Bakteriális lipopoliszachariddal kiváltott késői kardioprotekció az iszkémia/reperfúziós károsodással szemben = Late cardioprotection against ischemia-reperfusion injury induced by bacterial lipopolysaccharides

Hazánkban és a civilizált társadalmakban egyaránt, az iszkémiás szívbetegség az egyik leggyakoribb halálokok közé tartozik. A szivizom endogén adaptációs mechanizmusainak támogatása, illetve vizsgálata ezért nagy jelentőségű. Kísérleteink bizonyítják, hogy ez a mechanizmus emberben is hatásos védelmet jelent. Kimutattuk továbbá, hogy a farmakológiai prekondícionálás hatékonyan véd az iszkémiás károsodás ellen. A kis dózisú (0,5 mg/kg) bakteriális lipopoliszachariddal (LPS) előkezelt patkányok szíve 24 órával a kezelés után fokozottabb mértékben áll ellen az iszkémiát követő szívfunkciós károsodásnak. A 24 órás LPS előkezelés megnövelte a szívizom nitrogén-monoxid tartalmát az indukálható nitrogén oxid szintáz fokozott aktivitása révén. A LPS előkezelés hatására szignifikánsan emelkedett a szívizom szuperoxid termelése is a fokozott xantin oxidoreduktáz aktivitás miatt. A szérum nitrotirozin szintje ugyancsak emelkedett az LPS előkezelés hatására, jelezvén a NO és szuperoxid anion reakciója révén képződő peroxinitrit fokozott képződését a szervezetben. A LPS előkezelés növelte a miokardiális cGMP tartalmat is. Eredményeink szerint az oxidatív és nitrozatív stressz, valamint az NO-cGMP szignalizációs út fontos szerepet játszhat a szívizom farmakológiai prekondícionálásában. | Ischemic heart disease is one of the most frequent cause of death in the civilized societies, therefore investigation of cardioprotective mechanisms to prevent ischemic damage is essential. We have shown that this protective mechanism is effective in humans. We have also shown that pharmacological preconditioning effectively protects against ischemia/reperfuson injury. 24h pretreatment of rats with bacterial polysaccharides (LPS) has resulted in a protection against ischemia-induced deterioration of cardiac function. A 24h LPS pretreatment increased cardiac nitric oxide content via the induction of the inducible isoform of nitric oxide synthase. LPS pretreatment also increased cardiac superoxide production via an increase in the activity of xanthin oxidoreductase. LPS increased serum nitrotyrosine levels showing increased formation of peroxynitrite from NO and superoxide. In addition, LPS pretreatment increased myocardial cGMP level. Our results suggest, that both oxidative and nitrosative stress play an important role in pharmacological preconditioning of the heart

Metabolic syndrome influences cardiac gene expression pattern at the transcript level in male ZDF rats

Background: Metabolic syndrome (coexisting visceral obesity, dyslipidemia, hyperglycemia, and hypertension) is a prominent risk factor for cardiovascular morbidity and mortality, however, its effect on cardiac gene expression pattern is unclear. Therefore, we examined the possible alterations in cardiac gene expression pattern in male Zucker Diabetic Fatty (ZDF) rats, a model of metabolic syndrome. Methods: Fasting blood glucose, serum insulin, cholesterol and triglyceride levels were measured at 6, 16, and 25 wk of age in male ZDF and lean control rats. Oral glucose tolerance test was performed at 16 and 25 wk of age. At week 25, total RNA was isolated from the myocardium and assayed by rat oligonucleotide microarray for 14921 genes. Expression of selected genes was confirmed by qRT-PCR. Results: Fasting blood glucose, serum insulin, cholesterol and triglyceride levels were significantly increased, glucose tolerance and insulin sensitivity were impaired in ZDF rats compared to leans. In hearts of ZDF rats, 36 genes showed significant up-regulation and 49 genes showed down-regulation as compared to lean controls. Genes with significantly altered expression in the heart due to metabolic syndrome includes functional clusters of metabolism (e.g. 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 2; argininosuccinate synthetase; 2-amino-3ketobutyrate-coenzyme A ligase), structural proteins (e.g. myosin IXA; aggrecan1), signal transduction (e. g. activating transcription factor 3; phospholipase A2; insulin responsive sequence DNA binding protein-1) stress response (e.g. heat shock 70kD protein 1A; heat shock protein 60; glutathione S-transferase Yc2 subunit), ion channels and receptors (e.g. ATPase, (Na+)/K+ transporting, beta 4 polypeptide; ATPase, H+/K+ transporting, nongastric, alpha polypeptide). Moreover some other genes with no definite functional clusters were also changed such as e. g. S100 calcium binding protein A3; ubiquitin carboxy-terminal hydrolase L1; interleukin 18. Gene ontology analysis revealed several significantly enriched functional inter-relationships between genes influenced by metabolic syndrome. Conclusions: Metabolic syndrome significantly alters cardiac gene expression profile which may be involved in development of cardiac pathologies in the presence of metabolic syndrome

A szívizom sztressz-adaptációja: a peroxinitrit, a mátrix metalloproteinázok, és a hiperlipidémia szerepe = Stress adaptation of the myocardium: role of peroxynitrite, matrix metalloproteinases, and hyperlipidemia

A hiperlipidémia talaján kialakuló iszkémiás szívbetegség a leggyakoribb halálokok közé tartozik. A 4 éves project során a szívizom iszkémiának és az iszkémiás stressz adaptációs képességének (iszkémiás pre- és posztkondíció) celluláris mechanizmusait vizsgáltuk állatkísérletekben, különösképpen a peroxinitrit és celluláris targetjének, az MMP2-nek a szerepét. Új eredményeink közül néhányat emelünk ki. Kimutattuk, hogy hiperlipidemiában a szívben a peroxitrit képződés és ezáltal az MMP-2 aktivitása fokozódik, ami különösen hiperlipidmémiában jelentős, és ezt a folyamatot a prékondíció gátolja. DNA-chip vizsgálattal feltérképeztük hiperlipidémia hatására a génkifejeződés változásait a szívizomban. Kimutattuk, hogy az alacsony mértékű peroxinitrit képződés a stessz adaptáció kiváltásában igen fontos szerepet tölt be, hiszen olyan mechanizmusokat aktivál, melyek az iszkémiás stressz során túlzott mértékű peroxinitrit-MMP aktivitást csökkenti. Leírtuk, hogy nemspecifikus MMP gátlókkal az infarktus területe csökkenthető még hiperlipidémiás állatban is. Humán ApoB100 transzgenetikus eger modelleken megfigyeltük, hogy az oxidatív/nitrozatív stressz oka a hiperkoleszterinémia, és nem a hipertrigliceridémia. Kimutattuk továbbá, hogy a fiziológiás peroxinitrit szint, melyet a szívizom kapszaicin-érzékeny neuronjai szabályoznak, a normális szívizom relaxációt tartja fent. A project futamideje alatt az adott témában összesen 16 nemzetközi cikket (impakt faktor >70) közöltünk. | Ischemic heart disease developing due to hyperlipidemia is the number one killer in civilized societies. The present 4-year project was aiming at exploration of cellular mechanisms underlying stress adaptation of the myocardium, i.e. pre- and postconditioning, focusing on the role of peroxynitrite and its cellular target matrix metalloproteinase-2 (MMP2). Here we emphasize only some of the most important results of the project. We have shown that in hyperlipidemia, myocardial peroxynitrite formation and thereby MMP2 activity is increased, which is attenuated by preconditioning. We have mapped the changes in gene expression due to hyperlipidemia by the use of DNA-microarray assay. We have observed that moderate peroxynitrite formation is necessary to trigger the stress adaptation mechanisms, which in turn will decrease the pathological activation of the peroxynitrite-MMP2 signaling. We have shown that nonspecific MMP inhibitors are able to reduce infarct size even in the presence of hyperlipidemia. In human ApoB-100 transgene mice, we have observed that oxidative/nitrosative stress is due to hypercholesterolemia and not hypertriglyceridemia. Furthermore, we have shown that baseline physiological peroxynitrite formation, which is regulated by myocardial capsaicin-sensitive sensory nerves, plays an important role in the maintenance of normal relaxation of the myocardium. The present project yielded altogether 16 peer-reviewed papers (impact factor >70)

The combination of red palm oil and rooibos show anti-inflammatory effects in rats

BACKGROUND: Red palm oil (RPO) and rooibos have been shown to exhibit cardioprotective properties. RPO is rich in essential fatty acids and fat soluble antioxidants while rooibos contains polyphenolic compounds with a unique composition of flavonoids. They exert their biological effects in different cellular compartments. Therefore the combination of these two natural food compounds has the potential to enhance the spectrum of available dietary antioxidants in different cellular compartments, which could result in an enhanced protection against certain pathological conditions such as inflammation. METHODS: Male Wistar rats weighing 150-200 g were supplemented with RPO, rooibos or their combination for 28 days. The Langendorff system and the lipoposaccharide (LPS)-induced inflammatory model were used to establish if RPO and rooibos, when supplemented alone or in combination, will reverse the negative effects of LPS on cardiac function at baseline. The effect of dietary intervention was also investigated on modulation of pro-inflammatory and anti-inflammatory cytokines in plasma and myocardial tissue. RESULTS AND DISCUSSION: The LPS resulted in induction of systemic inflammation as evidenced by increased levels of IL-1beta in plasma of LPS-treated rats compared to their non-treated control counterparts. Dietary supplementation and LPS treatment did not have an effect on baseline cardiac functional parameters. However, the elevation of IL-1beta levels in plasma of LPS-induced rats consuming either RPO or rooibos alone were paralleled with increased levels of the anti-inflammatory cytokine, IL-10. The combination of rooibos and RPO was associated with enhanced endogenous production of myocardial IL-10 in LPS-induced rats. CONCLUSION: The results of this study indicate that RPO and rooibos when supplemented individually showed anti-inflammatory effect at systemic level while their combination exhibited an enhanced anti-inflammatory effect in the myocardial tissue. Therefore, the findings in the current study argue that the combination of these two natural food substances could be beneficial in clinically relevant conditions where inflammation plays a role

Nitrát tolerancia a szívizomban = Nitrate tolerance in the myocardium

Nagy dózisú nitroglicerin ismételt alkalmazásával vaszkuláris nitrát toleranciát váltottunk ki kísérleti állatokban. Kimutattuk hogy vaszkuláris nitrát toleranciában a szív és az aorta génexpressziós profilja eltérően változik. A szívizom génexpressziós változását egyéb szisztémás metabolikus állapotokban (metabolikus szindróma) is leírtuk. Kimutattuk, hogy az iszkémiás posztkondícionálás infarktus méretet csökkentő hatása jelentősen mérséklődik vaszkuláris nitrát tolerancia fennállása esetén. Továbbá azt tapasztaltuk, hogy ez a jelenség független a túlélő kinázok aktiválódásától. Kísérletes urémiában viszont a prekondícionálás kardioprotektív hatása megtartott marad. Izolált perfundált szívekben kimutattuk, hogy mind a koronária lekötés előtt, mind pedig a reperfúzió előtt megkezdett nitroglicerin perfúzió csökkenti a kialakuló infarktus méretét. Azonban a nitroglicerinnek ez a kardioprotektív hatása vaszkuláris nitrát tolerancia fennállása esetén nem volt megfigyelhető. Igazoltuk, hogy a NO-donor SNAP védi a szimulált iszkémia/reoxigenizációnak kitett primer szívizomsejt tenyészeteket a sejtelhalástól, valószínűleg részben a cGMP-PKG jelátviteli útvonal aktiválása révén. Továbbá kimutattuk, hogy a reperfúzió kezdetén alkalmazott szakaszos nagyfrekvenciás ingerléssel is kiváltható posztkondícionáló hatás a szívben. | We have induced vascular nitrate tolerance in rats by repeated administration of high dose nitroglycerin. We have found that the gene expression profile of the heart and the aorta was differentially altered in response to the development of vascular nitrate tolerance. Alterations in cardiac gene expression were shown in other systemic metabolic conditions (i.e. metabolic syndrome) as well. We have shown that the infarct size limiting effect of ischemic postconditioning is attenuated in the state of vascular nitrate tolerance. Moreover, we have found that this phenomenon is independent of survival kinase activation. Experimental uremia. however, did not lead to the loss of ischemic preconditioning. We have demonstrated in ex vivo hearts that nitroglycerin perfusion decreased infarct size when started before coronary occlusion and also when only started before reperfusion. This cardioprotective effect of nitroglycerin was diminished in the state of vascular nitrate tolerance. We have shown that the NO-donor molecule SNAP is able to protect primary cardiomyocyte cultures against simulated ischemia/reperfusion at least in part via activation of cGMP-PKG signaling. We have found that a postconditioning effect can be induced by applying short periods of ventricular overdrive pacing at the onset of reperfusion

Preconditioning protects the heart in a prolonged uremic condition

Metabolic diseases such as hyperlipidemia and diabetes attenuate the cardioprotective effect of ischemic preconditioning. In the present study, we examined whether another metabolic disease, prolonged uremia, affects ischemia/reperfusion injury and cardioprotection by ischemic preconditioning. Uremia was induced by partial nephrectomy in male Wistar rats. The development of uremia was verified 29 wk after surgery. Transthoracic echocardiography was performed to monitor cardiac function. At week 30, hearts of nephrectomized and sham-operated rats were isolated and subjected to a 30-min coronary occlusion followed by 120 min reperfusion with or without preceding preconditioning induced by three intermittent cycles of brief ischemia and reperfusion. In nephrectomized rats, plasma uric acid, carbamide, and creatinine as well as urine protein levels were increased as compared with sham-operated controls. Systolic anterior and septal wall thicknesses were increased in nephrectomized rats, suggesting the development of a minimal cardiac hypertrophy. Ejection fraction was decreased and isovolumic relaxation time was shortened in nephrectomized rats demonstrating a mild systolic and diastolic dysfunction. Infarct size was not affected significantly by nephrectomy itself. Ischemic preconditioning significantly decreased infarct size from 24.8 ± 5.2% to 6.6 ± 1.3% in the sham-operated group and also in the uremic group from 35.4 ± 9.5% to 11.9 ± 3.1% of the area at risk. Plasma ANG II and nitrotyrosine were significantly increased in the uremic rats. We conclude that although prolonged experimental uremia leads to severe metabolic changes and the development of a mild myocardial dysfunction, the cardioprotective effect of ischemic preconditioning is still preserved

Effect of a multivitamin preparation supplemented with phytosterol on serum lipids and infarct size in rats fed with normal and high cholesterol diet

BACKGROUND: Although complex multivitamin products are widely used as dietary supplements to maintain health or as special medical food in certain diseases, the effects of these products were not investigated in hyperlipidemia which is a major risk factor for cardiovascular diseases. Therefore, here we investigated if a preparation developed for human use containing different vitamins, minerals and trace elements enriched with phytosterol (VMTP) affects the severity of experimental hyperlipidemia as well as myocardial ischemia/reperfusion injury. METHODS: Male Wistar rats were fed a normal or cholesterol-enriched (2% cholesterol + 0.25% cholate) diet for 12 weeks to induce hyperlipidemia. From week 8, rats in both groups were fed with a VMTP preparation or placebo for 4 weeks. Serum triglyceride and cholesterol levels were measured at week 0, 8 and 12. At week 12, hearts were isolated, perfused according to Langendorff and subjected to a 30-min coronary occlusion followed by 120 min reperfusion to measure infarct size. RESULTS: At week 8, cholesterol-fed rats showed significantly higher serum cholesterol level as compared to normal animals, however, serum triglyceride level did not change. VMTP treatment significantly decreased serum cholesterol level in the hyperlipidemic group by week 12 without affecting triglyceride levels. However, VMTP did not show beneficial effect on infarct size. The inflammatory marker hs-CRP and the antioxidant uric acid were also not significantly different. CONCLUSIONS: This is the first demonstration that treatment of hyperlipidemic subjects with a VMTP preparation reduces serum cholesterol, the major risk factor for cardiovascular disease; however, it does not provide cardioprotection

Transcriptomic alterations in the heart of non-obese type 2 diabetic Goto-Kakizaki rats

BACKGROUND: There is a spectacular rise in the global prevalence of type 2 diabetes mellitus (T2DM) due to the worldwide obesity epidemic. However, a significant proportion of T2DM patients are non-obese and they also have an increased risk of cardiovascular diseases. As the Goto-Kakizaki (GK) rat is a well-known model of non-obese T2DM, the goal of this study was to investigate the effect of non-obese T2DM on cardiac alterations of the transcriptome in GK rats. METHODS: Fasting blood glucose, serum insulin and cholesterol levels were measured at 7, 11, and 15 weeks of age in male GK and control rats. Oral glucose tolerance test and pancreatic insulin level measurements were performed at 11 weeks of age. At week 15, total RNA was isolated from the myocardium and assayed by rat oligonucleotide microarray for 41,012 genes, and then expression of selected genes was confirmed by qRT-PCR. Gene ontology and protein-protein network analyses were performed to demonstrate potentially characteristic gene alterations and key genes in non-obese T2DM. RESULTS: Fasting blood glucose, serum insulin and cholesterol levels were significantly increased, glucose tolerance and insulin sensitivity were significantly impaired in GK rats as compared to controls. In hearts of GK rats, 204 genes showed significant up-regulation and 303 genes showed down-regulation as compared to controls according to microarray analysis. Genes with significantly altered expression in the heart due to non-obese T2DM includes functional clusters of metabolism (e.g. Cyp2e1, Akr1b10), signal transduction (e.g. Dpp4, Stat3), receptors and ion channels (e.g. Sln, Chrng), membrane and structural proteins (e.g. Tnni1, Mylk2, Col8a1, Adam33), cell growth and differentiation (e.g. Gpc3, Jund), immune response (e.g. C3, C4a), and others (e.g. Lrp8, Msln, Klkc1, Epn3). Gene ontology analysis revealed several significantly enriched functional inter-relationships between genes influenced by non-obese T2DM. Protein-protein interaction analysis demonstrated that Stat is a potential key gene influenced by non-obese T2DM. CONCLUSIONS: Non-obese T2DM alters cardiac gene expression profile. The altered genes may be involved in the development of cardiac pathologies and could be potential therapeutic targets in non-obese T2DM