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MyoD-positive myoblasts are present in mature fetal organs lacking skeletal muscle.

Gerhart J, Bast B, Neely C, Iem S, Amegbe P, Niewenhuis R, Miklasz S, Cheng PF, George-Weinstein M - J. Cell Biol. (2001)

Bottom Line: Practically all of the G8-positive cells from the intestine differentiated after purification by FACS.This population of ectopically located cells appears to be distinct from multipotential stem cells and myofibroblasts.They closely resemble quiescent, stably programmed skeletal myoblasts with the capacity to differentiate when placed in a permissive environment.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Philadelphia College of Osteopathic Medicine, Philadelphia, PA 19131, USA.

ABSTRACT
The epiblast of the chick embryo gives rise to the ectoderm, mesoderm, and endoderm during gastrulation. Previous studies revealed that MyoD-positive cells were present throughout the epiblast, suggesting that skeletal muscle precursors would become incorporated into all three germ layers. The focus of the present study was to examine a variety of organs from the chicken fetus for the presence of myogenic cells. RT-PCR and in situ hybridizations demonstrated that MyoD-positive cells were present in the brain, lung, intestine, kidney, spleen, heart, and liver. When these organs were dissociated and placed in culture, a subpopulation of cells differentiated into skeletal muscle. The G8 antibody was used to label those cells that expressed MyoD in vivo and to follow their fate in vitro. Most, if not all, of the muscle that formed in culture arose from cells that expressed MyoD and G8 in vivo. Practically all of the G8-positive cells from the intestine differentiated after purification by FACS. This population of ectopically located cells appears to be distinct from multipotential stem cells and myofibroblasts. They closely resemble quiescent, stably programmed skeletal myoblasts with the capacity to differentiate when placed in a permissive environment.

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Immunofluorescence localization of cell type–specific proteins and BrdU in G8-positive cells in vitro. Cells from fetal organs were prelabeled with the G8 antibody, cultured for 48 h, and then stained with a secondary antibody to G8 and antibodies to MyoD, Myf5, or sarcomeric myosin. Cells from the lung, liver, kidney, and intestine that were labeled with G8 (green) had MyoD and myosin (red). Double labeling for G8 (red) and Myf5 (green) was seen in lung and heart cells. G8-positive intestine cells (green) incorporated BrdU (red) indicating replication. In a second type of experiment, cells from the heart and brain were fixed directly after removal from the fetus, centrifuged onto slides, and stained with G8 (red in J; green in L) and antibodies to cardiac troponin I (green) and neurofilament protein (red), respectively. Cells labeled with G8 did not have detectable levels of cardiac troponin (J) or neurofilament protein (L). Cells with cardiac troponin (K) and neurofilament protein (L) were not labeled with G8. Bar, 9 μm.
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fig5: Immunofluorescence localization of cell type–specific proteins and BrdU in G8-positive cells in vitro. Cells from fetal organs were prelabeled with the G8 antibody, cultured for 48 h, and then stained with a secondary antibody to G8 and antibodies to MyoD, Myf5, or sarcomeric myosin. Cells from the lung, liver, kidney, and intestine that were labeled with G8 (green) had MyoD and myosin (red). Double labeling for G8 (red) and Myf5 (green) was seen in lung and heart cells. G8-positive intestine cells (green) incorporated BrdU (red) indicating replication. In a second type of experiment, cells from the heart and brain were fixed directly after removal from the fetus, centrifuged onto slides, and stained with G8 (red in J; green in L) and antibodies to cardiac troponin I (green) and neurofilament protein (red), respectively. Cells labeled with G8 did not have detectable levels of cardiac troponin (J) or neurofilament protein (L). Cells with cardiac troponin (K) and neurofilament protein (L) were not labeled with G8. Bar, 9 μm.

Mentions: The G8 antibody was used to mark living cells that express MyoD. Organs were dissociated, labeled in suspension with G8, and then placed in culture. Cells were stained with an IgM-specific secondary antibody conjugated with Alexa 488 to tag G8, the MF20 IgG antibody to sarcomeric myosin, and an IgG specific secondary antibody conjugated with rhodamine. All G8-positive cells contained MyoD protein, and all cells with MyoD were stained for G8 (Fig. 5, A, D, E, and G). Most of the G8-positive cells (∼95%) also expressed Myf5 protein, and all cells with Myf5 had G8 (Fig. 5, B and C).


MyoD-positive myoblasts are present in mature fetal organs lacking skeletal muscle.

Gerhart J, Bast B, Neely C, Iem S, Amegbe P, Niewenhuis R, Miklasz S, Cheng PF, George-Weinstein M - J. Cell Biol. (2001)

Immunofluorescence localization of cell type–specific proteins and BrdU in G8-positive cells in vitro. Cells from fetal organs were prelabeled with the G8 antibody, cultured for 48 h, and then stained with a secondary antibody to G8 and antibodies to MyoD, Myf5, or sarcomeric myosin. Cells from the lung, liver, kidney, and intestine that were labeled with G8 (green) had MyoD and myosin (red). Double labeling for G8 (red) and Myf5 (green) was seen in lung and heart cells. G8-positive intestine cells (green) incorporated BrdU (red) indicating replication. In a second type of experiment, cells from the heart and brain were fixed directly after removal from the fetus, centrifuged onto slides, and stained with G8 (red in J; green in L) and antibodies to cardiac troponin I (green) and neurofilament protein (red), respectively. Cells labeled with G8 did not have detectable levels of cardiac troponin (J) or neurofilament protein (L). Cells with cardiac troponin (K) and neurofilament protein (L) were not labeled with G8. Bar, 9 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2150848&req=5

fig5: Immunofluorescence localization of cell type–specific proteins and BrdU in G8-positive cells in vitro. Cells from fetal organs were prelabeled with the G8 antibody, cultured for 48 h, and then stained with a secondary antibody to G8 and antibodies to MyoD, Myf5, or sarcomeric myosin. Cells from the lung, liver, kidney, and intestine that were labeled with G8 (green) had MyoD and myosin (red). Double labeling for G8 (red) and Myf5 (green) was seen in lung and heart cells. G8-positive intestine cells (green) incorporated BrdU (red) indicating replication. In a second type of experiment, cells from the heart and brain were fixed directly after removal from the fetus, centrifuged onto slides, and stained with G8 (red in J; green in L) and antibodies to cardiac troponin I (green) and neurofilament protein (red), respectively. Cells labeled with G8 did not have detectable levels of cardiac troponin (J) or neurofilament protein (L). Cells with cardiac troponin (K) and neurofilament protein (L) were not labeled with G8. Bar, 9 μm.
Mentions: The G8 antibody was used to mark living cells that express MyoD. Organs were dissociated, labeled in suspension with G8, and then placed in culture. Cells were stained with an IgM-specific secondary antibody conjugated with Alexa 488 to tag G8, the MF20 IgG antibody to sarcomeric myosin, and an IgG specific secondary antibody conjugated with rhodamine. All G8-positive cells contained MyoD protein, and all cells with MyoD were stained for G8 (Fig. 5, A, D, E, and G). Most of the G8-positive cells (∼95%) also expressed Myf5 protein, and all cells with Myf5 had G8 (Fig. 5, B and C).

Bottom Line: Practically all of the G8-positive cells from the intestine differentiated after purification by FACS.This population of ectopically located cells appears to be distinct from multipotential stem cells and myofibroblasts.They closely resemble quiescent, stably programmed skeletal myoblasts with the capacity to differentiate when placed in a permissive environment.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Philadelphia College of Osteopathic Medicine, Philadelphia, PA 19131, USA.

ABSTRACT
The epiblast of the chick embryo gives rise to the ectoderm, mesoderm, and endoderm during gastrulation. Previous studies revealed that MyoD-positive cells were present throughout the epiblast, suggesting that skeletal muscle precursors would become incorporated into all three germ layers. The focus of the present study was to examine a variety of organs from the chicken fetus for the presence of myogenic cells. RT-PCR and in situ hybridizations demonstrated that MyoD-positive cells were present in the brain, lung, intestine, kidney, spleen, heart, and liver. When these organs were dissociated and placed in culture, a subpopulation of cells differentiated into skeletal muscle. The G8 antibody was used to label those cells that expressed MyoD in vivo and to follow their fate in vitro. Most, if not all, of the muscle that formed in culture arose from cells that expressed MyoD and G8 in vivo. Practically all of the G8-positive cells from the intestine differentiated after purification by FACS. This population of ectopically located cells appears to be distinct from multipotential stem cells and myofibroblasts. They closely resemble quiescent, stably programmed skeletal myoblasts with the capacity to differentiate when placed in a permissive environment.

Show MeSH