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Protection by the NO-Donor SNAP and BNP against Hypoxia/Reoxygenation in Rat Engineered Heart Tissue.

Görbe A, Eder A, Varga ZV, Pálóczi J, Hansen A, Ferdinandy P, Eschenhagen T - PLoS ONE (2015)

Bottom Line: H/R was accompanied by a small increase in LDH and non-significant increase in cTnI.SNAP and BNP showed small but significant protective effects during reoxygenation.The sensitivity of the model needs improvement.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Group, Department of Biochemistry, University of Szeged, Szeged, Hungary; Pharmahungary Group, Szeged, Hungary.

ABSTRACT
In vitro assays could replace animal experiments in drug screening and disease modeling, but have shortcomings in terms of functional readout. Force-generating engineered heart tissues (EHT) provide simple automated measurements of contractile function. Here we evaluated the response of EHTs to hypoxia/reoxygenation (H/R) and the effect of known cardiocytoprotective molecules. EHTs from neonatal rat heart cells were incubated for 24 h in EHT medium. Then they were subjected to 180 min hypoxia (93% N2, 7% CO2) and 120 min reoxygenation (40% O2, 53% N2, 7% CO2), change of medium and additional follow-up of 48 h. Time-matched controls (40% O2, 53% N2, 7% CO2) were run for comparison. The following conditions were applied during H/R: fresh EHT medium (positive control), the NO-donor S-nitroso-N-acetyl-D,L-penicillamine (SNAP, 10(-7), 10(-6), 10(-5) M) or the guanylate cyclase activator brain type natriuretic peptide (BNP, 10(-9), 10(-8), 10(-7) M). Frequency and force of contraction were repeatedly monitored over the entire experiment, pH, troponin I (cTnI), lactate dehydrogenase (LDH) and glucose concentrations measured in EHT medium. Beating activity of EHTs in 24 h-medium ceased during hypoxia, partially recovered during reoxygenation and reached time-control values during follow-up. H/R was accompanied by a small increase in LDH and non-significant increase in cTnI. In fresh medium, some EHTs continued beating during hypoxia and all EHTs recovered faster during reoxygenation. SNAP and BNP showed small but significant protective effects during reoxygenation. EHTs are applicable to test potential cardioprotective compounds in vitro, monitoring functional and biochemical endpoints, which otherwise could be only measured by using in vivo or ex vivo heart preparations. The sensitivity of the model needs improvement.

No MeSH data available.


Related in: MedlinePlus

Comparison of EHT contractile behavior in 24 h medium control (24 h MC) and SNAP treated groups (n = 6).(A) Beating pattern of EHTs. Grey boxes represent beating periods of EHTs whereas black boxes indicate non-beating phases. (B) Total number of beats is expressed as the percentage of baseline. (C) Rate force product of EHTs during reoxygenation. (D) Histological parameters of EHT sections. (E-F) cTnI, LDH release and glucose consumption are shown respectively. Data are expressed as mean ± SEM; **p<0.001 one-way ANOVA, followed by Dunett’s post hoc test, n = 6.
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pone.0132186.g004: Comparison of EHT contractile behavior in 24 h medium control (24 h MC) and SNAP treated groups (n = 6).(A) Beating pattern of EHTs. Grey boxes represent beating periods of EHTs whereas black boxes indicate non-beating phases. (B) Total number of beats is expressed as the percentage of baseline. (C) Rate force product of EHTs during reoxygenation. (D) Histological parameters of EHT sections. (E-F) cTnI, LDH release and glucose consumption are shown respectively. Data are expressed as mean ± SEM; **p<0.001 one-way ANOVA, followed by Dunett’s post hoc test, n = 6.

Mentions: The NO-donor SNAP or the B-type natriuretic peptide (BNP) were administered at 3 different concentrations to 24 h medium group before hypoxia/reoxygenation. Both had small, not clearly concentration-dependent effects on beating behavior of EHTs. None of the compounds caused significant protective effect during hypoxic phase (S2 and S3 Figs).In the presence of SNAP (10−7 M) more EHTs tended to beat during reoxygenation than in 24 h MC group (p = 0.06; Fig 4A). Beating activity (total number of beats; Fig 4B) was higher under SNAP 10−7 and 10−6 M, whereas the rate-force product during first stage of reoxygenation was not significantly changed, but during second stage of reoxygenation it was significantly higher under 10−7 and 10−5 M (Fig 4C). SNAP did not affect the N:C ratio (Fig 4D and S1 Fig) or biochemical markers (Fig 4E and 4F).


Protection by the NO-Donor SNAP and BNP against Hypoxia/Reoxygenation in Rat Engineered Heart Tissue.

Görbe A, Eder A, Varga ZV, Pálóczi J, Hansen A, Ferdinandy P, Eschenhagen T - PLoS ONE (2015)

Comparison of EHT contractile behavior in 24 h medium control (24 h MC) and SNAP treated groups (n = 6).(A) Beating pattern of EHTs. Grey boxes represent beating periods of EHTs whereas black boxes indicate non-beating phases. (B) Total number of beats is expressed as the percentage of baseline. (C) Rate force product of EHTs during reoxygenation. (D) Histological parameters of EHT sections. (E-F) cTnI, LDH release and glucose consumption are shown respectively. Data are expressed as mean ± SEM; **p<0.001 one-way ANOVA, followed by Dunett’s post hoc test, n = 6.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492769&req=5

pone.0132186.g004: Comparison of EHT contractile behavior in 24 h medium control (24 h MC) and SNAP treated groups (n = 6).(A) Beating pattern of EHTs. Grey boxes represent beating periods of EHTs whereas black boxes indicate non-beating phases. (B) Total number of beats is expressed as the percentage of baseline. (C) Rate force product of EHTs during reoxygenation. (D) Histological parameters of EHT sections. (E-F) cTnI, LDH release and glucose consumption are shown respectively. Data are expressed as mean ± SEM; **p<0.001 one-way ANOVA, followed by Dunett’s post hoc test, n = 6.
Mentions: The NO-donor SNAP or the B-type natriuretic peptide (BNP) were administered at 3 different concentrations to 24 h medium group before hypoxia/reoxygenation. Both had small, not clearly concentration-dependent effects on beating behavior of EHTs. None of the compounds caused significant protective effect during hypoxic phase (S2 and S3 Figs).In the presence of SNAP (10−7 M) more EHTs tended to beat during reoxygenation than in 24 h MC group (p = 0.06; Fig 4A). Beating activity (total number of beats; Fig 4B) was higher under SNAP 10−7 and 10−6 M, whereas the rate-force product during first stage of reoxygenation was not significantly changed, but during second stage of reoxygenation it was significantly higher under 10−7 and 10−5 M (Fig 4C). SNAP did not affect the N:C ratio (Fig 4D and S1 Fig) or biochemical markers (Fig 4E and 4F).

Bottom Line: H/R was accompanied by a small increase in LDH and non-significant increase in cTnI.SNAP and BNP showed small but significant protective effects during reoxygenation.The sensitivity of the model needs improvement.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Group, Department of Biochemistry, University of Szeged, Szeged, Hungary; Pharmahungary Group, Szeged, Hungary.

ABSTRACT
In vitro assays could replace animal experiments in drug screening and disease modeling, but have shortcomings in terms of functional readout. Force-generating engineered heart tissues (EHT) provide simple automated measurements of contractile function. Here we evaluated the response of EHTs to hypoxia/reoxygenation (H/R) and the effect of known cardiocytoprotective molecules. EHTs from neonatal rat heart cells were incubated for 24 h in EHT medium. Then they were subjected to 180 min hypoxia (93% N2, 7% CO2) and 120 min reoxygenation (40% O2, 53% N2, 7% CO2), change of medium and additional follow-up of 48 h. Time-matched controls (40% O2, 53% N2, 7% CO2) were run for comparison. The following conditions were applied during H/R: fresh EHT medium (positive control), the NO-donor S-nitroso-N-acetyl-D,L-penicillamine (SNAP, 10(-7), 10(-6), 10(-5) M) or the guanylate cyclase activator brain type natriuretic peptide (BNP, 10(-9), 10(-8), 10(-7) M). Frequency and force of contraction were repeatedly monitored over the entire experiment, pH, troponin I (cTnI), lactate dehydrogenase (LDH) and glucose concentrations measured in EHT medium. Beating activity of EHTs in 24 h-medium ceased during hypoxia, partially recovered during reoxygenation and reached time-control values during follow-up. H/R was accompanied by a small increase in LDH and non-significant increase in cTnI. In fresh medium, some EHTs continued beating during hypoxia and all EHTs recovered faster during reoxygenation. SNAP and BNP showed small but significant protective effects during reoxygenation. EHTs are applicable to test potential cardioprotective compounds in vitro, monitoring functional and biochemical endpoints, which otherwise could be only measured by using in vivo or ex vivo heart preparations. The sensitivity of the model needs improvement.

No MeSH data available.


Related in: MedlinePlus