Shiro Hoshida

Delayed effects of sublethal ischemia on the acquisition of tolerance to ischemia.

The infarct-limiting effect of ischemic preconditioning is believed to be a transient phenomenon. We examined the delayed effects of repetitive brief ischemia on limiting infarct size in an open-chest dog model by an occlusion (90 minutes) of the left anterior descending coronary artery (LAD) followed by reperfusion (5 hours). The dogs were preconditioned with four brief repeated ischemic episodes induced by 5-minute LAD occlusions with subsequent reperfusion. The size of infarcts initiated by a sustained occlusion immediately or 24 hours after preconditioning was significantly smaller when compared with infarcts in sham-operated dogs (for the immediate occlusion, 14.4 +/- 2.0% versus 39.0 +/- 3.7%, respectively [p < 0.01]; and for the delayed occlusion, 18.8 +/- 3.4% versus 35.1 +/- 4.6%, respectively [p < 0.05]); however, when the infarction was induced 3 hours (31.2 +/- 3.7% versus 37.5 +/- 4.2%, respectively) or 12 hours (25.4 +/- 4.8% versus 35.0 +/- 5.3%, respectively) after repetitive ischemia, the infarct size did not differ. No differences were seen in regional myocardial blood flow or rate-pressure products between the two groups. These results indicate that an infarct-limiting effect of brief repeated ischemia can be observed 24 hours after sublethal preconditioning.

Induction of manganese superoxide dismutase in rat cardiac myocytes increases tolerance to hypoxia 24 hours after preconditioning.

Manganese superoxide dismutase (Mn-SOD) is induced in ischemic hearts 24 h after ischemic preconditioning, when tolerance to ischemia is acquired. We examined the relationship between Mn-SOD induction and the protective effect of preconditioning using cultured rat cardiac myocytes. Exposure of cardiac myocytes to brief hypoxia (1 h) decreased creatine kinase release induced by sustained hypoxia (3 h) that follows when the sustained hypoxia was applied 24 h after hypoxic preconditioning (57% of that in cells without preconditioning). The activity and content of Mn-SOD in cardiac myocytes were increased 24 h after hypoxic preconditioning (activity, 170%; content, 139% compared with cells without preconditioning) coincidentally with the acquisition of tolerance to hypoxia. Mn-SOD mRNA was also increased 20-40 min after preconditioning. Antisense oligodeoxyribonucleotides corresponding to the initiation site of Mn-SOD translation inhibited the increases in the Mn-SOD content and activity and abolished the expected decrease in creatine kinase release induced by sustained hypoxia after 24 h of hypoxic preconditioning. Sense oligodeoxyribonucleotides did not abolish either Mn-SOD induction or tolerance to hypoxia. These results suggest that the induction of Mn-SOD in myocytes by preconditioning plays a pivotal role in the acquisition of tolerance to ischemia at a later phase (24 h) of ischemic preconditioning.

Whole-body hyperthermia provides biphasic cardioprotection against ischemia/reperfusion injury in the rat

BACKGROUND Hyperthermia increases cardiac tolerance to ischemia/reperfusion injury 24 hours after the heat stress. Free radicals and redox mechanisms have been implicated in such tolerance. However, the time course and its relation to the induction of antioxidative enzymes in the protection induced by whole-body hyperthermia against ischemia/reperfusion injury are unknown. METHODS AND RESULTS Hyperthermia was induced in anesthetized rats by placement in a temperature-controlled water bath. After the defined recovery interval(s) at room temperature, ischemia was induced by occlusion of the left coronary artery for 20 minutes, followed by reperfusion for 48 hours. The exposure to hyperthermia led to a recovery interval- dependent, biphasic reduction in the incidence of ventricular fibrillation during ischemia and in the size of the myocardial infarct as determined after 48 hours of reperfusion. The time course of the late-phase (24- to 96-hour recovery interval) but not the early-phase (0.5 hour) cardioprotection depended on the degree of hyperthermia. The time course of the increase in myocardial manganese superoxide dismutase (Mn-SOD) activity corresponded to that of the cardioprotective effects, although an increase in the content of Mn-SOD and of heat shock protein 72 corresponded only to the late-phase effects. Administration of an antioxidant before hyperthermia abolished the early- and late-phase cardioprotection and the increase in Mn-SOD activity. CONCLUSIONS THe activation of Mn-SOD mediated by free radical production during hyperthermia is important in the acquisition of early-phase and late-phase cardioprotection against ischemia/reperfusion injury in rats.

Relationship between effects of statins, aspirin and angiotensin II modulators on high-sensitive C-reactive protein levels

Statins, aspirin and angiotensin II modulators (A II-M: angiotensin-converting enzyme (ACE) inhibitors and angiotensin II type I receptor blockades) may have an anti-inflammatory effect, but the relationship between the effects of statins, aspirin and A II-M on high-sensitive C-reactive protein (hs-CRP) levels remains to be determined. We examined serum hs-CRP levels in consecutive patients with stable ischemic heart disease (IHD) (n=1231; 65+/-9 years; male/female, 927/304) and without IHD (n=226; 64+/-9 years; male/female, 117/109). Blood samples were collected on the day of catheterization. The hs-CRP levels were significantly higher in the IHD than in the non-IHD patients (0.32+/-0.52 vs. 0.24+/-0.29 mg/dl, P<0.05). Treatment with statins was associated with significantly lower hs-CRP levels in both groups (non-IHD, 0.17+/-0.14 vs. 0.26+/-0.31 mg/dl; IHD, 0.27+/-0.34 vs. 0.35+/-0.59 mg/dl; both P<0.05). hs-CRP levels were significantly lower only in IHD patients treated with A II-M than in those not treated with A II-M (0.28+/-0.34 vs. 0.34+/-0.58 mg/dl, P<0.05). Aspirin did not have any effect on the hs-CRP level in either group. The hs-CRP levels were significantly lower in IHD patients treated with statins and/or A II-M than those treated with neither statins nor A II-M (statin+/A II-M+, 0.28+/-0.29 mg/dl; statin+/A II-M-, 0.26+/-0.36 mg/dl; statin-/A II-M+, 0.28+/-0.37 mg/dl; statin-/A II-M-, 0.38+/-0.66 mg/dl; P<0.01). These results indicate that statins and A II-M, but not aspirin, in commonly used doses have an anti-inflammatory action as assessed by measurement of CRP levels in IHD patients.

The importance of manganese superoxide dismutase in delayed preconditioning: Involvement of reactive oxygen species and cytokines

It is clinically important to elucidate the mechanism underlying the delayed preconditioning against ischemia-reperfusion injury observed 24-72 h after sublethal stress such as brief ischemia, hyperthermia and exercise. The time course of induction of myocardial manganese-superoxide dismutase (Mn-SOD) and appearance of the ischemic tolerance coincide well, and the percent increase in Mn-SOD activity and percent reduction of infarct size are correlated well under various stresses. Furthermore, treatments with antisense oligodeoxynucleotides to Mn-SOD completely abolished the delayed preconditioning and any increase in Mn-SOD content. These results indicate that Mn-SOD induction plays a pivotal role in the late phase preconditioning afforded with brief ischemia, hyperthermia and exercise. We also showed that cytokines, e.g., tumor necrosis factor-alpha and interleukin-1beta, and reactive oxygen species are involved in the process of signal transduction for the Mn-SOD induction.

Increased Angiotensin-Converting Enzyme Activity in Coronary Artery Specimens From Patients With Acute Coronary Syndrome

Background —Angiotensin-converting enzyme (ACE) inhibitors are effective in the secondary prevention of ischemic heart disease, but they do not reduce the rate of restenosis. Vascular ACE activity in the culprit coronary lesions of these patients, however, has never been quantified. Methods and Results —We measured the ACE activity of vascular tissue obtained by directional coronary atherectomy in patients with acute coronary syndrome (n=17) and in patients with stable ischemic heart disease (n=36), with and without restenosis. The ACE activity of the culprit coronary lesions was significantly increased in patients with acute coronary syndrome (0.87±0.12 nmol · min–1 · mg protein–1;P <0.01) but not in patients with ischemic heart disease with restenosis (n=11, 0.19±0.05 nmol · min–1 · mg protein–1) when compared with those patients with ischemic heart disease without restenosis (n=25, 0.20±0.05 nmol · min–1 · mg protein–1). There was no difference between the ACE activity of the coronary tissue of the in-stent (n=5) and stent-unrelated (n=6) restenosis patients (0.24±0.10 versus 0.15±0.04 nmol · min–1 · mg protein–1). Serum ACE activity did not differ significantly among the patients. Conclusions —The present study demonstrates increased ACE activity in culprit lesions in acute coronary syndrome, indicating that enhanced ACE activity is related to the causative mechanism of active coronary lesions.

The involvement of cytokines in the second window of ischaemic preconditioning.

We utilized a rat model of myocardial infarction to investigate whether manganese superoxide dismutase (Mn‐SOD), an intrinsic radical scavenger, and tumour necrosis factor‐ α (TNF‐α) and/or interleukin‐1β (IL‐1β) are involved in the late phase of ischaemic preconditioning (IP). IP was induced in anaesthetized rats by four 3‐min left coronary artery (LCA) occlusions, each separated by 10 min of reperfusion. Twenty‐four hours after the repetitive brief ischaemia, the LCA was occluded for 20 min followed by reperfusion for 48 h. IP reduced the infarct size by approximately 46% as determined after 48 h of reperfusion. Antisense oligodeoxynucleotides to Mn‐SOD inhibited the increases in Mn‐SOD content and activity, and abolished the expected decrease in myocardial infarct size. Sense or scrambled oligodeoxynucleotides did not abolish either Mn‐SOD induction or tolerance to ischaemia/reperfusion. The simultaneous administration of the antibodies to TNF‐α (0.5 ml) and IL‐1β (0.5 mg) prior to IP abolished the cardioprotection and the increase in Mn‐SOD activity induced by IP. We conclude that the induction and activation of Mn‐SOD, mediated by TNF‐α and IL‐1β after IP, plays an important role in the acquisition of late‐phase cardioprotection against ischaemia/reperfusion injury in rats.

Repeated physiologic stresses provide persistent cardioprotection against ischemia-reperfusion injury in rats

OBJECTIVES We investigated the time course of myocardial tolerance to ischemia-reperfusion injury after repeated physiologic or pharmacologic stresses. BACKGROUND Sublethal stress provides cardiac tolerance to ischemia-reperfusion injury and increases the activity of manganese superoxide dismutase (Mn-SOD) in the myocardium in a biphasic manner. However, few studies have investigated the time course of the cardioprotective effects after repeated stresses. METHODS One or two episodes of the same physiologic or pharmacologic stress (exercise, whole-body hyperthermia, or tumor necrosis factor-alpha treatment), or a combination of two different types of stress, were induced after a 48-h interval. The rats were then subjected to 20 min of left coronary artery occlusion, followed by 48 h of reperfusion. The interval between the last stimulus and the induced ischemia was between 0.5 h and 168 h. The incidence of ventricular fibrillation during ischemia and the size of the myocardial infarct after reperfusion were then examined. RESULTS When two episodes of physiologic or pharmacologic stress were induced, the beneficial effects against ischemia-reperfusion injury were observed in a monophasic manner. These effects persisted for a period of 0.5 to 60 h. One episode of sublethal stress provoked the same beneficial effects, but in a biphasic manner. The increase in Mn-SOD activity in the cardiac tissue resembled the time course for cardioprotection against ischemia-reperfusion injury. CONCLUSIONS Two episodes of physiologic or pharmacologic stress can provide persistent cardioprotective effects against ischemia-reperfusion injury.

Amelioration by Quinapril of Myocardial Infarction Induced by Coronary Occlusion/Reperfusion in a Rabbit Model of Atherosclerosis Possible Mechanisms

BACKGROUND The increased severity of the myocardial injury produced by coronary occlusion-reperfusion in models of atherosclerosis is associated with an increase in leukocyte accumulation in the ischemic myocardium. Expression of P-selectin, an adhesion molecule involved in the interaction between leukocytes and endothelium, is increased in atherosclerotic vessels. Long-term angiotensin-converting enzyme (ACE) inhibition has been shown to reduce atherosclerotic vascular change in experimental models. METHODS AND RESULTS We examined changes in the size of the infarct resulting from coronary occlusion/reperfusion in normally fed and cholesterol-fed rabbits that were chronically treated with quinapril. Infarct size was significantly larger in the cholesterol-fed versus normally fed rabbits. ACE activity in the ischemic and nonischemic myocardium was significantly reduced by quinapril. Chronic quinapril administration significantly ameliorated the increased myocardial injury in cholesterol-fed rabbits. Quinapril administration markedly increased the myocardial cGMP content and reduced the myeloperoxidase activity in the border region of the ischemic myocardium in cholesterol-fed rabbits. The enhanced expression of P-selectin in myocardial tissue of cholesterol-fed rabbits was also effectively reduced by quinapril treatment. The above effects of quinapril were eliminated by blockade of bradykinin B2 receptors or inhibition of nitric oxide synthesis. CONCLUSIONS Chronic quinapril treatment ameliorated the severity of myocardial injury produced by coronary occlusion/reperfusion in cholesterol-fed rabbits, possibly because of reversal of the enhanced interaction between leukocytes and endothelium in the ischemic myocardium via a bradykinin-related pathway.

Vascular angiotensin-converting enzyme activity in cholesterol-fed rabbits : effects of enalapril

Many reports have shown inhibitory effects of angiotensin-converting enzyme (ACE) inhibitors on the progression of atherosclerotic plaque lesions in vascular tissue of experimental models. However, no report has shown alterations of ACE activity in vascular tissue during the process of atherosclerosis. We measured ACE activity in plasma and aortic tissue in rabbits fed a cholesterol-rich (1%) or normal diet for 10 weeks. We also evaluated the blood pressure response to angiotensin (Ang) I and II. These data were compared in untreated rabbits and in rabbits receiving chronic treatment with an ACE inhibitor, enalapril (3 mg/kg/day for 10 weeks). ACE activity in aortic tissue, but not in plasma, in cholesterol-fed rabbits was gradually but significantly increased compared with that in noncholesterol-fed rabbits even after the 4-week feeding period, when no atherosclerotic lesion was observed in the aortic tissue. Treatment with enalapril for 10 weeks, but not 4 weeks, significantly reduced the ACE activity in aortic tissue in association with the reductions in the elevated Ang II level and the atherosclerotic plaque area of the aortic tissue. These results indicated that ACE activity in aortic tissue was increased during the early phase of atherosclerotic process.