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Loss of the osteogenic differentiation potential during senescence is limited to bone progenitor cells and is dependent on p53.

Despars G, Carbonneau CL, Bardeau P, Coutu DL, Beauséjour CM - PLoS ONE (2013)

Bottom Line: Indeed, we show here that exposure to IR prevented the differentiation and mineralization functions of MSC, an effect we found was limited to this population as more differentiated OB-SC could still form mineralize nodules.This is in contrast to adipogenesis, which was inhibited in both IR-induced senescent MSC and 3T3-L1 pre-adipocytes.Furthermore, we demonstrate that IR-induced loss of osteogenic potential in MSC was p53-dependent, a phenotype that correlates with the inability to upregulate key osteogenic transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Centre de recherche du CHU Ste-Justine, Montréal, Québec, Canada.

ABSTRACT
DNA damage can lead to the induction of cellular senescence. In particular, we showed that exposure to ionizing radiation (IR) leads to the senescence of bone marrow-derived multipotent stromal cells (MSC) and osteoblast-like stromal cells (OB-SC), a phenotype associated with bone loss. The mechanism by which IR leads to bone dysfunction is not fully understood. One possibility involves that DNA damage-induced senescence limits the regeneration of bone progenitor cells. Another possibility entails that bone dysfunction arises from the inability of accumulating senescent cells to fulfill their physiological function. Indeed, we show here that exposure to IR prevented the differentiation and mineralization functions of MSC, an effect we found was limited to this population as more differentiated OB-SC could still form mineralize nodules. This is in contrast to adipogenesis, which was inhibited in both IR-induced senescent MSC and 3T3-L1 pre-adipocytes. Furthermore, we demonstrate that IR-induced loss of osteogenic potential in MSC was p53-dependent, a phenotype that correlates with the inability to upregulate key osteogenic transcription factors. These results are the first to demonstrate that senescence impacts osteogenesis in a cell type dependent manner and suggest that the accumulation of senescent osteoblasts is unlikely to significantly contribute to bone dysfunction in a cell autonomous manner.

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Exposure to IR abrogates adipogenesis independently of the stromal lineage potential.(A) MSC and 3T3-L1 cell populations were exposed (IR) or not (CTRL) to 10 Gy IR and one week later placed in adipogenic differentiation media. Representative photographs showing lipid accumulation stained with Oil Red O is shown for each population. (B) Quantification of lipid accumulation was determined by the extraction of Oil Red O staining and detection by spectrophotometry. (C–D) Expression of PPARγ was determined by quantitative real-time PCR using RNA extracted from control and IR-induced senescent MSC and 3T3-L1 populations cultured or not in adipogenic differentiation media. Mean ± standard error of at least three individual experiments is shown. p values were obtained by performing a Student’s t-test. *: p value < 0.05.
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pone-0073206-g002: Exposure to IR abrogates adipogenesis independently of the stromal lineage potential.(A) MSC and 3T3-L1 cell populations were exposed (IR) or not (CTRL) to 10 Gy IR and one week later placed in adipogenic differentiation media. Representative photographs showing lipid accumulation stained with Oil Red O is shown for each population. (B) Quantification of lipid accumulation was determined by the extraction of Oil Red O staining and detection by spectrophotometry. (C–D) Expression of PPARγ was determined by quantitative real-time PCR using RNA extracted from control and IR-induced senescent MSC and 3T3-L1 populations cultured or not in adipogenic differentiation media. Mean ± standard error of at least three individual experiments is shown. p values were obtained by performing a Student’s t-test. *: p value < 0.05.

Mentions: It is well known there is an increased adipocyte content in the bone marrow with age or following exposure to IR. We therefore investigated whether IR-induced senescence could skew the differentiation potential of multipotent MSC toward the adipogenic lineage. As such, control and one week post-irradiation MSC populations were cultured under adipogenic conditions and their differentiation measured. While control cultures of MSC differentiated into adipocytes, irradiated cell populations failed to do so (Figure 2A). This loss in the adipogenic potential was not unique to MSC progenitors as senescent 3T3-L1 pre-adipocytes also failed to differentiate under the same conditions (Figure 2A). Differentiation was evaluated by the ability to accumulate lipids as showed by quantification of Oil Red O staining (Figure 2B). Failure to differentiate into the adipogenic lineage was not due to the incapacity to upregulate the key adipogenic transcription factor PPARγ which we showed the expression was increased 100-1000 fold in both senescent MSC and 3T3-L1 populations cultured in adipogenic conditions. These results suggest that failure of IR-induced senescent cells to undergo adipogenic differentiation is independent of the multilineage potential and on the ability to activate PPARγ.


Loss of the osteogenic differentiation potential during senescence is limited to bone progenitor cells and is dependent on p53.

Despars G, Carbonneau CL, Bardeau P, Coutu DL, Beauséjour CM - PLoS ONE (2013)

Exposure to IR abrogates adipogenesis independently of the stromal lineage potential.(A) MSC and 3T3-L1 cell populations were exposed (IR) or not (CTRL) to 10 Gy IR and one week later placed in adipogenic differentiation media. Representative photographs showing lipid accumulation stained with Oil Red O is shown for each population. (B) Quantification of lipid accumulation was determined by the extraction of Oil Red O staining and detection by spectrophotometry. (C–D) Expression of PPARγ was determined by quantitative real-time PCR using RNA extracted from control and IR-induced senescent MSC and 3T3-L1 populations cultured or not in adipogenic differentiation media. Mean ± standard error of at least three individual experiments is shown. p values were obtained by performing a Student’s t-test. *: p value < 0.05.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3756945&req=5

pone-0073206-g002: Exposure to IR abrogates adipogenesis independently of the stromal lineage potential.(A) MSC and 3T3-L1 cell populations were exposed (IR) or not (CTRL) to 10 Gy IR and one week later placed in adipogenic differentiation media. Representative photographs showing lipid accumulation stained with Oil Red O is shown for each population. (B) Quantification of lipid accumulation was determined by the extraction of Oil Red O staining and detection by spectrophotometry. (C–D) Expression of PPARγ was determined by quantitative real-time PCR using RNA extracted from control and IR-induced senescent MSC and 3T3-L1 populations cultured or not in adipogenic differentiation media. Mean ± standard error of at least three individual experiments is shown. p values were obtained by performing a Student’s t-test. *: p value < 0.05.
Mentions: It is well known there is an increased adipocyte content in the bone marrow with age or following exposure to IR. We therefore investigated whether IR-induced senescence could skew the differentiation potential of multipotent MSC toward the adipogenic lineage. As such, control and one week post-irradiation MSC populations were cultured under adipogenic conditions and their differentiation measured. While control cultures of MSC differentiated into adipocytes, irradiated cell populations failed to do so (Figure 2A). This loss in the adipogenic potential was not unique to MSC progenitors as senescent 3T3-L1 pre-adipocytes also failed to differentiate under the same conditions (Figure 2A). Differentiation was evaluated by the ability to accumulate lipids as showed by quantification of Oil Red O staining (Figure 2B). Failure to differentiate into the adipogenic lineage was not due to the incapacity to upregulate the key adipogenic transcription factor PPARγ which we showed the expression was increased 100-1000 fold in both senescent MSC and 3T3-L1 populations cultured in adipogenic conditions. These results suggest that failure of IR-induced senescent cells to undergo adipogenic differentiation is independent of the multilineage potential and on the ability to activate PPARγ.

Bottom Line: Indeed, we show here that exposure to IR prevented the differentiation and mineralization functions of MSC, an effect we found was limited to this population as more differentiated OB-SC could still form mineralize nodules.This is in contrast to adipogenesis, which was inhibited in both IR-induced senescent MSC and 3T3-L1 pre-adipocytes.Furthermore, we demonstrate that IR-induced loss of osteogenic potential in MSC was p53-dependent, a phenotype that correlates with the inability to upregulate key osteogenic transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Centre de recherche du CHU Ste-Justine, Montréal, Québec, Canada.

ABSTRACT
DNA damage can lead to the induction of cellular senescence. In particular, we showed that exposure to ionizing radiation (IR) leads to the senescence of bone marrow-derived multipotent stromal cells (MSC) and osteoblast-like stromal cells (OB-SC), a phenotype associated with bone loss. The mechanism by which IR leads to bone dysfunction is not fully understood. One possibility involves that DNA damage-induced senescence limits the regeneration of bone progenitor cells. Another possibility entails that bone dysfunction arises from the inability of accumulating senescent cells to fulfill their physiological function. Indeed, we show here that exposure to IR prevented the differentiation and mineralization functions of MSC, an effect we found was limited to this population as more differentiated OB-SC could still form mineralize nodules. This is in contrast to adipogenesis, which was inhibited in both IR-induced senescent MSC and 3T3-L1 pre-adipocytes. Furthermore, we demonstrate that IR-induced loss of osteogenic potential in MSC was p53-dependent, a phenotype that correlates with the inability to upregulate key osteogenic transcription factors. These results are the first to demonstrate that senescence impacts osteogenesis in a cell type dependent manner and suggest that the accumulation of senescent osteoblasts is unlikely to significantly contribute to bone dysfunction in a cell autonomous manner.

Show MeSH
Related in: MedlinePlus