Phenotype and function of human hematopoietic cells engrafting immune- deficient CB17-severe combined immunodeficiency mice and nonobese diabetic-severe combined immunodeficiency mice after transplantation of human cord blood mononuclear cells

Abstract

In an attempt to understand better the regulation of stem cell function in chimeric immunodeficient mice transplanted with human cells, and the filiation between progenitor cells identified in vitro and in vivo, we assessed the different compartments of hematopoietic progenitors found in the marrow of CB17-severe combined immunodeficiency (SCID) mice (34 mice, 9 experiments) after intravenous injection of 2 to 3 x 10(7) cord blood mononuclear cells. On average 6.3 +/-4 x 10(5) human cells were detected per four long bones 4 to 6 weeks after the transplant predominantly represented by granulomonocytic (CD11b+) and B lymphoid (CD19+) cells. Twenty five percent of these human cells expressed the CD34 antigen, of which 90% coexpressed the CD38 antigen and 50% the CD19 antigen. Functional assessment of progenitor cells (both clonogenic and long-term culture-initiating cells [LTC-IC]) was performed after human CD34+ cells and CD34+/CD38- cells have been sorted from chimeric CB17-SCID marrow 3 to 10 weeks after intravenous (IV) injection of human cells. The frequency of both colony-forming cells and LTC-IC was low (4% and 0.4%, respectively in the CD34+ fraction) when compared with the frequencies of cells with similar function in CD34+ cells from the starting cord blood mononuclear cells (26% +/- 7% and 7.2% +/- 5%, respectively). More surprisingly, the frequency of LTC-IC was also low in the human CD34+ CD38- fraction sorted from chimeric mice. This observation might be partly accounted for by the expansion of the CD34+ CD19+ B-cell precursor compartment. Despite their decreased frequency and absolute numbers, the differentiation capability of these LTC-IC, assessed by their clonogenic progeny output after 5 weeks in coculture with murine stromal cells was intact when compared with that of input LTC-IC. Furthermore the ratio between clonogenic progenitor cells and LTC-IC was similar in severe combined immunodeficiency (SCID) mice studied 4 weeks after transplant and in adult marrow or cord blood suspensions. Results generated in experiments where nonobese diabetic (NOD)-SCID mice were used as recipients indicate a higher level of engraftment but no change in the distribution of clonogenic cells or LTC-IC. These results suggest that the hierarchy of hematopoietic differentiation classically defined in human hematopoietic tissues can be reconstituted in immunodeficient SCID or NOD-SCID mice.

Cobblestone Area‐Forming Cells in Human Cord Blood Are Heterogeneous and Differ from Long‐Term Culture‐Initiating Cells

“…Our estimates of the proportion of LTC-IC in CB using the LTC-IC readout suggest that, on average, 32/10 5 MNC or 43/10 3 CD34 + cells in CB are LTC-IC. These compare favorably with reported frequencies of 59/10 5 MNCs [27], 23/10 5 MNCs [28], and 71/10 3 CB CD34 + cells [29] using the LTC-IC readout after culture on normal bone marrow stroma feeder layers. The CAFC readout, although popular, seems to be fraught with problems.…”

Sustained proliferation, multi‐lineage differentiation and maintenance of primitive human haemopoietic cells in NOD/SCID mice transplanted with human cord blood

“…The discovery a few years ago that intravenously injected normal human haemopoietic cells can home into the marrow space of sublethally irradiated immunodeficient mice and remain viable there for many months opened a new era of investigation of the in vivo reconstituting properties of various types of human transplants (Dick, 1996). Recent studies using NOD/SCID mice as recipients (Larochelle et al, 1995(Larochelle et al, , 1996Pflumio et al, 1996;Lowry et al, 1996;Cashman et al, 1997) indicate the superiority of this latter model and suggest that it will be useful for addressing many outstanding questions about human haemopoietic stem cells with in vivo repopulating activity. One issue of current interest is the stem cell content and in vivo repopulating potential of human cord blood cells by comparison to adult marrow cells.…”

“…bone or fetal liver) previously implanted into mice with the severe combined immunodeficiency (SCID) mutation (the so-called SCID-hu mouse model) (McCune et al, 1988;Kyoizumi et al, 1992;Fraser et al, 1995), and by simple intravenous injection of various types of myeloablated immunodeficient mice (e.g. SCID mice, BEIGE-NUDE-Xid mice [bnx mice], and nonobese-diabetic-SCID [NOD/SCID] mice) (Kamel-Reid & Dick, 1988;Lapidot et al, 1992;Nolta et al, 1994;Lowry et al, 1996;Pflumio et al, 1996;Larochelle et al, 1996;Cashman et al, 1997). These latter studies have shown that conditions prevailing within the marrow microenvironment of several species, including mice, are able to sustain many aspects of human haemopoiesis in spite of the known species-restricted activity of some of the molecules used to stimulate these cells in vitro.…”

“…These latter studies have shown that conditions prevailing within the marrow microenvironment of several species, including mice, are able to sustain many aspects of human haemopoiesis in spite of the known species-restricted activity of some of the molecules used to stimulate these cells in vitro. Although each of the xenogeneic transplant models described offers certain advantages, the NOD/SCID model has general appeal for studies of human stem cells because it can support both human B-lymphopoiesis and human myelopoiesis at higher levels than any other murine host thus far examined (Pflumio et al, 1996;Cashman et al, 1997). In addition, this model is readily applicable to largescale quantitative studies.…”

SCID Mouse Models of Human Stem Cell Engraftment

“…The initial report on the increased engraftment of human stem cells in NOD-scid mice was rapidly confirmed in a number of laboratories [105][106][107]. The enhanced ability of NODscid mice to support human stem cell engraftment was further demonstrated by comparing the levels of human cell engraftment to those observed in BALB/c-scid mice expressing transgenes for SCF, IL-3, and GM-CSF [77].…”