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Redox proteomics identification of oxidatively modified myocardial proteins in human heart failure: implications for protein function.

Brioschi M, Polvani G, Fratto P, Parolari A, Agostoni P, Tremoli E, Banfi C - PLoS ONE (2012)

Bottom Line: The levels of carbonylated proteins were significantly higher in the HF patients than in the controls (p<0.01).Exposure of cardiomyocytes to angiotensin II and norepinephrine led to ROS generation and M-CK carbonylation with loss of its enzymatic activity.Our findings indicate that protein carbonylation is increased in the myocardium during HF and that these oxidative changes may help to explain the decreased CK activity and consequent defects in energy metabolism observed in HF.

View Article: PubMed Central - PubMed

Affiliation: Centro Cardiologico Monzino IRCCS, Milan, Italy.

ABSTRACT
Increased oxidative stress in a failing heart may contribute to the pathogenesis of heart failure (HF). The aim of this study was to identify the oxidised proteins in the myocardium of HF patients and analyse the consequences of oxidation on protein function. The carbonylated proteins in left ventricular tissue from failing (n = 14) and non-failing human hearts (n = 13) were measured by immunoassay and identified by proteomics. HL-1 cardiomyocytes were incubated in the presence of stimuli relevant for HF in order to assess the generation of reactive oxygen species (ROS), the induction of protein carbonylation, and its consequences on protein function. The levels of carbonylated proteins were significantly higher in the HF patients than in the controls (p<0.01). We identified two proteins that mainly underwent carbonylation: M-type creatine kinase (M-CK), whose activity is impaired, and, to a lesser extent, α-cardiac actin. Exposure of cardiomyocytes to angiotensin II and norepinephrine led to ROS generation and M-CK carbonylation with loss of its enzymatic activity. Our findings indicate that protein carbonylation is increased in the myocardium during HF and that these oxidative changes may help to explain the decreased CK activity and consequent defects in energy metabolism observed in HF.

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Intracellular generation of ROS in cardiomyocytes exposed to various compounds.(A) Relative DCF fluorescence of ROS generation. (B) Percentage increase in ROS generation vs control cells after 5 min. *p<0.01 vs control cells; n = 5. PE, phenylephrine; ISO, isoprotenerol; NE, norepinephrine; AngII, angiotensin II; ET-1, endothelin-1, TNFα, tumor necrosis factor α.
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pone-0035841-g003: Intracellular generation of ROS in cardiomyocytes exposed to various compounds.(A) Relative DCF fluorescence of ROS generation. (B) Percentage increase in ROS generation vs control cells after 5 min. *p<0.01 vs control cells; n = 5. PE, phenylephrine; ISO, isoprotenerol; NE, norepinephrine; AngII, angiotensin II; ET-1, endothelin-1, TNFα, tumor necrosis factor α.

Mentions: In order to identify the factors responsible for the increased formation of carbonylated proteins, in vitro cultured HL-1 cardiomyocytes were exposed to the β1 and β2 agonist isoprotenerol (0.1–1 µmol/L), the α1 agonist phenylephrine (10–100 µmol/L), the α1, α2 and β1 agonist norepinephrine (10–100 µmol/L), angiotensin II (10–100 nmol/L), endothelin-1 (10–100 nmol/L), and TNFα (1–10 ng/mL). Intracellular H2O2 formation measured on the basis of the intensity of DCF emission was monitored for 1 h. All of the tested compounds induced the rapid intracellular generation of H2O2, which subsequently gradually decreased to baseline levels (Fig. 3).


Redox proteomics identification of oxidatively modified myocardial proteins in human heart failure: implications for protein function.

Brioschi M, Polvani G, Fratto P, Parolari A, Agostoni P, Tremoli E, Banfi C - PLoS ONE (2012)

Intracellular generation of ROS in cardiomyocytes exposed to various compounds.(A) Relative DCF fluorescence of ROS generation. (B) Percentage increase in ROS generation vs control cells after 5 min. *p<0.01 vs control cells; n = 5. PE, phenylephrine; ISO, isoprotenerol; NE, norepinephrine; AngII, angiotensin II; ET-1, endothelin-1, TNFα, tumor necrosis factor α.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3351458&req=5

pone-0035841-g003: Intracellular generation of ROS in cardiomyocytes exposed to various compounds.(A) Relative DCF fluorescence of ROS generation. (B) Percentage increase in ROS generation vs control cells after 5 min. *p<0.01 vs control cells; n = 5. PE, phenylephrine; ISO, isoprotenerol; NE, norepinephrine; AngII, angiotensin II; ET-1, endothelin-1, TNFα, tumor necrosis factor α.
Mentions: In order to identify the factors responsible for the increased formation of carbonylated proteins, in vitro cultured HL-1 cardiomyocytes were exposed to the β1 and β2 agonist isoprotenerol (0.1–1 µmol/L), the α1 agonist phenylephrine (10–100 µmol/L), the α1, α2 and β1 agonist norepinephrine (10–100 µmol/L), angiotensin II (10–100 nmol/L), endothelin-1 (10–100 nmol/L), and TNFα (1–10 ng/mL). Intracellular H2O2 formation measured on the basis of the intensity of DCF emission was monitored for 1 h. All of the tested compounds induced the rapid intracellular generation of H2O2, which subsequently gradually decreased to baseline levels (Fig. 3).

Bottom Line: The levels of carbonylated proteins were significantly higher in the HF patients than in the controls (p<0.01).Exposure of cardiomyocytes to angiotensin II and norepinephrine led to ROS generation and M-CK carbonylation with loss of its enzymatic activity.Our findings indicate that protein carbonylation is increased in the myocardium during HF and that these oxidative changes may help to explain the decreased CK activity and consequent defects in energy metabolism observed in HF.

View Article: PubMed Central - PubMed

Affiliation: Centro Cardiologico Monzino IRCCS, Milan, Italy.

ABSTRACT
Increased oxidative stress in a failing heart may contribute to the pathogenesis of heart failure (HF). The aim of this study was to identify the oxidised proteins in the myocardium of HF patients and analyse the consequences of oxidation on protein function. The carbonylated proteins in left ventricular tissue from failing (n = 14) and non-failing human hearts (n = 13) were measured by immunoassay and identified by proteomics. HL-1 cardiomyocytes were incubated in the presence of stimuli relevant for HF in order to assess the generation of reactive oxygen species (ROS), the induction of protein carbonylation, and its consequences on protein function. The levels of carbonylated proteins were significantly higher in the HF patients than in the controls (p<0.01). We identified two proteins that mainly underwent carbonylation: M-type creatine kinase (M-CK), whose activity is impaired, and, to a lesser extent, α-cardiac actin. Exposure of cardiomyocytes to angiotensin II and norepinephrine led to ROS generation and M-CK carbonylation with loss of its enzymatic activity. Our findings indicate that protein carbonylation is increased in the myocardium during HF and that these oxidative changes may help to explain the decreased CK activity and consequent defects in energy metabolism observed in HF.

Show MeSH
Related in: MedlinePlus