CLPs find their inner T-cell potential

It has recently been debated whether CLPs possess significant amounts of thymocyte progenitor activity. In this issue of Blood, Serwold and colleagues demonstrate that Flk2+CD27+CLP are the major T-cell source.

The clonogenic common lymphoid progenitor (CLP) was originally defined as a population of cells that are Lin(-)IL-7R(+)Thy-1(-)Sca-1(lo)c-Kit(lo) and possess both B-lymphocyte and T-lymphocyte potential.¹  However, the T-cell potential for CLPs has progressively been challenged on the grounds of several observations that led to the alternative hypothesis that CLPs are mostly B-cell committed, and that T-cell progenitors can develop independent of CLPs through routes directly from a subset of multipotent progenitors (MPPs). Evidence has come from initial studies demonstrating longer T-cell output from the MPPs than from CLPs and intact thymopoiesis in ikaros-deficient mice which lack phenotypic CLPs.²  Recently, the ability of early thymic DN1 progenitors to demonstrate myeloid potential in a culture system^3,4  has also been cited as evidence that CLPs cannot account for thymic DN1 progenitors, since CLPs, unlike MPPs, lack myeloid potential. While these studies have suggested alternative routes for T-cell development, they did not factor in the developmental relationship between MPPs and CLPs or the purity of CLP isolation.

A fundamental tenant of the hierarchical development of blood cells is that the stem cell has the greatest proliferative potential, and that each successive progenitor population loses some of that potential and becomes more restricted in lineage development. The hematopoietic stem cell (HSC) is therefore capable of producing more of any particular blood cell lineage over time than a more lineage-restricted progenitor because it can self-renew and provide sustained progenitor pools. In erythroid development, the common myeloid progenitor (CMP) and the megakaryocyte-erythroid progenitor (MEP) have this type of relationship. The MEP possesses the predominant short-term day 8 CFU-S activity and radioprotection potential whereas the CMP can sustain long-term myelopoiesis. Likewise, thymocyte progenitors can arise from both MPPs and CLPs, but the relationship between these progenitor pools dictates that every MPP is capable of producing more CLPs. Therefore, it would be expected that the long-term total output of T cells from transplanted MPPs would be greater than for the CLPs on a per cell basis. However, greater T-cell output from MPPs does not prove that MPPs produce T cells independent of a CLP intermediate.

The important study by Serwold et al has addressed the recent challenges to the origin of thymocyte progenitors by keeping in mind the hierarchical relationship and relative abundance of MPPs and CLPs. The study carefully examined the contribution of both MPPs and CLPs to thymic-seeding ability and T-cell development. Injection of highly purified cells was based on the donor equivalent ratio rather than on the cell number following sorting so as not to artificially bias toward the MPPs, which have greater proliferative potential. Furthermore, the analysis of short-term time points was critical since MPPs would require additional time to differentiate into the CLP. Earlier time points would thus afford the best chance for observing MPP and CLP potential for rapid thymus-seeding capacity.

The study further demonstrates that the purification quality of CLP is important, especially for analyses of T-cell development. The original phenotypic CLP has recently been refined by addition of positivity for Flk2⁵  and now a smaller population of cells including CD27 are shown to contain the T-cell potential. A specific criticism of the CLP as the thymus-seeding cell has been the lack of evidence that CLP can be detected in thymus. The study further challenged the MPP-seeding hypothesis by identifying the presence of a low level of circulating Flk2+CD27+CLP, thus formally demonstrating that CLPs can be detected in transit, presumably to the thymus. Since CLPs have not yet been detected in thymus, likely due to rapid adoption of DN1 phenotype, this is an important step forward by identifying a missing link. Overall, the study provides important information about the inner potential of the originally defined CLP and offers additional markers to help all labs standardize their CLP isolations. Let's hope that these advances will help to resolve the thymic progenitor debate.