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Decreased myocardial injury and improved contractility after administration of a peptide derived against the alpha-interacting domain of the L-type calcium channel.

Viola HM, Jordan MC, Roos KP, Hool LC - J Am Heart Assoc (2014)

Bottom Line: Activation of the channel also alters mitochondrial function.In search of the mechanism for the effect, we found that intracellular calcium ([Ca(2+)]i, Fura-2), superoxide production (dihydroethidium fluorescence), mitochondrial membrane potential (Ψm, JC-1 fluorescence), reduced nicotinamide adenine dinucleotide production, and flavoprotein oxidation (autofluorescence) are decreased after application of AID-TAT peptide.Application of AID-TAT peptide significantly decreased infarct size and supported contractility up to 12 weeks postcoronary artery occlusion as a result of a decrease in metabolic demand during reperfusion.

View Article: PubMed Central - PubMed

Affiliation: School of Anatomy, Physiology and Human Biology, The University of Western Australia, Crawley, WA, Australia (H.M.V., L.C.H.).

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Effect of AID‐TAT peptide on cellular superoxide in vitro. A, Representative traces of DHE recorded in a myocyte before and after 5 minutes exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase and a myocyte before and after 5 minutes of exposure to 0 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase. Vertical arrows indicate when drugs were added. B, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), caused a 93.6±1.9% decrease in the rate of DHE signal. C, Representative traces of DHE recorded in myocytes before and after 5 minutes of exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase in the presence of either 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide as indicated. Vertical arrow indicates when drugs were added. D, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: Mean±SEM of changes in DHE for myocytes before and 30 and 60 minutes after exposure to 10 μmol/L of AID‐TAT peptide as indicated. Statistical significance was determined using the Kruskal‐Wallis test followed by Dunn's multiple comparison test. AID indicates alpha‐interacting domain; DHE, dihydroethidium; TAT, transactivator of transcription.
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fig04: Effect of AID‐TAT peptide on cellular superoxide in vitro. A, Representative traces of DHE recorded in a myocyte before and after 5 minutes exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase and a myocyte before and after 5 minutes of exposure to 0 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase. Vertical arrows indicate when drugs were added. B, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), caused a 93.6±1.9% decrease in the rate of DHE signal. C, Representative traces of DHE recorded in myocytes before and after 5 minutes of exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase in the presence of either 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide as indicated. Vertical arrow indicates when drugs were added. D, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: Mean±SEM of changes in DHE for myocytes before and 30 and 60 minutes after exposure to 10 μmol/L of AID‐TAT peptide as indicated. Statistical significance was determined using the Kruskal‐Wallis test followed by Dunn's multiple comparison test. AID indicates alpha‐interacting domain; DHE, dihydroethidium; TAT, transactivator of transcription.

Mentions: We have shown previously that 5 minutes of exposure of myocytes to H2O2 results in a significant further increase in superoxide assessed as changes in dihydroethidium (DHE) fluorescence in guinea pig ventricular myocytes.10,30 The further increase in superoxide occurred as a result of increased calcium uptake into mitochondria, increased reduction of NAD+, and increased electron flow into mitochondrial complex II.30 We assessed the effect of AID‐TAT peptide on DHE fluorescence in ventricular myocytes from C57BL mice. Application of 40 μmol/L of H2O2 resulted in 70.3±25.1% further increase in DHE signal that was completely attenuated with nisoldipine (Figure 4A and 4B). Application of the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), completely abolished DHE signal, confirming specificity of the indicator for superoxide (Figure 4B, inset right). Application of 1 μmol/L of AID‐TAT peptide had no effect on the H2O2‐induced increase in DHE signal, but 10 μmol/L of AID‐TAT peptide significantly attenuated the increase in DHE signal (Figure 4C and 4D). AID‐TAT peptide had no effect on the DHE signal in the absence of H2O2, indicating that basal production of superoxide was unaffected by the peptide (Figure 4D, inset right).


Decreased myocardial injury and improved contractility after administration of a peptide derived against the alpha-interacting domain of the L-type calcium channel.

Viola HM, Jordan MC, Roos KP, Hool LC - J Am Heart Assoc (2014)

Effect of AID‐TAT peptide on cellular superoxide in vitro. A, Representative traces of DHE recorded in a myocyte before and after 5 minutes exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase and a myocyte before and after 5 minutes of exposure to 0 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase. Vertical arrows indicate when drugs were added. B, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), caused a 93.6±1.9% decrease in the rate of DHE signal. C, Representative traces of DHE recorded in myocytes before and after 5 minutes of exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase in the presence of either 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide as indicated. Vertical arrow indicates when drugs were added. D, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: Mean±SEM of changes in DHE for myocytes before and 30 and 60 minutes after exposure to 10 μmol/L of AID‐TAT peptide as indicated. Statistical significance was determined using the Kruskal‐Wallis test followed by Dunn's multiple comparison test. AID indicates alpha‐interacting domain; DHE, dihydroethidium; TAT, transactivator of transcription.
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fig04: Effect of AID‐TAT peptide on cellular superoxide in vitro. A, Representative traces of DHE recorded in a myocyte before and after 5 minutes exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase and a myocyte before and after 5 minutes of exposure to 0 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase. Vertical arrows indicate when drugs were added. B, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), caused a 93.6±1.9% decrease in the rate of DHE signal. C, Representative traces of DHE recorded in myocytes before and after 5 minutes of exposure to 40 μmol/L of H2O2 followed by 5 minutes of exposure to 10 U/mL of catalase in the presence of either 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide as indicated. Vertical arrow indicates when drugs were added. D, Mean±SEM of changes in DHE for all myocytes exposed to 40 μmol/L of H2O2, 10 U/mL of catalase, 1 or 10 μmol/L of AID(S)‐TAT or AID‐TAT peptide, and 10 μmol/L of nisoldipine (Nisol) as indicated. Inset above: Mean±SEM of changes in DHE for myocytes before and 30 and 60 minutes after exposure to 10 μmol/L of AID‐TAT peptide as indicated. Statistical significance was determined using the Kruskal‐Wallis test followed by Dunn's multiple comparison test. AID indicates alpha‐interacting domain; DHE, dihydroethidium; TAT, transactivator of transcription.
Mentions: We have shown previously that 5 minutes of exposure of myocytes to H2O2 results in a significant further increase in superoxide assessed as changes in dihydroethidium (DHE) fluorescence in guinea pig ventricular myocytes.10,30 The further increase in superoxide occurred as a result of increased calcium uptake into mitochondria, increased reduction of NAD+, and increased electron flow into mitochondrial complex II.30 We assessed the effect of AID‐TAT peptide on DHE fluorescence in ventricular myocytes from C57BL mice. Application of 40 μmol/L of H2O2 resulted in 70.3±25.1% further increase in DHE signal that was completely attenuated with nisoldipine (Figure 4A and 4B). Application of the superoxide scavenger, N‐tert‐butyl‐α‐phenyl‐nitrone (PBN), completely abolished DHE signal, confirming specificity of the indicator for superoxide (Figure 4B, inset right). Application of 1 μmol/L of AID‐TAT peptide had no effect on the H2O2‐induced increase in DHE signal, but 10 μmol/L of AID‐TAT peptide significantly attenuated the increase in DHE signal (Figure 4C and 4D). AID‐TAT peptide had no effect on the DHE signal in the absence of H2O2, indicating that basal production of superoxide was unaffected by the peptide (Figure 4D, inset right).

Bottom Line: Activation of the channel also alters mitochondrial function.In search of the mechanism for the effect, we found that intracellular calcium ([Ca(2+)]i, Fura-2), superoxide production (dihydroethidium fluorescence), mitochondrial membrane potential (Ψm, JC-1 fluorescence), reduced nicotinamide adenine dinucleotide production, and flavoprotein oxidation (autofluorescence) are decreased after application of AID-TAT peptide.Application of AID-TAT peptide significantly decreased infarct size and supported contractility up to 12 weeks postcoronary artery occlusion as a result of a decrease in metabolic demand during reperfusion.

View Article: PubMed Central - PubMed

Affiliation: School of Anatomy, Physiology and Human Biology, The University of Western Australia, Crawley, WA, Australia (H.M.V., L.C.H.).

Show MeSH
Related in: MedlinePlus