2,3-dehydrosilybin (DHS) is a minor component of silymarin, Silybum marianum seed extract, used in some dietary supplements. One of the most promising activities of this compound is its anticancer and cardioprotective activity that results, at least partially, from its cytoprotective, antioxidant, and chemopreventive properties. The present study investigated the cardioprotective effects of DHS in myocardial ischemia and reperfusion injury in rats. Isolated hearts were perfused by the Langendorff technique with low dose DHS (100 nM) prior to 30 min of ischemia induced by coronary artery occlusion. After 60 min of coronary reperfusion infarct size was determined by triphenyltetrazolium staining, while lactatedehydrogenase activity was evaluated in perfusate samples collected at several timepoints during the entire perfusion procedure. Signalosomes were isolated from a heart tissue after reperfusion and involved signalling proteins were detected. DHS reduced the extent of infarction compared with untreated control hearts at low concentration; infarct size as proportion of ischemic risk zone was 7.47 ± 3.1% for DHS versus 75.3 ± 4.8% for ischemia. This protective effect was comparable to infarct limitation induced by ischemic preconditioning (22.3 ± 4.5%). Selective inhibition of Src-family kinases with PP2 (4-Amino-3-(4-chlorophenyl)-1-(t-butyl)-1H-pyrazolo[3,4-d]pyrimidine) abrogated the protection afforded by DHS. This study provides experimental evidence that DHS can mediate Src-kinase-dependent cardioprotection against myocardial damage produced by ischemia/reperfusion injury.
- MeSH
- infarkt myokardu farmakoterapie MeSH
- kardiotonika farmakologie MeSH
- krysa rodu rattus MeSH
- potkani Wistar MeSH
- reperfuzní poškození farmakoterapie MeSH
- silymarin farmakologie MeSH
- srdce účinky léků MeSH
- techniky in vitro MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
2,3-dehydrosilybin (DHS) is a minor flavonolignan component of Silybum marianum seed extract known for its hepatoprotective activity. Recently we identified DHS as a potentially cardioprotective substance during hypoxia/reoxygenation in isolated neonatal rat cardiomyocytes. This is the first report of positive inotropic effect of DHS on perfused adult rat heart. When applied to perfused adult rat heart, DHS caused a dose-dependent inotropic effect resembling that of catecholamines. The effect was apparent with DHS concentration as low as 10 nM. Suspecting direct interaction with β-adrenergic receptors, we tested whether DHS can trigger β agonist-dependent gene transcription in a model cell line. While DHS alone was unable to trigger β agonist-dependent gene transcription, it enhanced the effect of isoproterenol, a known unspecific β agonist. Further tests confirmed that DHS could not induce cAMP accumulation in isolated neonatal rat cardiomyocytes even though high concentrations (≥ 10 μM) of DHS were capable of decreasing phosphodiesterase activity. Pre-treatment of rats with reserpine, an indole alkaloid which depletes catecholamines from peripheral sympathetic nerve endings, abolished the DHS inotropic effect in perfused hearts. Our data suggest that DHS causes the inotropic effect without acting as a β agonist. Hence we identify DHS as a novel inotropic agent.
- MeSH
- buněčné linie MeSH
- kardiomyocyty fyziologie MeSH
- kardiotonika farmakologie MeSH
- kontrakce myokardu účinky léků MeSH
- krysa rodu rattus MeSH
- ostropestřec mariánský MeSH
- potkani Wistar MeSH
- reserpin farmakologie MeSH
- rostlinné přípravky farmakologie MeSH
- silymarin farmakologie MeSH
- srdce účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Ischemic postconditioning and remote conditioning are potentially useful tools for protecting ischemic myocardium. This study tested the hypothesis that 2,3-dehydrosilybin (DHS), a flavonolignan component of Silybum marianum, could attenuate cardiomyocyte damage following hypoxia/reoxygenation by decreasing the generation of reactive oxygen species (ROS). After 5-6 days of cell culture in normoxic conditions the rat neonatal cardiomyocytes were divided into four groups. Control group (9 h at normoxic conditions), hypoxia/reoxygenation group (3 h at 1 % O₂, 94 % N₂and 5 % CO₂followed by 10 min of 10 micromol·l⁻¹DHS and 6 h of reoxygenation in normoxia) and postconditioning group (3 h of hypoxia, three cycles of 5 min reoxygenation and 5 min hypoxia followed by 6 h of normoxia). Cell viability assessed by propidium iodide staining was decreased after DHS treatment consistent with increased levels of lactatedehydrogenase (LDH) after reoxygenation. LDH leakage was significantly reduced when cardiomyocytes in the H/Re group were exposed to DHS. DHS treatment reduced H₂O₂production and also decreased the generation of ROS in the H/Re group as evidenced by a fluorescence indicator. DHS treatment reduces reoxygenation-induced injury in cardiomyocytes by attenuation of ROS generation, H₂O₂and protein carbonyls levels. In addition, we found that both the postconditioning protocol and the DHS treatment are associated with restored ratio of phosphorylated/total protein kinase C epsilon, relative to the H/Re group. In conclusion, our data support the protective role of DHS in hypoxia/reperfusion injury and indicate that DHS may act as a postconditioning mimic.
- MeSH
- antioxidancia farmakologie MeSH
- cytoprotekce MeSH
- fosforylace MeSH
- ischemické přivykání MeSH
- karbonylace proteinů účinky léků MeSH
- kardiomyocyty účinky léků metabolismus patologie MeSH
- kultivované buňky MeSH
- L-laktátdehydrogenasa metabolismus MeSH
- novorozená zvířata MeSH
- oxidační stres účinky léků MeSH
- peroxid vodíku metabolismus MeSH
- potkani Wistar MeSH
- proteinkinasa C-epsilon metabolismus MeSH
- reperfuzní poškození myokardu metabolismus patologie prevence a kontrola MeSH
- signální transdukce účinky léků MeSH
- silymarin farmakologie MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Reactive oxygen species (ROS) originating from mitochondria are perceived as a factor contributing to cell aging and means have been sought to attenuate ROS formation with the aim of extending the cell lifespan. Silybin and dehydrosilybin, two polyphenolic compounds, display a plethora of biological effects generally ascribed to their known antioxidant capacity. When investigating the cytoprotective effects of these two compounds in the primary cell cultures of neonatal rat cardiomyocytes, we noted the ability of dehydrosilybin to de-energize the cells by monitoring JC-1 fluorescence. Experiments evaluating oxygen consumption and membrane potential revealed that dehydrosilybin uncouples the respiration of isolated rat heart mitochondria albeit with a much lower potency than synthetic uncouplers. Furthermore, dehydrosilybin revealed a very high potency in suppressing ROS formation in isolated rat heart mitochondria with IC(50) = 0.15 μM. It is far more effective than its effect in a purely chemical system generating superoxide or in cells capable of oxidative burst, where the IC(50) for dehydrosilybin exceeds 50 μM. Dehydrosilybin also attenuated ROS formation caused by rotenone in the primary cultures of neonatal rat cardiomyocytes. We infer that the apparent uncoupler-like activity of dehydrosilybin is the basis of its ROS modulation effect in neonatal rat cardiomyocytes and leads us to propose a hypothesis on natural ischemia preconditioning by dietary polyphenols.
- MeSH
- analýza rozptylu MeSH
- benzimidazoly MeSH
- fluorescenční barviva MeSH
- inhibiční koncentrace 50 MeSH
- karbocyaniny MeSH
- kardiomyocyty metabolismus MeSH
- krysa rodu rattus MeSH
- mitochondrie metabolismus MeSH
- molekulární struktura MeSH
- potkani Wistar MeSH
- reaktivní formy kyslíku metabolismus MeSH
- rotenon toxicita MeSH
- silymarin chemie farmakologie MeSH
- spotřeba kyslíku účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Background: Discovery of uncoupling protein 2 (UCP2) in 1997 and demonstration of its wide tissue expressionhas triggered an important question about controlled oxidative phosphorylation uncoupling and the physiologicalfunction of this process. Uncoupling protein 2 (UcP2) is a mitochondrial protein that can infl uence the mitochondrialmembrane potential and hence the production of reactive oxygen species by mitochondria. It is also thought to beinvolved in apoptotic signaling pathways and it has been suggested to be important in cardio- and neuroprotection.Methods and results: We examined the recent literature (2003–2007) in the MedLine database for evidence linkingp38, one of the stress-related protein kinases, with modulation of UCP2 expression in the heart. While two reportsclearly demonstrate p38 as down-regulating UcP2 expression, only circumstantial evidence exists for cardiomyocytes.Confl icting results on p38-regulated cardiomyocyte survival after ischemia leave an open venue for hypotheses on thediff erential regulation of protein expression, including UCP2. Conclusions: Reviewing the evidence connecting UCP2 and its cytoprotective activities, we propose a tissue specifi clink that may explain the variable infl uence of p38 via modulation of UCP2 expression.
- MeSH
- financování organizované MeSH
- iontové kanály MeSH
- ischemická choroba srdeční enzymologie MeSH
- kardiomyocyty metabolismus účinky léků MeSH
- kardiotonika farmakologie terapeutické užití MeSH
- lidé MeSH
- medicína založená na důkazech MeSH
- MEDLINE využití MeSH
- mitochondriální proteiny MeSH
- mitogenem aktivované proteinkinasy p38 MeSH
- oxidativní fosforylace MeSH
- reaktivní formy kyslíku chemie metabolismus MeSH
- rozpřahující látky chemie MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
Cell death-inducing DFF[DNA fragmentation factor]-like effector-a (CIDEa), may initiate apoptosis by disrupting a complex consisting of 40-kDa caspase-3-activated nuclease (DFF40/CAD) and its 45-kDa inhibitor (DFF45/ICAD). CIDEa, however, was found to be localized in mitochondria. We have performed immunodetection of CIDEa in whole cells and subcellular fractions of HeLa cells adapted for a tetracycline-inducible CIDEa expression. Using immunocytochemistry we observed redistribution, enhanced upon treatment with camptothecin or valinomycin, of CIDEa to nucleus. Similarly, CIDEa content increased in the nuclear fraction but decreased in cytosolic fraction in cells treated to initiate apoptosis. We hypothesize that CIDEa is sequestered in mitochondria while transfer of this potentially dangerous protein from mitochondria into nucleus intensifies or even initiates apoptosis.
