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MUSCLE CELL BIOLOGY AND CELL MOTILITY
1Laboratorio di Biologia Vascolare e Terapia Genica, Centro Cardiologico Fondazione Monzino, Milan; and 2Laboratorio di Patologia Vascolare, Istituto Dermopatico dell'Immacolata, Rome, Italy
Submitted 24 August 2007 ; accepted in final form 16 December 2007
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ABSTRACT |
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 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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adult tissues; ectopic skeletal muscle progenitors; spontaneous myogenesis
Satellite cells can be easily isolated from adult skeletal muscle by enzymatic digestion and cultured in vitro. However, different isolation/purification procedures have led to the identification of a heterogeneous population of spontaneously differentiating myogenic cells, which can be distinguished by differential expression of some surface antigens, such as Sca-1, CD34 and/or by the lack versus presence of early myogenic markers such as Pax3, Pax7, Myf5, as well as different adhesive and proliferative properties (24, 27, 31). Whether all these cells are derived from or represent progenitor of sublaminal satellite cells is not clear. Nonetheless, although they may initially display several differences, these cells acquire a very similar phenotype when cultured in vitro and undergo spontaneous skeletal muscle differentiation in low serum conditions. Moreover, when transplanted in vivo into damaged muscle, probably due to their intrinsic myogenicity, they exhibit a high regenerative potential and contribute to muscle fiber formation as well as to the replenishment of the satellite cells pool (3, 19, 24).
Several studies indicate that cells derived from nonmuscle adult tissues are also able to activate the myogenic program. Such cells, which include bone marrow-derived cells, side population cells, and mesenchymal cells as well as circulating progenitors, can be directed toward a myogenic phenotype when in contact with differentiating primary myoblasts in vitro or in vivo (2, 4, 11, 16, 23, 28, 30). However, these cells do not possess an intrinsic myogenic potential, do not spontaneously form skeletal myotubes and, in some cases, although they can fuse with preexisting myotubes, are unable to generate new muscle fibers de novo (27) or even to activate a complete skeletal muscle program (18).
The detection of skeletal muscle progenitors, termed mesoangioblasts, in the dorsal aorta of the mouse embryo (6) has led to the suggestive hypothesis that adult organs, especially those that are highly vascularized, may serve as a reservoir of muscle stem cells. Mesoangioblast-like cells have been isolated from vascularized muscle biopsies of postnatal dogs (26). In addition, pericytes obtained from human postnatal muscle biopsies have been reported to acquire a myogenic phenotype (7). However, the possibility to obtain fully competent myogenic cells from the vascular or any other compartment of adult muscle-devoid organs has, up to now, remained unproven.
In the present study we report, for the first time, the identification of spontaneously differentiating myogenic precursors in the pancreas and in other muscle-devoid organs of the adult mouse. Such cells, referred to as autonomously myogenic cells (AMCs), may be isolated from Flk-1+ progenitors and spontaneously form multinucleated, contractile myotubes when cultured in vitro. Individual clones of myogenic cells from nonmuscle organs are morphologically and functionally indistinguishable from skeletal muscle-derived primary myoblasts and are able to participate to muscle regeneration in vivo.
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MATERIALS AND METHODS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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For Langerhans islets isolation, pancreas was digested with 1 mg/ml collagenase P. Islets were picked by hand under a dissection microscope, cleaned from any surrounding exocrine and ductal tissue, and transferred on uncoated tissue chamber slides in GM.
To isolate myogenic clones, cell suspensions obtained by collagenase digestion were seeded on uncoated culture dishes in GM. After 8 h, the supernatant was transferred on fibronectin-coated plates in GM supplemented with 5 ng/ml basic FGF (bFGF). Well-isolated clones were picked up with the aid of cloning cylinders and transferred to new plates.
For the Matrigel assay, proliferating AMCs were seeded on growth factor-reduced Matrigel (BD) in DMEM supplemented with 5% FBS in the presence or absence of 50 ng/ml VEGF. Quantification of capillary-like structures formation was calculated as the mean number of branching points in five different fields in three independent experiments. AMCs arranged in capillary-like structures were extracted from Matrigel with dispase (1 mg/ml in PBS) and were replated on fibronectin-coated dishes in GM supplemented with 50 ng/ml VEGF. After an overnight culture, cells were either incubated with acetylated LDL (Ac-LDL) and then fixed and stained for TnT or were fixed and stained for both TnT and von Willebrand factor. Endothelial cells derived from digested aorta samples were used as controls.
Streptozotocin treatment. Two-month-old Swiss CD1 mice were treated with a single intraperitoneal injection of 200 mg/kg streptozotocin (STZ; Sigma) in 0.1 M citrate buffer, pH 4.5, which induced pancreatic damage and secondary hyperglycemia. Control animals were injected with an equivalent volume of citrate buffer. Studies in a test group of animals indicated that this procedure assured the development of hyperglycemia (i.e., blood glucose >350 mg/dl) in 100% of injected mice 10 days after treatment. Morphological assessment of pancreatic sections stained with hematoxylin and eosin and for insulin showed that pancreatic damage by STZ was associated with the destruction of pancreatic islets at day 10. Animals with a plasma glucose concentration >400 mg/dl 10 days after treatment were used for all experiments.
Patch-clamp recordings.
Cells were bathed in normal external solution containing (in mM) 140 NaCl, 2.8 KCl, 2 CaCl2, 2 MgCl2, 10 HEPES, and 10 glucose at pH 7.3. Whole cell currents were recorded at room temperature (23°–26°C), using an Axopatch 200B amplifier (Axon Instruments) and a gravity-driven fast perfusion system (MSC-200, Bio-Logic, Claix, France). Data were sampled and analyzed using pCLAMP software (version 9.0, Axon Instruments). Patch pipettes of borosilicate glass (WPI) had a tip resistance of 2–4 M
(compensated by 70–80%) and were filled with a solution containing (in mM) 140 KCl, 11 EGTA, 1 CaCl2, 2 MgCl2, 10 HEPES, and 2 ATP-Mg, buffered with KOH at pH 7.3.
ACh current-voltage relation curves were constructed by measuring the cell responses elicited by ACh (10 µM) at different test potentials, holding the cell at –70 mV between applications. ACh currents from each cell were normalized taking the response at –70 mV as 100%. ACh was applied at different concentrations to each cell, with a 60-s interval between applications (holding potential –70 mV). The amplitude of ACh currents was normalized to the values obtained at 100 µM ACh in each cell and averaged. Voltage-gated currents were studied by delivering 200-ms-long depolarizing pulses from a resting potential of –70 mV. Membrane capacitance (Cm) was measured from capacitive transients evoked by a 10-mV depolarizing step (Vstep). The total charge mobilized by the voltage step (Qstep) was derived from the transient integral, and then Cm = Qstep/Vstep was calculated.
RT-PCR. Contractile clones were harvested by the aid of cloning cylinders. Total RNA was extracted using RNeasy mini kit (Qiagen) according to manufacturer instructions. RNA was reverse transcribed with M-MLV Reverse Transcriptase (Promega) using random examers. PCR from RT samples was performed to exclude genomic DNA contamination.
Immunofluorescence. Cultures were fixed with 4% paraformaldehyde in PBS, permeabilized with 0.1% Triton X-100, and stained with antibodies against myogenin (gift from Dr. F. Martelli), troponin T (TnT; Santa Cruz), insulin, smooth muscle (SM) actin, and von Willebrand factor (Abcam). Positive cells were detected with Texas Red or FITC-conjugated anti-mouse IgG antibody (Vector). Nuclei were visualized with Hoechst.
Immunomagnetic sorting. Single-cell suspensions were incubated with anti-mouse Flk-1 phycoerythrin-conjugated monoclonal antibody (BD Pharmingen) and sorted with miniMACS (Miltenyi) or Easy-Sep (Stem Cell) kits, according to the manufacturers' procedures.
AMC transplantation. All experimental procedures complied with the Guidelines of the Italian National Institutes of Health and were approved by the Institutional Animal Care and Use Committee.
All mice were anesthetized with 2.5% Avertin. Hindlimb ischemia was induced by femoral artery removal as previously reported (9). AMCs were obtained from the pancreas and periaortic tissue of green fluorescent protein (GFP)-positive mice (20). Cells (2.5 x 105; passages IV–VI) were delivered by five injections (5 x 104/injection in 10 µl PBS) in the left adductor muscle of GFP-negative syngenic mice immediately after femoral artery removal. Ischemic limbs were processed for immunohistochemistry 1 wk after injection. A rabbit polyclonal antibody against GFP (Abcam) was used to visualize donor-derived fibers.
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RESULTS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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-skeletal actin) specific skeletal muscle markers (Fig. 1B, lane 2), whereas these markers were absent in uncultured pancreas-derived cells (Fig. 1B, lane 1). Conversely, expression of the pancreatic lineage specific marker Pdx1, as well as of the SM-specific myosin heavy chain (SM-MHC), was detected in the uncultured total cell population but not in pancreas-derived contractile clones. Expression of skeletal muscle-specific protein by a subpopulation of pancreas-derived cells was confirmed by immunofluorescence staining with antibodies against myogenin (Myog, Fig. 1, C and D) and TnT (Fig. 1, E and F). The presence of both immature mononucleated myoblasts and more differentiated multinucleated myotubes in each contractile clone, as revealed by TnT staining, suggests that pancreas-derived myogenic precursors undergo several rounds of proliferation before the onset of differentiation. Approximately 0.01% (0.0084 ± 0.00138; n = 10) of total plated cells form contractile clones. However, the number of myotubes in each clone and therefore the total number of myotubes varied between 0.1% and 1% in different experiments and was inversely correlated to cell density. Switching to DM accelerated the appearance but was not required for the formation of TnT-positive cells, since, as it happens for muscle-derived myogenic precursors, high cell density was sufficient to induce myotube formation and terminal differentiation.
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Pancreatic myogenic cells are associated with Langerhans islets. To establish which pancreas compartment contains myogenic cells, the exocrine and endocrine pancreatic tissues were cultured separately. Pancreas was briefly digested with collagenase P, and either isolated Langerhans islets or little fragments of islet-free exocrine tissue were independently picked and transferred to uncoated culture dishes in GM. Exocrine tissue immediately assumed a flattened morphology, whereas islets maintained their roundish appearance for several days. After 1 wk, both tissues generated a monolayer of cells with various morphology. By 2 wk, spontaneously contracting elongated cells started to appear in islet cultures, but they were never detected in exocrine tissue cultures. The formation of skeletal myotubes by pancreatic islets-associated cells was confirmed by double immunofluorescence with antibodies against insulin and TnT. As shown in Fig. 2, insulin-expressing cells (green) are confined to the islet, whereas TnT-expressing cells (red) are found in the insulin-negative cell population that had migrated out of the islet. Approximately 25% of cultured islets gave rise to TnT-positive cells. Moreover, the number of TnT-positive cells obtained from each islet was very variable, ranging from 10 to 1,000 as determined from 15 experiments. As in the case of total pancreas cultures described above, switching to DM accelerated the appearance but was not required for the formation of TnT-positive cells. Exocrine tissue cultures were instead negative for both endocrine (insulin and glucagone) and skeletal muscle markers, as confirmed by RT-PCR analysis (Fig. 2D).
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70% of total islet cells, and it has been shown that under conditions similar to those of the present study, they can undergo an epithelial-mesenchymal transition and assume a phenotype characterized by the absence of hormone production, concomitantly with the expression of mesenchymal markers such as vimentin and SM actin (14). SZT is a chemical compound that is commonly used to induce diabetes in experimental animals and that is highly cytotoxic for cells expressing the glucose transporter Glut-2, a protein expressed in β-cells. To test whether myogenic cells could be related to Glut-2/insulin-producing cells, the presence of TnT-positive cells was examined in cultures of pancreatic cells from mice that had been previously treated with STZ and in which the ablation of β-cells was documented by determination of blood glucose concentration. Formation of contractile, TnT-positive cells was observed in both STZ-treated and control mice (Fig. S1; the online version of this article contains supplemental data). Moreover, the number of TnT-positive myotubes was comparable in STZ-treated and saline-treated mice (n = 8). Altogether, these results show that pancreatic myogenic precursors are associated with the endocrine pancreas compartment but also that they are not derived from the population of insulin-producing cells.
Pancreas-derived and skeletal muscle-derived myotubes have comparable electrophysiological properties. To test the electrophysiological properties of pancreas-derived myotubes, patch-clamp recordings were performed on multinucleated myotubes obtained from pancreatic islet cultures. Myotubes generated from skeletal-muscle derived primary satellite cells isolated from age-matched mice were used for comparison.
The resting potential measured in pancreas-derived myotubes corresponded to –50.3 ± 7.2 mV (n = 3) and was very similar to that found in skeletal-muscle-derived myotubes (–50.2 ± 7.1 mV; n = 3).
ACh currents were observed in all tested samples. As in skeletal myotubes, the current amplitude depended on ACh concentration and on membrane test potential. With an ACh concentration of 10 µM at –70 mV, the average response was –10.0 ± 2.0 nA (n = 4), ranging between –5.2 and –14.6 nA (Fig. 3A). The voltage dependence of ACh-evoked responses showed an average reverse potential of 2.5 ± 2.3 mV (n = 4). Elicited ACh currents were directly proportional to ACh concentration (Fig. 3B). Experiments performed with satellite cell-derived myotubes produced totally comparable results: on exposure to 10 µM ACh, currents had an average response of –13.7 ± 2.1 nA (n = 7) ranging between –4.6 and –18.5 nA. ACh-evoked responses corresponded to 2.5 ± 1.7 mV (n = 5), and ACh currents were directly proportional to ACh concentration (data not shown).
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These results show that pancreas-derived myotubes express functional ACh receptors and that their electrophysiological behavior is very similar to that of satellite cell-derived myotubes, thus demonstrating that pancreas-associated cells can give rise to terminally differentiated, mature, and functional skeletal muscle cells.
Spontaneously differentiating myogenic cells can be found in a variety of adult mouse tissues of both endodermal and mesodermal origin and can be isolated as individual clones.
In additional experiments, it was examined whether AMCs are pancreas specific, or, alternatively, can be found in other adult organs. Tissues of both endodermal (i.e., stomach, liver, and lung) and mesodermal (i.e., spleen, heart, and aorta) origin were screened for the ability to generate skeletal myotubes. As described for pancreas, single-cell suspensions obtained by collagenase digestion of the different organs were plated on fibronectin-coated dishes, were kept in GM for 1 wk, and were then switched to DM for an additional 2 days. Contractile myotubes were visible in spleen- and stomach-derived cell cultures, whereas they were never detected in culture from heart, liver, or lungs (n = 6 for each sample). Surprisingly, AMCs were not detected in samples derived from the aorta. However, the periaortic adipose tissue was particularly enriched for such cells. The differentiation toward a striated muscle phenotype was confirmed by TnT staining (Fig. 4). RT-PCR analysis of myogenic markers revealed that, as expected, contractile cells were positive for a whole set of skeletal muscle markers (Myf5, MyoD, and myogenin), whereas they were negative for the cardiac muscle-specific markers Nkx2.5, Tbx5, and -MHC (data not shown). Similarly to that observed for pancreatic islets, uncultured digested tissue samples were instead always negative for all skeletal muscle marker tested, including Pax3 and Pax7. Owing to the marked differences in cell composition, adhesion properties, and plating efficiency of the various cell populations, it is very difficult to provide a meaningful estimate of the percentage of myogenic progenitors present in each tissue. However, the number of TnT-positive cells varied among the different organ sources and was higher in pancreas and periaortic tissue (0.0051 ± 0.0011% and 0.0053 ± 0.001% of total plated cells, respectively), compared with spleen and stomach (0.00084 ± 0.0001% and 0.0002 ± 0.00007%, respectively) (n = 6). Patch-clamp experiments performed on multinucleated myotubes demonstrated that, as in the case of pancreas-derived myotubes, the electrophysiological behavior of differentiated AMCs from spleen, stomach, and periaortic tissue was comparable to that of satellite cell-derived myotubes (data not shown).
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AMCs derive from Flk-1-positive progenitors. RT-PCR analysis indicates that AMCs express Flk-1. Differently from endothelial cells, AMCs did not respond to VEGF with a robust proliferative response. In fact, the addition of VEGF instead of bFGF in the GM did not induce myogenic clone expansion. Nonetheless, VEGF stimulated the formation of capillary-like structures in an in vitro Matrigel assay (Fig. 6), although in such assay, AMCs maintained their myogenic identity and did not exhibit characteristics of functional endothelial cells such as von Willebrand factor or Ac-LDL uptake (Fig. S2).
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0.01% of Flk-1+ cells (0.089 ± 0.0022%, n = 6 and 0.09 ± 0.0026%, n = 5, respectively), whereas in the case of spleen the frequency was lower (0.005 ± 0.002%, n = 4). The presence of AMCs was easily detected because of their characteristic morphology and was always confirmed by the ability to form contractile, multinucleated myotubes upon switching to low serum. Phenotypical analysis of nonmyogenic clones identified most of them as endothelial, since they expressed the endothelial markers Flk-1, CD31, and vascular endothelial-cadherin (data not shown) and were positive in an Ac-LDL uptake assay, whereas they did not express the myogenic marker TnT (Fig. 7B). As already shown in Fig. 5, AMCs expressed endothelial markers but differently from endothelial cells, failed to uptake Ac-LDL (Fig. 7A). As established by RT-PCR (Fig. 1) and confirmed by immunostaining (supplemental data Fig. S3) myogenic clones were negative for the SM markers SM actin and SM-MHC.
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AMCs cells participate in skeletal muscle regeneration in vivo.
To establish their in vivo myogenic potential, AMCs were assayed for the capacity to contribute to muscle regeneration in a mouse model of hindlimb ischemia. Pancreas-derived AMC cell clones were isolated from GFP-positive mice, expanded in vitro for 20 passage doublings, and then injected into the adductor muscle of GFP-negative syngenic mice immediately after femoral artery removal. Engrafted GFP-expressing cells were visualized 1 wk after injection. Representative images from control PBS-injected and AMC-injected hindlimbs are shown in Fig. 8, A and B, respectively. GFP-positive fibers represented 20% ± 4% (n = 6) of the total area of adductor muscle sections from treated hindlimbs, whereas no GFP staining was observed in control sections from PBS-injected limbs. Analogous results were obtained with periaortic tissue-derived cells (data not shown), suggesting that AMCs derived from different organs have similar in vivo regenerative abilities. These data show that AMCs cells can be efficiently incorporated into skeletal muscle fibers in vivo and may effectively contribute to skeletal muscle regeneration.
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DISCUSSION |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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The presence of AMCs in adult nonmuscle tissues has been reported in only two cases. One is represented by the thymic myoid cells, a restricted population of committed myogenic progenitors of unknown derivation present in the thymic medulla (21), and the other consists of a subsets of adipose tissue-associated cells, recently described in our laboratory (9).
In the present study we report, for the first time, the identification of AMCs in the pancreas as well as in several other muscle-devoid organs of both endodermal and mesodermal origin, namely stomach, spleen, and periaortic tissue of the adult mouse.
Similarly to embryonic mesoangioblasts, adult AMCs closely resemble muscle satellite cells, express a number of myogenic and endothelial markers, and differentiate spontaneously into contractile skeletal myotubes. Recently, the isolation of mesoangioblasts from biopsies of postnatal (p15) skeletal muscle and their regenerative ability in a canine model of muscular dystrophy have been reported (26). However, at variance with their mouse embryonic counterpart and with the AMCs described in the present study, postnatal mesoangioblasts do not spontaneously give rise to skeletal myotubes but do so only in the presence of predetermined myogenic cells. The same group has also reported the isolation from human adult muscle biopsies of pericyte-derived cells endowed with a certain degree of spontaneous myogenic potential (7). Whether pericytes isolated from sources outside of muscle tissue also possess myogenic potential has not been determined. Nonetheless, proliferating pericyte-derived cells do not express any myogenic or endothelial marker and therefore must represent a distinct subset of myogenesis competent cells compared with AMCs.
Similarly to muscle-derived myogenic precursors, AMCs belong to a slow-adherent population distinct from the fast-adhering mesenchymal compartment, from which they can be partially separated by preplating techniques. Individual clones of myogenic cells from nonmuscle organs can be induced to proliferate in vitro and are morphologically indistinguishable from skeletal muscle-derived primary satellite cells. Moreover, as suggested by transplantation experiments into a damaged muscle, AMCs cells can significantly participate in skeletal muscle regeneration, further demonstrating that they can be considered as bona fide muscle precursors.
Differently from embryonal brain, chick fetal organs, and the thymic medulla, which contain cells expressing skeletal muscle differentiation markers, but similarly to the embryonic dorsal aorta, adult tissues from nonmuscle sources do not express markers of myogenic commitment before cell dissociation and culturing. Freshly dissected organs examined by RT-PCR were negative for a whole panel of myogenic markers, including Pax3, Pax7, Myf-5, and MyoD. Immunostaining for TnT of pancreatic islet sections gave negative results as well. This suggests that AMCs are not originally committed to the myogenic phenotype but could represent a more immature or different phenotype, which becomes myogenic under specific culture conditions.
The strict similarity among AMCs isolated from different sources would suggest that they represent a unique cell type derived from a common cellular compartment, probably of mesodermal origin, present in embryologically unrelated organs. Immunomagnetic selection procedures have allowed us to establish that AMCs from different organs all derive from Flk-1+ cells, suggesting that AMCs progenitors may be associated with the endothelial layer of the vasculature. In this respect, AMCs can be detected in cultures of pancreatic islets, which are highly vascularized, and display a very high density of Flk-1+ cell, but not of exocrine pancreas, which has a much lower vessels content (1). On the other side, we were unable to isolate AMCs from heart, liver, or lung, which are also highly vascularized. This may be due either to the fact that the digestion procedure has to be optimized for each different tissue to allow the release of myogenesis-competent cells or, more plausibly, that the myogenic potential can be retained only in particular niches or vascular districts. It has been reported that when injected into a damaged muscle, embryonal lung-derived endothelial cells can be incorporated into skeletal muscle fibers, while postnatal lung cells do that with a very limited efficiency (5). However, in agreement with the results described in the present study, there was no experimental evidence that postnatal or embryonal lung-derived endothelial cells can spontaneously differentiate into skeletal muscle.
The fact that the adult aorta is devoid of AMCs, while they are found in the surrounding periaortic adipose tissue, suggests that in the adult animal, myogenesis-competent cells may be restricted to the microvascular compartment. Further studies will be needed to examine whether AMCs originate from an undifferentiated Flk-1+ mesodermal progenitor or, alternatively, by transdifferentiation of a terminally differentiated endothelial cell.
Similarly to what we have previously observed for adipose tissue-derived myogenic cells (9), attempts to prospectively isolate AMCs by mean of immunomagnetic selection with antibodies against antigens commonly associated with muscle progenitors, such as CD34 and Sca1, have produced negative results, since AMCs are invariably associated with the CD34-depleted or Sca1-depleted fraction. This indicates that AMCs belong to a CD34–/Sca1– cell population and that, similarly to what happens for muscle-specific markers, Sca1 and CD34 start to be expressed only upon culturing.
Most skeletal muscles derive from embryonic somites, and recent studies have established that the majority of the satellite cells are derived from Pax3+/Pax7+ progenitors, also located in the somite (15, 25). Several lines of evidence indicate that somites contain progenitors for both the skeletal muscle and the vascular lineages. With the use of retroviral insertion as a lineage marking method, it has been demonstrated that myogenic and endothelial cells in the limbs are derived from a common somitic precursor (17). Notably, in addition to dorsal aorta, myogenic cells can be isolated from limb buds of c-met mutant mouse embryos, which lack appendicular muscles but have a normal vascular system (6). Moreover, a retrospective clonal analysis has indicated that cells in the dorsal aorta and skeletal muscle share a common clonal somitic progenitor (10). These results are consistent with the hypothesis that somite-derived cells of the embryonic vasculature may retain myogenic potential. Here we show that adult Flk-1+ cells residing outside muscle organs also retain myogenic potential. However, it is worth noting that while the dorsal aorta and the limb vasculature receive contributions from the somites, the endothelium (22), as well as other mesodermal compartments of internal organs, are embryologically derived from the splanchnopleura and do not have any lineage relationship with somitic cells. Therefore, to imply that all AMCs have a somitic origin, it has to be assumed that they have risen from somite-derived tissues and have successively been distributed to internal organs, perhaps through the circulation. Alternatively, it can be hypothesized that AMCs from internal organs are derived from progenitors of nonsomitic origin.
In conclusion, we show for the first time that fully competent myogenic progenitors derived from the Flk-1+ cell compartment can be isolated from several adult tissues that are embryologically unrelated to skeletal muscle. Such results provide additional proof for the concept that cells with an unexpected degree of plasticity do actually exist in adult mammalian organs.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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