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A differential response to newt regeneration extract by C2C12 and primary mammalian muscle cells.

Kawesa S, Vanstone J, Tsilfidis C - Skelet Muscle (2015)

Bottom Line: We isolated extract from early newt forelimb regenerates and assessed its effects on differentiation of proliferating primary and C2C12 myoblasts.We have confirmed the results obtained in C2C12 cells and expanded these studies to also examine the effects of newt regeneration extracts on primary muscle cells.However, unlike C2C12 cells, primary muscle cells do not re-enter the cell cycle in response to treatment with newt extracts.

View Article: PubMed Central - PubMed

Affiliation: Ottawa Hospital Research Institute, Vision Research/Regenerative Medicine Program, 501 Smyth Road, Box 307, Ottawa, Ontario K1H 8L6 Canada ; Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5 Canada.

ABSTRACT

Background: Dedifferentiation, a process whereby differentiated cells lose their specialized characteristics and revert to a less differentiated state, plays a key role in the regeneration process in urodele amphibians such as the red spotted newt, Notophthalmus viridescens. Dedifferentiation of fully mature tissues is generally absent in mammalian cells. Previous studies have shown that mouse C2C12 multinucleated myotubes treated with extract derived from regenerating newt forelimbs can re-enter the cell cycle, fragment into mononucleated cells, and proliferate. However, this response has been difficult to replicate.

Methods: We isolated extract from early newt forelimb regenerates and assessed its effects on differentiation of proliferating primary and C2C12 myoblasts. We also treated fully differentiated primary and C2C12 myotube cultures with extract and assessed cell cycle re-entry and myotube fragmentation.

Results: We have confirmed the results obtained in C2C12 cells and expanded these studies to also examine the effects of newt regeneration extracts on primary muscle cells. Newt extract can block differentiation of both C2C12 and primary myoblasts. Once differentiation is induced, treatment with newt extract causes cell cycle re-entry and fragmentation of C2C12 myotubes. Downregulation of p21 and muscle-specific markers is also induced. Primary myotubes also fragment in response to extract treatment, and the fragmented cells remain viable for long periods of time in culture. However, unlike C2C12 cells, primary muscle cells do not re-enter the cell cycle in response to treatment with newt extracts.

Conclusions: Dedifferentiation of fully mature muscle occurs during regeneration in the newt forelimb to contribute cells to the regeneration process. Our study shows that extracts derived from regenerating newt forelimbs can induce dedifferentiation, cell cycle re-entry, and fragmentation of mouse C2C12 cells but can only induce fragmentation in primary muscle cells.

No MeSH data available.


Related in: MedlinePlus

p21 expression is downregulated by treatment with newt extract (see also Table 1). In control cells, p21 expression is seen in myoblasts, as they exit the cell cycle, and in MHC-expressing myocytes. In extract-treated cultures, the extensive p21 expression in mononucleated cells appears to be lost. However, larger myotubes retain p21 expression
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Fig6: p21 expression is downregulated by treatment with newt extract (see also Table 1). In control cells, p21 expression is seen in myoblasts, as they exit the cell cycle, and in MHC-expressing myocytes. In extract-treated cultures, the extensive p21 expression in mononucleated cells appears to be lost. However, larger myotubes retain p21 expression

Mentions: C2C12 myoblasts were allowed to differentiate for 5 days. Newt extract in growth medium (GM) was added to the differentiated cells for 3 days followed by immunostaining for p21. A significant decrease in p21 expression was seen in mononucleated cells in treated cultures (Fig. 6 and Table 1, p = 0.015). There was also a significant increase (p = 0.048) in the p21-/MHC- mononucleated cells (Fig. 6 and Table 1). This suggests that newt extract may block differentiation by blocking p21 expression or may have the ability to downregulate p21 expression in myocytes and thus prevent differentiation and MHC expression.Fig. 6


A differential response to newt regeneration extract by C2C12 and primary mammalian muscle cells.

Kawesa S, Vanstone J, Tsilfidis C - Skelet Muscle (2015)

p21 expression is downregulated by treatment with newt extract (see also Table 1). In control cells, p21 expression is seen in myoblasts, as they exit the cell cycle, and in MHC-expressing myocytes. In extract-treated cultures, the extensive p21 expression in mononucleated cells appears to be lost. However, larger myotubes retain p21 expression
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4471912&req=5

Fig6: p21 expression is downregulated by treatment with newt extract (see also Table 1). In control cells, p21 expression is seen in myoblasts, as they exit the cell cycle, and in MHC-expressing myocytes. In extract-treated cultures, the extensive p21 expression in mononucleated cells appears to be lost. However, larger myotubes retain p21 expression
Mentions: C2C12 myoblasts were allowed to differentiate for 5 days. Newt extract in growth medium (GM) was added to the differentiated cells for 3 days followed by immunostaining for p21. A significant decrease in p21 expression was seen in mononucleated cells in treated cultures (Fig. 6 and Table 1, p = 0.015). There was also a significant increase (p = 0.048) in the p21-/MHC- mononucleated cells (Fig. 6 and Table 1). This suggests that newt extract may block differentiation by blocking p21 expression or may have the ability to downregulate p21 expression in myocytes and thus prevent differentiation and MHC expression.Fig. 6

Bottom Line: We isolated extract from early newt forelimb regenerates and assessed its effects on differentiation of proliferating primary and C2C12 myoblasts.We have confirmed the results obtained in C2C12 cells and expanded these studies to also examine the effects of newt regeneration extracts on primary muscle cells.However, unlike C2C12 cells, primary muscle cells do not re-enter the cell cycle in response to treatment with newt extracts.

View Article: PubMed Central - PubMed

Affiliation: Ottawa Hospital Research Institute, Vision Research/Regenerative Medicine Program, 501 Smyth Road, Box 307, Ottawa, Ontario K1H 8L6 Canada ; Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5 Canada.

ABSTRACT

Background: Dedifferentiation, a process whereby differentiated cells lose their specialized characteristics and revert to a less differentiated state, plays a key role in the regeneration process in urodele amphibians such as the red spotted newt, Notophthalmus viridescens. Dedifferentiation of fully mature tissues is generally absent in mammalian cells. Previous studies have shown that mouse C2C12 multinucleated myotubes treated with extract derived from regenerating newt forelimbs can re-enter the cell cycle, fragment into mononucleated cells, and proliferate. However, this response has been difficult to replicate.

Methods: We isolated extract from early newt forelimb regenerates and assessed its effects on differentiation of proliferating primary and C2C12 myoblasts. We also treated fully differentiated primary and C2C12 myotube cultures with extract and assessed cell cycle re-entry and myotube fragmentation.

Results: We have confirmed the results obtained in C2C12 cells and expanded these studies to also examine the effects of newt regeneration extracts on primary muscle cells. Newt extract can block differentiation of both C2C12 and primary myoblasts. Once differentiation is induced, treatment with newt extract causes cell cycle re-entry and fragmentation of C2C12 myotubes. Downregulation of p21 and muscle-specific markers is also induced. Primary myotubes also fragment in response to extract treatment, and the fragmented cells remain viable for long periods of time in culture. However, unlike C2C12 cells, primary muscle cells do not re-enter the cell cycle in response to treatment with newt extracts.

Conclusions: Dedifferentiation of fully mature muscle occurs during regeneration in the newt forelimb to contribute cells to the regeneration process. Our study shows that extracts derived from regenerating newt forelimbs can induce dedifferentiation, cell cycle re-entry, and fragmentation of mouse C2C12 cells but can only induce fragmentation in primary muscle cells.

No MeSH data available.


Related in: MedlinePlus