Abstract-The concept that bone marrow (BM)-derived cells participate in cardiac regeneration remains highly controversial and the identity of the specific cell type(s) involved remains unknown. In this study, we report that the postnatal BM contains a mobile pool of cells that express early cardiac lineage markers (Nkx2.5/Csx, GATA-4, and MEF2C). These cells are present in significant amounts in BM harvested from young mice but their abundance decreases with age; in addition, the responsiveness of these cells to gradients of motomorphogens SDF-1, HGF, and LIF changes with age. FACS analysis, combined with analysis of early cardiac markers at the mRNA and protein levels, revealed that cells expressing these markers reside in the nonadherent, nonhematopoietic CXCR4 ϩ /Sca-1 ϩ /lin Ϫ /CD45 Ϫ mononuclear cell (MNC) fraction in mice and in the CXCR4 ϩ /CD34 ϩ /AC133 ϩ /CD45 Ϫ BMMNC fraction in humans. These cells are mobilized into the peripheral blood after myocardial infarction and chemoattracted to the infarcted myocardium in an SDF-1-CXCR4 -, HGF-c-Met-, and LIF-LIF-R-dependent manner. To our knowledge, this is the first demonstration that the postnatal BM harbors a nonhematopoietic population of cells that express markers for cardiac differentiation. We propose that these potential cardiac progenitors may account for the myocardial regenerative effects of BM. The present findings provide a novel paradigm that could reconcile current controversies and a rationale for investigating the use of BM-derived cardiac progenitors for myocardial regeneration. Key Words: stem cells Ⅲ bone marrow Ⅲ myocardial infarction Ⅲ myocardial regeneration Ⅲ CXCR4-SDF-1 axis S everal recent reports suggest that bone marrow (BM)-derived hematopoietic stem cells (HSCs), 1,2 mesenchymal stem cells (MSCs), 3,4 and mononuclear cells (MNCs) 5,6 exhibit a high degree of differentiation plasticity and thus could potentially be used to regenerate infarcted myocardium in humans. These reports, however, have been challenged vigorously and the concept of transdifferentiation of adult HSCs has recently been called into question. 7,8 Furthermore, the identity of the specific BM-derived cellular subset(s) capable of regenerating cardiac tissue remains elusive.These uncertainties are part of an ongoing intense debate regarding the mechanism responsible for the functional benefits observed after cellular transplantation. Two basic possibilities have been proposed: cell fusion 9,10 and transdifferentiation. In this report, we propose a third possibility, which may reconcile the apparently discordant results obtained by the proponents of cell fusion and transdifferentiation. We have previously demonstrated that the adult murine BM harbors CXCR4 ϩ tissuecommitted stem cells (TCSCs) for various tissues, including skeletal muscle, liver, and neural tissue [11][12][13] and have proposed that these TCSCs contribute to the organ/tissue chimerism observed after transplantation of BM-or mobilized peripheral blood (PB)-derived cells in earlier reports. In this...

Bone marrow transplantation offers great promise for treating a number of disease states. However, the widespread application of this approach is dependent upon the development of less toxic methods to establish chimerism and avoid graft-versus-host disease (GVHD). CD8+/TCR− facilitating cells (FCs) have been shown to enhance engraftment of hematopoietic stem cells (HSCs) in allogeneic recipients without causing GVHD. In the present studies, we have identified the main subpopulation of FCs as plasmacytoid precursor dendritic cells (p-preDCs). FCs and p-preDCs share many phenotypic, morphological, and functional features: both produce IFN-α and TNF-α, both are activated by toll-like receptor (TLR)-9 ligand (CpG ODN) stimulation, and both expand and mature after Flt3 ligand (FL) treatment. FL-mobilized FCs, most of which express a preDC phenotype, significantly enhance engraftment of HSCs and induce donor-specific tolerance to skin allografts. However, p-preDCs alone or p-preDCs from the FC population facilitate HSC engraftment less efficiently than total FCs. Moreover, FCs depleted of preDCs completely fail to facilitate HSC engraftment. These results are the first to define a direct functional role for p-preDCs in HSC engraftment, and also suggest that p-preDCs need to be in a certain state of maturation/activation to be fully functional.

The use of tolerogenic cells as an approach to induce tolerance to solid organ allografts is being aggressively pursued. A major limitation to the clinical application of cell-based therapies has been the ability to obtain sufficient numbers and also preserve their tolerogenic state. We previously reported that small numbers of bone marrow-derived CD8+/TCR− graft facilitating cells (FC) significantly enhance hemopoietic stem cell (HSC) engraftment in allogeneic and syngeneic recipients. Although the majority of FC resemble precursor plasmacytoid dendritic cells (p-preDC), p-preDC do not replace FC in facilitating function. In the present studies, we investigated the mechanism of FC function. We show for the first time that FC significantly enhance HSC clonogenicity, increase the proportion of multipotent progenitors, and prevent apoptosis of HSC. These effects require direct cell:cell contact between FC and HSC. Separation of FC from HSC by transwell membranes completely abrogates the FC effect on HSC. p-preDC FC do not replace FC total in these effects on HSC function. FC produce TNF-α, and FC from TNF-α-deficient mice exhibit impaired facilitation in vivo and loss of the in vitro effects on HSC. Neutralizing TNF-α in FC similarly blocks the FC effect. The antiapoptotic effect of FC is associated with up-regulation of Bcl-3 transcripts in HSC and blocking of TNF-α is associated with abrogation of up-regulation of Bcl-3 transcripts. These data demonstrate a critical role for TNF-α in mediating FC function. FC may have a significant impact upon the safe use of chimerism to establish tolerance to transplanted organs and tissue.

The mechanism by which mixed chimerism reverses autoimmunity in type 1 diabetes has not been defined. NOD mice have a well-characterized defect in the production of myeloid progenitors that is believed to contribute significantly to the autoimmune process. We therefore investigated whether chimerism induces a correction of this defect.

The events that regulate engraftment and long-term repopulating ability of hematopoietic stem cells (HSCs) after transplantation are not well defined. We report for the first time that major histocompatibility complex ( IntroductionHematopoietic stem cells (HSCs) are responsible for the continuous generation of lineage-committed progenitor cells, which in turn give rise to mature blood cells. Successful bone marrow transplantation requires that HSCs with the ability to self-renew survive in the transplant recipient. After bone marrow transplantation, some HSCs retain self-renewal, while others retain only short-term repopulating potential and terminally differentiate to produce progeny. The mechanism controlling HSC fate has not been defined but is clearly influenced by the hematopoietic microenvironment. In the present studies, we have evaluated the influence of the major histocompatibility complex (MHC) on HSC engraftment.Transplantation of purified HSCs across MHC barriers encounters greater resistance than whole bone marrow cells. 1,2 If the HSC donor and recipient are MHC matched, irrespective of minor antigen matching, long-term engraftment of HSCs occurs reliably. 1,3 In contrast, if donor and recipient are MHC disparate, purified HSCs engraft less readily and much higher cell doses are required. In addition, often only short-term radioprotection is observed, even when syngeneic marrow is coadministered concomitantly with the allogeneic HSCs. 3 This graft failure has been attributed by some to natural killer (NK)-mediated rejection. [4][5][6] However, NK cells do not directly reject purified HSCs,7,8 and the fact that a similar outcome is observed in recipients deficient in perforin and granzyme B, as well as those deficient in Fas and Fas ligand, strongly suggests that cytotoxic mechanisms are not responsible for failure of durable HSC engraftment. [9][10][11] The kinetic for graft failure of MHC-disparate purified HSCs also differs significantly from the relatively rapid NK-mediated rejection observed in bone marrow transplants from class I-deficient donors. 4,12 NK cell activity is regulated by MHC molecules through activating or inhibiting receptors. 1,2 In the mouse, 2 types of receptors have been identified: the Ly49 and the CD94/NKG2 families, 13 which are both expressed on the same cell. As a result, NK cells can be cytolytic under certain conditions and inhibitory under others.We have previously shown that CD8 ϩ /T-cell receptor-negative (TCR Ϫ ) facilitating cells (FCs) enhance engraftment of purified HSCs in allogeneic recipients. 1 To function, FCs must be MHC matched with the HSCs. In the present study, we therefore evaluated the role of MHC class I and class II molecules in the engraftment of purified HSCs in allogeneic recipients. We demonstrate for the first time that matching between recipient and donor HSCs at class I K is critical to durable HSC engraftment. Strikingly, mice deficient in production of NK cells or depleted of NK cells in vivo engraft MHC class I K disparate HSCs, implic...