Osteoblasts and bone marrow mesenchymal stromal cells control hematopoietic stem cell migration and proliferation in 3D in vitro model.
BACKGROUND
Migration, proliferation, and differentiation of hematopoietic stem cells (HSCs) are dependent upon a complex three-dimensional (3D) bone marrow microenvironment. Although osteoblasts control the HSC pool, the subendosteal niche is complex and its cellular composition and the role of each cell population in HSC fate have not been established. In vivo models are complex and involve subtle species-specific differences, while bidimensional cultures do not reflect the 3D tissue organization. The aim of this study was to investigate in vitro the role of human bone marrow-derived mesenchymal stromal cells (BMSC) and active osteoblasts in control of migration, lodgment, and proliferation of HSCs.


RESULTS
A complex mixed multicellular spheroid in vitro model was developed with human BMSC, undifferentiated or induced for one week into osteoblasts. A clear limit between the two stromal cells was established, and deposition of extracellular matrix proteins fibronectin, collagens I and IV, laminin, and osteopontin was similar to the observed in vivo. Noninduced BMSC cultured as spheroid expressed higher levels of mRNA for the chemokine CXCL12, and the growth factors Wnt5a and Kit ligand. Cord blood and bone marrow CD34(+) cells moved in and out the spheroids, and some lodged at the interface of the two stromal cells. Myeloid colony-forming cells were maintained after seven days of coculture with mixed spheroids, and the frequency of cycling CD34(+) cells was decreased.


CONCLUSIONS
Undifferentiated and one-week osteo-induced BMSC self-assembled in a 3D spheroid and formed a microenvironment that is informative for hematopoietic progenitor cells, allowing their lodgment and controlling their proliferation.
de Barros
Ana Paula D N
AP
Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
Takiya
Christina M
CM
Garzoni
Luciana R
LR
Leal-Ferreira
Mona Lisa
ML
Dutra
Hélio S
HS
Chiarini
Luciana B
LB
Meirelles
Maria Nazareth
MN
Borojevic
Radovan
R
Rossi
Maria Isabel D
MI
D000328:Adult; D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis; D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism; D018929:Cell Culture Techniques; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D054377:Chemokine CXCL12 / Q000235:genetics; D018920:Coculture Techniques; D005312:Fetal Blood / Q000166:cytology; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007231:Infant, Newborn; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism; D008854:Microscopy, Electron; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018874:Spheroids, Cellular / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D017154:Stromal Cells / Q000166:cytology* / Q000378:metabolism
D018952:Antigens, CD34; D054377:Chemokine CXCL12