Abstract
Current in vitro hematopoiesis models fail to demonstrate the cellular diversity and complex functions of living bone marrow; hence, most translational studies relevant to the hematologic system are conducted in live animals. Here we describe a method for fabricating 'bone marrow–on–a–chip' that permits culture of living marrow with a functional hematopoietic niche in vitro by first engineering new bone in vivo, removing it whole and perfusing it with culture medium in a microfluidic device. The engineered bone marrow (eBM) retains hematopoietic stem and progenitor cells in normal in vivo–like proportions for at least 1 week in culture. eBM models organ-level marrow toxicity responses and protective effects of radiation countermeasure drugs, whereas conventional bone marrow culture methods do not. This biomimetic microdevice offers a new approach for analysis of drug responses and toxicities in bone marrow as well as for study of hematopoiesis and hematologic diseases in vitro.
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References
Sacchetti, B. et al. Self-renewing osteoprogenitors in bone marrow sinusoids can organize a hematopoietic microenvironment. Cell 131, 324–336 (2007).
Chan, C.K.F. et al. Endochondral ossification is required for haematopoietic stem-cell niche formation. Nature 457, 490–494 (2009).
Méndez-Ferrer, S. Mesenchymal and haematopoietic stem cells form a unique bone marrow niche. Nature 466, 829–834 (2010).
Orkin, S.H. & Zon, L.I. Hematopoiesis: an evolving paradigm for stem cell biology. Cell 132, 631–644 (2008).
Wang, L.D. & Wagers, A.J. Dynamic niches in the origination and differentiation of haematopoietic stem cells. Nat. Rev. Mol. Cell Biol. 12, 643–655 (2011).
Di Maggio, N. et al. Toward modeling the bone marrow niche using scaffold-based 3D culture systems. Biomaterials 32, 321–329 (2011).
Takagi, M. Cell processing engineering for ex-vivo expansion of hematopoietic cells. J. Biosci. Bioeng. 99, 189–196 (2005).
Nichols, J.E. et al. In vitro analog of human bone marrow from 3D scaffolds with biomimetic inverted colloidal crystal geometry. Biomaterials 30, 1071–1079 (2009).
Cook, M.M. et al. Micromarrows-three-dimensional coculture of hematopoietic stem cells and mesenchymal stromal cells. Tissue Eng. Part C Methods 18, 319–328 (2012).
Csaszar, E. et al. Rapid expansion of human hematopoietic stem cells by automated control of inhibitory feedback signaling. Cell Stem Cell 10, 218–229 (2012).
Boitano, A.E. et al. Aryl hydrocarbon receptor antagonists promote the expansion of human hematopoietic stem cells. Science 329, 1345–1348 (2010).
Cao, X. et al. Irradiation induces bone injury by damaging bone marrow microenvironment for stem cells. Proc. Natl. Acad. Sci. USA 108, 1609–1614 (2011).
Askmyr, M., Quach, J. & Purton, L.E. Effects of the bone marrow microenvironment on hematopoietic malignancy. Bone 48, 115–120 (2011).
Greenberger, J.S. & Epperly, M. Bone marrow-derived stem cells and radiation response. Semin. Radiat. Oncol. 19, 133–139 (2009).
Meads, M.B., Hazlehurst, L.A. & Dalton, W.S. The bone marrow microenvironment as a tumor sanctuary and contributor to drug resistance. Clin. Cancer Res. 14, 2519–2526 (2008).
Scotti, C. et al. Engineering of a functional bone organ through endochondral ossification. Proc. Natl. Acad. Sci. USA 110, 3997–4002 (2013).
Lee, J. et al. Implantable microenvironments to attract hematopoietic stem/cancer cells. Proc. Natl. Acad. Sci. USA 109, 19638–19643 (2012).
Reddi, A.H. & Huggins, C. Biochemical sequences in the transformation of normal fibroblasts in adolescent rats. Proc. Natl. Acad. Sci. USA 69, 1601–1605 (1972).
Krupnick, A.S., Shaaban, S., Radu, A. & Flake, A.W. Bone marrow tissue engineering. Tissue Eng. 8, 145–155 (2002).
Chen, B. et al. Homogeneous osteogenesis and bone regeneration by demineralized bone matrix loading with collagen-targeting bone morphogenetic protein-2. Biomaterials 28, 1027–1035 (2007).
Schwartz, Z. et al. Differential effects of bone graft substitutes on regeneration of bone marrow. Clin. Oral Implants Res. 19, 1233–1245 (2008).
Ekelund, A., Brosjö, O. & Nilsson, O.S. Experimental induction of heterotopic bone. Clin. Orthop. Relat. Res. 263, 102–112 (1991).
Naveiras, O. et al. Bone-marrow adipocytes as negative regulators of the haematopoietic microenvironment. Nature 460, 259–263 (2009).
Xie, Y. et al. Detection of functional haematopoetic stem cell niche using real-time imaging. Nature 457, 97–101 (2009).
Calvi, L.M. et al. Osteoblastic cells regulate the haematopoietic stem cell niche. Nature 425, 841–846 (2003).
Ding, L. & Morrison, S.J. Haematopoietic stem cells and early lymphoid progenitors occupy distinct bone marrow niches. Nature 495, 231–235 (2013).
Zou, Y.-R. et al. Function of the chemokine receptor CXCR4 in haematopoiesis and in cerebellar development. Nature 393, 595–599 (1998).
Peled, A. et al. The chemokine SDF-1 stimulates integrin-mediated arrest of CS34+ cells on vascular endothelium under shear flow. J. Clin. Invest. 104, 1199–1211 (1999).
Olaharski, A.J. et al. In vitro to in vivo concordance of a high throughput assay of bone marrow toxicity across a diverse set of drug candidates. Toxicol. Lett. 188, 98–103 (2009).
Hoeksema, K.A. et al. Systematic in-vitro evaluation of the NCI/NIH Developmental Therapeutics Program Approved Oncology Drug Set for the identification of a candidate drug repertoire for MLL-rearranged leukemia. OncoTargets and Therapy 4, 149–168 (2011).
Dexter, T.M., Wright, E.G., Krizsa, F. & Lajtha, L.G. Regulation of haemopoietic stem cells proliferation in long term bone marrow cultures. Biomedicine 27, 344–349 (1977).
Bryder, D. & Jacobsen, E.W. Interleukin-3 supports expansion of long-term multilineage repopulating activity after multiple stem cell divisions in vitro. Blood 96, 1748–1755 (2000).
Miller, C.L. & Eaves, C.J. Expansion in vitro of adult murine hematopoietic stem cells with transplantable lympho-myeloid reconstituting ability. Proc. Natl. Acad. Sci. USA 94, 13648–13653 (1997).
Williams, J.P. et al. Animal models for medical countermeasures to radiation exposure. Radiat. Res. 173, 557–578 (2010).
Cary, L.H., Ngudiankama, B.F., Salber, R.E., Ledney, G.D. & Whitnall, M.H. Efficacy of radiation countermeasures depends on radiation quality. Radiat. Res. 177, 663–675 (2012).
Huh, D. et al. Microengineered physiological biomimicry: organs-in-chips. Lab Chip 12, 2156–2164 (2012).
Schwartz, R.M., Palsson, B.O. & Emerson, S.G. Rapid medium perfusion rate significantly increase the productivity and longevity of human bone marrow cultures. Proc. Natl. Acad. Sci. USA 88, 6760–6764 (1991).
Wendt, D., Stroebel, S., Jakob, M., John, G.T. & Martin, I. Uniform tissues engineered by seeding and culturing cells in 3D scaffolds under perfusion at defined oxygen tensions. Biorheology 43, 481–488 (2006).
Hérodin, F. & Drouet, M. Cytokine-based treatment of accidentally irradiated victims and new approaches. Exp. Hematol. 33, 1071–1080 (2005).
Acknowledgements
We thank G.Q. Daley for guidance and helpful discussions and P.L. Wenzel, N. Arora, E. Jiang, A. Jiang, B. Mosadegh, D. Huh, A. Bahinski and G.A. Hamilton for their technical assistance and advice. This work was supported by the Wyss Institute for Biologically Inspired Engineering at Harvard University, the Defense Advanced Research Projects Agency under Cooperative Agreement Number W911NF-12-2-0036, and the US Food and Drug Administration (FDA) HHSF223201310079C.
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Y.-s.T., C.S.S. and D.E.I. conceived of the experiments; Y.-s.T. and C.S.S. performed the experiments, designed research and analyzed data with assistance from T.M., A.M., J.C.W., T.T., J.J.C. and D.E.I. Y.-s.T., C.S.S. and D.E.I. wrote the manuscript.
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Torisawa, Ys., Spina, C., Mammoto, T. et al. Bone marrow–on–a–chip replicates hematopoietic niche physiology in vitro. Nat Methods 11, 663–669 (2014). https://doi.org/10.1038/nmeth.2938
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DOI: https://doi.org/10.1038/nmeth.2938
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