Muscle-derived hematopoietic stem cells are hematopoietic in origin - PubMed (original) (raw)

Muscle-derived hematopoietic stem cells are hematopoietic in origin

Shannon L McKinney-Freeman et al. Proc Natl Acad Sci U S A. 2002.

Abstract

It has recently been shown that mononuclear cells from murine skeletal muscle contain the potential to repopulate all major peripheral blood lineages in lethally irradiated mice, but the origin of this activity is unknown. We have fractionated muscle cells on the basis of hematopoietic markers to show that the active population exclusively expresses the hematopoietic stem cell antigens Sca-1 and CD45. Muscle cells obtained from 6- to 8-week-old C57BL/6-CD45.1 mice and enriched for cells expressing Sca-1 and CD45 were able to generate hematopoietic but not myogenic colonies in vitro and repopulated multiple hematopoietic lineages of lethally irradiated C57BL/6-CD45.2 mice. These data show that muscle-derived hematopoietic stem cells are likely derived from the hematopoietic system and are a result not of transdifferentiation of myogenic stem cells but instead of the presence of substantial numbers of hematopoietic stem cells in the muscle. Although CD45-negative cells were highly myogenic in vitro and in vivo, CD45-positive muscle-derived cells displayed only very limited myogenic activity and only in vivo.

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Figures

Figure 1

Figure 1

Sca-1-positive and -negative and CD45-positive muscle cells display in vitro hematopoietic progenitor activity_._ (A) Freshly isolated muscle-derived cells were stained with monoclonal antibodies recognizing CD45 and Sca-1 analyzed by flow cytometry. (B) Sca-1-positive and -negative muscle-derived cells were plated into methylcellulose and assessed for colony number 9 days after plating (a). CD45-positive and -negative cells were also plated into methylcellulose and assessed for colony number 9 days after plating (b). (C) Methylcellulose cultures of CD45-positive cells (a). Cultures of CD45-negative cells (b). The arrowhead indicates an actively twitching myofiber. Unfractionated muscle-derived colonies were picked, cytospun, and stained with Wright–Giemsa (c and d).

Figure 2

Figure 2

In vivo hematopoietic activity of Sca-1 and CD45-sorted populations of muscle-derived cells. Muscle was isolated from C57BL/6-CD45.1 mice and magnetically enriched for Sca-1 or CD45 expression. Positive and negative populations were transplanted into lethally irradiated C57BL/6-CD45.2 recipients along with 2 × 105 C57BL/6-CD45.2 WBM cells. PB of recipients was analyzed via FACS for CD45.1 and hematopoietic lineage markers at the indicated time points after transplantation. In the representative Sca-1 transplant shown, Thy.1 staining was not performed at 20 weeks after transplant and therefore does not appear.

Figure 3

Figure 3

In vivo myogenic potential of muscle-derived cells. Muscle-derived cells were purified from C57BL/6-ROSA26 (B–F) or C57/M_lacZ_ (H, I) mice by flow cytometry or magnetically followed by flow cytometry. Test populations were injected into the TA muscles of non-ROSA26 animals that had been injected with cardiotoxin 24 h earlier. The TA muscles were removed from recipients 2–3 weeks later, cryosectioned, and stained with X-Gal for β-galactosidase expression. (Bar = 50 μM.) (A) Injured muscle injected with HBSS. (B) Muscle injected with unfractionated muscle-derived cells. The large blue tracts are fibers regenerated from Rosa26-derived cells. (C) Incorporation of Sca-1-positive muscle cells. (D) Incorporation of Sca-1-negative muscle cells. (E) Incorporation of Sca-1+/CD45+ cells. (F) Incorporation of Sca-1+/CD45− cells. (G) Section of unmanipulated C57/M_lacZ_ muscle. Note the nuclear localization of lacZ at the edges of the fibers shown in cross section. (H) CD45-positive C57/M_lacZ_-derived cells injected into regenerating muscle and sectioned longitudinally. (I) CD45-negative C57/M_lacZ_ incorporation.

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