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Characterisation of nuclear architectural alterations during in vitro differentiation of human stem cells of myogenic origin.

Rozwadowska N, Kolanowski T, Wiland E, Siatkowski M, Pawlak P, Malcher A, Mietkiewski T, Olszewska M, Kurpisz M - PLoS ONE (2013)

Bottom Line: Additionally, almost all of the chromosomal centromeres in the differentiated cells preferentially localised near the nuclear periphery when compared to the undifferentiated cells.The exceptions were chromosomes 7 and 11, in which we were unable to confirm the centromere repositioning.Based on these data we can conclude that the myogenic differentiation with global gene expression changes is accompanied by the spatial repositioning of chromosomes and chromatin remodelling, which are important processes that regulate cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Reproductive Biology and Stem Cells, Institute of Human Genetics, Polish Academy of Sciences, Poznan, Poland.

ABSTRACT
Cell differentiation is based on a synchronised orchestra of complex pathways of intrinsic and extrinsic signals that manifest in the induced expression of specific transcription factors and pivotal genes within the nucleus. One cannot ignore the epigenetic status of differentiating cells, comprising not only histones and DNA modifications but also the spatial and temporal intranuclear chromatin organisation, which is an important regulator of nuclear processes. In the present study, we investigated the nuclear architecture of human primary myoblasts and myocytes in an in vitro culture, with reference to global changes in genomic expression. Repositioning of the chromosomal centromeres, along with alterations in the nuclear shape and volume, was observed as a consequence of myotube formation. Moreover, the microarray data showed that during in vitro myogenesis cells tend to silence rather than induce gene expression. The creation of a chromosome map marked with gene expression changes that were at least 2-fold confirmed the observation. Additionally, almost all of the chromosomal centromeres in the differentiated cells preferentially localised near the nuclear periphery when compared to the undifferentiated cells. The exceptions were chromosomes 7 and 11, in which we were unable to confirm the centromere repositioning. In our opinion, this is the first reported observation of the movement of chromosomal centromeres along differentiating myogenic cells. Based on these data we can conclude that the myogenic differentiation with global gene expression changes is accompanied by the spatial repositioning of chromosomes and chromatin remodelling, which are important processes that regulate cell differentiation.

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Immunofluorescence analysis of desmin (DES), myosin heavy chain (MYH) and alpha-actinin (ACTN) in myoblasts (Mb24 hrs) and myocytes after 7 days of differentiation (Mc7d).(Magnification in the left and middle columns-200x; in the right column 630x).
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pone-0073231-g003: Immunofluorescence analysis of desmin (DES), myosin heavy chain (MYH) and alpha-actinin (ACTN) in myoblasts (Mb24 hrs) and myocytes after 7 days of differentiation (Mc7d).(Magnification in the left and middle columns-200x; in the right column 630x).

Mentions: We evaluated the expression of myosin heavy chain (MYH), desmin (DES) and actinin (ACTN) in different populations of myoblasts and myocytes (Figure 3). The human proliferating myoblasts showed a high expression of desmin filaments, confirming their myogenic origin, and this expression was further maintained following cell differentiation. Some of the myoblasts also expressed MYH, although the signal intensity was lower compared to myocytes, most likely due to a more diffused distribution. On the contrary, ACTN was present only in the myocytes. Surprisingly, we observed actinin-positive myoblasts, which were most likely correlated with changes in the cell shape and spontaneous differentiation. The fraction of actinin-positive cells within the myoblast cell population was negligible.


Characterisation of nuclear architectural alterations during in vitro differentiation of human stem cells of myogenic origin.

Rozwadowska N, Kolanowski T, Wiland E, Siatkowski M, Pawlak P, Malcher A, Mietkiewski T, Olszewska M, Kurpisz M - PLoS ONE (2013)

Immunofluorescence analysis of desmin (DES), myosin heavy chain (MYH) and alpha-actinin (ACTN) in myoblasts (Mb24 hrs) and myocytes after 7 days of differentiation (Mc7d).(Magnification in the left and middle columns-200x; in the right column 630x).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3760906&req=5

pone-0073231-g003: Immunofluorescence analysis of desmin (DES), myosin heavy chain (MYH) and alpha-actinin (ACTN) in myoblasts (Mb24 hrs) and myocytes after 7 days of differentiation (Mc7d).(Magnification in the left and middle columns-200x; in the right column 630x).
Mentions: We evaluated the expression of myosin heavy chain (MYH), desmin (DES) and actinin (ACTN) in different populations of myoblasts and myocytes (Figure 3). The human proliferating myoblasts showed a high expression of desmin filaments, confirming their myogenic origin, and this expression was further maintained following cell differentiation. Some of the myoblasts also expressed MYH, although the signal intensity was lower compared to myocytes, most likely due to a more diffused distribution. On the contrary, ACTN was present only in the myocytes. Surprisingly, we observed actinin-positive myoblasts, which were most likely correlated with changes in the cell shape and spontaneous differentiation. The fraction of actinin-positive cells within the myoblast cell population was negligible.

Bottom Line: Additionally, almost all of the chromosomal centromeres in the differentiated cells preferentially localised near the nuclear periphery when compared to the undifferentiated cells.The exceptions were chromosomes 7 and 11, in which we were unable to confirm the centromere repositioning.Based on these data we can conclude that the myogenic differentiation with global gene expression changes is accompanied by the spatial repositioning of chromosomes and chromatin remodelling, which are important processes that regulate cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Reproductive Biology and Stem Cells, Institute of Human Genetics, Polish Academy of Sciences, Poznan, Poland.

ABSTRACT
Cell differentiation is based on a synchronised orchestra of complex pathways of intrinsic and extrinsic signals that manifest in the induced expression of specific transcription factors and pivotal genes within the nucleus. One cannot ignore the epigenetic status of differentiating cells, comprising not only histones and DNA modifications but also the spatial and temporal intranuclear chromatin organisation, which is an important regulator of nuclear processes. In the present study, we investigated the nuclear architecture of human primary myoblasts and myocytes in an in vitro culture, with reference to global changes in genomic expression. Repositioning of the chromosomal centromeres, along with alterations in the nuclear shape and volume, was observed as a consequence of myotube formation. Moreover, the microarray data showed that during in vitro myogenesis cells tend to silence rather than induce gene expression. The creation of a chromosome map marked with gene expression changes that were at least 2-fold confirmed the observation. Additionally, almost all of the chromosomal centromeres in the differentiated cells preferentially localised near the nuclear periphery when compared to the undifferentiated cells. The exceptions were chromosomes 7 and 11, in which we were unable to confirm the centromere repositioning. In our opinion, this is the first reported observation of the movement of chromosomal centromeres along differentiating myogenic cells. Based on these data we can conclude that the myogenic differentiation with global gene expression changes is accompanied by the spatial repositioning of chromosomes and chromatin remodelling, which are important processes that regulate cell differentiation.

Show MeSH
Related in: MedlinePlus