Dendroaspis natriuretic peptide protects the post-ischemic myocardial injury

The present study used isolated rat hearts to investigate whether (1) Dendroaspis natriuretic peptide (DNP) is protective against post-ischemic myocardial dysfunction, and (2) whether the cardioprotective effects of DNP is related to alteration of Bcl-2 family protein levels. The excised hearts of Sprague-Dawley rats were perfused on a Langendorff apparatus with Krebs-Henseleit solution with a gas mixture of 95% O2 and 5% CO2. Left ventricular end-diastolic pressure (LVEDP, mmHg), left ventricular developed pressure (LVDP, mmHg) and coronary flow (CF, ml/min) were continuously monitored. In the presence of 50 nM DNP, all hearts were perfused for a total of 100 min consisting of a 20 min pre-ischemic period followed by a 30 min global ischemia and 50 min reperfusion. Lactate dehydrogenase (LDH) activity in the effluent was measured during reperfusion. Treatment with DNP alone improved the pre-ischemic LVEDP and post-ischemic LVEDP significantly comparing with the untreated control hearts during reperfusion. However, DNP did not affect the LVDP, heart rate (HR, beats/min), and CF. Bcl-2, an anti-apoptotic protein expressed in ischemic myocardium of DNP+ischemia/reperfusion (I/R) group, was higher than that in I/R alone group. Bax, a pro-apoptotic protein expressed in ischemic myocardium of DNP+I/R group, has no significant difference compared with I/R alone group. These results suggest that the protective effects of DNP against I/R injury would be mediated, at least in part, through the increased ratio of Bcl-2 to Bax protein after ischemia-reperfusion.     

Free radical scavenger depletion in post-ischemic reperfusion brain damage

In the present study the influence of pretreatment with various GSH depletors such as buthionine sulfoximine (BSO) and diethylmaleate (DEM) was investigated in rats following cerebral postischemic reperfusion. Moreover, the effect of diethyldithiocarbamic acid (DDC), inhibitor of endogenous Cu,Zn-SOD, was evaluated. A significant depletion (40% of control value) of GSH levels was observed 24 h after DEM administration; after 48 h the value reached control levels. BSO showed maximal GSH depletion (59%) 24 h after administration and it was constant for almost 48 h. DDC administration caused a marked decrease (60%) of Cu,Zn-SOD activity 4 h after the injection and induced a marked decrease in percentage of survival with respect to control (untreated, ischemic) rats, when administered 4 h before ischemia. BSO and DEM prolonged the survival time of animals when administered 24 h before ischemia. This last paradoxical effect is unclear at present, but it might be due to an influence on glutamate cascade.     

Beneficial effects of carbon monoxide-releasing molecules on post-ischemic myocardial recovery

There is increasing evidence corroborating a protective role of carbon monoxide releasing molecules (CORMs) in injured tissues. Carbon monoxide (CO) carriers have been recently developed as a pharmacological tool to simulate the effect of heme oxygenase-1-derived CO. The effects of CORM-3, a water-soluble CO releaser, on the incidence of reperfusion-induced ventricular fibrillation (VF) and tachycardia (VT) were studied in isolated rat hearts. Hearts were treated with different doses of CORM-3 before the induction of 30 min global ischemia followed by 120 min reperfusion. We found that at concentrations of 25 μM and 50 μM of CORM-3 promoted a significant reduction in the incidence of VF and VT. Thus, the incidence of VF was reduced by 67% (p < 0.05) and 92% (p < 0.05) with 25 μM and 50 μM of CORM-3, respectively. The protective effect of CORM-3 on the incidence of VT followed the same pattern. The antiarrhythmic protection was associated with a marked attenuation in infarct size, significant decreases in cellular Na⁺ and Ca²⁺ gains and K⁺ loss. Consequently, the recovery of post-ischemic function was significantly improved. In conclusion, CORM-3 exerts beneficial effects against ischemia/reperfusion-induced injury through its abilities to release CO which mediates a cardioprotective action by regulating tissue Na⁺, K⁺, and Ca²⁺ levels.     

Spin-trapping evidence that graded myocardial ischemia alters post-ischemic superoxide production

The spin trapping agent 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) was used to investigate oxy-radical production in post-ischemic rat hearts previously exposed to 20, 30, or 40 minutes of global ischemia. A hydroxyl spin adduct (DMPO-OH) was identified in coronary effluent during the initial seconds of reperfusion by Electron Spin Resonance (ESR) Spectroscopy. The intensity of the ESR signal in post-ischemic effluent increased as ischemic duration was prolonged; however, regardless of the duration of ischemia, maximal spin adduct detection occurred 3 minutes after initiation of reperfusion. Superoxide dismutase inhibited the formation of DMPO-OH, suggesting that superoxide anion was initially generated and is the principle source for the production on the hydroxyl adduct. Our investigations indicate that superoxide anion is produced during the early moments of reperfusion and that its production in the post-ischemic heart is related to the severity of ischemia.     

Thromboxane receptor blockade with SQ 30,741 improves post-ischemic myocardial function in anesthetized dogs

This study was conducted to determine whether the thromboxane A2 receptor antagonist SQ 30,741 can improve post-ischemic recovery of cardiac function in anesthetized dogs. Saline or SQ 30,741 was infused throughout a 15-min coronary occlusion and 5 hr of reperfusion. Ischemic regional cardiac function was determined using subendocardial ultrasonic crystals. Despite no differences in collateral blood flow or reperfusion flow, SQ 30,741 significantly improved ventricular segmental shortening at all times measured during reperfusion. At 5 hr after the initiation of reperfusion, segmental shortening was 3 +/- 16 and 44 +/- 10% of baseline values for saline and SQ 30,741 groups, respectively. These results implicate thromboxane receptor activation in the pathogenesis of myocardial stunning, and thromboxane antagonists may be useful in mitigating this functional deficit.     

Possible contribution of free radicals and lipid peroxidations on pathogenesis of post-ischemic brain damage.

It has been widely accepted concept that peroxidate decomposition of membrane lipids contribute cellular damage in cerebral ischemia, and varieties of putative cascading reactions has been proposed in this regard as the underlying detrimental processes. But little is known about existence and significance of the blood born chemical mediator. In this communication, the authors tried to emphasize the prospective role of leukocyte invasion to the ischemic brain tissue. Effectiveness of some clinically applicable antioxidants against ischemic brain damage was also discussed.     

Adjunctive pharmacologic treatment for elective stenting of the extracranial carotid arteries

Endovascular stenting of the extracranial carotid arteries is a viable treatment option to carotid endarterectomy in selected patients. Patients undergoing this procedure must be treated with aspirin 325 mg at least 24 hours prior to the procedure and clopidogrel 75 mg oral, three to five days prior to the procedure (or 300 mg at least six hours prior to stenting). During the intervention, heparin is given to maintain an activated clotting time of at least 300 seconds. Although there are theoretical advantages on the use of a platelet IIb/IIIa inhibitor, their routine use during elective carotid stenting is not recommended until their clinical benefits have been clearly demonstrated in this patient population. Hemodynamic instability during balloon inflation and stent deployment should be treated with intravenous fluids, atropine, and an alpha agonist such as neosynephrine. All the oral antihypertensive must be discontinued after the procedure and if there is persistence of hypotension making discontinuation of neosynephrine difficult, an oral alpha agonist such as midodrine (2.5 to 5 mg two to three times daily) is helpful. In the majority of patients this medication can be tapered off three to five days following stenting.     

Effect of ursolic acid treatment on apoptosis and DNA damage in isoproterenol-induced myocardial infarction

The present study was designed to evaluate the protective effect of ursolic acid (UA) against isoproterenol-induced myocardial infarction. Myocardial infarction was induced by subcutaneous injection of isoproterenol hydrochloride (ISO) (85 mg/kg BW), for two consecutive days. ISO-induced rats showed elevated levels of cardiac troponins T (cTn T) and I (cTn I) and increased activity of creatine kinase-MB (CK-MB) in serum. Lipid peroxidative markers (thiobarbituric acid reactive substances (TBARS), conjugated dienes (CD) and lipid hydroperoxides (HP)) elevated in the plasma and heart tissue whereas decreased activities of enzymatic antioxidants (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione-S-transferase (GST) and glutathione reductase (GR)) in erythrocytes and heart tissue of ISO-induced rats. Non-enzymatic antioxidants (vitamin C, vitamin E and reduced glutathione (GSH)) levels were decreased significantly in the plasma and heart tissue of ISO-induced rats. Furthermore, ISO-induced rats showed increased DNA fragmentation, upregulations of myocardial pro-apoptotic B-cell lymphoma-2 associated-x (Bax), caspase-3, -8 and -9, cytochrome c, tumor necrosis factor-α (TNF-α), Fas and down-regulated expressions of anti-apoptotic B-cell lymphoma-2 (Bcl-2) and B-cell lymphoma-extra large (Bcl-xL). UA-administered rats showed decreased levels/activity of cardiac markers, DNA fragmentation and the levels of lipid peroxidative markers in the plasma and heart tissue. Activities of enzymatic antioxidants were increased significantly in the erythrocytes and heart tissue and also non-enzymatic antioxidants levels were increased significantly in the plasma and heart tissue in UA-administered rats. UA influenced decreased DNA fragmentation and an apoptosis by upregulation of anti-apoptotic proteins such as Bcl-2, Bcl-xL and down-regulation of Bax, caspase-3, -8 and -9, cytochrome c, TNF-α, Fas through mitochondrial pathway. Histopathological observations were also found in line with biochemical parameters. Thus, results of the present study demonstrated that the UA has anti-apoptotic properties in ISO-induced rats.     

Permeability of the post-ischemic rat aorta.

The time course of the vascular permeability disorder following upon acute hypoxia and the fate of plasma substances entering the vessel wall in consequence of increased permeability were studied on the rat abdominal aorta rendered hypoxic for one hour by double ligature and recirculated for periods ranging from one hour to 30 days. The distribution and quantity of mural plasma inhibition were determined histochemically by means of a colloidal iron tracer and Prussian blue reaction and by photometric analysis, respectively. Plasma imbibition reached its maximum after recirculation for 24 to 48 hours and fell to an almost normal level after 10 days. Administration of the colloidal iron tracer on the second day of recirculation, when the permeability disorder was at its peak, showed plasma imbibition in every layer of the vessel wall. At seven days it was restricted to the outer third of the media and the adventitia. The endothelium is acting as the main barrier to mural plasma imbibition and in the case of a permeability disorder only the elastic lamellae constitute a temporary mechanical obstacle to the ingression of plasma.     

Prevention of damage in acute spinal cord injury by peptides and pharmacologic agents

Cats were used as models of traumatic spinal cord injury. Each experimental animal received a 500 g-cm force to the exposed dura at the level of thoracic fourth vertebra. Somatosensory evoked potentials (SEPs), carotid arterial blood pressure (BP), and abdominal aorta blood flow in the treated groups were compared with those of the control group. The three treated groups received naloxone (5 mg/kg), TRH (5 mg/kg), and a combination of methyl-prednisolone sodium succinate (MP, 35 mg/kg) and epsilon-aminocaproic acid (EACA, 350 mg/kg). The SEPs which were done only in the naloxone treated group approached "normalcy" 24-26 hours after trauma as compared with the absence of SEPs in traumatized untreated group. In all three groups, the treatment increased the blood flow in abdominal aorta significantly. Morphine sulfate increased substance P (SP) immunoreactivity in the dorsal and ventral gray matter. Naloxone not only reversed this effect, it depleted SP below the saline control level. In order to establish that lipid free radicals are responsible for damage to biological membranes, their effects were also investigated in vitro: 14C-GABA uptake by mouse cortical slices which had decreased by 33% in the presence of superoxide (. O-2) generating system, horseradish peroxidase (HRP), was reduced only by 9% when superoxide dismutase was added to the medium. The latter also protected the nerve endings from damage by (. O-2) as examined by electron microscopy. It is concluded that the agents used in this study produce their ameliorating effects by virtue of their anti-inflammatory, anti-oxidant, and membrane stabilizing properties in addition to their effect on enhancing the regional microcirculation. The release of SP by naloxone may be responsible for the increase in blood flow. The consequences of traumatic injury as depicted in Fig. 1 are discussed at length.     

Lipid peroxidation and myocardial contractile function in the post-ischemic period depending on the level of hypothermic protection of the heart]

The experiments on rats isolated hearts showed lipid peroxidation state to depend on myocardial cooling level in ischemic period. Cooling to 8-12 degrees C does not induce significant impairment in the system of lipid peroxidation/antioxidant activity, thus preventing the development of reperfusion impairment in cardiac activity restoration. Temperature decrease to 4-6 degrees C during the ischemic period results in lipid peroxidation, antioxidant cell system exhaustion and impairment of contractive myocardial function in reperfusion.     

Correction of catecholamine-induced myocardial damage with ionol

Experiments on male rats exhibiting both high and low resistance to hypoxia have shown that ionol acts as a cardioprotective agent in the adrenalin-induced myocardial dystrophy. This effect is realized through the depression of the lipid peroxidation activity.     

Insulin inhibits myocardial ischemia-induced apoptosis and alleviates chronic adverse changes in post-ischemic cardiac structure and function

Insulin has been shown to possess significant anti-apoptotic effect in myocardial ischemia/reperfusion (MI/R). However, the contribution by this protection of insulin to the prolonged cardiac function in rats subjected to ischemia remains unclear. The present study attempted to test whether early insulin treatment influences adverse prolonged post-ischemic cardiac structural and functional changes. Adult male rats were subjected to left anterior descending coronary artery occlusion and were randomized to receive one of the following treatments: saline (4 ml/kg/h i.v. injection beginning 10 min before the ischemia and continuing for 2 h), insulin (60 U/l, i.v. injection following the same routine, and hypodermic injection of insulin (0.5 U/ml, 1 ml/kg/d) for 3 days after the ischemia surgery) or insulin plus wortmannin (15 μg/kg i.v. injection 15 min before each insulin administration). Treatment with insulin significantly reduced infarct size, decreased plasma creatine kinase and lactate dehydrogenase activities, decreased apoptosis index and caspase-3 activity (all P < 0.01 vs. saline), and improved cardiac function 24 h after ischemia. Importantly, at the end of 4 weeks after the ischemia surgery, MI rats receiving insulin treatment showed smaller left ventricle (LV) cavity and thicker systolic interventricular septum, and increased cardiac ejection fraction and LV fractional shortening (all P < 0.05 vs. saline). Inhibition of insulin signaling with wortmannin not only blocked insulin’s anti-apoptotic effect, but also almost completely abolished effects of insulin on cardiac structure and function. These data indicate that inhibition of apoptosis by early insulin treatment alleviates chronic adverse changes in post-ischemic cardiac structure and function. Wenjuan Xing and Wenjun Yan contributed equally to this study.     

The role of intracellular calcium in the ischemic myocardial damage

The isolated, perfused ventricles of guinea-pig and rat hearts stimulated at the rate of 60/min were equilibrated for 60 min with 45Ca containing solution. Thereafter some of them were perfused for the last 10 min of experiment with deoxygenated (pO2 = 35 not equal to 7 mm Hg) radioactive solution. Hypoxia resulted in decrease of exchangeable calcium (45Ca) content by 0.90 mmol/kg w.w. in guinea-pig and by 0.26 mmol/kg w.w. in the rat. The amount of 15Ca lost by guinea-pig ventricles is equal to the content of rate-dependent fraction Ca2 described in the previous papers [Pytkowski et al., 1983; Lewartowski et al., 1984]. The isolated papillary muscles of the right ventricles of guinea-pig and rat hearts were subjected to 90 min of ischemia simulated by immersion in the warm, deoxygenated paraffin oil. Some of the guinea-pig muscles were deprived of Ca2 fraction by means of prolonged rest (20 min) immediately prior to ischemia. All the preparations were quiescent during ischemia. The guinea-pig muscles deprived of fraction Ca2 and the rat muscles developed much weaker contracture during ischemia and showed better recovery of phasic contraction upon reperfusion than the guinea-pig muscles containing Ca2 fraction prior to ischemia. We propose that Ca2 fraction is released from its binding sites at the early phases of ischemia contributing to the disturbances in Ca homeostasis and to mechanism of damage of ischemic cardiac muscle.     

Determination of susceptibility to pharmacologic treatment in Graves'-Basedow disease]

The authors present the results of research on the usefulness of clinical data, thyroid hormones, TSH, antithyroid antibody, HLA estimations and a type of behavior, for prognosing the susceptibility to pharmacological therapy in thyrotoxicosis, specially in Graves disease. The highest chance of predictability was found in patients with small goiter, HLA-DR5 positive with onset of the disease at the age before 40. In general it is difficult to predict the reaction to therapy in individual case with high probability.     

Ultrastructure of isoproterenol-induced myocardial damage in chick embryos

Isoproterenol is known to cause severe myocardial lesions when given in toxic doses to adult homoiotherms. Previous studies on chick embryos revealed myocardial damage with scattered necroses in the outer layer of ventricular myocardium. The present ultrastructural study, performed on embryos 6 to 20 days old, has shown various types of cellular lesions; mainly cellular oedema, mitochondrial swelling, necroses of isolated cardiac muscle cells, fatty degeneration, accumulation of glycogen, and signs of increased proteosynthesis in the surviving muscle cells. Morphological features of the lesions differed from those which are known to be induced by isoproterenol in adult animals and seemed to depend on the stage of embryonic development.     

Prevention of myocardial reperfusion injury by increasing artificial trans-membrane sodium gradient in "calcium paradox" and post-ischemic reperfusion

An effect of the high sodium gradient during "calcium paradox" and postischemic reperfusion has been studied. A decrease of Na/Ca exchange by high sodium gradient (200 mM NaCl in the perfusion solution) resulted in the reduction of myoglobin release from the heart during "calcium paradox". High sodium concentration solution (200 mM) increased protective effect of ATP during "calcium paradox". Exogenous phosphocreatine (100 mumol/mol) increased myoglobin release from the heart. During perfusion of the heart by high sodium concentration, phosphocreatine efficiently decreased myoglobin release from the heart during "calcium paradox". Exogenous ATP (as Na-pump activator) and high Na+ concentration solution (180 mM) prevented the LDH release from the myocardium, decreased ATP hydrolysis, inhibited Ca influx, maintained total adenine nucleotides, phosphate potential, energy charge of the cardiomyocytes.