Biology subjects

Santiago-Moreno, J.

Publications and source records attributed to Santiago-Moreno, J..

3 recordsLinked to original sources

bioRxiv2026

NANOG is dispensable for hypoblast but essential for embryonic disc development during conceptus elongation in sheep

NANOG is a conserved regulator of epiblast identity and is essential for hypoblast development and embryo survival after the blastocyst stage in mice. However, its role during post-blastocyst development remains poorly understood in non-rodent mammals. Here, we investigated the function of NANOG during ovine embryogenesis using CRISPR/Cas9-mediated gene ablation. NANOG ablation did not affect blastocyst formation or the initial development of the epiblast. However, NANOG-deficient blastocysts contained significantly fewer SOX17+ hypoblast cells by day (D) 8, followed by reduced hypoblast migration during early post-hatching stages in D12 in vitro embryos. These defects were transient, as hypoblast development was comparable between NANOG-deficient and wild-type conceptuses during subsequent in vivo conceptus elongation from embryonic day (E) 11 onwards. In contrast, although extraembryonic membranes development and conceptus elongation proceeded normally, NANOG-deficient conceptuses lacked SOX2+ epiblast cells and failed to form an embryonic disc (ED) from E11 onwards. These findings demonstrate that NANOG is dispensable for hypoblast development but is required for epiblast maintenance and ED formation during conceptus elongation in sheep.

developmental biology↗

bioRxiv2026

A Single-Cell Transcriptomic Atlas of Symmetry Breaking Across Eutherian Mammals

The mammalian body plan is established during symmetry breaking and gastrulation. In ungulates and primates, these events occur within a flat embryonic disc and coincide with the developmental window most susceptible to pregnancy loss. Here, we generated a single-cell RNA sequencing atlas of in vivo sheep embryos that defines the timing and molecular programs underlying lineage segregation, as well as the inter-lineage signalling mediating anterior visceral hypoblast specification (AVH) and primitive streak formation. By integrating sheep data with stage-matched datasets from cow, pig, rabbit, mouse, and marmoset, we constructed a cross-species atlas of anterior-posterior axis formation and identified conserved anterior and posterior hypoblast markers. Despite major differences in embryo morphology and developmental tempo, core signalling pathways NODAL, WNT, BMP, and FGF were broadly conserved. However, species-specific features emerged, including divergent BMP and FGF ligand usage and distinct BMP sources: only the mouse relied on trophectoderm-derived BMPs, whereas in other mammals BMPs originated primarily from the hypoblast. Functional experiments showed that NODAL is dispensable for early epiblast specification but essential for maintaining epiblast and AVH survival during symmetry breaking in sheep. These findings establish sheep as a powerful model for understanding human peri-implantation development and improving stem cell-based embryo models.

developmental biology↗

bioRxiv2025

Tailoring a glycerol-free cryopreservation protocol with anti-freeze (glycol)proteins for commercial and native breeds of chicken

This study evaluated the effects of antifreeze proteins AFPI, AFPIII, and AFGP during fresh processing, and freezing-thawing of chicken semen from a commercial broiler breed (BB) and the local breed Yellow Hungarian (YH), using 0.6 mol/L DMA as cryoprotectant. AFPI had small but significant (p<0.05) positive effects on post-thaw sperm viability and motility in BB. For YH semen, three freezing protocols with different dilution rates (4x, 2.3x, 2.1x) were compared. Protocol 1 (dilution rate 4x) gave the best post-thaw viability (P<0.001) and was used further. Fertility rates (FR) of pre-freezing steps were tested by inseminating WL hens (4 AI/hen in 2 weeks, 100 x 10L sp/dose). FRs for A) raw semen, B) chilled semen, C) B + DMA, D) B + AFPI, and E) B + DMA + AFPI were 48.2, 3.7, 31.4, 21.4, and 16.1%, respectively. B was lower than A (p<0.001), while C-E were higher than B (p<0.005). AFPI during freezing gave no advantage compared with DMA, except for improved post-thaw DNA integrity. Inseminations with frozen-thawed semen (8 AI/hen in 3 weeks, 100 x 10L sp/dose) gave low FR with YH semen, regardless of hen type. FR with YH semen was 0% without AFPI and 1.5% with AFPI. The oviduct embryo mortality (18.0%{+/-}1.4) observed across all the different hen groups suggested insufficient number of spermatozoa inseminated. In conclusion, AFPI improved some post-thaw traits, but fertility outcomes remain inconclusive. Preventing pre-freezing fertility loss and increasing sperm dose concentration are required.

cell biology↗