Benjamin P. Chen

Hematopoietic progenitor cells of lymphocytes and dendritic cells.

Lymphocytes and dendritic cells (DCs) are critical for immune responses, yet how they develop from pluripotent hematopoietic stem cells is poorly defined. In humans and mice, it is possible to isolate phenotypically defined subsets of bone marrow (BM) cells that represent intermediate progenitors without long-term repopulating characteristics but with specific lineage differentiation properties. For instance, murine BM CD34+ CD45RA+ cells are progenitors for B and T lymphocytes with no in vivo repopulation activity. In human BM, a small subset (5%) of cells having the phenotype CD34+ Lin- CD10+ CD45RA+ CD38+ Thy-1- c-kit- represents a new class of hematopoietic progenitor cells that gives rise to lymphocytes [T, B, and natural killer (NK) cells] and to DCs but does not produce myeloid or erythroid cells. The identification of such progenitor cells provides the opportunity to define the differentiation and growth requirements for the production of lymphocytes and DCs. Genes involved in lineage specification can also be studied. Altogether, these studies have fundamental implications for understanding the biology of pivotal lineages of immune cells. This understanding could be used to treat a variety of immunodeficiencies and to design novel immunotherapies particularly in the context of hematopoietic cell transplantation.

Delineation of the human hematolymphoid system : potential applications of defined cell populations in cellular therapy

Summary: Hematopoietic stem cells (HSC) have the capacity to reconstitute ail the blood cells in the body HSC are rare, representing on average 0.0 5% of the mononuclear cells present in healthy human bone marrow. Due to their capacity for self–renewal and their pluripotent, long–term reconstituting potential. HSC are considered ideal for transplantation to reconstitute the hematopoietic system after treatment for various hematologic disorders or as a target for the delivery of therapeutic genes. Human HSC also have potential applications in restoring the immune system in autoimmune diseases and in the induction of tolerance for allogeneic solid organ transplantation. With the increased interest in human HSC for clinical applications, technology for the isolation of candidate HSC and knowledge of human hematopoiesis have been growing rapidly. In this article, we discuss the functional characterization of a human CD34+ Thy‐1+ HSC population which is essentially free of residual disease, our efforts to generate alternate monoclonal antibodies for the isolation of clinically useful stem or progenitor cell populations, and the identification of a novel lymphoid progenitor as part of an exploration towards defining progenitors with potential application as adjuncts to HSC–based cellular therapy.

LONG-TERM MULTILINEAGE DEVELOPMENT FROM HUMAN UMBILICAL CORD BLOOD STEM CELLS IN A NOVEL SCID-HU GRAFT

Although human hematopoietic stem cells can be isolated based on cell surface antigen expression, their proliferative and developmental capacity must still be defined by their ability to produce multiple hematopoietic cell lineages for prolonged times in vivo. In order to derive a long-term in vivo multilineage SCID-hu graft we transplanted a human fetal bone and spleen adjacent to an HLA class I mismatched fetal thymus fragment in immunodeficient SCID mice (SCID-hu BTS). Grafts were analyzed at various times post transplant for cells expressing specific lineage markers. The bone marrow of SCID-hu BTS grafts maintained B cells and myeloid cells for at least 36 weeks post transplant. Analysis for progenitor content within grafts revealed that CD34+ cells, CFU-G/M, and CFU-GEMM were maintained in 94%(109/116), 100%(66/66) and 79%(52/66) up to 28 weeks. HSC (CD34hiThy-l+Lin-) sorted from 20 week old SCID-hu BTS grafts demonstrated potent secondary multilineage reconstituting potential when injected into HLA mismatched SCID-hu bone grafts. In addition both immature and mature T-cells, derived from progenitors within the fetal bone, were found in 87%(101/116) of grafts analyzed up to 36 weeks in vivo. Injection of irradiated SCID-hu BTS grafts with CD34+Thy-1+Lin-umbilical cord blood cells produced B-cells, myeloid cells, T-cells and CD34+ cells in individual grafts when analyzed 8 weeks post reconstitution, further demonstrating the multipotential nature of these HSC populations. This model is currently being used to define soluble factors or cells that are capable of enhancing human HSC engraftment.

Detection of Human Hematopoietic Stem Cells in SCID-hu Mice

Totipotential hematopoietic stem cells (HSC) have an extremely high proliferative potential, therefore they can differentiate into all hematopoietic lineages, generating a high number of hematopoietic progenitors and providing a long-lasting supply of blood-borne cells.1–3 Thus, a number of in vitro assays identify HSC on the basis of their capacity to derive multiple hematopoietic lineages and of their high proliferative potential in culture (reviewed in ref. 4). It is however commonly accepted that in vitro assays do not accurately reflect the global and physiological development of hematopoiesis and thus are not ideal for preclinical testing. Research on murine HSC has strongly benefited from in vivo experimentation which is not possible with human HSC for ethical reasons. To overcome these limitations, several xenogeneic models have been designed to support the in vivo development of human hematopoietic cells in surrogate animals.