Engineering the Human Thymic Microenvironment to Support Thymopoiesis In Vivo

Abstract

A system that allows manipulation of the human thymic microenvironment is needed both to elucidate the extrinsic mechanisms that control human thymopoiesis, and to develop potential cell therapies for thymic insufficiency. In this report, we developed an implantable thymic microenvironment composed of two human thymic stroma populations critical for thymopoiesis; thymic epithelial cells (TECs) and thymic mesenchyme (TM). TECs and TM from postnatal human thymi were cultured in specific conditions, allowing cell expansion and manipulation of gene expression, prior to re-aggregation into a functional thymic unit. Human CD34+ hematopoietic stem and progenitor cells (HSPC) differentiated into T cells in the aggregates in vitro and in vivo following inguinal implantation of aggregates in immune deficient mice. Cord blood HSPC previously engrafted into murine bone marrow, migrated to implants and differentiated into human T cells with a broad T cell receptor repertoire. Furthermore, lentiviral-mediated expression of vascular endothelial growth factor in TM enhanced implant size and function, and significantly increased thymocyte production. These results demonstrate an in vivo system for the generation of T cells from human HSPC, and represent the first model to allow manipulation of gene expression and cell composition in the microenvironment of the human thymus.

Generation of mature T cells from human hematopoietic stem and progenitor cells in artificial thymic organoids

“…A specific role for 3D structure in T cell positive selection is consistent with the reported ability of FTOCs and reaggregated primary thymic stromal organoids to permit T cell maturation, albeit at low efficiencies 10, 11, 12 . 3D interactions may support positive selection by increasing the valence and/or duration of contact between T cell precursors and selective ligands (such as pMHC); facilitating crosstalk between stromal and hematopoietic cells; or exerting developmental signals on T cell precursors through mechanical forces and/or metabolic gradients not otherwise possible in 2D.…”

Thymus: the next (re)generation

“…In trials (136,138,141,(193)(194)(195)(196)(197) IL-12 Thymocytes Preclinical (198,199) Preclinical (187,190,191,212,213) Thymus bioengineering Mature T cells Preclinical (192,214) IND, Investigational New Drug; HSPC, Hematopoietic Stem/Progenitor Cells; Flt3L, FMS-like tyrosine kinase 3 ligand.…”

Enabling allogeneic therapies: CIRM-funded strategies for immune tolerance and immune evasion

“…A similar approach of using TEC for the purpose of inducing tolerance was taken by Dr Gay Crooks at UCLA, as part of a CIRM‐funded project in which she generated organoids composed of human fetal TEC and thymic mesenchyme cells that also are capable of supporting T‐cell differentiation (Table 1). 15 Dr Crooks' technology has further matured to the point that her artificial thymic organoid can support efficient and scalable production of T cells in vitro from either primary HSC 16 or reprogrammed PSC 17 . This technology represents a significant advance in the ability to manufacture T cell‐based immunotherapies.…”