TY - JOUR
T1 - Identification of factors promoting ex vivo maintenance of mouse hematopoietic stem cells by long-term single-cell quantification
AU - Kokkaliaris, Konstantinos D.
AU - Drew, Erin
AU - Endele, Max
AU - Loeffler, Dirk
AU - Hoppe, Philipp S.
AU - Hilsenbeck, Oliver
AU - Schauberger, Bernhard
AU - Hinzen, Christoph
AU - Skylaki, Stavroula
AU - Theodorou, Marina
AU - Kieslinger, Matthias
AU - Lemischka, Ihor
AU - Moore, Kateri
AU - Schroeder, Timm
N1 - Publisher Copyright:
© 2016 by The American Society of Hematology.
PY - 2016/9/1
Y1 - 2016/9/1
N2 - The maintenance of hematopoietic stem cells (HSCs) during ex vivo culture is an important prerequisite for their therapeutic manipulation. However, despite intense research, culture conditions for robust maintenance of HSCs are still missing. Cultured HSCs are quickly lost, preventing their improved analysis and manipulation. Identification of novel factors supporting HSC ex vivo maintenance is therefore necessary. Coculture with the AFT024 stroma cell line is capable of maintaining HSCs ex vivo longterm, but the responsible molecular players remain unknown. Here, we use continuous long-termsingle-cell observation to identify the HSC behavioral signature under supportive or nonsupportive stroma cocultures. We report early HSC survival as amajor characteristic of HSC-maintaining conditions. Behavioral screening after manipulation of candidate molecules revealed that the extracellular matrix protein dermatopontin (Dpt) is involved in HSCmaintenance. DPT knockdown in supportive stroma impaired HSC survival, whereas ectopic expression of the Dpt gene or protein in nonsupportive conditions restored HSC survival. Supplementing definedstroma- andserum-free cultureconditions with recombinant DPT protein improved HSC clonogenicity. These findings illustrate a previously uncharacterized role of Dpt in maintaining HSCs ex vivo.
AB - The maintenance of hematopoietic stem cells (HSCs) during ex vivo culture is an important prerequisite for their therapeutic manipulation. However, despite intense research, culture conditions for robust maintenance of HSCs are still missing. Cultured HSCs are quickly lost, preventing their improved analysis and manipulation. Identification of novel factors supporting HSC ex vivo maintenance is therefore necessary. Coculture with the AFT024 stroma cell line is capable of maintaining HSCs ex vivo longterm, but the responsible molecular players remain unknown. Here, we use continuous long-termsingle-cell observation to identify the HSC behavioral signature under supportive or nonsupportive stroma cocultures. We report early HSC survival as amajor characteristic of HSC-maintaining conditions. Behavioral screening after manipulation of candidate molecules revealed that the extracellular matrix protein dermatopontin (Dpt) is involved in HSCmaintenance. DPT knockdown in supportive stroma impaired HSC survival, whereas ectopic expression of the Dpt gene or protein in nonsupportive conditions restored HSC survival. Supplementing definedstroma- andserum-free cultureconditions with recombinant DPT protein improved HSC clonogenicity. These findings illustrate a previously uncharacterized role of Dpt in maintaining HSCs ex vivo.
UR - http://www.scopus.com/inward/record.url?scp=85016325794&partnerID=8YFLogxK
U2 - 10.1182/blood-2016-03-705590
DO - 10.1182/blood-2016-03-705590
M3 - Article
C2 - 27365423
AN - SCOPUS:85016325794
SN - 0006-4971
VL - 128
SP - 1181
EP - 1192
JO - Blood
JF - Blood
IS - 9
ER -