Ischaemic preconditioning reduces infarct size following global ischaemia in the murine myocardium

Ischaemic preconditioning has not been demonstrated to reduce infarct size following global ischaemia in murine myocardium. Eighteen mouse hearts were isolated, perfused in the Langendorff mode, and randomised to control or preconditioning groups. Preconditioned hearts received four periods of five minutes global ischaemia with five minutes of intervening reperfusion. Control hearts were perfused normally during this period. Both groups were then subjected to 30 minutes global ischaemia followed by 30 minutes reperfusion. Infarct size, contractile recovery and LDH leakage were assessed. Mean infarct size was reduced from 57% of ventricular volume (controls) to 33% in preconditioned hearts (P=0.003). A small improvement in contractile recovery was also observed (6.3% of baseline in controls, 15.5% in preconditioned hearts; P=0.004). Release of LDH did not differ significantly between groups. This study confirms for the first time that ischaemic preconditioning can delay the onset of myocardial necrosis following global ischaemia in the isolated mouse heart. Received: 26 January 1998, Returned for 1. revision: 16 February 1998, 1. Revision received: 16 April 1998, Accepted: 14 May 1998     

Ischaemic preconditioning reduces spinal cord injury in transient ischaemia

OBJECTIVE: Paraplegia remains a devastating complication after thoracic and thoracoabdominal aortic surgery for coarctations, dissections or aneurysms. Since the advent of ischaemic preconditioning of the myocardium, attention has been directed to the nervous system.This study was designed to evaluate the acute protective effect of ischaemic preconditioning on the spinal cord. MEDHODS AND RESULTS: Thirty-six New Zealand white rabbits were randomly assigned to one of three groups. The preconditioning group had 5 minutes of aortic occlusion, 25 minutes reperfusion and 20 minutes of ischaemia, whereas the controls had only 20 minutes of ischaemia.The sham group was anaesthetized and subjected to laparotomy without aortic occlusion. Physiological parameters and somatosensory evoked potentials were monitored during the experiment. Neurological outcome was clinically evaluated up to 48 hour after ischaemia and motor function was scored.Then the animals were sacrificed.Their spinal cord, abdominal aorta and its branches were removed and processed for histopathological examination. Histhopathological changes of the gray matter in the lumbosacral segments were scored from 0 to 6 according to a semi-quantitative scala. The changes in amplitudes of evoked potentials during ischaemia and recovery periods were similar in preconditioning and control groups.The average motor function score was significantly higher in the preconditioning group than the control group at 24 and 48 hours after the ischaemic event (p < 0.05). Histological observations were consistent with the neurological findings.The histopathological scores in the control group and the preconditioning group were 3.2 (1.4-5.2) and 2.4 (0.8-4.4), respectively (p < 0.05). CONCLUSIONS: The results suggest that ischaemic preconditioning reduces the spinal cord injury and improves neurological outcome in transient ischaemia in rabbits.This protective mechanism is rapidly invoked within only 25 minutes interval between the preconditioning stimulus and the ischaemic insult.     

Bound inorganic phosphate and early contractile failure in global ischaemia

Inorganic phosphate (P_i) accumulates extremely rapidly in ischaemic heart muscle and intracellular binding of this metabolite may account for the precipitous loss of function seen at the onset of severe ischaemia. We have used³¹P-NMR spectroscopy to measure the free cytosolic [P_i] and chemical assay techniques to measure total tissue P_i at 0, 1, 2, 3, 5, and 12 min of complete global ischaemia in the isolated isovolumic rat heart. At zero time, the P_i assayed chemically was 30.77±5.52 mol/g dry wt (mean±SD, n=7) whilst P_i assayed by NMR was 3.39±1.21 mol/g dry wt (n=15). Thus, 27.38 mol/g dry wt of P_i was bound at a cytosolic [P_i] of 0.82 mM. After 12 min of ischaemia, 49.88 mol/g dry wt of P_i was bound at a cytosolic [P_i] of 4.11 mM. When all data were fitted, using a non-linear, least squares fit (p<0.05), to the binding isotherm: Bound P_i=B_max. [P_i]/(K_d+[P_i]), the apparent binding parameters K_d and B_max were estimated to be 1.1±0.6 mM and 64.0±10.2 mol/g dry wt respectively. During the first minute of global ischaemia when the rate-pressure product had decreased by 79% of its pre-ischaemic value, bound P_i had increased by 58% and free cytosolic [P_i] by 162%. When functional and metabolite changes were expressed as a fraction of the total change which occurred during the 12-min ischaemic period, bound P_i had the profile most similar to the rate-pressure product. Both the amount of bound P_i and free cytosolic [P_i] correlated with loss of contractile function as the ischaemic period progressed. The results snow that during ischaemia, P_i is bound progressively as free cytosolic [P_i] is increased as the result of high energy phosphate hydrolysis. While these results are consistent with the possibility that P_i binding may contribute to ischaemic contractile failure, no molecular explanation for the possible effect of bound P_i on contraction has been propsed.     

Ischaemic preconditioning protects against myocardial dysfunction caused by ischaemia in isolated hypertrophied rat hearts

Although ischaemic preconditioning (PC) has been shown to protect normal hearts from a subsequent ischaemic insults, its protective effect on the hypertrophied myocardium has not been widely studied. This study was designed to investigate whether ischaemic preconditioning protects hearts with hypertrophy (HYP). Cardiac HYP was produced in rats by suprarenal abdominal aortic constriction of 5 weeks' duration, and was defined as left ventricular weight: body weight [LVW: BW (mg/g)] ratio over 3.0. Isolated rat hearts were perfused with a modified Krebs-Henseleit buffer at 37°C in a Langendorff preparation. Hearts from sham-operated animals (NORM) and those with HYP underwent a PC protocol consisting of 3 min of global zero flow ischaemia, 5 min of reperfusion followed by 5 min of ischaemia and 5 min of reperfusion. This was followed by 20 min ischaemia and 45 min reperfusion. Control hearts in the HYP and NORM groups were not subjected to the PC protocol. There were, thus, four experimental groups: NORM control (n=9), NORM, PC (n=9), HYP control (n=9), HYP, PC (n=11). The recovery of function after ischaemia was evaluated by recovery of left ventricular developed pressure (LVDP) expressed as % of the initial value (LVDP%). The LVW: BW ratio for the HYP groups was 3.4 (SEM 0.08). LVDP% was higher (p<0.01) in preconditioned groups as compared with controls. In NORM control recovery was 49.3 (6.1), NORM, PC 76.5 (3.4), HYP control 39.8 (4.6) HYP, PC 70.1 (4.1). These data indicate that the ability of preconditioning to protect against ischanemic ventricular dysfunction is preserved in this model of cardiac hypertrophy.     

Glucose homeostasis. Comparison between hypothermic and normothermic cardiopulmonary bypass.

STUDY OBJECTIVE: Disturbance in blood glucose homeostasis during cardiac surgery may cause visceral and metabolic alterations. Hypothermic CPB induces glucose and hormonal changes. As normothermic CPB is used at some institutions, a comparison of blood glucose and plasma hormones between hypothermic and normothermic CPB was performed. DESIGN: Prospective nonrandomized study. SETTING: University cardiac center. PATIENTS: Twenty-two nondiabetic adults undergoing elective coronary bypass and/or valvular surgery. INTERVENTIONS: Group 1 (n = 12) underwent hypothermic CPB (25 degrees C) and group 2 (n = 10) normothermic CPB (37 degrees C). In both groups nonpulsatile CPB was achieved with a membrane oxygenator and dextrose-free crystalloid priming. Dextrose was not administered during surgery but was infused postoperatively (125 mg/kg/h). MEASUREMENTS AND RESULTS: Eight blood samples were drawn during the period of arrival in the operating room (control) to the third postoperative hour. During hypothermic CPB in group 1, blood glucose level increased to 154 +/- 20 mg/dl (mean +/- SD) associated with a decrease in plasma insulin and an increase in epinephrine, despite a decrease in cortisol and growth hormone. During rewarming, the blood glucose value continued to increase (to 197 +/- 35 mg/dl) associated with an increase in glucagon, growth hormone and catecholamines, despite a 374 percent increase in insulin. During CPB in group 2, insulin, glucagon, cortisol and catecholamines were significantly higher than during hypothermic CPB so that the blood glucose level was not significantly different between the two groups during CPB. Blood glucose value was higher in group 1 than in group 2 at closure of the chest (208 +/- 30 vs 175 +/- 19 mg/dl, respectively, p less than 0.02) and at the third postoperative hour (271 +/- 30 vs 221 +/- 51 mg/dl, p less than 0.01). In both groups, however, the postoperative increase in blood glucose was accompanied by a similar increase in insulin, cortisol and catecholamines but glucagon was lower after hypothermic CPB. CONCLUSIONS: Hyperglycemia occurred perioperatively in cardiac surgery with dextrose-free priming both during hypothermic and normothermic CPB but normothermic CPB resulted in a slow and steady increase in both glucose and insulin concentrations without the major perturbations that occurred with hypothermic CPB. Postoperatively, higher blood glucose was observed in the hypothermic CPB group.     

Cardioprotective effects of mioflazine during 1 h normothermic global ischaemia in the canine heart

The cardioprotective effects of mioflazine, a recently developed cardiovascular drug, were investigated in 41 anaesthetised open chest Beagle dogs subjected to 1 h normothermic global myocardial ischaemia. The severity of the model is evidenced by the finding that only one out of 20 control dogs could be weaned from extracorporeal bypass. Oral pretreatment with mioflazine (2.5 mg X kg-1) resulted in complete functional recovery in 17 out of 20 animals. Biochemical analysis of left ventricular biopsies taken before, during and after aortic cross clamping showed a preservation of purines and a better recovery of ATP, ATP/ADP X Pi ratio and energy charge (p less than 0.05) in the pretreated animals. Morphological and cytochemical assessment of the myocardium demonstrated that the ultrastructure of the sarcolemma and its calcium binding capacity is remarkably well preserved in the drug treated animals. These results indicate a strong cardioprotective effect of mioflazine. The biochemical, cytochemical and ultrastructural findings suggest an interaction of the drug with the sarcolemma.     

常温和中低温体外循环对甲状腺素变化的影响

目的探讨常温及低温体外循环心脏直视手术对甲状腺素的影响。方法选择先天性心脏病患者80例,随机分为常温组及低温组,各40例,分别于术晨、体外循环结束时及术后6、12、24、48h抽取患者动脉血,测定血浆T3、T4和TSH。结果两组术前各项检查指标差异无统计学意义。常温体外循环下甲状腺素T3升高水平低于中低温,恢复时间比中低温明显缩短。结论常温体外循环心脏直视手术对甲状腺素的影响显著轻于低温组,因而术后机体的恢复优于低温方法。     

Ischaemic preconditioning limits infarct size in the rat heart.

OBJECTIVE: One of the mechanisms by which ischaemic preconditioning is thought to protect against later prolonged ischaemia is via a reduction in ATP utilisation during ischaemia. The ATP "wastage" that occurs during ischaemia is thought to be due to mitochondrial ATPase activity, which may be prevented in ischaemic preconditioning by the binding of a specific inhibitor protein. As the rat is known to have less inhibitor protein than other species, this study was designed to determine whether the rat heart could be ischaemically preconditioned. METHODS: Rats were anaesthetised with pentobarbitone, the chest opened and the hearts ischaemically preconditioned with a 5 min occlusion of the left main coronary artery followed by 10 min reperfusion. The hearts were then subjected to a 45 min occlusion followed by 3 h reperfusion. Control hearts were treated identically but without ischaemic preconditioning. Infarct size was measured using triphenyl tetrazolium and expressed as a percentage of the region at risk, measured with fluorescent particles. RESULTS: Infarct size as a percent of the risk area in the ischaemically preconditioned group (n = 8) was 31.4(SEM 6.1)%, versus 61.0(4.8)% in control hearts (n = 8) (p < 0.005). CONCLUSIONS: These results show that rat hearts can be ischaemically preconditioned and suggest that the protective mechanism involved in this phenomenon is not mediated through the endogenous inhibition of mitochondrial ATPase. An overall reduction in mitochondrial ATP "wastage" may not be the sole mechanism in the protection seen in ischaemic preconditioning.     

Ischaemic preconditioning: from molecular characterisation to clinical application--part II

Ischaemic preconditioning was originally described in animal hearts as histological infarct-size limitation by a previous brief episode of ischaemia. In humans, ischaemic preconditioning has been demonstrated in several in vitro and in vivo models, including coronary artery bypass grafting and percutaneous transluminal coronary angiograplasty, using surrogate markers of ischaemia and reperfusion injury. Increasing knowledge of the molecular signalling pathways mediating protection by ischaemic preconditioning has provided rational targets for pharmacological intervention. Several widely used drugs are able to mimic ischaemic preconditioning (e.g. adenosine, adenosine-uptake inhibitors, ACE inhibitors, angiotensin II antagonists, statins, opioids, volatile anaesthetics and ethanol), whereas others inhibit ischaemic preconditioning-induced protection (e.g. sulphonylureas and adenosine antagonists). The present review focuses on these different classes of drugs. Prudent use or avoidance of these drugs in patients who are at risk for myocardial infarction could theoretically limit ischaemia and reperfusion injury.     

Ischaemic preconditioning: from molecular characterisation to clinical application--part I

Ischaemic preconditioning is defined as an increased tolerance to ischaemia and reperfusion induced by a previous sublethal period of ischaemia. Since this is the most powerful mechanism for limiting infarct size, other than timely reperfusion, an overwhelming number of studies have addressed the way in which this form of protection occurs. During the short preconditioning period of ischaemia, several trigger substances are released (adenosine, bradykinin, norepinephrine, opioids). By activation of membrane-bound receptors, these substances activate a complex intracellular signalling cascade, which converges on mitochondrial end-effectors, including the ATP-sensitive potassium channel and the mitochondrial permeability transition pore. Activation of this pathway protects cardiomyocytes against both necrosis and apoptosis during a subsequent more prolonged ischaemic episode. The protection afforded by preconditioning lasts only two to three hours, but reappears 24 hours after the preconditioning stimulus. This 'delayed preconditioning' requires synthesis of new proteins, including inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and heat shock proteins. Additionally, preconditioning is not confined to one organ, but can also limit infarct size in remote, non-preconditioned organs ('remote preconditioning'). Knowledge of these mechanisms mediating ischaemic preconditioning is essential to understand which drugs are able to mimic preconditioning or interfere with pre-conditioning in patients at risk for myocardial ischaemia. This review aims to summarise current knowledge regarding the different forms and mechanisms of ischaemic preconditioning.     

Blood gas and plasma ion concentrations of asphyxiated hypothermic and normothermic rats

• 1.1. Cardiac punctures were made in four groups of rats during and after the time they were subjected to the procedures described for the induction of hypothermia with subsequent cardiac arrest.
• 2.2. The absolute amounts of oxygen were low and the carbon dioxide was high in the arterial and mixed venous blood. Even under these situations a large A-V oxygen and carbon-dioxide difference remained.
• 3.3. Although animals in all groups were acidotic, fibrillation was not observed on the electrocardiographic recordings during immersion of the animals of Groups AAH (ambient asphyxiation followed by hypothermia) and HAH (hypothermic asphyxiation followed by hypothermia) in the ice baths.
• 4.4. A decreased buffering capacity of the blood, as evidenced by the BHCO₃/H₂CO₃ ratio, was prominent especially in those groups ambiently asphyxiated.
• 5.5. Plasma K⁺ increased at least 80 per cent in all groups. However, plasma sodium exhibited a significant decrease only in Groups HA (hypothermic asphyxiation) and HAH.
• 6.6. It is suggested that one of the limiting factors for the induction of hypothermia with cardiac arrest is the absolute amount of oxygen available to the animal after respiratory arrest.
     

Ischaemic preconditioning and a mitochondrial KATP channel opener both produce cardioprotection accompanied by F1F0-ATPase inhibition in early ischaemia

Ischaemic preconditioning gives powerful protection against prolonged ischaemia affecting several intracellular regulatory and messenger pathways, although their mutual importance is far from established. Protective, preconditioning-like effects have been reported for K_ATP channel openers, and most of the evidence points to the mitochondrial K_ATP channels. We show here that the K_ATP channel opener diazoxide, which acts selectively on the mitochondrial channel, causes potentiation of ischaemic inhibition of mitochondrial ATP synthase (F₁F₀-ATPase) along with cardioprotection. These effects are comparable with that of ischaemic preconditioning. The administration of diazoxide did not affect the cellular energy state as monitored with ³¹P NMR. The actions of both diazoxide and ischaemic preconditioning were prevented by 5-hydroxydecanoate, a specific inhibitor of the mitochondrial K_ATP channel. Thus mitochondrial K_ATP channel opening and ischaemic preconditioning must share common mechanisms of action involving mitochondrial F₁F₀-ATPase, although involvement of the energy state in protection could not be proved. Received: 2 December 2002, Returned for revision: 9 January 2003, Revision received: 26 March 2003, Accepted: 8 April 2003, Published online: 9 May 2003 RID="*" ID="*"Publishers Note In the “online-first” publication of this article figure 3 and 4 were erroneously swapped. This mistake was corrected in the printed version and in the final online version on the authors request. The publisher apologizes for this oversight. Correspondence to: I. Hassinen     

Evidence that continuous normothermic blood cardioplegia offers better myocardial protection than intermittent hypothermic cardioplegia.

OBJECTIVES--To compare transmyocardial ischaemia and oxidative stress, as well as non-infarction myocardial injury, in patients randomised to intermittent hypothermic cardioplegia or continuous normothermic blood-potassium cardioplegia. DESIGN--Prospective randomised trial. SETTING--Tertiary cardiac referral centre. METHODS--24 patients undergoing elective coronary artery bypass surgery were randomised to hypothermic (13 patients, mean (SEM) age 59.5 (2.6) years) or normothermic (11 patients, mean (SEM) age 59.7 (3.3) years) cardioplegia. Transmyocardial oxidative stress and ischaemia were assessed by the difference in plasma concentrations of oxidised glutathione and lactate respectively, from samples taken simultaneously from the coronary sinus and aortic root. Blood samples were taken just before cross clamp application and at intervals up to 15 min after cross clamp release. Non-infarction myocardial injury was assessed by measurement of creatine kinase MB isoenzyme activity from peripheral venous blood taken 2 and 18 h after surgery. RESULTS--Intermittent hypothermic cardioplegia resulted in a significant increase in transmyocardial ischaemia (P < 0.001) and oxidative stress (P < 0.001). Evidence of significantly increased myocyte damage was also present (P < 0.01). No significant corresponding changes were present with normothermic cardioplegia. CONCLUSIONS--Normothermic blood cardioplegia seems to avoid significant changes in myocardial ischaemic status and consequent oxidative stress. This study provides direct evidence that normothermic cardioplegia offers enhanced myocardial protection compared with that of hypothermic cardioplegia. Certain subsets of patients may derive more benefit from normothermic cardioplegia, although it is unclear whether this would be the case for all patients.