Biology subjects

Forde, N.

Publications and source records attributed to Forde, N..

At least 19 recordsLinked to original sources

bioRxiv2025

The complete genome of a songbird

Bird genomes are the smallest among amniotes, but remain challenging to assemble due to their structural complexity. This study presents the first fully phased, diploid, telomere-to-telomere (T2T) reference genome for the zebra finch (Taeniopygia guttata), a model organism for neuroscience and evolutionary genomics. Combining multiple sequencing strategies resulted in closing nearly all gaps, adding [~]90 Mbp of previously missing sequence (7.8%). This includes T2T assemblies for all microchromosomes, including dot chromosomes, and the previously almost entirely missing chr16. The T2T genome is comprehensively annotated for genes, repeats, structural variants, and long-read methylation calls. Complete centromeric structures were assembled and annotated along with kinetochore binding sites. Relative to the previous high-quality reference of the Vertebrate Genomes Project, 2,778 (8.51%) previously unassembled or unannotated genes were identified, of which 9% overlap with segmental duplications. This first complete genome of a songbird, now the new public reference, illuminates avian genome architecture and function.

genomics↗

bioRxiv2025

Mammal placental phenotypes are predictable from microRNA repertoires.

Similar placental morphologies evolved multiple times independently in the history of mammal evolution1,2. Yet the genetic architecture that repeatedly guides distinct mammal lineages towards similar complex placental phenotypes has remained elusive. MicroRNAs (miRNAs), despite their diversity in mammals3-7 and known roles as developmental regulators8-10, remain under-examined as drivers of morphological innovation. We identified presence-absence patterns for 429 miRNA gene families across 398 mammalian genomes and discovered that placental phenotype is highly predictable from genomic miRNA repertoires (classification accuracy 74.5-95.8%). We identified 42 miRNA gene families significantly associated with placentation type, whose gene targets are enriched for developmental processes. Notably, convergent placental morphologies consistently involve identical miRNA families, revealing that evolution of this trait is constrained to predictable genetic pathways. We demonstrate that MIR-11986, uniquely associated with cotyledonary placentation, has tissue-specific expression in key reproductive tissues. MiRNA-mediated regulation therefore constrains placental morphological diversification into reproducible programs, offering insights into how genetic architecture shapes the predictability of convergent evolution. This striking pattern reveals a fundamental principle of evolution: that the miRNA regulatory networks available to control and guide complex placental morphological innovation are constrained and predictable.

genomics↗

bioRxiv2025

An evolutionarily conserved microRNA, miR-185, regulates key pathways that may contribute to implantation failure.

Recurrent implantation failure (RIF) is defined after three or more good quality embryo transfers following in vitro fertilisation without a successful pregnancy outcome. Many factors contribute to RIF however, the endometrial contribution remains unclear. Previous work by our group has identified a micro-RNA (miRNA) miR-185-5p as conserved across placental mammal irrespective of implantation strategies. We tested the hypothesis that miR-185-5p and the pathways it regulates, may be disrupted in the endometria of women with RIF. A human endometrial epithelial cells line (Ishikawa cells) was transfected with mimics or inhibitors for miR-185-5p for 24 (for implantation assay) or 48 hr (for proteomic analysis) along with non-targeting controls. There was a significant different in percentage attachment of BeWoW spheroids to cells transfected with miR-185-5p mimic compared to inhibitor (P<0.05). Transfection of epithelial cells with miR-185-5p altered expression of 1450 (mimic alone) and 509 (inhibitor alone) proteins respective of which, 146 were modified by both. Comparison of predicted targets of miR-185-5p, proteins modified by this study, and key endometrial genes from the literature determined genes and proteins associated with CNP family were further investigated in biopsies from individuals with (n=10) and without RIF (n=9) with CSNK1D expression significantly lower (p<0.05) in individuals with RIF. Collectively these data demonstrate that miR-185-5p modifies pathways that are important for successful implantation in humans.

cell biology↗

bioRxiv2024

Bovine endometrial organoids: A new tool to study conceptus-maternal interactions in mammals

In cattle, communication between endometrium and conceptus during the peri-implantation period is crucial for successful pregnancy. Understanding these interactions is vital as most early pregnancy loss occurs during this time. A major challenge in understanding uterine function and early pregnancy is lack of appropriate in-vitro models. Two-dimensional models are available, but do not recapitulate the endometriums complex multicellular structure. Here, we describe a hormonally responsive organoid model of the bovine endometrium, developed as a tool for studying endometrial function and early pregnancy. Bovine glandular epithelial cells were isolated from reproductive tracts and cultured in an extracellular matrix hydrogel (Cultrex 2) at 38.5{degrees}C, 5% CO (n=3). RNA was extracted and qPCR confirmed the presence of gland markers: leukemia inhibitory factor, mucin-1, insulin-like growth factor binding protein-1, kruppel-like factor-5 and forkhead box protein-A2. Organoids were imaged at specific time-points to monitor growth and passaged 3 times in 1:2 or 1:3 ratios after growing for a minimum of 10 days per passage. Morphologically, organoids were spherical and fast-growing at passages 0 and 1, but this declines following passage 2. Bovine endometrial organoids (n=3, passage 0) were treated with 1000 ng/ml recombinant ovine Interferon Tau (IFNT) or 10 g/ml progesterone (P4) for 24 hours and analysed by RNASeq to assess hormone responsiveness. Differential expression analysis by DeSeq2 negative binomial distribution model followed by Wald test and Benjamini-Hochberg correction identified 373 transcripts significantly upregulated (padj<0.05 or log2fold change >0.05) in response to P4 treatment, with downstream analysis showing significant overrepresentation (FDR<0.05) of genes associated with positive regulation of protein localisation to plasma membrane and cell periphery. Of the 240 genes significantly downregulated by P4 these were significantly overrepresented (FDR<0.05) in biological processes of cilium and cytoskeleton organisation. IFNT treatment resulted in significant upregulation of 414 genes and downregulation of 119 genes. The largest cluster associated with differentially expressed genes in response to IFNT is defence to virus and interferon signalling. There were 30 genes altered by both P4 treatment and IFNT treatment. Organoids were also shown to express conserved microRNAs, and it was possible to culture them in a microfluidics device - making them a useful model for a multitude of potential investigations. This model provides a tool to investigate bovine endometrial function and peri-implantation communication, subsequently allowing species comparison to understand diversity in reproductive strategies.

cell biology↗

bioRxiv2024

Lifespan Normative Models of White Matter Fractional Anisotropy: Applications to Early Psychosis

This study presents large-scale normative models of white matter (WM) organization across the lifespan, using diffusion MRI data from over 25,000 healthy individuals aged 0-100 years. These models capture lifespan trajectories and inter-individual variation in fractional anisotropy (FA), a marker of white matter integrity. By addressing non-Gaussian data distributions, race, and site effects, the models offer reference baselines across diverse ages, ethnicities, and scanning conditions. We applied these FA models to the HCP Early Psychosis cohort and performed a multivariate analysis to map symptoms onto deviations from multimodal normative models using multi-view sparse canonical correlation analysis (msCCA). Our results reveal extensive white matter heterogeneity in psychosis, which is not captured by group-level analyses, with key regions identified, including the right uncinate fasciculus and thalami. These normative models offer valuable tools for individualized WM deviation identification, improving precision in psychiatric assessments. All models are publicly available for community use. TeaserLifespan models of white matter offer insights into brain health, providing tools for tracking individual deviations across ages.

neuroscience↗

bioRxiv2024

A 3D endometrium-on-a-chip reveals the role of conceptus-derived factors CAPG and PDI in conceptus-endometrial communication

Early embryo loss affects all mammalian species, including humans and agriculturally important food-producing mammals such as cattle. The developing conceptus (embryo and extra-embryonic membranes) secretes factors which modify the endometrium and can be critical for early pregnancy processes such maternal recognition of pregnancy (MRP) and enhancing uterine receptivity to implantation. For example, a competent bovine conceptus secretes IFNT to initiate MRP. The bovine conceptus also secretes other proteins at the time of MRP, including CAPG and PDI, which are highly conserved among placental mammals. We have previously shown that these proteins act upon the endometrium to modulate receptivity, embryo development, and implantation in species with different implantation strategies (humans and cattle). We hypothesise that developing a novel 3D bovine endometrium on a chip system will enhance our understanding of the role of conceptus-derived factors in altering the endometrium and/or ULF secretion. Here we have developed a 3D bovine endometrium on a chip system, comprising both stromal and epithelial cell culture combined with culture medium flow better mimics the in vivo endometrium and exposure to conceptus-derived factors than conventional 2D endometrial cell culture. We have demonstrated that the conceptus-derived proteins CAPG and PDI modulate the endometrial transcriptome and secretory response to promote pathways associated with early pregnancy and alter ULF composition. This work highlights the critical need for more robust and in vivo -like culture systems to study endometrial-conceptus interactions in vitro to further investigate the role of conceptus derived factors for pregnancy success. SIGNIFICANCE STATEMENTWe have developed an in vitro 3D bovine endometrium-on-a-chip system comprising both primary stromal cells under static conditions and epithelial cells under flow conditions to mimic the in vivo endometrial environment from the conceptuses perspective. The secretome of the 3D endometrium-on-a-chip was characterised, was found to contain proteins associated with cell adhesion and tissue development, and contained proteins previously identified in in vivo uterine luminal fluid. PDI and CAPG (previously identified conceptus-derived factors) altered the transcriptome and secretome of cells within the system. Exposure to CAPG or PDI altered the secretome of proteins previously identified in pregnant uterine luminal fluid or associated with early pregnancy, and exposure to CAPG or PDI also altered the transcriptome to support processes such as immune response, secretion, proliferation, and adhesion related pathways. This data supports previously published works and highlights the need for the use of more in vivo-like in vitro models to study conceptus-endometrial interactions.

molecular biology↗

bioRxiv2024

RNAseq analysis reveals transcriptome changes associated with the progression of non-alcoholic liver disease in livers from Efcab4b knockout mice

EFCAB4B is an evolutionarily conserved protein that encodes for the Rab GTPase Rab46, and the CRAC channel modulator, CRACR2A. Previous genome wide association studies have demonstrated the association of EFCAB4B variants in the progression of non-alcoholic liver disease (NAFLD). In this study we show that mice with global depletion of Efcab4b-/- have significantly larger livers than their wild-type (WT) counterparts. We performed RNA-sequencing (RNA-seq) analysis of liver tissues to investigate differential global gene expression among Efcab4b-/-and WT mice. Of the 69 differentially expressed genes (DEGs), analyses of biological processes found significant enrichment in liver and bile development, with 6 genes (Pck1, Aacs, Onecut1, E2f8, Xbp1, and Hes1) involved in both processes. Specific consideration of possible roles of DEGs or their products in NAFLD progression to (NASH) and hepatocarcinoma (HCC), demonstrated DEGs in the livers of Efcab4b-/- mice had roles in molecular pathways including lipid metabolism, inflammation, ER stress and fibrosis. The results in this study provide additional insights into molecular mechanisms responsible for increasing susceptibility of liver injuries associated with EFCAB4B.

genetics↗

bioRxiv2024

The embryo-derived protein PDI is highly conserved among placental mammals and alters the function of the endometrium in species with different implantation strategies.

Pregnancy establishment in mammals requires a complex sequence of events, including bi-lateral embryo-maternal communication, leading up to implantation. This is the time when most pregnancy loss occurs in mammals (including humans and food production species) and dysregulation in embryo-maternal communication contributes to pregnancy loss. Embryo-derived factors modify the function of the endometrium for pregnancy success. We hypothesise that these previously unexplored conceptus-derived proteins may be involved in altering the function of the endometrium to facilitate early pregnancy events in mammals with different early pregnancy phenotypes. Here, we show that protein disulphide-isomerase (PDI) is a highly conserved protein among mammals, and provide evidence for a species-specific roles for PDI in endometrial function in mammals with different implantation strategies. We show how PDI alters the endometrial transcriptome in human and bovine in vitro in a species-specific manner, and using a microfluidic approach we demonstrate that it alters the secretome capability of the endometrium. We also provide evidence from in vitro assays using human-derived cells that MNS1, a transcript commonly downregulated in response to PDI in human and bovine endometrial epithelial cells, may be involved in the attachment (but not invasion) phase of implantation. We propose that the trophoblast-derived protein PDI, is involved in supporting the modulation of the uterine luminal fluid secreted by the endometrium to support conceptus nourishment, and also in the process of embryo attachment to the uterine lumen for pregnancy success in mammals. SIGNIFICANCE STATEMENTWe provide evidence that a highly conserved protein (PDI) alters the endometrial transcriptome in a species- and cell-specific manner. Exposure of endometrial epithelia to PDI altered genes belonging to immune modulatory, pro-inflammatory, and adhesion-pathways. One transcript, MNS1, was commonly downregulated in endometrial epithelia from species with superficial (bovine) and invasive (human) implantation morphologies. Knockdown of MNS1 expression in humans epithelia altered the ability of human trophoblast BeWo spheroids to attach suggesting a mechanism by which PDI affects implantation in human and bovine. In addition, using a microfluidics approach we have shown that PDI alters the secretome in a species-specific manner demonstrating PDI alters a key function of the endometrium in mammals.

developmental biology↗

bioRxiv2023

Biosensor capability of the endometrium is mediated in part, by altered miRNA cargo from conceptus-derived extracellular vesicles.

We tested the hypothesis that the biosensor capability of the endometrium is mediated in part, by the effect of different cargo contained in the extracellular vesicles secreted by the conceptus during the peri-implantation period of pregnancy. We transferred Bos taurus taurus embryos of different origin: In vivo (high developmental potential (IV)), in vitro (intermediate developmental potential (IVF)), or cloned (low developmental potential (NT)), into Bos taurus indicus recipients. Extracellular vesicles (EVs) recovered from Day 16 conceptus conditioned medium were characterized and their microRNA (miRNA) cargo sequenced alongside RNA sequencing of their respective endometria. There were substantial differences in the endometrial response to in vivo Vs in vitro and in vivo Vs cloned conceptuses (1153 and 334DEGs respectively) with limited differences between in vitro Vs cloned conceptuses (36 DEGs). miRNA cargo was similar between all three groups (426 common cargo) differences between in vivo and cloned (8 miRNAs), and in vivo and in vitro (6 miRNAs) observed. Treatment of endometrial epithelial cells with mimic or inhibitors for miR-128 and miR-1298 changes to the proteomic content of target cells (96, and 85 respectively) of which mRNAs are altered in the endometrium in vivo (PLXDC2, COPG1, HSPA12A, MCM5, TBL1XR1, and TTF). In conclusion, we have determined that the biosensor capability of the endometrium is mediated in part, by its response to different EVs miRNA cargo produced by the conceptus during the peri-implantation period of pregnancy. SIGNIFICANCE STATEMENTDuring the peri-implantation period of pregnancy in mammals, the endometrium acts as a biosensor for the developmental competency of the embryo. However, the mechanism by which biosensor capability of the endometrium is established, remains elusive. In this study, we show that embryos of different developmental competencies have distinct microRNA cargo contained in their extracellular vesicles (EVs). Exposure of the endometrium to these conceptuses alters the transcriptional response of the endometrium during the process of pregnancy recognition. This differential response is mediated in part, by the delivery and action of the these differentially abundant microRNAs into EVs. Here we propose differences in EV-mediated miRNA cargo are responsible in part for this biosensor capability of the endometrium.

developmental biology↗

bioRxiv2023

Differences in binding preferences for XIST partners are observed in mammals with different early pregnancy morphologies

In comparison to protein-coding genes, long non-coding RNAs (lncRNAs) are poorly conserved between species at the sequence level yet they often perform conserved and essential regulatory roles. The lncRNA XIST mediates X chromosome inactivation (XCI) through the interaction of numerous proteins, most of which bind to its repeat regions. Whilst XIST is present across placental mammals, its sequence is divergent. In addition, the timing and mechanistic details of the process of XCI also vary across placental mammals. Here, we sought to determine whether XIST interactor proteins previously identified in a model mammalian species (mouse) also bind in other mammals which exhibit divergent timing of XCI (human and bovine) and differing pre-implantation embryo morphology. We show that XIST and its putative protein partners are coordinately expressed in the endometria of mammals with different pre-implantation embryo morphologies. RNA immunoprecipitations revealed that human WTAP, SPEN, hnRNPK and CIZ1 proteins bind human XIST. CIZ1-XIST binding was also detected in bovine. In both human and cow, we found CIZ1 binds to the repeat E element of XIST. However, human CIZ1 protein is not able to bind bovine repeat E, indicating a species-specific binding interaction. Together these data indicate that several of the key RNA-protein interactions with XIST are common across placental mammals, even though we observe divergence in both the RNA sequence of XIST, and early embryo morphologies. This work sheds light upon the evolution of RNA-protein binding interactions, revealing that the binding events can be conserved even when the precise mechanism of binding has changed. Statements and DeclarationsAuthors have no financial or non-financial interests that are directly or indirectly related to the work submitted for publication.

developmental biology↗

bioRxiv2023

Objective QC for diffusion MRI data: artefact detection using normative modelling

Diffusion MRI is a neuroimaging modality used to evaluate brain structure at a microscopic level and can be exploited to map white matter fibre bundles and microstructure in the brain. One common issue is the presence of artefacts, such as acquisition artefacts, physiological artefacts, distortions or image processing-related artefacts. These may lead to problems with other downstream processes and can bias subsequent analyses. In this work we use normative modelling to create a semi-automated pipeline for detecting diffusion imaging artefacts and errors by modelling 24 white matter imaging derived phenotypes from the UK Biobank dataset. The considered features comprised 4 microstructural features (from models with different complexity such as fractional anisotropy and mean diffusivity from a diffusion tensor model and parameters from neurite orientation, dispersion and density models), each within six pre-selected white matter tracts of various sizes and geometrical complexity (corpus callosum, bilateral corticospinal tract and uncinate fasciculus and fornix). Our method was compared to two traditional quality control approaches: a visual quality control protocol performed on 500 subjects and quantitative quality control using metrics derived from image pre-processing. The normative modelling framework proves to be comprehensive and efficient in detecting diffusion imaging artefacts arising from various sources (such as susceptibility induced distortions or motion), as well as outliers resulting from inaccurate processing (such as erroneous spatial registrations). This is an important contribution by virtue of this methods ability to identify the two problem sources (i) image artefacts and (ii) processing errors, which subsequently allows for a better understanding of our data and informs on inclusion/exclusion criteria of participants.

neuroscience↗

bioRxiv2022

Successful reproduction of a large EEG study across software packages

As an active field of research and with the development of state-of-the-art algorithms to analyze EEG datasets, the parametrization of Electroencephalography (EEG) analysis workflows has become increasingly flexible and complex, with a great variety of methodological options and tools to be selected at each step. This high analytical flexibility can be problematic as it can yield to variability in research outcomes. Therefore, growing attention has been recently paid to understand the potential impact of different methodological decisions on the reproducibility of results. In this paper, we aim to examine how sensitive the results of EEG analyses are to variations in preprocessing with different software tools. We reanalyzed the shared EEG data (N=500) from (Williams et al. 2021) using three of the most commonly used EEG software tools: EEGLAB, Brainstorm and FieldTrip. After reproducing the same original preprocessing workflow in each software, the resulting evoked-related potentials (ERPs) were qualitatively and quantitatively compared in order to examine the degree of consistency/discrepancy between softwares. Our findings show a good degree of convergence in terms of the general profile of ERP waveforms, peak latencies and effect size estimates related to specific signal features. However, considerable variability was also observed in the magnitude of the absolute voltage observed with each software package as reflected by the similarity values and observed statistical differences at particular channels and time instants. In conclusion, we believe that this study provides valuable clues to better understand the impact of the software tool on the analysis of EEG results.

neuroscience↗

bioRxiv2022

Sex-bias in utero alters ovarian reserve but not uterine capacity in female offspring

Environmental stressors to which a foetus is exposed, affect a range of physiological functions in post-natal offspring. Such stressors include disproportionate steroid hormone concentrations in the uterine environment. We aimed to determine the in-utero effect of steroid hormones on reproductive potential of female offspring using a porcine model. Hypothesising that an in-utero sex bias will influence ovarian reserve and endometrial morphology in the breeding gilt. Reproductive tracts of pigs from female-biased litters (>65% female, n=15), non-biased litters (45-54.9% female, n=15), and male-biased litters (<35% females, n=9) were collected at slaughter (95-115 kg). Ovaries and uterine horns were processed for histological approaches and stained using H&E or IHC techniques. All measurements were conducted in QuPath (Bankhead et al, 2017). Variability of data within groups was analysed with a Levenes test, whilst data was analysed using linear models in R. In the ovarian reserve, there was a significant interaction between the birth weight and the sex ratio of a litter from which a pig originated (p=.015), with low-birth-weight pigs from male-biased litters having a higher number of primordial follicles and the opposite trend seen in pigs from female-biased litters. This was not reflected in recruited, nor atretic follicles. In the uterine horn sex bias held no effect on development as seen in this study. Birth weight held more effects on the gilts. A lower BW decreased the proportion of glands found in the endometrium (p=.045). BW was found to be far more variable in both male-biased and female-biased litters (p=.026). The variability of primordial follicles from male-biased litters was greater than non-and female-biased litters (p=.014). Similarly, endometrial stromal nuclei had a greater range in male- and female-biased litters than non-biased litters (p=.028). There was a greater effect on both ovarian reserve and uterine development of piglet BW than the litter bias. There seems a benefit of being androgenised on ovarian reserve whilst no effects were found for the morphology or endometrial gland proliferation of the uterine horns. However, a crucial finding was in the variability of the data. Both primordial follicles in the male-biased ovary, and stromal nuclei in the male- and female-biased uterine horns had a wider spread in numbers than non-biased litters. This could be inflating the variability of reproductive success seen in animals form male-biased litters by two means. Firstly, by a higher likelihood of insufficient primordial pools. Secondly, through a potential impact on stromal-derived growth factors or insufficient support of the underlying implantation structures, leading to an increased variability in uterine implantation capabilities, and thus survival of the embryo.

developmental biology↗

bioRxiv2022

Placental mammal derived microRNAs alter pathways in the endometrial epithelia important for endometrial function.

We tested the hypothesis that a panel of placental mammal-specific miRNAs and their targets play important to establish receptivity to implantation and their dysregulated expression may be a feature in women with early pregnancy loss. Relative expression levels of miR-340-5p, -542-3p, and -671-5p all increased following treatment of Ishikawa cells with progesterone (10 g/ml) for 24 hrs (p < 0.05). RNA sequencing of these P4-treated cells identified co-ordinate changes to 6,367 transcripts of which 1713 were predicted targets of miR-340-5p, 670 of miR-542-3p, and 618 of miR-671-5p. Quantitative proteomic analysis of Ishikawa cells transfected with mimic or inhibitor (48 hrs: n=3 biological replicates) for each of the P4-regulated miRNAs was carried out to identify targets of these miRNAs. Excluding off target effects, mir-340-5p mimic altered 1,369 proteins while inhibition changed expression of 376 proteins (p < 0.05) of which, 72 were common to both treatments. A total of 280 proteins were identified between predicted (mirDB) and confirmed (in vitro) targets. In total, 171 proteins predicted to be targets by mirDB were altered in vitro by treatment with miR-340-5p mimic or inhibitor and were also altered by treatment of endometrial epithelial cells with P4. In vitro targets of miR-542-3p identified 1,378 proteins altered by mimic while inhibition altered 975 a core of 200 proteins were changed by both. 100 protein targets were predicted and only 46 proteins were P4 regulated. miR-671-mimic altered 1,252 proteins with inhibition changing 492 proteins of which 97 were common to both, 95 were miDB predicted targets and 46 were also P4-regulated. All miRNAs were detected in endometrial biopsies taken from patients during the luteal phase of their cycle, irrespective of prior or future pregnancy outcomes Expression of mir-340-5p showed an overall increase in patients who had previously suffered a miscarriage and had a subsequent miscarriage, as compared to those who had infertility or previous miscarriage and subsequently went on to have a life birth outcome. The regulation of these miRNAs and their protein targets regulate the function of transport and secretion, and adhesion of the endometrial epithelia required for successful implantation in humans. Dysfunction of these miRNAs (and therefore the targets they regulate) may contribute to endometrial-derived recurrent pregnancy loss in women.

cell biology↗

bioRxiv2021

Excitatory/inhibitory imbalance in autism: the role of glutamate and GABA gene-sets in symptoms and cortical brain structure

BackgroundThe excitatory/inhibitory (E/I) imbalance hypothesis posits that an imbalance between excitatory (glutamatergic) and inhibitory (GABAergic) mechanisms underlies the behavioral characteristics of autism spectrum disorder (autism). However, how E/I imbalance arises and how it may differ across autism symptomatology and brain regions is not well understood. MethodsWe used innovative analysis methods - combining competitive gene-set analysis and gene-expression profiles in relation to cortical thickness (CT)- to investigate the relationship between genetic variance, brain structure and autism symptomatology of participants from the EU-AIMS LEAP cohort (autism=360, male/female=259/101; neurotypical control participants=279, male/female=178/101) aged 6 to 30 years. Competitive gene-set analysis investigated associations between glutamatergic and GABAergic signaling pathway gene-sets and clinical measures, and CT. Additionally, we investigated expression profiles of the genes within those sets throughout the brain and how those profiles relate to differences in CT between autistic and neurotypical control participants in the same regions. ResultsThe glutamate gene-set was associated with all autism symptom severity scores on the Autism Diagnostic Observation Schedule-2 (ADOS-2) and the Autism Diagnostic Interview-Revised (ADI-R) within the autistic group, while the GABA set was associated with sensory processing measures (using the SSP subscales) across all participants. Brain regions with greater gene expression of both glutamate and GABA genes showed greater differences in CT between autistic and neurotypical control participants. ConclusionsOur results suggest crucial roles for glutamate and GABA genes in autism symptomatology as well as CT, where GABA is more strongly associated with sensory processing and glutamate more with autism symptom severity.

neuroscience↗

bioRxiv2021

Insulin exposed endometrial epithelial cells cultured in a microfluidic device alters transcripts involved in translation that may contribute to reduced implantation capacity of the endometrium.

Obesity is a rapidly growing public health issue among women of reproductive age. It is also associated with decreased reproductive function including implantation failure. Implantation failure can result from a myriad of factors including impaired gametes and endometrial dysfunction. The mechanisms of how obesity-related hyperinsulinaemia disrupts endometrial function and implantation are poorly understood. Our study aims to investigate potential mechanisms by which insulin alters endometrial transcript expression, which may affect endometrial receptivity. Ishikawa cells mimicking human endometrial epithelium were seeded into a microfluidics organ-on-chip device to produce an in vitro endometrium. Syringe pump was attached to the microfluidics device to deliver three varying treatments into Ishikawa cells: 1) media control 2) vehicle control (PBS acidified to pH3 with acetic acid) 3) Insulin (2mg/mL) at a constant flow rate of 1uL/min for 24 hours to mimic secretion in vivo. Three biological replicates were obtained. Insulin-induced transcriptomic response of the in vitro endometrium was quantified via RNA sequencing, and subsequently analysed using DAVID and Webgestalt to identify Gene Ontology (GO) terms and signalling pathways. A Total of 29 transcripts showed differential expression levels across two comparison groups (control v vehicle control; vehicle control v insulin). There were nine transcripts significantly differentially expressed in vehicle control v insulin group (p<0.05). Functional annotation analysis of transcripts altered by insulin (n=9) identified three significantly enriched GO terms: SRP-dependent cotranslational protein targeting to membrane, poly(A) binding, and RNA binding (p<0.05). Over-representation analysis found three significantly enriched signalling pathways relating to insulin-induced transcriptomic response: protein export, glutathione metabolism, and ribosome pathways (p<0.05). Insulin-induced dysregulation of biological functions and pathways highlight potential mechanisms by which high insulin concentrations within maternal circulation may perturb endometrial receptivity.

physiology↗

bioRxiv2021

An ancestral burst of regulatory and protein innovation drives divergent implantation in eutherian mammals.

Summary paragraphThe origin of live birth in mammals [~]148 million years ago was a dramatic shift in reproductive strategy, yet the molecular changes that established mammal viviparity are largely unknown. Although progesterone receptor signalling predates the origin of mammals andis highly conserved in, and critical for, successful mammal pregnancy, it alone cannot explain the origin and subsequent diversity of implantation strategies throughout the placental mammal radiation. MiRNAs are known to be flexible and dynamic regulators with a well established role in the pathophysiology of mammal placenta. We propose that a dynamic core microRNA (miRNA) network originated early in placental mammal evolution, responds to conserved mammal pregnancy cues (e.g. progesterone), and facilitates species-specific responses. Here we identify 13 miRNAs that arose at the origin of placental mammals and were subsequently retained in all descendent lineages. The expression of these 13 miRNAs in response to early pregnancy molecules is regulated in a species-specific manner in endometrial epithelia of species with extremes of implantation strategies. Furthermore, these 13 miRNAs preferentially target 84 proteins under positive selective pressure on ancestral eutherian. Discovery of this core "live-birth" toolkit and specifically adapted proteins helps explain the origin and evolution of the placenta in mammals.

evolutionary biology↗

bioRxiv2020

Endometrium-on-a-chip reveals the endometrial transcriptome, and protein content of secretome are altered by changes in circulating concentrations of insulin and glucose in vitro.

The molecular interactions between the maternal environment and developing embryo that are key for early pregnancy success are known to be influenced by factors such as the metabolic status. We are, however, limited in our understanding of the mechanism by which these individual nutritional stressors alter endometrial function and the in utero environment for early pregnancy success. Here we report for the first time the use of endometrium-on-a-chip microfluidics approach to produce a multi-cellular endometrium in vitro, that is exposed to glucose and insulin concentrations associated with maternal metabolic stressors. Following isolation of endometrial cells (epithelial and stromal) from the uteri of non-pregnant cows in early-luteal phase (Day 4-7 approximately) epithelial cells were seeded into the upper chamber (4-6 104 cells/mL) and stromal cells seeded in the lower chamber (1.5-2 104 cells/mL). Three different concentration of glucose 1) 0.5 mM 2) 5.0 mM or 3) 50 mM or insulin 1) Vehicle, 2) 1 ng/mL or 3) 10 ng/mL were performed in the endometrial cells at a flow rate of 1{micro}L/min for 72 hr to mimic the rate of secretion in vivo. Quantitative differences in the transcriptomic response of the cells and the secreted proteome of in vitro-derived uterine luminal fluid (ULF) were determined by RNA-sequencing and TMT respectively. Changes in maternal glucose altered 21 and 191 protein coding genes in epithelial and stromal cells respectively (p<0.05). While there was a dose-dependent quantitative change in protein secretome (1 and 23 proteins). Insulin resulted in limited transcriptional changes including insulin-like binding proteins that were cell specific (5, 12, and 20) but altered the quantitative secretion of 196 proteins including those involved in extracellular matrix-receptor interaction and proteoglycan signaling in cancer. Collectively, these highlight the potential mechanism by which changes to maternal glucose and insulin alter uterine function.

molecular biology↗