A monoclonal antibody, A7R34, that recognizes the high-affinity interleukin 7 receptor (IL-7Ra) and blocks the binding between IL-7 and IL-7Ra has been produced. Cell surface staining with A7R34 demonstrated that IL-7Ra is expressed in both B-and T-cell lineages. In the bone marrow, immature B-lineage cells that do not express surface IgM were IL-7Ra+. In the thymus, IL-7Ra was detected in CD4-8-T cells and also in CD4 or CD8 single-positive cells but not in CD4+8+ double-positive cells. In the peripheral lymphoid tissues, both CD4 and CD8 single-positive cells were the manor cell types that express IL-7Ra. Addition of A7R34 to a long-term B-precursor-cell culture inhibited proliferation of the B-lineage cells, indicating that IL-7 is an absolute requirement for in vitro B-cell genesis. Consistent with this in vitro result, continuous nijection of A7R34 into an adult mouse resulted in a decrease of B-precursor cells and also of thymocytes, whereas a considerable fraction of mature B and T cells in the peripheral tissues persisted over 2 weeks of the experiment. When A7R34 iniection is started from day 14 of gestation, it is possible to produce mice that lack B cells. These results indicate that IL-7 is an essential molecule for generation of both B and T cells in murine bone marrow and thymus, respectively. Moreover, IL-7Ra would be the sole receptor system regulating these processes.B-cell genesis in the adult mouse is regulated by a meshwork of stromal-cell components in the bone marrow (1, 2). Establishment of the stromal-cell lines that can support B-cell genesis from multipotent hematopoietic stem cells facilitated identification of the molecules that are required for this process (3-6). Among a number of molecules that are expressed in the stromal-cell lines, interleukin 7 (IL-7) is the first molecule that has been shown to be able to induce proliferation of B-cell precursors (7,8). Initial use of recombinant IL-7 demonstrated that IL-7 can induce proliferation of pre-B cells (8)(9)(10). Subsequent studies further demonstrated that IL-7, when used in combination with the stromal cell lines or the ligand for c-kit, can act on earlier stages of B precursors (11)(12)(13)(14)(15). Besides the proliferative activity on B-precursor cells, it was reported that IL-7 induces proliferation of both mature and immature mouse T cells (16)(17)(18)(19)(20)(21)(22). In fact, studies of mice that received continuous IL-7 injections or bore the IL-7 transgene indicate that elevation of IL-7 results in enormous expansion of both T and B cells (23)(24)(25)(26). Despite all such positive evidence suggesting the role of IL-7 in lymphopoiesis, whether IL-7 is a functional requirement for in vivo lymphocyte production is yet to be determined. To address this question, we have produced an antagonistic monoclonal antibody (mAb) to the high-affinity IL-7 receptor (IL-7Ra) (27) and have investigated its expression and function. Our study demonstrates that IL-7Ra is The publication costs of this article were defrayed in part b...

The gamma chain of the interleukin-2 (IL-2) receptor is shared with the functional IL-4 receptor and is causatively related to X-linked severe combined immunodeficiency (XSCID), which is ascribed to a profound T cell defect. Studies with monoclonal antibodies specific for the IL-2 receptor gamma chain showed that the gamma chain participates in the functional high-affinity receptor complexes for IL-7 that are involved in the differentiation of T and B cells. Participation of the gamma subunit in more than one receptor may enable the elucidation of the mechanisms of XSCID development and lymphocyte differentiation.

Interactions between the cell cycle machinery and transcription factors play a central role in coordinating terminal differentiation and proliferation arrest. We here show that cyclin-dependent kinase 6 (Cdk6) is specifically expressed in proliferating hematopoietic progenitor cells, and that Cdk6 inhibits transcriptional activation by Runx1, but not C/EBPalpha or PU.1. Cdk6 inhibits Runx1 activity by binding to the runt domain of Runx1, interfering with Runx1 DNA binding and Runx1-C/EBPalpha interaction. Cdk6 expression increased myeloid progenitor proliferation, and inhibited myeloid lineage-specific gene expression and terminal differentiation in vitro and in vivo. These effects of Cdk6 did not require Cdk6 kinase activity. Cdk6-mediated inhibition of granulocytic differentiation could be reversed by excess Runx1, consistent with Runx1 being the major target for Cdk6. We propose that Cdk6 downregulation in myeloid progenitors releases Runx1 from Cdk6 inhibition, thereby allowing terminal differentiation. Since Runx transcription factors play central roles in hematopoietic, neuronal and osteogenic lineages, this novel, noncanonical Cdk6 function may control terminal differentiation in multiple tissues and cell types.

Primitive macrophages emerged in the sinusoidal lumen of the fetal mouse liver at 10 days of gestation before the initiation of hepatic hematopoiesis and matured into fetal macrophages. In the culture of cell suspensions from the fetal liver with LP3-conditioned medium, monocyte colonies were formed, but monocytopoiesis was poor in the early stage of hepatic hematopoiesis in vivo. In the culture of cell suspensions obtained from the fetal liver at 10 days of gestation on the monolayer of a mouse bone marrow stromal cell line, ST2, primitive/fetal macrophage colonies were formed before the development of monocyte/macrophage colonies and showed differentiation of primitive macrophages into fetal macrophages without passing through the stage of promonocytes and monocytes. At this time, the fetal cardiovascular system was connected with the vitelline vein just before the formation of the liver. With the progress of gestation, a monocytic cell series was observed to develop and form a monocyte/macrophage population. This was confirmed by in vitro studies with an LP3-conditioned medium and on a monolayer of ST2. Thus, it appears that there exist two different macrophage populations, a primitive/fetal macrophage population and a monocyte/macrophage population in hepatic hematopoiesis. It also appears that fetal macrophages are differentiated from primitive macrophages which are colonized into the fetal liver from the yolk sac or which develop in loco, presumably from hematopoietic stem cells.

The reeler gene (Reln(rl), formerly rl) product Reelin controls neuronal migration and positioning and thereby plays a key role in brain development. Mutation of Reln leads to widespread disruption of laminar cortical regions and ectopia in some brainstem nuclei. In the embryonic striatum of normal mice, a substantial expression of reelin mRNA has been documented; however, the anomalous positioning of neurons in the basal ganglia of reeler mice remains to be studied. We provide first evidence for a potential role of Reelin in the developmental formation of the substantia nigra. In reeler mutant mice lacking Reelin, dopaminergic neurons destined for the substantia nigra fail to migrate laterally and become anomalously clustered just lateral to the ventral tegmental area. Their axons appear to project to striatal patches forming "dopamine islands." Results from the normal mice show that, at the midembryonic stage, Reelin identified with CR-50 is highly concentrated in the ventral mesencephalon, where nigral dopaminergic neurons are in progress to migrate laterally to their eventual position of the adult brain. A combination of CR-50 labeling and anterograde axonal tracing provided evidence that embryonic striatal neurons may supply the ventral portion of the mesencephalon with Reelin through their axonal projections. We hypothesize that Reelin plays a role in the positioning of nigral dopaminergic neurons and that it can act as an environmental cue at a remote site far from its birthplace via a transaxonal delivery system.