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Myostatin negatively regulates satellite cell activation and self-renewal.

McCroskery S, Thomas M, Maxwell L, Sharma M, Kambadur R - J. Cell Biol. (2003)

Bottom Line: BrdU labeling in vivo revealed that, among the Myostatin-deficient satellite cells, higher numbers of satellite cells are activated as compared with wild type.Cell cycle analysis confirms that Myostatin up-regulated p21, a Cdk inhibitor, and decreased the levels and activity of Cdk2 protein in satellite cells.Taken together, these results suggest that Myostatin is a potent negative regulator of satellite cell activation and thus signals the quiescence of satellite cells.

View Article: PubMed Central - PubMed

Affiliation: Animal Genomics, AgResearch, Hamilton 2015, New Zealand.

ABSTRACT
Satellite cells are quiescent muscle stem cells that promote postnatal muscle growth and repair. Here we show that myostatin, a TGF-beta member, signals satellite cell quiescence and also negatively regulates satellite cell self-renewal. BrdU labeling in vivo revealed that, among the Myostatin-deficient satellite cells, higher numbers of satellite cells are activated as compared with wild type. In contrast, addition of Myostatin to myofiber explant cultures inhibits satellite cell activation. Cell cycle analysis confirms that Myostatin up-regulated p21, a Cdk inhibitor, and decreased the levels and activity of Cdk2 protein in satellite cells. Hence, Myostatin negatively regulates the G1 to S progression and thus maintains the quiescent status of satellite cells. Immunohistochemical analysis with CD34 antibodies indicates that there is an increased number of satellite cells per unit length of freshly isolated Mstn-/- muscle fibers. Determination of proliferation rate suggests that this elevation in satellite cell number could be due to increased self-renewal and delayed expression of the differentiation gene (myogenin) in Mstn-/- adult myoblasts. Taken together, these results suggest that Myostatin is a potent negative regulator of satellite cell activation and thus signals the quiescence of satellite cells.

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Myostatin is expressed in adult myoblasts derived from satellite cells. (A) Agarose gel electrophoresis of myostatin ORF amplified in a combined RT-PCR reaction using total RNA from adult myoblasts (lane 2). Amplicons are not detected in the absence of template (negative control) (lane 3). 1-kb plus DNA ladder is shown in lane 1. (B) Western analysis from proteins isolated from adult myoblasts cultured for 48 h showing both the full-length (52 kD) and LAP (38 kD) peptides of Myostatin. (C) Representative immunofluorescence showing the presence of Myostatin in wild-type adult myoblasts (i) and absence in myostatin- (Mstn−/−) myoblasts (iii). DAPI staining of the nuclei is shown in the corresponding fields (ii and iv).
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fig3: Myostatin is expressed in adult myoblasts derived from satellite cells. (A) Agarose gel electrophoresis of myostatin ORF amplified in a combined RT-PCR reaction using total RNA from adult myoblasts (lane 2). Amplicons are not detected in the absence of template (negative control) (lane 3). 1-kb plus DNA ladder is shown in lane 1. (B) Western analysis from proteins isolated from adult myoblasts cultured for 48 h showing both the full-length (52 kD) and LAP (38 kD) peptides of Myostatin. (C) Representative immunofluorescence showing the presence of Myostatin in wild-type adult myoblasts (i) and absence in myostatin- (Mstn−/−) myoblasts (iii). DAPI staining of the nuclei is shown in the corresponding fields (ii and iv).

Mentions: Three independent techniques, RT-PCR, Western blotting, and immunocytochemistry, were used to ascertain the expression of myostatin in adult myoblasts. As shown in Fig. 3 A, the PCR reaction amplified the expected size fragment of 1.2-kb myostatin ORF. In addition, anti-Myostatin antibodies specifically recognized Myostatin on the Western blot performed on the protein extracts derived from the adult myoblasts (Fig. 3 B). To further confirm the expression of Myostatin, primary cultures were immunostained with Myostatin antibodies. As shown in Fig. 3 C (i and ii), virtually all the cultured myoblasts showed staining with Myostatin antibodies. Thus, it is clear from these results that Myostatin is expressed in satellite cells and in the adult myoblasts derived from satellite cells.


Myostatin negatively regulates satellite cell activation and self-renewal.

McCroskery S, Thomas M, Maxwell L, Sharma M, Kambadur R - J. Cell Biol. (2003)

Myostatin is expressed in adult myoblasts derived from satellite cells. (A) Agarose gel electrophoresis of myostatin ORF amplified in a combined RT-PCR reaction using total RNA from adult myoblasts (lane 2). Amplicons are not detected in the absence of template (negative control) (lane 3). 1-kb plus DNA ladder is shown in lane 1. (B) Western analysis from proteins isolated from adult myoblasts cultured for 48 h showing both the full-length (52 kD) and LAP (38 kD) peptides of Myostatin. (C) Representative immunofluorescence showing the presence of Myostatin in wild-type adult myoblasts (i) and absence in myostatin- (Mstn−/−) myoblasts (iii). DAPI staining of the nuclei is shown in the corresponding fields (ii and iv).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172861&req=5

fig3: Myostatin is expressed in adult myoblasts derived from satellite cells. (A) Agarose gel electrophoresis of myostatin ORF amplified in a combined RT-PCR reaction using total RNA from adult myoblasts (lane 2). Amplicons are not detected in the absence of template (negative control) (lane 3). 1-kb plus DNA ladder is shown in lane 1. (B) Western analysis from proteins isolated from adult myoblasts cultured for 48 h showing both the full-length (52 kD) and LAP (38 kD) peptides of Myostatin. (C) Representative immunofluorescence showing the presence of Myostatin in wild-type adult myoblasts (i) and absence in myostatin- (Mstn−/−) myoblasts (iii). DAPI staining of the nuclei is shown in the corresponding fields (ii and iv).
Mentions: Three independent techniques, RT-PCR, Western blotting, and immunocytochemistry, were used to ascertain the expression of myostatin in adult myoblasts. As shown in Fig. 3 A, the PCR reaction amplified the expected size fragment of 1.2-kb myostatin ORF. In addition, anti-Myostatin antibodies specifically recognized Myostatin on the Western blot performed on the protein extracts derived from the adult myoblasts (Fig. 3 B). To further confirm the expression of Myostatin, primary cultures were immunostained with Myostatin antibodies. As shown in Fig. 3 C (i and ii), virtually all the cultured myoblasts showed staining with Myostatin antibodies. Thus, it is clear from these results that Myostatin is expressed in satellite cells and in the adult myoblasts derived from satellite cells.

Bottom Line: BrdU labeling in vivo revealed that, among the Myostatin-deficient satellite cells, higher numbers of satellite cells are activated as compared with wild type.Cell cycle analysis confirms that Myostatin up-regulated p21, a Cdk inhibitor, and decreased the levels and activity of Cdk2 protein in satellite cells.Taken together, these results suggest that Myostatin is a potent negative regulator of satellite cell activation and thus signals the quiescence of satellite cells.

View Article: PubMed Central - PubMed

Affiliation: Animal Genomics, AgResearch, Hamilton 2015, New Zealand.

ABSTRACT
Satellite cells are quiescent muscle stem cells that promote postnatal muscle growth and repair. Here we show that myostatin, a TGF-beta member, signals satellite cell quiescence and also negatively regulates satellite cell self-renewal. BrdU labeling in vivo revealed that, among the Myostatin-deficient satellite cells, higher numbers of satellite cells are activated as compared with wild type. In contrast, addition of Myostatin to myofiber explant cultures inhibits satellite cell activation. Cell cycle analysis confirms that Myostatin up-regulated p21, a Cdk inhibitor, and decreased the levels and activity of Cdk2 protein in satellite cells. Hence, Myostatin negatively regulates the G1 to S progression and thus maintains the quiescent status of satellite cells. Immunohistochemical analysis with CD34 antibodies indicates that there is an increased number of satellite cells per unit length of freshly isolated Mstn-/- muscle fibers. Determination of proliferation rate suggests that this elevation in satellite cell number could be due to increased self-renewal and delayed expression of the differentiation gene (myogenin) in Mstn-/- adult myoblasts. Taken together, these results suggest that Myostatin is a potent negative regulator of satellite cell activation and thus signals the quiescence of satellite cells.

Show MeSH
Related in: MedlinePlus