Distinct roles of IL-7 and stem cell factor in the OP9-DL1 T-cell differentiation culture system.
OBJECTIVE
The OP9-DL1 culture system is an in vitro model for T-cell development in which activation of the Notch pathway by Delta-like 1 promotes differentiation of mature T cells from progenitors. The roles of specific cytokines in this culture system have not been well defined, and controversy regarding the role of IL-7 has recently emerged. We examined the roles played by IL-7, Flt3 ligand, and stem cell factor (SCF) in differentiation of adult bone marrow cells in the OP9-DL1 culture system.


METHODS
Hematopoietic progenitor cells isolated from mouse bone marrow were cultured with OP9 or OP9-DL1 stromal cells and evaluated for T and B lymphocyte differentiation using immunofluorescent staining.


RESULTS
IL-7 provided both survival/proliferation and differentiation signals in a dose-dependent manner. T-cell development from the CD4/CD8 double-negative (DN) stage to the CD4/CD8 double-positive (DP) stage required IL-7 provided by the stromal cells, while differentiation from the DP to the CD8 single-positive (SP) stage required addition of exogenous IL-7. SCF favored the proliferation of DN lymphoid progenitors and inhibited differentiation to the DP stage in a dose-dependent manner. Conversely, blocking the function of SCF expressed endogenously by OP9-DL1 cells inhibited proliferation of lymphoid progenitors and accelerated T-lineage differentiation. Flt3 ligand promoted proliferation without affecting differentiation.


CONCLUSIONS
These results validate the OP9-DL1 model for the analysis of T-cell development from bone marrow-derived progenitor cells, and demonstrate specific roles of SCF, IL-7, and Flt3L in promoting efficient T-lineage differentiation.
Wang
Hongfang
H
Department of Pathology, The University of Utah, Salt Lake City, Utah 84132-2022, USA.
Pierce
L Jeanne
LJ
Spangrude
Gerald J
GJ
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001402:B-Lymphocytes / Q000276:immunology; D001549:Benzamides; D001854:Bone Marrow Cells / Q000166:cytology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000502:physiology*; D000068877:Imatinib Mesylate; D015851:Interleukin-7 / Q000502:physiology*; D008565:Membrane Proteins / Q000502:physiology; D051379:Mice; D020297:Mice, Congenic; D008810:Mice, Inbred C57BL; D010879:Piperazines / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D013194:Staining and Labeling; D019089:Stem Cell Factor / Q000037:antagonists & inhibitors / Q000502:physiology*; D017154:Stromal Cells / Q000166:cytology / Q000502:physiology; D013601:T-Lymphocytes / Q000276:immunology*
D000906:Antibodies; D001549:Benzamides; D015851:Interleukin-7; D008565:Membrane Proteins; D010879:Piperazines; D011743:Pyrimidines; D019089:Stem Cell Factor; C084881:flt3 ligand protein; D000068877:Imatinib Mesylate