An in vitro neurogenetics platform for precision disease modeling in the mouse.

The power and scope of disease modeling can be markedly enhanced through the incorporation of broad genetic diversity. The introduction of pathogenic mutations into a single inbred mouse strain sometimes fails to mimic human disease. We describe a cross-species precision disease modeling platform that exploits mouse genetic diversity to bridge cell-based modeling with whole organism analysis. We developed a universal protocol that permitted robust and reproducible neural differentiation of genetically diverse human and mouse pluripotent stem cell lines and then carried out a proof-of-concept study of the neurodevelopmental gene DYRK1A. Results in vitro reliably predicted the effects of genetic background on Dyrk1a loss-of-function phenotypes in vivo. Transcriptomic comparison of responsive and unresponsive strains identified molecular pathways conferring sensitivity or resilience to Dyrk1a1A loss and highlighted differential messenger RNA isoform usage as an important determinant of response. This cross-species strategy provides a powerful tool in the functional analysis of candidate disease variants identified through human genetic studies.

Scalable Generation of Pre-Vascularized and Functional Human Beige Adipose Organoids.

Obesity and type 2 diabetes are becoming a global sociobiomedical burden. Beige adipocytes are emerging as key inducible actors and putative relevant therapeutic targets for improving metabolic health. However, in vitro models of human beige adipose tissue are currently lacking and hinder research into this cell type and biotherapy development. Unlike traditional bottom-up engineering approaches that aim to generate building blocks, here a scalable system is proposed to generate pre-vascularized and functional human beige adipose tissue organoids using the human stromal vascular fraction of white adipose tissue as a source of adipose and endothelial progenitors. This engineered method uses a defined biomechanical and chemical environment using tumor growth factor ß (TGFß) pathway inhibition and specific gelatin methacryloyl (GelMA) embedding parameters to promote the self-organization of spheroids in GelMA hydrogel, facilitating beige adipogenesis and vascularization. The resulting vascularized organoids display key features of native beige adipose tissue including inducible Uncoupling Protein-1 (UCP1) expression, increased uncoupled mitochondrial respiration, and batokines secretion. The controlled assembly of spheroids allows to translate organoid morphogenesis to a macroscopic scale, generating vascularized centimeter-scale beige adipose micro-tissues. This approach represents a significant advancement in developing in vitro human beige adipose tissue models and facilitates broad applications ranging from basic research to biotherapies.

Cell Aggregate Assembly through Microengineering for Functional Tissue Emergence.

Compared to cell suspensions or monolayers, 3D cell aggregates provide cellular interactions organized in space and heterogeneity that better resume the real organization of native tissues. They represent powerful tools to narrow down the gap between in vitro and in vivo models, thanks to their self-evolving capabilities. Recent strategies have demonstrated their potential as building blocks to generate microtissues. Developing specific methodologies capable of organizing these cell aggregates into 3D architectures and environments has become essential to convert them into functional microtissues adapted for regenerative medicine or pharmaceutical screening purposes. Although the techniques for producing individual cell aggregates have been on the market for over a decade, the methodology for engineering functional tissues starting from them is still a young and quickly evolving field of research. In this review, we first present a panorama of emerging cell aggregates microfabrication and assembly technologies. We further discuss the perspectives opened in the establishment of functional tissues with a specific focus on controlled architecture and heterogeneity to favor cell differentiation and proliferation.