Table 1.

Characteristic features, applications, advantages, and limitations of engineering approaches for T lymphopoesis

Engineering approaches	Applications	Advantages (+) and limitations (−)	Representative and notable references
1) Reconstruction of TSC’s 3-D network
• Fetal thymic organ culture (FTOC) • Reaggregate thymus organ culture (RTOC) • Artificial scaffolds • Decellularized thymic scaffolds	• To study T cell tolerance and MHC restriction in vitro • To study thymopoiesis in vivo upon grafting into an ectopic locations of athymic mice	• Simple and straightforward design (+) • Absolute dependency on biopsy and isolation of thymus or thymic cells (−) • Limited culture sizes of 3-D platforms (−) • Limited number of T cells that can be generated in vitro (−)	• FTOC [27, 31] • RTOC [38] • Grafting in ectopic locations [32, 36] • Artificial scaffolds [40, 41, 44] • Decellularized scaffolds [45]
2) Cellular Engineering
• Differentiation of stem cells into TSCs • Genetic introduction of effector molecules that define TSC functions to cell lines or somatic cells • Cell reprogramming	• To use human pluripotent stem cells for regeneration of thymus or induction of immune tolerance • To generate T cell precursors and functional T cells using robust 2-D culture platforms in vitro	• Use of clinically relevant, endogenous stem cell sources (+) • Use of readily available 2-D culture platforms for recapitulation of T lymphopoesis in vitro (+) • Potential xenogenic cross-contamination (OP9-DL1) (−) • Ineffective positive selection of CD4+ T cells (OP9-DL1) (−) • Need for complex genetic modifications and related risk of viral contamination (−)	• mESC to TEPC [50, 51] • hESC to TEC [53, 54] • OP9-DL1 [59] • Clinical usage of OP9-DL1 platform [67–69] • Cell reprogramming [49]
3) Biomaterials-driven artificial presentation of developmental signaling molecules
• Plate- or bead-bound Notch ligands for differentiation of T precursors from various stem cells • Use of pMHC tetramer to induce antigen specificity on developing T cells	• To generate T precursors from various stem cells in vitro, which later can be employed in adoptive cell therapies. • To induce or selectively expand antigen-specific T cells	• Potential realization of purely biomaterial-based T lymphopoesis ex vivo (+) • Requirement for expensive recombinant proteins (−) • Generation of potentially self-reactive T cells due to lack of negative selection (−) • Limited T cell expansion (−)	• Notch ligands [74, 75, 77, 79] • pMHC tetramer [80, 81]