Limits...
Resveratrol Induced Premature Senescence Is Associated with DNA Damage Mediated SIRT1 and SIRT2 Down-Regulation.

Kilic Eren M, Kilincli A, Eren Ö - PLoS ONE (2015)

Bottom Line: Interestingly, at concentrations where resveratrol induced premature senescence we show a significant decrease in SIRT1 and SIRT2 levels by Western Blot and quantitative RT-PCR analysis.Interestingly DNA damaging agent doxorubicin also induced senescence in BJ fibroblasts associated with decreased SIRT1/2 levels.In conclusion our data reveal that resveratrol induced premature senescence is associated with SIRT1 and SIRT2 down regulation in human dermal fibroblasts.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biology, Adnan Menderes University Medical School, Aydın, Turkey; ADU-BILTEM (Adnan Menderes University, Science and Technology Research and Application Center), Aydin, Turkey.

ABSTRACT
The natural polyphenolic compound resveratrol (3,4,5-trihydroxy-trans-stilbene) has broad spectrum health beneficial activities including antioxidant, anti-inflammatory, anti-aging, anti-cancer, cardioprotective, and neuroprotective effects. Remarkably, resveratrol also induces apoptosis and cellular senescence in primary and cancer cells. Resveratrol's anti-aging effects both in vitro and in vivo attributed to activation of a (NAD)-dependent histone deacetylase family member sirtuin-1 (SIRT1) protein. In mammals seven members (SIRT1-7) of sirtuin family have been identified. Among those, SIRT1 is the most extensively studied with perceptive effects on mammalian physiology and suppression of the diseases of aging. Yet no data has specified the role of sirtuins, under conditions where resveratrol treatment induces senescence. Current study was undertaken to investigate the effects of resveratrol in human primary dermal fibroblasts (BJ) and to clarify the role of sirtuin family members in particular SIRT1 and SIRT2 that are known to be involved in cellular stress responses and cell cycle, respectively. Here, we show that resveratrol decreases proliferation of BJ cells in a time and dose dependent manner. In addition the increase in senescence associated β-galactosidase (SA-β-gal) activity and methylated H3K9-me indicate the induction of premature senescence. A significant increase in phosphorylation of γ-H2AX, a surrogate of DNA double strand breaks, as well as in levels of p53, p21CIP1 and p16INK4A is also detected. Interestingly, at concentrations where resveratrol induced premature senescence we show a significant decrease in SIRT1 and SIRT2 levels by Western Blot and quantitative RT-PCR analysis. Conversely inhibition of SIRT1 and SIRT2 via siRNA or sirtinol treatment also induced senescence in BJ fibroblasts associated with increased SA-β-gal activity, γ-H2AX phosphorylation and p53, p21CIP1 and p16INK4A levels. Interestingly DNA damaging agent doxorubicin also induced senescence in BJ fibroblasts associated with decreased SIRT1/2 levels. In conclusion our data reveal that resveratrol induced premature senescence is associated with SIRT1 and SIRT2 down regulation in human dermal fibroblasts. Here we suggest that the concomitant decline in SIRT1/2 expression in response to resveratrol treatment may be a cause for induction of senescence, which is most likely mediated by a regulatory mechanism activated by DNA damage response.

Show MeSH

Related in: MedlinePlus

Inhibition of SIRT1/2 by sirtinol induces senescence in BJ fibroblasts.BJ fibroblasts were treated with 50 and 100 μM sirtinol for three days and subsequently (A) analysed for expression of SIRT1, SIRT2, p53, p21CIP1, p16INK4A and and full length (p32) and cleaved (p20) Caspase-3 levels by WB. β-actin was used as loading control (B) stained for SA-β-galactivity and γ-H2AX foci formation. Dapi was used to counterstain nuclei.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4414559&req=5

pone.0124837.g009: Inhibition of SIRT1/2 by sirtinol induces senescence in BJ fibroblasts.BJ fibroblasts were treated with 50 and 100 μM sirtinol for three days and subsequently (A) analysed for expression of SIRT1, SIRT2, p53, p21CIP1, p16INK4A and and full length (p32) and cleaved (p20) Caspase-3 levels by WB. β-actin was used as loading control (B) stained for SA-β-galactivity and γ-H2AX foci formation. Dapi was used to counterstain nuclei.

Mentions: Upon finding that genetic knock down of SIRT1 /2 induces senescence we asked whether or not chemical inhibitors of sirtuin family members show similar effects. We used a well-known chemical inhibitor, namely sirtinol in order to repress SIRT1/2 activity as suggested in previous reports [6]. As shown in “Fig 9A” 100 μM sirtinol treatment induced senescence in BJ fibroblasts as evidenced by increased SA-βgal activity (Fig 9A). Consistent with previous reports [36,37] we detected a slight decrease in SIRT1/2 expressions in BJ fibroblasts in response to sirtinol treatment suggesting SIRT1/2 activity might also play a role in regulation of sirtinol induced senescence. Additionally, increased levels of p53, p21CIP1 and p16 INK4A expressions were also detected by sirtinol treatment. More importantly 100 μM of sirtinol induced γ -H2A.X foci formation indicating to the activation of DNA damage response (Fig 9B). However no cleaved caspase-3 expression was detected with 100 μM of sirtinol treatment indicating apoptosis is not induced at this concentration in BJ fibroblasts (Fig 9A).


Resveratrol Induced Premature Senescence Is Associated with DNA Damage Mediated SIRT1 and SIRT2 Down-Regulation.

Kilic Eren M, Kilincli A, Eren Ö - PLoS ONE (2015)

Inhibition of SIRT1/2 by sirtinol induces senescence in BJ fibroblasts.BJ fibroblasts were treated with 50 and 100 μM sirtinol for three days and subsequently (A) analysed for expression of SIRT1, SIRT2, p53, p21CIP1, p16INK4A and and full length (p32) and cleaved (p20) Caspase-3 levels by WB. β-actin was used as loading control (B) stained for SA-β-galactivity and γ-H2AX foci formation. Dapi was used to counterstain nuclei.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4414559&req=5

pone.0124837.g009: Inhibition of SIRT1/2 by sirtinol induces senescence in BJ fibroblasts.BJ fibroblasts were treated with 50 and 100 μM sirtinol for three days and subsequently (A) analysed for expression of SIRT1, SIRT2, p53, p21CIP1, p16INK4A and and full length (p32) and cleaved (p20) Caspase-3 levels by WB. β-actin was used as loading control (B) stained for SA-β-galactivity and γ-H2AX foci formation. Dapi was used to counterstain nuclei.
Mentions: Upon finding that genetic knock down of SIRT1 /2 induces senescence we asked whether or not chemical inhibitors of sirtuin family members show similar effects. We used a well-known chemical inhibitor, namely sirtinol in order to repress SIRT1/2 activity as suggested in previous reports [6]. As shown in “Fig 9A” 100 μM sirtinol treatment induced senescence in BJ fibroblasts as evidenced by increased SA-βgal activity (Fig 9A). Consistent with previous reports [36,37] we detected a slight decrease in SIRT1/2 expressions in BJ fibroblasts in response to sirtinol treatment suggesting SIRT1/2 activity might also play a role in regulation of sirtinol induced senescence. Additionally, increased levels of p53, p21CIP1 and p16 INK4A expressions were also detected by sirtinol treatment. More importantly 100 μM of sirtinol induced γ -H2A.X foci formation indicating to the activation of DNA damage response (Fig 9B). However no cleaved caspase-3 expression was detected with 100 μM of sirtinol treatment indicating apoptosis is not induced at this concentration in BJ fibroblasts (Fig 9A).

Bottom Line: Interestingly, at concentrations where resveratrol induced premature senescence we show a significant decrease in SIRT1 and SIRT2 levels by Western Blot and quantitative RT-PCR analysis.Interestingly DNA damaging agent doxorubicin also induced senescence in BJ fibroblasts associated with decreased SIRT1/2 levels.In conclusion our data reveal that resveratrol induced premature senescence is associated with SIRT1 and SIRT2 down regulation in human dermal fibroblasts.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biology, Adnan Menderes University Medical School, Aydın, Turkey; ADU-BILTEM (Adnan Menderes University, Science and Technology Research and Application Center), Aydin, Turkey.

ABSTRACT
The natural polyphenolic compound resveratrol (3,4,5-trihydroxy-trans-stilbene) has broad spectrum health beneficial activities including antioxidant, anti-inflammatory, anti-aging, anti-cancer, cardioprotective, and neuroprotective effects. Remarkably, resveratrol also induces apoptosis and cellular senescence in primary and cancer cells. Resveratrol's anti-aging effects both in vitro and in vivo attributed to activation of a (NAD)-dependent histone deacetylase family member sirtuin-1 (SIRT1) protein. In mammals seven members (SIRT1-7) of sirtuin family have been identified. Among those, SIRT1 is the most extensively studied with perceptive effects on mammalian physiology and suppression of the diseases of aging. Yet no data has specified the role of sirtuins, under conditions where resveratrol treatment induces senescence. Current study was undertaken to investigate the effects of resveratrol in human primary dermal fibroblasts (BJ) and to clarify the role of sirtuin family members in particular SIRT1 and SIRT2 that are known to be involved in cellular stress responses and cell cycle, respectively. Here, we show that resveratrol decreases proliferation of BJ cells in a time and dose dependent manner. In addition the increase in senescence associated β-galactosidase (SA-β-gal) activity and methylated H3K9-me indicate the induction of premature senescence. A significant increase in phosphorylation of γ-H2AX, a surrogate of DNA double strand breaks, as well as in levels of p53, p21CIP1 and p16INK4A is also detected. Interestingly, at concentrations where resveratrol induced premature senescence we show a significant decrease in SIRT1 and SIRT2 levels by Western Blot and quantitative RT-PCR analysis. Conversely inhibition of SIRT1 and SIRT2 via siRNA or sirtinol treatment also induced senescence in BJ fibroblasts associated with increased SA-β-gal activity, γ-H2AX phosphorylation and p53, p21CIP1 and p16INK4A levels. Interestingly DNA damaging agent doxorubicin also induced senescence in BJ fibroblasts associated with decreased SIRT1/2 levels. In conclusion our data reveal that resveratrol induced premature senescence is associated with SIRT1 and SIRT2 down regulation in human dermal fibroblasts. Here we suggest that the concomitant decline in SIRT1/2 expression in response to resveratrol treatment may be a cause for induction of senescence, which is most likely mediated by a regulatory mechanism activated by DNA damage response.

Show MeSH
Related in: MedlinePlus