Engineering The Human Thymic Microenvironment To Support Thymopoiesis

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. Implantation of intact fetal thymic fragments into immune deficient mice can be used to model human thymopoiesis in vivo, but this approach has limited capacity for cell or molecular manipulation for either experimental or translational application. 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, were able to migrate to the implants and differentiate into functional human T cells with a broad T cell receptor repertoire. Furthermore, lentiviral mediated expression of vascular endothelial growth factor in TM enhanced TEC survival and function, resulting in significantly greater thymocyte production relative to control implants. These results demonstrate an efficient 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.