Bruno Canque

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

ABSTRACT 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. In this study, we provide several lines of evidence suggesting that intracellular storage of intact virions does not contribute to HIV transmission. We show that endocytosis-defective DC-SIGN molecules enhance T-cell infection as efficiently as their wild-type counterparts, indicating that DC-SIGN-mediated HIV internalization is dispensable for trans-enhancement. Furthermore, using immature DCs that are genetically resistant to infection, we demonstrate that several days after viral uptake, HIV transfer from DCs to T cells requires viral fusion and occurs exclusively through DC infection and transmission of newly synthesized viral particles. Importantly, our results suggest that DC-SIGN participates in this process by cooperating with the HIV entry receptors to facilitate cis-infection of immature DCs and subsequent viral transfer to T cells. We suggest that such a mechanism, rather than intracellular storage of incoming virus, accounts for the long-term transfer of HIV to CD4+ T cells and may contribute to the spread of infection by DCs.

IL-4 and CD40 ligation affect differently the differentiation, maturation, and function of human CD34+ cell-derived CD1a+CD14- and CD1a-CD14+ dendritic cell precursors in vitro.

We examined the effect of interleukin (IL)‐4 or CD40 ligation on the differentiation and maturation of CD1a+CD14– and CD1a–CD14+ dendritic cell (DC) precursors. Cord blood CD34+ cells were cultured with granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) and tumor necrosis factor α (TNF‐α), to which stem cell factor and Flt‐3 ligand were added for 5 days. Phenotypic analysis of DC precursors on culture day 7 showed that CD1a+CD14– cells expressed higher CD11c and CD80 levels and lower CD116/GM‐CSFR and CCR‐5 levels than their CD1a–CD14+ counterparts. Culturing CD1a+CD14– precursors with GM‐CSF and TNF‐α resulted in DC with heterogeneous CD1a, HLA;SMDR (DR), CD11b, and CD83 expression, 10% of which acquired CD14. IL‐4 and CD40 ligation affected their differentiation in contrasting ways: IL‐4 induced CD1ahiCD14– DRloCD11b+CD83–S100+ DC with reduced MLR‐stimulating capacity, whereas CD40 ligation led to CD1alo/‐CD14–CD40–DRhiCD11b–CD83+S100+/– DC with stronger MLR‐stimulating capacity. Also, both IL‐4 and CD40 ligation promoted RelB expression and nuclear translocation. When CD1a–CD14+ precursors were maintained in only the presence of GM‐CSF and TNF‐α, this led to mixed populations of adherent macrophages and nonadherent CD1a–CD14+ monocytes, and of CD1a+CD14– and CD1a+CD14+ DC, which were DRloCD11b+‐CD83–S100–. IL‐4 or CD40 ligation prevented their differentiation into macrophages and resulted in DC with phenotypes close to those issued from CD1a+CD14– precursors, with only a minority staying CD14+ but most being S100–; their MLR‐stimulating capacity also increased but remained lower than that of DC differentiated from CD1a+CD14– precursors. Thus, IL‐4 or CD40 ligation induced CD1a+CD14– and CD1a–CD14+ DC precursors to differentiate into phenotypically close but functionally different DC populations, suggesting that DC function is primarily determined by their origin. The heterogeneity of DC should then be related to different developmental pathways and to different stages of maturation/activation. J. Leukoc. Biol. 64: 235–244; 1998.

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 differentiatein 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. According to their morphology, functions, and surface staining they could be separated into two distinct subpopulations: mature HLA‐DRhi, mostly interleukin‐3‐R (CD123)‐negative cells, associated with thymocytes, some apoptotic, and expressed myeloid and activation markers but no lymphoid markers. In contrast, immature HLA‐DR+ CD123hi CD36+ cells with monocytoid morphology lacked activation and myeloid antigens but expressed lymphoid antigens. The latter express pTα mRNA, which is also found in CD34+ thymocytes and in blood CD123hi DC further linking this subset to lymphoid DC. However, the DC generated from CD34+ thymic progenitors under standard conditions were pTα‐negative. Thymic lymphoid DC showed similar phenotype and cytokine production profile as blood/tonsillar lymphoid DC but responded to GM‐CSF, and at variance with them produced no or little type I interferon upon infection with viruses and did not induce a strict polarization of naive T cells into TH2 cells. Their function in the thymus remains therefore to be elucidated.

Persistent Infection with Primate Foamy Virus Type 1 Increases Human Immunodeficiency Virus Type 1 Cell Binding via a Bet-Independent Mechanism

ABSTRACT 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. Given that both viruses infect similar animal species, the issue of whether coinfection with primate foamy viruses interferes with the natural course of lentivirus infections in nonhuman primates should be considered.

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. However, any DC can ultimately present antigens in either an immunogenic or tolerogenic manner according to whether they are more or less or not at all activated towards maturation, depending on the condition under which they encountered antigen. Hence, DC either induce the appropriate immune response to pathogens or prevent autoimmune reactivity. Thus, besides default programming, which should be necessary to face the challenges of their usual setting, each type of DC can also display functions that are similar, in an instructive mode, to elicit immune responses deemed necessary for unexpected stimuli. In such a system, DC provide enough flexibility and sufficient redundancy to ensure that an essential function of the immune system, i.e., passing information from its innate to adaptive arms and affecting the latter’s responses, occurs under optimal conditions. Working on the basis of such a unified theory of DC diversity should be useful for learning to adequately manipulate the immune system for the development of cellular immunotherapy.

Special Susceptibility to Apoptosis of CD1a+ Dendritic Cell Precursors Differentiating from Cord Blood CD34+ Progenitors

We analyzed the effect of tumor necrosis factor (TNF)‐α on the differentiation and viability of dendritic cells (DC) generated from cord blood CD34+ progenitors cultured for five days with GM‐CSF, Flt‐3 ligand (FL), and stem cell factor (SCF), and then with GM‐CSF only [TNF(−) cultures]. Adding TNF‐α from the start [TNF(+) cultures] potentiated progenitor cell proliferation and promoted early differentiation of CD1a+ DC precursors without affecting differentiation of CD14+ cells, which comprise bipotent precursors of DC and macrophages, nor of CD15+ granulocytic cells. Use of TNF‐α was associated with increased cell mortality, which peaked on culture day 10 and mainly involved CD1a+ DC. Selective apoptosis of CD1a+ DC precursors was confirmed by showing that survival of day‐7‐sorted CD1a+CD14− cells from TNF(+) cultures was lower than that of CD1a−CD14+ cells. That similar findings were noted for sorted CD1a+CD14− cells of TNF(−) cultures, further cultured with GM‐CSF without or with TNF‐α, indicates that apoptosis of CD1a+ DC precursors was not induced by TNF‐α. Apoptosis of CD1a+ DC precursors occurred after the cells had lost the capacity to incorporate bromodeoxyuridin. Finally, using higher GM‐CSF concentrations or adding interleukin 3 (IL‐3) improved viability of CD1a+ cells. Other cytokines, such as IL‐4 and transforming growth factor (TGF)‐β1, were ineffective in this respect, though they promoted differentiation of CD1a+ DC. These results indicate that TNF‐α promotes the differentiation of CD1a+ DC precursors, which display a high susceptibility to apoptosis that can be prevented by high concentrations of GM‐CSF or use of IL‐3, without affecting the differentiation of the CD14+ DC precursors.

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

&NA; 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. Whereas CD127− ELPs comprised precursors of T cells, marginal zone B cells, and natural killer (NK) and innate lymphoid cells (ILCs), CD127+ ELPs supported production of all NK cell, ILC, and B cell populations but lacked T potential. On the basis of these results, we propose a “two‐family” model of human lymphoid development that differs from the prevailing model of hematopoiesis. Graphical Abstract Figure. No caption available. Highlights:Human lymphopoiesis displays a bipartite architectureCD127− and CD127+ ELPs differentiate independently from LMDPsCD127− ELPs generate T cells, marginal zone B cells, NK cells, and ILCsCD127+ ELPs generate marginal zone and follicular B cells, NK cells, and ILCs &NA; The development pathways of human lymphoid cells remain unclear. Alhaj Hussen et al. show that human lymphoid cells stem from two functionally specialized populations of CD127− and CD127+ early lymphoid progenitors. They propose a “two‐family” model of human lymphopoiesis that differs from the standard model of hematopoiesis established in mice.

Developmental Biology of Mammalian T-Cell Progenitors: From Early Lymphoid Progenitors to Thymus-Colonizing Cells

This review gives a general overview of the recent advances in the field of fetal hematopoiesis and T-cell development. Although it is now well recognized that early lymphoid progenitors first emerge in the fetal liver (FL) where active B lymphopoiesis take places, the identity of early T-cell precursors, as well as the mechanisms of thymus colonization, has long been controversial. Here we discuss the experimental evidence supporting the concept of prothymocyte in both human and mouse species, as well as its major implications regarding lineage relationships among the immune system and general hematopoietic organization.

Les cellules dendritiques: un système cellulaire complexe

Dendritic cells (DC) are the most potent antigen-presenting cells. Thus, ex vivo antigen-pulsed DC are a potentially powerful tool to induce in vivo immunity against tumor-associated or viral antigens. Therefore, culture methods to generate high numbers of DC from bone marrow or blood CD34+ hematopoietic progenitor cells have recently been developed. These methods, which use different combinations of growth factor--mainly granulocyte/macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor (TNF)-alpha and interleukin (IL)-4--make the characterization of DC obtained from CD34+ cells of different origins easier and allow to assess whether DC relate to a unique or distinct differentiation pathways. Monocytes and even macrophages can also directly differentiate into DC in the presence of GM-CSF and IL-4. This has to be reconciled with evidence supporting earlier branching off of the macrophage and DC lineages, and raises questions as to the identity of the latter lineage. Apart from DC of myeloid origin, DC may also originate from lymphoid progenitors. Because the capacity of DC to capture, process and present antigens is known to vary according to their differentiation stage, and lymphoid DC might behave differently from lymphoid DC in this respect, the definition of which type of DC to use for immunotherapy must be more precise, in order to avoid detrimental side effects or results. From a practical point of view, it is also necessary to define the most appropriate cytokine combinations and schedules thereof to optimize proliferation and differentiation of DC from different origins. These conditions should then be applied to generated DC for their efficient and safe use for clinical immunotherapy.

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. Consistently, lentivirus-mediated overexpression of TMEM131L in human CD34+ hematopoietic progenitor cells leads to decreased susceptibility to Notch1 ligation and defective commitment toward the T lineage. These results show that TMEM131L corresponds to an evolutionary conserved regulator of the Notch signaling pathway.