Development of a three-dimensional long-term bone marrow culture system for drug testing

Bone marrow is the hemopoietic tissue and a primary lymphoid organ. Its intricate, three-dimensional architecture facilitates cell-cell and cell-matrix interactions and provides a microenvironment that supports self-renewal and multilineal differentiation. Traditional in vitro culture systems (flask cultures) are limited by the absence of normal marrow spatial organization and cellular interactions with the extracellular matrix. They deviate from the three-dimensional marrow structure, inducing artificial lipogenesis and cell flattening, and produce only limited cell lineages. Our hypotheses are: 1) the marrow’s function in promoting multilineal hemopoiesis depends on its three-dimensional structure and 2) this function can be (substantially) reproduced in vitro only if its three-dimensional structure is preserved. We have developed a novel human ex vivo bone marrow model that mimics bone marrow both structurally and functionally. By providing an artificial scaffolding, we were able to obtain a three-dimensional growth configuration with high cell density and intimate physical contact between hemopoietic and stromal cells. Characterization of the stroma developed demonstrated that an anatomical tissue-like structure was obtained. More importantly, the three-dimensional culture system supported multilineal hemopoiesis, including the development of myeloid, erythroid, and lymphoid lineages. In conclusion, the ability to engineer a structural and functional human bone marrow mimicry ex vivo provides a more physiologically meaningful approach for delineating the hemopoietic microenvironment, identifying the cellular and molecular signals important in hemopotiesis, and could potentially lead to novel bone marrow technologies.