Embryology of the fat-tailed dunnart (Sminthopsis crassicaudata): A marsupial model for comparative mammalian developmental and evolutionary biology.

BACKGROUND: Marsupials are a diverse and unique group of mammals, but remain underutilized in developmental biology studies, hindering our understanding of mammalian diversity. This study focuses on establishing the fat-tailed dunnart (Sminthopsis crassicaudata) as an emerging laboratory model, providing reproductive monitoring methods and a detailed atlas of its embryonic development. RESULTS: We monitored the reproductive cycles of female dunnarts and established methods to confirm pregnancy and generate timed embryos. With this, we characterized dunnart embryo development from cleavage to birth, and provided detailed descriptions of its organogenesis and heterochronic growth patterns. Drawing stage-matched comparisons with other species, we highlight the dunnarts accelerated craniofacial and limb development, characteristic of marsupials. CONCLUSIONS: The fat-tailed dunnart is an exceptional marsupial model for developmental studies, where our detailed practices for reproductive monitoring and embryo collection enhance its accessibility in other laboratories. The accelerated developmental patterns observed in the Dunnart provide a valuable system for investigating molecular mechanisms underlying heterochrony. This study not only contributes to our understanding of marsupial development but also equips the scientific community with new resources for addressing biodiversity challenges and developing effective conservation strategies in marsupials.

Detrimental actions of obesity-associated advanced glycation end-products on endometrial epithelial cell proliferation are alleviated by antioxidants.

RESEARCH QUESTION: Advanced glycation end-products (AGE) are elevated in the uterine environment of obese infertile women. Can the detrimental effects of AGE on endometrial epithelial cells be mitigated with therapeutics, and recapitulated in a more physiologically relevant primary model (organoids)? DESIGN: Human endometrial epithelial cells (ECC-1) were exposed to AGE at concentrations physiologically representative of uterine fluid in lean or obese individuals, and three potential therapeutics: 25 nmol/l receptor for AGE (RAGE) antagonist FPS-ZM1, 100 µmol/l metformin, or a combination of antioxidants (10 µmol/l N-acetyl-l-cysteine, 10 µmol/l N-acetyl-l-carnitine and 5 µmol/l α-lipoic acid). Real-time cell analysis (xCELLigence, ACEA Biosciences) determined the rate of adhesion and proliferation. The proliferation of organoid-derived cells and secretion of cytokines from organoids was characterized in the presence of AGE (n = 5). The uterine fluid of women undergoing assisted reproduction was profiled for AGE-associated inflammatory markers (n = 77). RESULTS: ECC-1 proliferation was reduced by AGE from obese versus lean conditions and vehicle control (P = 0.04 and P < 0.001, respectively), and restored to a proliferation corresponding to lean conditions by antioxidants. AGE influenced organoid derived primary endometrial epithelial cell proliferation in a donor-dependent manner. AGE increased the organoid secretion of the proinflammatory cytokine CXCL16 (P = 0.006). Clinically, CXCL16 correlated positively to maternal body mass index (R = 0.264, P = 0.021) and intrauterine glucose concentration (R = 0.736, P < 0.0001). CONCLUSIONS: Physiologically relevant concentrations of AGE alter endometrial epithelial cell function. Antioxidants restore the rate of proliferation of AGE-treated endometrial epithelial (ECC-1) cells. Primary endometrial epithelial cells, cultured as organoids, demonstrate altered proliferation and CXCL16 secretion in the presence of AGE equimolar with the uterine fluid from obese individuals.

A microenvironment of high lactate and low pH created by the blastocyst promotes endometrial receptivity and implantation.

RESEARCH QUESTION: Is the blastocyst's idiosyncratic metabolic production of lactate, and creation of a specialized microenvironment at the implatation site, an important mediator of maternal-fetal signalling to promote endometrial receptivity and implantation? DESIGN: Hormonally primed ECC-1 and Ishikawa cells were used to assess functional changes to the endometrial epithelium after exposure to lactic acid (LA), LA with neutralized pH (nLA) or acidic pH (pHL). Tight junction integrity (transepithelial resistance [TER]), cellular proliferation or changes to gene expression by RT-PCR were analysed. The effect of LA on Endometrial stromal cells decidualization and migratory capacity, and HUVEC endothelial tube formation and angiogenesis, were also assessed. RESULTS: Treatment of ECC-1 cells with 2.5 mM (P = 0.0037), 5 mM (P = 0.0044), 7.5 mM and 10 mM (P = 0.003) (P = 0.0021) LA significantly decreased the rate of cellular proliferation while TER was decreased with exposure to 2.5 mM LA (P = 0.024), 5 mM LA (P = 0.021) and 7.5 mM LA (P = 0.033). Exposure to nLA or pHL had no effect on proliferation or TER. Upregulation of GLUT4 (P = 0.002), GPR81 (P = 0.048), VEGF, SNAI1 (both P < 0.001) and RELA (P = 0.023) mRNA expression was observed after exposure of Ishikawa cells to combined LA plus pHL. Lactic acid increased the migratory capacity of decidualized stromal cells (P = 0.047) without changing the extent of decidualization. HUVEC tube formation was significantly increased by 5 mM LA exposure (P = 0.009). CONCLUSIONS: The identification of LA as an important mediator in the maternal-fetal dialogue underpinning implantation is supported. Further examination of the role of LA within the infertile or compromised endometrium could improve natural and assisted pregnancy success and needs further investigation.

Cyclical endometrial repair and regeneration.

Uniquely among adult tissues, the human endometrium undergoes cyclical shedding, scar-free repair and regeneration during a woman's reproductive life. Therefore, it presents an outstanding model for study of such processes. This Review examines what is known of endometrial repair and regeneration following menstruation and parturition, including comparisons with wound repair and the influence of menstrual fluid components. We also discuss the contribution of endometrial stem/progenitor cells to endometrial regeneration, including the importance of the stem cell niche and stem cell-derived extracellular vesicles. Finally, we comment on the value of endometrial epithelial organoids to extend our understanding of endometrial development and regeneration, as well as therapeutic applications.

Advanced glycation end products present in the obese uterine environment compromise preimplantation embryo development.

RESEARCH QUESTION: Proinflammatory advanced glycation end products (AGE), highly elevated within the uterine cavity of obese women, compromise endometrial function. Do AGE also impact preimplantation embryo development and function? DESIGN: Mouse embryos were cultured in AGE equimolar to uterine fluid concentrations in lean (1-2 µmol/l) or obese (4-8 µmol/l) women. Differential nuclear staining identified cell allocation to inner cell mass (ICM) and trophectoderm (TE) (day 4 and 5 of culture). Cell apoptosis was examined by terminal deoxynucleotidyl transferase-mediated dUDP nick-end labelling assay (day 5). Day 4 embryos were placed on bovine serum albumin/fibronectin-coated plates and embryo outgrowth assessed 93 h later as a marker of implantation potential. AGE effects on cell lineage allocation were reassessed following pharmacological interventions: either 12.5 nmol/l AGE receptor (RAGE) antagonist; 0.1 nmol/l metformin; or combination of 10 µmol/l acetyl-l-carnitine, 10 µmol/l N-acetyl-l-cysteine, and 5 µmol/l alpha-lipoic acid. RESULTS: 8 µmol/l AGE reduced: hatching rates (day 5, P < 0.01); total cell number (days 4, 5, P < 0.01); TE cell number (day 5, P < 0.01), and embryo outgrowth (P < 0.01). RAGE antagonism improved day 5 TE cell number. CONCLUSIONS: AGE equimolar with the obese uterine environment detrimentally impact preimplantation embryo development. In natural cycles, prolonged exposure to AGE may developmentally compromise embryos, whereas following assisted reproductive technology cycles, placement of a high-quality embryo into an adverse 'high AGE' environment may impede implantation success. The modest impact of short-term RAGE antagonism on improving embryo outcomes indicates preconception AGE reduction via pharmacological or dietary intervention may improve reproductive outcomes for overweight/obese women.

Differential growth of bowel commensal Bacteroides species on plant xylans of differing structural complexity.

Alterations to the composition of the bowel microbiota (dysbioses) are associated with particular diseases and conditions of humans. There is a need to discover new, indigestible polysaccharides which are selective growth substrates for commensal bowel bacteria. These substrates (prebiotics) could be added to food in intervention studies to correct bowel dysbiosis. A collection of commensal bacteria was screened for growth in culture using a highly-branched xylan produced by New Zealand flax. Two, Bacteroides ovatus ATCC 8483 and Bacteroides xylanisolvens DSM 18836 grew well on this substrate. The utilisation of the xylan was studied chromatographically and by constituent sugar analysis. The two closely related species utilised the xylan in different ways, and differently from their use of wheat arabinoxylan. The growth of Bacteroides species on other plant xylans having differing chemical structures was also investigated. Novel xylans expand the choice of potential prebiotics that could be used to correct bowel dysbioses.