RRC ID 64905
Author Mohtashami M, Zarin P, Zúñiga-Pflücker JC.
Title Induction of T Cell Development In Vitro by Delta-Like (Dll)-Expressing Stromal Cells.
Journal Methods Mol Biol
Abstract Recreating the thymic microenvironment in vitro poses a great challenge to immunologists. Until recently, the only approach was to utilize the thymic tissue in its three-dimensional form and to transfer the hematopoietic progenitors into this tissue to generate de novo T cells. With the advent of OP9-DL cells (bone marrow-derived cells that are transduced to express Notch ligand, Delta-like), hematopoietic stem cells (HSC) could be induced to differentiate into T cells in culture for the first time outside of the thymic tissue on a monolayer. We, as well as others, asked whether the ability to support T cell development in vitro in a monolayer is unique to BM-derived OP9 cells, and showed that provision of Delta-like expression to thymic epithelial cells and fibroblasts also allowed for T cell development. This provides the opportunity to design an autologous coculture system where the supportive stromal and the hematopoietic components are both derived from the same individual, which has obvious clinical implications. In this chapter, we describe methods for establishing a primary murine dermal fibroblast cell population that is transduced to express Delta-like 4, and describe the conditions for its coculture with HSCs to support T cell lineage initiation and expansion, while comparing it to the now classic OP9-DL coculture.
Volume 1323
Pages 159-67
Published 2016-1-1
DOI 10.1007/978-1-4939-2809-5_14
PMID 26294407
MeSH Animals Cell Communication* Cell Differentiation* Cell Line Cell Separation Coculture Techniques Fibroblasts Gene Expression* Hematopoietic Stem Cells / cytology Hematopoietic Stem Cells / metabolism Intracellular Signaling Peptides and Proteins / genetics* Membrane Proteins / genetics* Mice Stromal Cells / metabolism* T-Lymphocytes / cytology* T-Lymphocytes / metabolism*
Human and Animal Cells OP9(RCB1124)