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Journal Article
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Generating human bone marrow organoids for disease modeling and drug discovery.
Nature Protocols 2024 March 27
The bone marrow supports and regulates hematopoiesis, responding to physiological requirements for blood cell production over ontogeny and during pathological challenges. Interactions between hematopoietic cells and niche components are challenging to study mechanistically in the human context, but are important to delineate in order to explore the pathobiology of blood and bone marrow disorders. Organoids are proving transformative in many research settings, but an accurate human bone marrow model incorporating multiple hematopoietic and stromal elements has been lacking. This protocol describes a method to generate three-dimensional, multilineage bone marrow organoids from human induced pluripotent stem cells (hiPSCs), detailing the steps for the directed differentiation of hiPSCs using a series of cytokine cocktails and hydrogel embedding. Over 18 days of differentiation, hiPSCs yield the key lineages that are present in central myelopoietic bone marrow, organized in a well-vascularized architecture that resembles native hematopoietic tissues. This presents a robust, in vitro system that can model healthy and perturbed hematopoiesis in a scalable three-dimensional microenvironment. Bone marrow organoids also support the growth of immortalized cell lines and primary cells from healthy donors and patients with myeloid and lymphoid cancers, including cell types that are poorly viable in standard culture systems. Moreover, we discuss assays for the characterization of organoids, including interrogation of pathogenic remodeling using recombinant TGF-ß treatment, and methods for organoid engraftment with exogenous cells. This protocol can be readily adapted to specific experimental requirements, can be easily implemented by users with tissue culture experience and does not require access to specialist equipment.
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