Perturbation of single hematopoietic stem cell fates in artificial niches.
Hematopoietic stem cells (HSCs) are capable of extensive self-renewal in vivo and are successfully employed clinically to treat hematopoietic malignancies, yet are in limited supply as in culture this self-renewal capacity is lost. Using an approach at the interface of stem cell biology and bioengineering, here we describe a novel platform of hydrogel microwell arrays for assessing the effects of either secreted or tethered proteins characteristic of the in vivo microenvironment, or niche, on HSC fate in vitro. Time-lapse microscopic analyses of single cells were crucial to overcoming inevitable heterogeneity of FACS-enriched HSCs. A reduction in proliferation kinetics or an increase in asynchronous division of single HSCs in microwells in response to specific proteins (Wnt3a and N-Cadherin) correlated well with subsequent serial long-term blood reconstitution in mice in vivo. Single cells that divided once in the presence of a given protein were capable of in vivo reconstitution, providing evidence of self-renewal divisions of HSCs in vitro. These results validate the hydrogel microwell platform as a broadly applicable paradigm for dissecting the regulatory role of specific signals within a complex stem cell niche.
Lutolf
Matthias P
MP
Department of Microbiology and Immunology, BioX and Stem Cell Institute, Stanford University School of Medicine, CA 94305, USA.
Doyonnas
Regis
R
Havenstrite
Karen
K
Koleckar
Kassie
K
Blau
Helen M
HM
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation*; D002469:Cell Separation / Q000295:instrumentation*; D002478:Cells, Cultured; D004867:Equipment Design; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D046228:Microarray Analysis / Q000295:instrumentation*