Putative myogenic and endothelial (myo-endothelial) cell progenitors were identified in the interstitial spaces of murine skeletal muscle by immunohistochemistry and immunoelectron microscopy using CD34 antigen. Enzymatically isolated cells were characterized by fluorescence-activated cell sorting on the basis of cell surface antigen expression, and were sorted as a CD34+ and CD45− fraction. Cells in this fraction were ∼94% positive for Sca-1, and mostly negative (<3% positive) for CD14, 31, 49, 144, c-kit, and FLK-1. The CD34+/45− cells formed colonies in clonal cell cultures and colony-forming units displayed the potential to differentiate into adipocytes, endothelial, and myogenic cells. The CD34+/45− cells fully differentiated into vascular endothelial cells and skeletal muscle fibers in vivo after transplantation. Immediately after sorting, CD34+/45− cells expressed only c-met mRNA, and did not express any other myogenic cell-related markers such as MyoD, myf-5, myf-6, myogenin, M-cadherin, Pax-3, and Pax-7. However, after 3 d of culture, these cells expressed mRNA for all myogenic markers. CD34+/45− cells were distinct from satellite cells, as they expressed Bcrp1/ABCG2 gene mRNA (Zhou et al., 2001). These findings suggest that myo-endothelial progenitors reside in the interstitial spaces of mammalian skeletal muscles, and that they can potentially contribute to postnatal skeletal muscle growth.

IntroductionMaintenance of the hematopoietic system requires continual replenishment of mature blood cells from a rare population of bone marrow residing hematopoietic stem cells (HSCs). The alteration of the homeostatic control of hematopoiesis is considered to be a major culprit of drastic increase in pathologic incidences, such as bone marrow failure, anemia, and myeloid leukemia during aging. 1 However, the underlying mechanisms of pathogenesis of hematologic malignancy in elderly population remain poorly understood.Mounting evidence supports the idea that the accumulation of somatic DNA damage is a main cause of aging in multicellular organisms. [2][3][4][5] Mice with mutations in various DNA repair genes exhibit accelerated aging in the hematopoietic system because of the premature exhaustion of HSCs, indicating that DNA repair is crucial for the maintenance of HSC self-renewal and hematopoietic function. 6,7 DNA damage can directly result from genotoxic treatment such as ionizing radiation (IR), or may simply occur as a consequence of genome duplication infidelity or of genotoxic effects of reactive oxygen species (ROS). ROS, such as superoxide anions and hydrogen peroxide, are byproducts of normal oxidative metabolism in eukaryotic cells and are involved in many signaling process. However, they can be harmful to cellular components, including DNA. 3,8,9 An uncontrolled elevation of intracellular ROS levels is believed to contribute to cellular aging and the senescence process. 3 In fact, an abnormal elevation of intracellular ROS levels has been implicated in the pathogenesis of various diseases, such as ataxia telangiectasia and Fanconi anemia. 3 In that sense, the maintenance of ROS levels, through highly regulated mechanisms, is essential for cellular homeostasis. 10 Being continuously exposed to oxidants produced during metabolic activity and to external oxidants or oxidant-inducers through normal cellular physiology, DNAs within cells inevitably suffer the oxidative damage. Therefore, an accelerated proliferation of hematopoietic cells, which is expected to occur after clinical HSC transplantation, might lead to DNA damage through overexposure to oxidative stress generated on each cell cycle. Indeed, a hyperproliferation caused an accumulation of oxidative stress and resulted in functional exhaustion of murine HSCs, as shown by the failure to reconstitute hematopoiesis after serial transplantations. 11 Taken together, we hypothesize that the continuous production of ROS during long-term repopulation induces an accumulation of genomic damage that leads to exhaustion of human HSCs. We have previously developed a strategy that enables to examine the multipotency of a single human HSC using a reliable surrogate system. 12,13 By determining the in vivo repopulating dynamics of individual human HSCs, we demonstrated that the repopulating potential of the majority of human HSCs progressively deteriorated as they underwent extensive repopulation process. Furthermore, the self-renewing long-term repopulating...

NK cells and dendritic cells (DCs) are both important in the innate host defense. However, the role of DCs in NK cell-mediated cytotoxicity is unclear. In this study, we designed two culture systems in which human cord blood CD34+ cells from the same donor were induced to generate NK cells and DCs, respectively. Coculture of the NK cells with DCs resulted in significant enhancement of NK cell cytotoxicity and IFN-γ production. However, NK cell cytotoxicity and IFN-γ production were not increased when NK cells and DCs were grown together separated by a transwell membrane. Functional studies demonstrated that 1) concanamycin A, a selective inhibitor of perforin/granzyme B-based cytolysis, blocked DC-stimulated NK cytotoxicity against K562 cells; and 2) neutralizing mAb against Fas ligand (FasL) significantly reduced DC-stimulated NK cytotoxicity against Fas-positive Jurkat cells. In addition, a marked increase of FasL mRNA and FasL protein expression was observed in DC-stimulated NK cells. The addition of neutralizing mAb against IL-18 and IL-12 significantly suppressed DC-stimulated NK cell cytotoxicity. Neutralizing IFN-γ Ab almost completely inhibited NK cell cytotoxicity against Jurkat cells. These observations suggest that DCs enhance NK cell cytotoxicity by up-regulating both perforin/granzyme B- and FasL/Fas-based pathways. Direct interaction between DCs and NK cells is necessary for DC-mediated enhancement of NK cell cytotoxicity. Furthermore, DC-derived IL-18 and IL-12 were involved in the up-regulation of NK cell cytotoxicity, and endogenous IFN-γ production plays an important role in Fas-mediated cytotoxicity.

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