next-generation sequencing technology that targets the V5-V8 regions of the 16S rRNA gene and compared to speciation using qPCR. We also performed re-analysis of published data using alternate variable regions of the 16S rRNA gene. In this analysis, we explore next-generation sequencing of clinical genital tract isolates as a method for high throughput identification to species-level of key Lactobacillus sp. Data revealed that characterization of genital tract taxa is hindered by a lack of a consensus protocol and 16S rRNA gene region target allowing comparison between studies.

Analysis of mitochondrial regulatory transcripts in publicly available datasets with validation in placentae from pre-term, post-term and fetal growth restriction pregnancies.

INTRODUCTION: The human placenta has a defined lifespan and placental aging is a key feature as pregnancy progresses. Placental aging and mitochondrial dysfunction are known to play a key role in pregnancy pathophysiology. Premature aging of the placenta has also been linked with placental dysfunction resulting in poor fetal development and premature birth. METHODS: The expression of key mitochondrial-related genes were analysed in a series of publicly available databases then expression changes were validated in placental samples collected from term, pre-term, post-term pregnancies and pregnancies complicated by fetal growth restriction (FGR). Gene and protein expression levels of MFN1, MFN2, TFAM, TOMM20, OPA3 and SIRT4 were measured in placental tissues via qPCR and western blotting. RESULTS: Initial analysis found that key mitochondrial transcripts related to biogenesis, bioenergetics and mitophagy clustered by pregnancy trimester. A refined list of 13 mitochondrial-related genes were investigated in additional external datasets of pregnancy complications. In the new cohort, protein expression of MFN1 was decreased in FGR and MFN2 is decreased in post-term placenta. Analysis of placental tissues revealed that TOMM20 gene and protein expression was altered in FGR and post-term placenta. DISCUSSION: MFN1 and MFN2 play a major role in mitochondrial dynamics, and alterations in these markers have been highlighted in early unexplained miscarriage. TOMM20 is an importer protein that plays a major role in mitophagy and changes have also been identified in age-related diseases. Significant changes in MFN1, MFN2 and TOMM20 indicate that mitochondrial regulators play a critical role in placental aging and placental pathophysiology.

The Placental Ferroxidase Zyklopen Is Not Essential for Iron Transport to the Fetus in Mice.

BACKGROUND: The ferroxidase zyklopen (Zp) has been implicated in the placental transfer of iron to the fetus. However, the evidence for this is largely circumstantial. OBJECTIVES: This study aimed to determine whether Zp is essential for placental iron transfer. METHODS: A model was established using 8- to 12-wk-old pregnant C57BL/6 mice on standard rodent chow in which Zp was knocked out in the fetus and fetal components of the placenta. Zp was also disrupted in the entire placenta using global Zp knockout mice. Inductively coupled plasma MS was used to measure total fetal iron, an indicator of the amount of iron transferred by the placenta to the fetus, at embryonic day 18.5 of gestation. Iron transporter expression in the placenta was measured by Western blotting, and the expression of Hamp1, the gene encoding the iron regulatory hormone hepcidin, was determined in fetal liver by real-time PCR. RESULTS: There was no change in the amount of iron transferred to the fetus when Zp was disrupted in either the fetal component of the placenta or the entire placenta. No compensatory changes in the expression of the iron transport proteins transferrin receptor 1 or ferroportin were observed, nor was there any change in fetal liver Hamp1 mRNA. Hephl1, the gene encoding Zp, was expressed mainly in the maternal decidua of the placenta and not in the nutrient-transporting syncytiotrophoblast. Disruption of Zp in the whole placenta resulted in a 26% increase in placental size (P < 0.01). CONCLUSIONS: Our data indicate that Zp is not essential for the efficient transfer of iron to the fetus in mice and is localized predominantly in the maternal decidua. The increase in placental size observed when Zp is knocked out in the entire placenta suggests that this protein may play a role in placental development.

Modulation of Placental Gene Expression in Small-for-Gestational-Age Infants.

Small-for-gestational-age (SGA) infants are fetuses that have not reached their genetically programmed growth potential. Low birth weight predisposes these infants to an increased risk of developing cardiovascular, metabolic and neurodevelopmental conditions in later life. However, our understanding of how this pathology occurs is currently incomplete. Previous research has focused on understanding the transcriptome, epigenome and bacterial signatures separately. However, we hypothesise that interactions between moderators of gene expression are critical to understanding fetal growth restriction. Through a review of the current literature, we identify that there is evidence of modulated expression/methylation of the placental genome and the presence of bacterial DNA in the placental tissue of SGA infants. We also identify that despite limited evidence of the interactions between the above results, there are promising suggestions of a relationship between bacterial signatures and placental function. This review aims to summarise the current literature concerning fetal growth from multiple avenues and propose a novel relationship between the placental transcriptome, methylome and bacterial signature that, if characterised, may be able to improve our current understanding of the placental response to stress and the aetiology of growth restriction.