A new model of human lymphopoiesis across development and aging.

Over the past decade our research has implemented a multimodal approach to human lymphopoiesis, combining clonal-scale mapping of lymphoid developmental architecture with the monitoring of dynamic changes in the pattern of lymphocyte generation across ontogeny. We propose that lymphopoiesis stems from founder populations of CD127/interleukin (IL)7R or CD127/IL7R early lymphoid progenitors (ELPs) polarized respectively toward the T-natural killer (NK)/innate lymphoid cell (ILC) or B lineages, arising from newly characterized CD117 multi-lymphoid progenitors (MLPs). Recent data on the lifelong lymphocyte dynamics of healthy donors suggest that, after birth, lymphopoiesis may become increasingly oriented toward the production of B lymphocytes.

Distinct subsets of multi-lymphoid progenitors support ontogeny-related changes in human lymphopoiesis.

Changes in lymphocyte production patterns occurring across human ontogeny remain poorly defined. In this study, we demonstrate that human lymphopoiesis is supported by three waves of embryonic, fetal, and postnatal multi-lymphoid progenitors (MLPs) differing in CD7 and CD10 expression and their output of CD127 early lymphoid progenitors (ELPs). In addition, our results reveal that, like the fetal-to-adult switch in erythropoiesis, transition to postnatal life coincides with a shift from multilineage to B lineage-biased lymphopoiesis and an increase in production of CD127 ELPs, which persists until puberty.

Modeling Human Fetal Hematopoiesis in Humanized Mice.

Due to difficulties to access primary human bone marrow samples and age or donor effects, human hematopoiesis has long remained far less well characterized than in the mouse. Despite recent progresses in single-cell RNA profiling only little is known as to phenotype, function and developmental trajectories of human lymphomyeloid progenitors and precursors. This is especially true regarding the developmental architecture of the lymphoid lineage which has been the subject of persistent controversies over the past decades.

CD4CD8 T-Lymphocytes in Xenogeneic and Human Graft-versus-Host Disease.

Mechanisms driving acute graft-versus-host disease (aGVHD) onset in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT) are still poorly understood. To provide a detailed characterization of tissue-infiltrating T lymphocytes (TL) and search for eventual site-specific specificities, we developed a xenogeneic model of aGVHD in immunodeficient mice. Phenotypic characterization of xenoreactive T lymphocytes (TL) in diseased mice disclosed a massive infiltration of GVHD target organs by an original CD4CD8 TL subset.

[Bipartite organization of human lymphopoiesis].

Due to difficulties to access primary bone marrow samples, human hematopoiesis has long remained far less characterized than in the mouse. Using an in vivo modeling approach of fetal hematopoiesis in humanized mice, we recently showed that human lymphoid cells stem from two functionally specialized populations of CD127 and CD127 early lymphoid progenitors (ELP) that differentiate independently, respond differently to growth factors, undergo divergent modes of lineage restriction and generate distinct lymphoid populations. Our results demonstrate that, conversely to the mouse, human lymphopoiesis displays a bipartite developmental architecture.

Molecular and Functional Characterization of Lymphoid Progenitor Subsets Reveals a Bipartite Architecture of Human Lymphopoiesis.

The classical model of hematopoiesis established in the mouse postulates that lymphoid cells originate from a founder population of common lymphoid progenitors. Here, using a modeling approach in humanized mice, we showed that human lymphoid development stemmed from distinct populations of CD127 and CD127 early lymphoid progenitors (ELPs). Combining molecular analyses with in vitro and in vivo functional assays, we demonstrated that CD127 and CD127 ELPs emerged independently from lympho-mono-dendritic progenitors, responded differently to Notch1 signals, underwent divergent modes of lineage restriction, and displayed both common and specific differentiation potentials.

Evolutionary conservation of Notch signaling inhibition by TMEM131L overexpression.

Human KIAA0922/TMEM131L encodes a transmembrane protein, TMEM131L, that regulates the canonical Wnt/β-catenin signaling pathway by eliciting the lysosome-dependent degradation of phosphorylated LRP6 co-receptor. Here, we use a heterospecific Drosophila transgenic model to examine the potential evolutionary conservation of TMEM131L function. Analysis of TMEM131L transgenic flies shows that TMEM131L interference with the Wnt pathway results primarily from a Notch-dependent decrease in Wingless production.

Identification of TMEM131L as a novel regulator of thymocyte proliferation in humans.

In this study, we identify transmembrane protein 131-like (TMEM131L) as a novel regulator of thymocyte proliferation and demonstrate that it corresponds to a not as yet reported inhibitor of Wnt signaling. Short hairpin RNA-mediated silencing of TMEM131L in human CD34(+) hematopoietic progenitors, which were then grafted in NOD-SCID/IL-2rγ(null) mice, resulted in both thymocyte hyperproliferation and multiple pre- and post-β-selection intrathymic developmental defects. Consistent with deregulated Wnt signaling, TMEM131L-deficient thymocytes expressed Wnt target genes at abnormally high levels, and they displayed both constitutive phosphorylation of Wnt coreceptor LRP6 and β-catenin intranuclear accumulation.

Dynamics of human prothymocytes and xenogeneic thymopoiesis in hematopoietic stem cell-engrafted nonobese diabetic-SCID/IL-2rγnull mice.

To model the developmental pattern of human prothymocytes and thymopoiesis, we used NOD-scid/γc(-/-) mice grafted with human umbilical cord blood CD34(+) hematopoietic progenitor cells (HPCs). Human prothymocytes developed in the murine bone marrow (BM) from multipotent CD34(++)CD38(lo)lineage(-) HPCs to CD34(++)CD7(+)CD2(-) pro-T1 cells that progressed in a Notch-dependent manner to CD34(+)CD7(++)CD2(+) pro-T2 cells, which migrated to the thymus. BM prothymocyte numbers peaked 1 mo after graft, dropped at mo 2, and persisted at low levels thereafter, with only a few CD34(+)CD7(lo) prothymocytes with limited T potential being detected by mo 5.

AF1q/MLLT11 regulates the emergence of human prothymocytes through cooperative interaction with the Notch signaling pathway.

The mechanisms regulating the emergence of BM prothymocytes remain poorly characterized. Genome-wide transcriptome analyses looking for genes expressed in human prothymocytes led to the identification of AF1q/MLLT11 as a candidate gene conceivably involved in this process. Analysis of AF1q protein subcellular localization and intracellular trafficking showed that despite pronounced karyophily, it was subjected to constitutive nuclear export followed by ubiquitin-mediated degradation in the centrosomal area.

Infection of dendritic cells (DCs), not DC-SIGN-mediated internalization of human immunodeficiency virus, is required for long-term transfer of virus to T cells.

The C-type lectin DC-SIGN expressed on immature dendritic cells (DCs) captures human immunodeficiency virus (HIV) particles and enhances the infection of CD4+ T cells. This process, known as trans-enhancement of T-cell infection, has been related to HIV endocytosis. It has been proposed that DC-SIGN targets HIV to a nondegradative compartment within DCs and DC-SIGN-expressing cells, allowing incoming virus to persist for several days before infecting target cells.

Dynamics of thymus-colonizing cells during human development.

Here, we identify fetal bone marrow (BM)-derived CD34hiCD45RAhiCD7+ hematopoietic progenitors as thymus-colonizing cells. This population, virtually absent from the fetal liver (FL), emerges in the BM by development weeks 8-9, where it accumulates throughout the second trimester, to finally decline around birth. Based on phenotypic, molecular, and functional criteria, we demonstrate that CD34hiCD45RAhiCD7+ cells represent the direct precursors of the most immature CD34hiCD1a- fetal thymocytes that follow a similar dynamics pattern during fetal and early postnatal development.

Human immunodeficiency virus type 1 KK26-27 matrix mutants display impaired infectivity, circularization and integration but not nuclear import.

We analyzed the role of human immunodeficiency virus (HIV)-1 matrix protein (MA) during the virus replication afferent phase. Single-round infection of H9 T lymphocytes showed that the combined mutation of MA Lys residues 26-27 in MA reported nuclear localization signal (NLS)-1 impaired infectivity, abrogated 2-LTR-circle formation and significantly reduced integration. However, the mutation did not affect viral DNA docking to chromatin in either interphasic or mitotic cells, indicating that MA N-terminal basic domain should not represent a major determinant of HIV-1 nuclear import in T lymphocytes.

Cell cycle regulation of human immunodeficiency virus type 1 integration in T cells: antagonistic effects of nuclear envelope breakdown and chromatin condensation.

We examined the influence of mitosis on the kinetics of human immunodeficiency virus type 1 integration in T cells. Single-round infection of cells arrested in G1b or allowed to synchronously proceed through division showed that mitosis delays virus integration until 18-24 h postinfection, whereas integration reaches maximum levels by 15 h in G1b-arrested cells. Subcellular fractionation of metaphase-arrested cells indicated that, while nuclear envelope disassembly facilitates docking of viral DNA to chromatin, chromosome condensation directly antagonizes and therefore delays integration.

Persistent infection with primate foamy virus type 1 increases human immunodeficiency virus type 1 cell binding via a Bet-independent mechanism.

We report that human T cells persistently infected with primate foamy virus type 1 (PFV-1) display an increased capacity to bind human immunodeficiency virus type 1 (HIV-1), resulting in increased cell permissiveness to HIV-1 infection and enhanced cell-to-cell virus transmission. This phenomenon is independent of HIV-1 receptor, CD4, and it is not related to PFV-1 Bet protein expression. Increased virus attachment is specifically inhibited by heparin, indicating that it should be mediated by interactions with heparan sulfate glycosaminoglycans expressed on the target cells.

Molecular characterization of early human T/NK and B-lymphoid progenitor cells in umbilical cord blood.

The early stages of human lymphopoiesis are poorly characterized. Here, we compared the lymphoid potential of a novel umbilical cord blood CD34(+)CD45RA(hi)CD7(+) hematopoietic progenitor cell (HPC) population with that of CD34(+)CD45RA(hi)Lin(-)CD10(+) HPCs, previously proposed as candidate common lymphoid progenitors. Limiting-dilution and clonal analysis, fetal thymic organ cultures, and culture onto Notch ligand Delta-like-1-expressing OP9 cells, showed that although CD34(+)CD45RA(hi)CD7(+) HPCs could generate cells of the 3 lymphoid lineages, their potential was skewed toward the T/natural killer (T/NK) lineages.

Characterization of a novel hematopoietic marker expressed from early embryonic hematopoietic stem cells to adult mature lineages.

A novel membrane protein has been identified in the course of screening for differentially expressed cDNAs in human embryonic hematopoietic sites. This 37- to 38-kDa molecule, designated KLIP-1 (killer lineage protein), consisting of 350 amino acids and containing five transmembrane domains, is encoded by the 5093-bp KLIP-1 gene, composed of nine exons and located on chromosome 6 (6p21.1-6p21.

Dendritic cells: a complex simplicity.

Dendritic cells (DC) are essential antigen-presenting cells that initiate and regulate adaptive immune responses. There are distinct DC populations of diverse origins, which develop from hematopoietic progenitors already committed to the lymphoid or the myeloid lineages and, in the latter case, even from terminally differentiated macrophages. One may assume that DC of lymphoid origin are dedicated to the adaptive immune system, along which they have phylogenetically co-evolved, whereas myeloid DC would be more involved as an interface between the innate and adaptive immune systems.

The maturation of dendritic cells results in postintegration inhibition of HIV-1 replication.

Maturation of dendritic cells (DC) is known to result in decreased capacity to produce HIV due to postentry block of its replicative cycle. In this study, we compared the early phases of this cycle in immature DC (iDC) and mature DC (mDC) generated from monocytes cultured with GM-CSF and IL-4, trimeric CD40 ligand (DC(CD40LT)), or monocyte-conditioned medium (DC(MCM)) being added or not from day 5. Culture day 8 cells exposed to X4 HIV-1(LAI) or R5 HIV-1(Ba-L) were analyzed by semiquantitative R-U5 PCR, which detects total HIV DNA.

Identification of mature and immature human thymic dendritic cells that differentially express HLA-DR and interleukin-3 receptor in vivo.

We have previously shown that thymic CD34+ cells have a very limited myeloid differentiation capacity and differentiate in vitro mostly into CD1a+-derived but not CD14+-derived dendritic cells (DC). Herein we characterized the human neonatal thymic DC extracted from the organ in relationship with the DC generated from CD34+ cells in situ. We show that in vivo thymic DC express E cadherin, CLA, CD4, CD38, CD40, CD44, and granulocyte-macrophage colony-stimulating factor-R (GM-CSF-R; CD116) but no CD1a.

Characterization of dendritic cell differentiation pathways from cord blood CD34(+)CD7(+)CD45RA(+) hematopoietic progenitor cells.

To better characterize human dendritic cells (DCs) that originate from lymphoid progenitors, the authors examined the DC differentiation pathways from a novel CD7(+)CD45RA(+) progenitor population found among cord blood CD34(+) cells. Unlike CD7(-)CD45RA(+) and CD7(+)CD45RA(-) progenitors, this population displayed high natural killer (NK) cell differentiation capacity when cultured with stem cell factor (SCF), interleukin (IL)-2, IL-7, and IL-15, attesting to its lymphoid potential. In cultures with SCF, Flt3 ligand (FL), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor (TNF)-alpha (standard condition), CD7(+)CD45RA(+) progenitors expanded less (37- vs 155-fold) but yielded 2-fold higher CD1a(+) DC percentages than CD7(-)CD45RA(+) or CD7(+)CD45RA(-) progenitors.