Establishing a physiologically relevant in vitro model of blood development

The goal of this project is the establishment of a faithful 3D model of human embryonic blood development from pluripotent stem cells (PSCs), which recapitulates emergence of successive waves of blood progenitors and the specification of haematopoietic stem cells (HSCs), to replace embryo usage. Such a platform will enable systematic mechanistic understanding of HSC formation at a throughput that is not possible in embryo studies. It will pave the way for efficient and scalable production of HSC and their derivatives. And it will provide access to leukaemia transformation events which occur in utero to improve prognosis of babies with leukaemia.

Although PSCs (embryonic and induced) have been extensively used to model blood formation, they commonly fail to produce blood with adult characteristics. Production of HSCs has been rarely reported, and is either dependent on extensive genetic engineering or on unique culture conditions not easily transferrable or scalable for widespread use. Furthermore, protocols that generate HSCs do not achieve physiological stepwise reconstitution of the different embryonic haematopoietic niches and the successive waves of progenitor cell production, preventing mechanistic studies of developmental haematopoiesis, normal and leukaemic. Mechanistic understanding remains dependent on animal embryos and human abortion material, which raise ethical considerations and lack the throughput to enable systematic studies.

Three-dimensional models of embryogenesis have sought to capture early development in vitro. Amongst these, gastruloids successfully mirrored symmetry breaking, axis formation, and key events like somitogenesis with temporal and spatial congruency. Gastruloids lack developed extra-embryonic structures and cannot generate viable implantable embryos, constituting a widely-acceptable ethical approach to replacing embryo usage.

We recently developed a mouse PSC-based haemogenic gastruloid (hGx) which captures successive haematopoietic waves, and produces cells with transcriptional characteristics of HSCs capable of at least transient haematopoietic establishment in vivo. Herein, we propose to establish a human hGx model from induced PSCs, which can be used for mechanistic understanding of blood formation, and optimised for robust generation of HSCs. We will make use of available single-cell omics data on embryonic blood and HSC emergence to inform hGx development and systematically test cellular trajectories and gene regulatory networks against in vivo data and in response to chemical signals. We will add mechanical cues such as stiffness and shear stress to further optimise niche reconstitution.

Successful generation of a faithful in vitro model of developmental haematopoiesis will fully replace embryo usage for investigational and regenerative applications in blood and HSC production.