Automated, High-Throughput Phenotypic Screening and Analysis Platform to Study Pre- and Post-Implantation Morphogenesis in Stem Cell-Derived Embryo-Like Structures.

Combining high-throughput generation and high-content imaging of embryo models will enable large-scale screening assays in the fields of (embryo) toxicity, drug development, embryogenesis, and reproductive medicine. This study shows the continuous culture and in situ (i.e., in microwell) imaging-based readout of a 3D stem cell-based model of peri-implantation epiblast (Epi)/extraembryonic endoderm (XEn) development with an expanded pro-amniotic cavity (PAC) (E3.5 E5.5), namely XEn/EPiCs. Automated image analysis and supervised machine learning permit the identification of embryonic morphogenesis, tissue compartmentalization, cell differentiation, and consecutive classification. Screens with signaling pathway modulators at different time windows provide spatiotemporal information on their phenotypic effect on developmental processes leading to the formation of XEn/EPiCs. Exposure of the biological model in the microwell platform to pathway modulators at two time windows, namely 0-72 h and 48-120 h, show that Wnt and Fgf/MAPK pathway modulators affect Epi differentiation and its polarization, while modulation of BMP and Tgfß/Nodal pathway affects XEn specification and epithelialization. Further, their collective role is identified in the timing of the formation and expansion of PAC. The newly developed, scalable culture and analysis platform, thereby, provides a unique opportunity to quantitatively and systematically study effects of pathway modulators on early embryonic development.

From Mice to Men: Generation of Human Blastocyst-Like Structures In Vitro.

Advances in the field of stem cell-based models have in recent years lead to the development of blastocyst-like structures termed blastoids. Blastoids can be used to study key events in mammalian pre-implantation development, as they mimic the blastocyst morphologically and transcriptionally, can progress to the post-implantation stage and can be generated in large numbers. Blastoids were originally developed using mouse pluripotent stem cells, and since several groups have successfully generated blastocyst models of the human system. Here we provide a comparison of the mouse and human protocols with the aim of deriving the core requirements for blastoid formation, discuss the models' current ability to mimic blastocysts and give an outlook on potential future applications.

From Snapshots to Development: Identifying the Gaps in the Development of Stem Cell-based Embryo Models along the Embryonic Timeline.

In recent years, stem cell-based models that reconstruct mouse and human embryogenesis have gained significant traction due to their near-physiological similarity to natural embryos. Embryo models can be generated in large numbers, provide accessibility to a variety of experimental tools such as genetic and chemical manipulation, and confer compatibility with automated readouts, which permits exciting experimental avenues for exploring the genetic and molecular principles of self-organization, development, and disease. However, the current embryo models recapitulate only snapshots within the continuum of embryonic development, allowing the progression of the embryonic tissues along a specific direction. Hence, to fully exploit the potential of stem cell-based embryo models, multiple important gaps in the developmental landscape need to be covered. These include recapitulating the lesser-explored interactions between embryonic and extraembryonic tissues such as the yolk sac, placenta, and the umbilical cord; spatial and temporal organization of tissues; and the anterior patterning of embryonic development. Here, it is detailed how combinations of stem cells and versatile bioengineering technologies can help in addressing these gaps and thereby extend the implications of embryo models in the fields of cell biology, development, and regenerative medicine.

Disparate effects of PEG or albumin based surface modification on the uptake of nano- and micro-particles.

Surface modification of particulate systems is a commonly employed strategy to alter their interaction with proteins and cells. Past studies on nano-particles have shown that surface functionalization with polyethylene glycol (PEG) or proteins such as albumin increases circulation times by reducing their phagocytic uptake. However, studies on surface functionalized micro-particles have reported contradictory results. Here, we investigate the effects of surface functionalization using polystyrene particles with 4 different diameters ranging from 30 nm to 2.6 µm and coating them with either albumin or PEG. Our results show that with increasing particle size, surface functionalization has less to no effect on altering phagocytic uptake. The data also suggest that these differences are observed with a dense arrangement of molecules on the surface (dense brush conformation for PEG conjugation), appear to be independent of the serum proteins adsorbing on particle surfaces, and are independent of the endocytic uptake pathway. These results provide insight into the differences in the ability of surface modified nano- and micro-particles to avoid phagocytic uptake.