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Intermittent high glucose implements stress-induced senescence in human vascular endothelial cells: role of superoxide production by NADPH oxidase.

Maeda M, Hayashi T, Mizuno N, Hattori Y, Kuzuya M - PLoS ONE (2015)

Bottom Line: Impaired glucose tolerance characterized by postprandial hyperglycemia, which occurs frequently in elderly persons and represents an important preliminary step in diabetes mellitus, poses an independent risk factor for the development of atherosclerosis.Interestingly, in intermittent high glucose, this effect was more pronounced as well as increase of p21 and p16INK4a , senescence related proteins with DNA damage.However, telomerase was not activated and telomere length was not shortened, thus stress-induced senescence was shown.

View Article: PubMed Central - PubMed

Affiliation: Department of Geriatrics, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.

ABSTRACT
Impaired glucose tolerance characterized by postprandial hyperglycemia, which occurs frequently in elderly persons and represents an important preliminary step in diabetes mellitus, poses an independent risk factor for the development of atherosclerosis. Endothelial cellular senescence is reported to precede atherosclerosis. We reported that continuous high glucose stimulus causes endothelial senescence more markedly than hypertension or dyslipidemia stimulus. In the present study, we evaluated the effect of fluctuating glucose levels on human endothelial senescence. Constant high glucose increased senescence-associated-β-galactosidase (SA-β-gal) activity, a widely used marker for cellular senescence. Interestingly, in intermittent high glucose, this effect was more pronounced as well as increase of p21 and p16INK4a , senescence related proteins with DNA damage. However, telomerase was not activated and telomere length was not shortened, thus stress-induced senescence was shown. However, constant high glucose activated telomerase and shortened telomere length, which suggested replicative senescence. Intermittent but not constant high glucose strikingly up-regulated the expression of p22phox, an NADPH oxidase component, increasing superoxide. The small interfering RNA of p22phox undermined the increase in SA-β-gal activity induced by intermittent high glucose. Conclusively, intermittent high glucose can promote vascular endothelial senescence more than constant high glucose, which is in partially dependent on superoxide overproduction.

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Research Design.The endothelial cells were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM:NG); (2) constant high glucose medium (22 mM: HG); and (3) alternating normal and high glucose media every 12 h (N/HG).
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pone.0123169.g001: Research Design.The endothelial cells were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM:NG); (2) constant high glucose medium (22 mM: HG); and (3) alternating normal and high glucose media every 12 h (N/HG).

Mentions: Human umbilical venous endothelial cells (HUVECs) were purchased from Lonza (Walkersville, MD, USA) and were cultured in endothelial cell growth medium-2 until the start of the experiment. To exclude the insulin effect [6], the cells were cultured in modified endothelial cell growth medium-2 that lacked insulin-like growth factor-1 but contained 2% fetal bovine serum during the experimental period. According to our previous study [3], five- to seven-passage subconfluent cells were used in the experiments. The cells were harvested at subconfluence and were seeded into six-well plates. Subsequently, they were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM); (2) constant high glucose medium (22 mM); and (3) alternating normal and high glucose media every 12 h (Fig 1). In the constant normal or high glucose group, each fresh medium was provided every 24 h. Mannitol was used to rule out the effect of osmotic pressure [6]. In measurements of telomere length, we continued the culture with every 3 days changing the tissue culture medium up to 28 days.


Intermittent high glucose implements stress-induced senescence in human vascular endothelial cells: role of superoxide production by NADPH oxidase.

Maeda M, Hayashi T, Mizuno N, Hattori Y, Kuzuya M - PLoS ONE (2015)

Research Design.The endothelial cells were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM:NG); (2) constant high glucose medium (22 mM: HG); and (3) alternating normal and high glucose media every 12 h (N/HG).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400006&req=5

pone.0123169.g001: Research Design.The endothelial cells were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM:NG); (2) constant high glucose medium (22 mM: HG); and (3) alternating normal and high glucose media every 12 h (N/HG).
Mentions: Human umbilical venous endothelial cells (HUVECs) were purchased from Lonza (Walkersville, MD, USA) and were cultured in endothelial cell growth medium-2 until the start of the experiment. To exclude the insulin effect [6], the cells were cultured in modified endothelial cell growth medium-2 that lacked insulin-like growth factor-1 but contained 2% fetal bovine serum during the experimental period. According to our previous study [3], five- to seven-passage subconfluent cells were used in the experiments. The cells were harvested at subconfluence and were seeded into six-well plates. Subsequently, they were exposed to the experimental condition for 3 days. They were grouped as follows: (1) constant normal glucose medium (5.5 mM); (2) constant high glucose medium (22 mM); and (3) alternating normal and high glucose media every 12 h (Fig 1). In the constant normal or high glucose group, each fresh medium was provided every 24 h. Mannitol was used to rule out the effect of osmotic pressure [6]. In measurements of telomere length, we continued the culture with every 3 days changing the tissue culture medium up to 28 days.

Bottom Line: Impaired glucose tolerance characterized by postprandial hyperglycemia, which occurs frequently in elderly persons and represents an important preliminary step in diabetes mellitus, poses an independent risk factor for the development of atherosclerosis.Interestingly, in intermittent high glucose, this effect was more pronounced as well as increase of p21 and p16INK4a , senescence related proteins with DNA damage.However, telomerase was not activated and telomere length was not shortened, thus stress-induced senescence was shown.

View Article: PubMed Central - PubMed

Affiliation: Department of Geriatrics, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.

ABSTRACT
Impaired glucose tolerance characterized by postprandial hyperglycemia, which occurs frequently in elderly persons and represents an important preliminary step in diabetes mellitus, poses an independent risk factor for the development of atherosclerosis. Endothelial cellular senescence is reported to precede atherosclerosis. We reported that continuous high glucose stimulus causes endothelial senescence more markedly than hypertension or dyslipidemia stimulus. In the present study, we evaluated the effect of fluctuating glucose levels on human endothelial senescence. Constant high glucose increased senescence-associated-β-galactosidase (SA-β-gal) activity, a widely used marker for cellular senescence. Interestingly, in intermittent high glucose, this effect was more pronounced as well as increase of p21 and p16INK4a , senescence related proteins with DNA damage. However, telomerase was not activated and telomere length was not shortened, thus stress-induced senescence was shown. However, constant high glucose activated telomerase and shortened telomere length, which suggested replicative senescence. Intermittent but not constant high glucose strikingly up-regulated the expression of p22phox, an NADPH oxidase component, increasing superoxide. The small interfering RNA of p22phox undermined the increase in SA-β-gal activity induced by intermittent high glucose. Conclusively, intermittent high glucose can promote vascular endothelial senescence more than constant high glucose, which is in partially dependent on superoxide overproduction.

Show MeSH
Related in: MedlinePlus