Reactive oxygen species are critical mediators of coronary collateral development in a canine model

Recent evidence suggests that reactive oxygen species (ROS) promote proliferation and migration of vascular smooth muscle (VSMC) and endothelial cells (EC). We tested the hypothesis that ROS serve as crucial messengers during coronary collateral development. Dogs were subjected to brief (2 min), repetitive coronary artery occlusions (1/h, 8/day, 21 day duration) in the absence (occlusion, n = 8) or presence of N-acetylcysteine (NAC) (occlusion + NAC, n = 8). A sham group (n = 8) was instrumented identically but received no occlusions. In separate experiments, ROS generation after a single 2-min coronary artery occlusion was assessed with dihydroethidium fluorescence. Coronary collateral blood flow (expressed as a percentage of normal zone flow) was significantly increased (71 +/- 7%) in occlusion dogs after 21 days but remained unchanged (13 +/- 3%) in sham dogs. Treatment with NAC attenuated increases in collateral blood flow (28 +/- 8%). Brief coronary artery occlusion and reperfusion caused ROS production (256 +/- 33% of baseline values), which was abolished with NAC (104 +/- 12%). Myocardial interstitial fluid produced tube formation and proliferation of VSMC and EC in occlusion but not in NAC-treated or sham dogs. The results indicate that ROS are critical for the development of the coronary collateral circulation.     

Autologous vascular smooth muscle cell-based myocardial gene therapy to induce coronary collateral growth

For therapeutic angiogenesis to achieve clinical relevance, it must be effective, with minimal side effects to other end organ systems. We developed a cardiac-specific gene delivery mechanism by transfecting autologous vascular smooth muscle cells (VSMC) with VEGF and administering these cells via intracoronary injection. We evaluated the efficacy of this protocol by its ability to stimulate angiogenesis in the presence of a subthreshold stimulus for collateralization. A modified canine repetitive coronary occlusion model was utilized in these experiments with left anterior descending coronary artery occlusions for 2 min every 2 h four times per day for 21 days. An intramyocardial catheter in the perfusion territory of the left anterior descending coronary artery measured proteins in the myocardial interstitial fluid. VSMC from jugular vein explants were isolated, amplified in culture for 3 wk, and transfected with a plasmid expressing VEGF-165 and/or enhanced green fluorescent protein. Cells were injected before commencement of occlusions. VEGF levels in myocardial interstitial fluid were significantly higher in VEGF-transfected animals than in sham (repetitive occlusions without cell transplantation) and control (repetitive occlusions with enhanced green fluorescent protein-transfected cells) animals at the onset of occlusions (P < 0.05). In the VEGF group, collateral flow was increased at day 7 and remained higher than in sham and control groups thereafter. We found that intracoronary administration of VEGF-transfected autologous VSMC effectively promotes collateral development. This approach may provide a way to confine delivery of a gene to a specified organ, thus minimizing complications related to gene transfection in nontargeted organ systems.     

Effects of exercise on collateral development in myocardial ischemia in pigs

To study the effects of exercise on collateral development in myocardial ischemia, we induced coronary arterial stenosis of the left circumflex coronary artery (LCCA) in 18 of 30 pigs. During that surgery, we identified the coronary bed at risk. Nine of these pigs were then subjected to 5 mo of exercise training on a treadmill. After exercise training, we determined regional collateral and myocardial blood flow using radiolabeled microspheres. At autopsy, all animals had complete occlusion of the LCCA. Infarct size in the exercise-trained pigs was significantly less than in the sedentary pigs (5.9 +/- 1.0 vs. 11.7 +/- 1.0% of the left ventricle). The exercise-trained animals had a greater increase in collateral flow, 35.1 +/- 3.0 vs. 28.7 +/- 4.1 ml X min-1 X 100 g-1, in the noninfarcted jeopardized zone of the LCCA bed. The major findings of the study were the following: 1) chronic coronary artery stenosis progressing to occlusion stimulated development of the collateral circulation and salvaged tissue in the jeopardized myocardium of an animal model with sparse collaterals; 2) development of the collateral circulation and tissue salvage is increased by exercise training; 3) collaterals develop primarily in or near the ischemic zone; and 4) all collateral beds develop a circumferential flow gradient following occlusion.     

Optimal reactive oxygen species concentration and p38 MAP kinase are required for coronary collateral growth

Reactive oxygen species (ROS) are implicated in coronary collateral growth (CCG). We evaluated the requirement for ROS in human coronary artery endothelial cell (HCAEC) tube formation, CCG in vivo, and signaling (p38 MAP kinase) by which ROS may stimulate vascular growth. The flavin-containing oxidase inhibitor diphenyleneiodonium (DPI) or the superoxide dismutase inhibitor diethyldithiocarbamate (DETC) blocked vascular endothelial growth factor-induced HCAEC tube formation in Matrigel. We assessed the effect of DPI and DETC on CCG in a rat model of repetitive ischemia (RI) (40 s left anterior descending coronary artery occlusion every 20 min for 2 h 20 min, 3 times/day, 10 days). DPI or DETC was given intraperitoneally, or the NAD(P)H oxidase inhibitor apocynin was given in drinking water. Collateral-dependent flow (measured by using microspheres) was expressed as a ratio of normal and ischemic zone flows. In sham-operated rats, collateral flow in the ischemic zone was 18 +/- 6% of normal zone; in the RI group, collateral flow in the ischemic zone was 83 +/- 5% of normal zone. DPI prevented the increase in collateral flow after RI (25 +/- 4% of normal zone). Similar results were obtained with apocynin following RI (32 +/- 7% of that in the normal zone). DETC achieved similar results (collateral flow after RI was 21 +/- 2% of normal zone). DPI and DETC blocked RI-induced p38 MAP kinase activation in response to vascular endothelial growth factor and RI. These results demonstrate a requirement for optimal ROS concentration in HCAEC tube formation, CCG, and p38 MAP kinase activation. p38 MAP kinase inhibition prevented HCAEC tube formation and partially blocked RI-induced CCG (42 +/- 7% of normal zone flow), indicating that p38 MAP kinase is a critical signaling mediator of CCG.     

Coronary sinus occlusion enhances coronary collateral flow and reduces subendocardial ischemia

On the hypothesis that coronary sinus occlusion (CSO) may reduce myocardial ischemia, we examined the effects of CSO on coronary collateral blood flow and on the distribution of regional myocardial blood flow (RMBF) in dogs. Thirty-eight anesthetized dogs underwent occlusion of the left anterior descending coronary artery with or without CSO and intact vasomotor tone. We measured RMBF and intramyocardial pressure (IMP) in the subendocardium (Endo) and subepicardium (Epi) separately. With intact vasomotor tone, CSO during ischemia significantly increased RMBF in the ischemic region (IR), particularly in Endo from 0.17 +/- 0.03 to 0.33 +/- 0.05 ml x min(-1) x g(-1) (P < 0.05), and increased the Endo/Epi from 0.59 +/- 0.10 to 1.15 +/- 0.15 (P < 0.01). These effects of CSO were partially abolished by adenosine. However, the Endo/Epi was still increased from 0.90 +/- 0.13 to 2.09 +/- 0.30 (P < 0.01). The changes in RMBF in IR were significantly correlated with the peak CS pressure during CSO. The Endo/Epi of IMP in IR was significantly decreased during CSO. In conclusion, CSO potentially enhances coronary collateral flow, and preserves the ischemic myocardium, especially in Endo.     

Ischemic postconditioning during reperfusion activates Akt and ERK without protecting against lethal myocardial ischemia-reperfusion injury in pigs

Transient episodes of ischemic preconditioning (PC) render myocardium protected against subsequent lethal injury after ischemia and reperfusion. Recent studies indicate that application of short, repetitive ischemia only during the onset of reperfusion after the lethal ischemic event may obtain equivalent protection. We assessed whether such ischemic postconditioning (Postcon) is cardioprotective in pigs by limiting lethal injury. Pentobarbital sodium-anesthetized, open-chest pigs underwent 30 min of complete occlusion of the left anterior descending coronary artery and 3-h reflow. PC was elicited by two cycles of 5-min occlusion plus 10-min reperfusion before the 30-min occlusion period. Postcon was elicited by three cycles of 30-s reperfusion, followed by 30-s reocclusion, after the 30-min occlusion period and before the 3-h reflow. Infarct size (%area-at-risk using triphenyltetrazolium chloride macrochemistry; means +/- SE) after 30 min of ischemia was 26.5 +/- 5.2% (n = 7 hearts/treatment group). PC markedly limited myocardial infarct size (2.8 +/- 1.2%, n = 7 hearts/treatment group, P < 0.05 vs. controls). However, Postcon had no effect on infarct size (37.8 +/- 5.1%, n = 7 hearts/treatment group). Within the subendocardium, Postcon increased phosphorylation of Akt (74 +/- 12%) and ERK1/2 (56 +/- 10%) compared with control hearts subjected only to 30-min occlusion and 15-min reperfusion (P < or = 0.05), and these changes were not different from the response triggered by PC (n = 5 hearts/treatment group). Phosphorylation of downstream p70S6K was also equivalent in PC and Postcon groups. These data do not support the hypothesis that application of 30-s cycles of repetitive ischemia during reperfusion exerts a protective effect on pig hearts subjected to lethal ischemia, but this is not due to a failure to phosphorylate ERK and Akt during early reperfusion.     

犬心肌缺血早期血小板聚集功能和冠脉侧支循环的改变

本实验在18只麻醉开胸犬观察了急性心肌缺血早期血小板聚集功能和冠脉侧支循环功能的变化。实验结果如下:阻断冠脉后心肌缺血区血液中血小板聚集率(PAgR)增大,血小板计数(PC)减少。缺血50min时,PAgR增大58.7±5.6％,PC减少39.5±23.6％,与对照值有明显差异(均为P<0.01)。与此同时,在控制血压条件下,心肌缺血早期单位压力差下冠脉侧支血流量的变化与对照值无明显差异,而根据Wyatt等公式计算的流经缺血区末梢血管的有效侧支血流量明显降低,缺血50min时较对照值降低23.5±9.7％(P<0.05)。PAgR变化与有效侧支血流量改变呈明显负相关(r=-0.887,P<0.01);冠脉侧支指数与梗塞范围呈明显负相关(r=-0.847,P<0.01)。阻断冠脉前静脉注射血小板聚集功能抑制剂阿斯匹林,可明显减轻上述各项参数的异常变化。这些结果提示,心肌缺血早期血小板聚集功能的异常变化虽然对冠脉侧支血管的血流阻力影响较小,但却使流经缺血区末梢血管的有效侧支血流量明显减小,进而扩大梗塞范围。     

Effects of exercise training on coronary collateralization and control of collateral resistance

Coronary collateral vessels serve as a natural protective mechanism to provide coronary flow to ischemic myocardium secondary to critical coronary artery stenosis. The innate collateral circulation of the normal human heart is typically minimal and considerable variability occurs in extent of collateralization in coronary artery disease patients. A well-developed collateral circulation has been documented to exert protective effects upon myocardial perfusion, contractile function, infarct size, and electrocardiographic abnormalities. Thus therapeutic augmentation of collateral vessel development and/or functional adaptations in collateral and collateral-dependent arteries to reduce resistance into the ischemic myocardium represent a desirable goal in the management of coronary artery disease. Tremendous evidence has provided documentation for the therapeutic benefits of exercise training programs in patients with coronary artery disease (and collateralization); mechanisms that underlie these benefits are numerous and multifaceted, and currently under investigation in multiple laboratories worldwide. The role of enhanced collateralization as a major beneficial contributor has not been fully resolved. This topical review highlights literature that examines the effects of exercise training on collateralization in the diseased heart, as well as effects of exercise training on vascular endothelial and smooth muscle control of regional coronary tone in the collateralized heart. Future directions for research in this area involve further delineation of cellular/molecular mechanisms involved in effects of exercise training on collateralized myocardium, as well as development of novel therapies based on emerging concepts regarding exercise training and coronary artery disease.     

Effects of autonomic blockers on linear and nonlinear indexes of blood pressure and heart rate in SHR

Endothelium-derived hyperpolarizing factor (EDHF) is released in response to agonists such as ACh and bradykinin and regulates vascular smooth muscle tone. Several studies have indicated that ouabain blocks agonist-induced, endothelium-dependent hyperpolarization of smooth muscle. We have demonstrated that epoxyeicosatrienoic acids (EETs), cytochrome P-450 metabolites of arachidonic acid, function as EDHFs. To further test the hypothesis that EETs represent EDHFs, we have examined the effects of ouabain on the electrical and mechanical effects of 14,15- and 11,12-EET in bovine coronary arteries. These arteries are relaxed in a concentration-dependent manner to 14,15- and 11,12-EET (EC(50) = 6 x 10(-7) M), bradykinin (EC(50) = 1 x 10(-9) M), sodium nitroprusside (SNP; EC(50) = 2 x 10(-7) M), and bimakalim (BMK; EC(50) = 1 x 10(-7) M). 11,12-EET-induced relaxations were identical in vessels with and without an endothelium. Potassium chloride (1-15 x 10(-3) M) inhibited [(3)H]ouabain binding to smooth muscle cells but failed to relax the arteries. Ouabain (10(-5) to 10(-4) M) increased basal tone and inhibited the relaxations to bradykinin, 11,12-EET, and 14,15-EET, but not to SNP or BMK. Barium (3 x 10(-5) M) did not alter EET-induced relaxations and ouabain plus barium was similar to ouabain alone. Resting membrane potential (E(m)) of isolated smooth muscle cells was -50.2 +/- 0.5 mV. Ouabain (3 x 10(-5) and 1 x 10(-4) M) decreased E(m) (-48.4 +/- 0.2 mV), whereas 11,12-EET (10(-7) M) increased E(m) (-59.2 +/- 2.2 mV). Ouabain inhibited the 11,12-EET-induced increase in E(m). In cell-attached patch clamp studies, 11,12-EET significantly increased the open-state probability (NP(o)) of a calcium-activated potassium channel compared with control cells (0.26 +/- 0.06 vs. 0.02 +/- 0.01). Ouabain did not change NP(o) but blocked the 14,15-EET-induced increase in NP(o). These results indicate that: 1) EETs relax coronary arteries in an endothelium-independent manner, 2) unlike EETs, potassium chloride does not relax the coronary artery, and 3) ouabain inhibits bradykinin- and EET-induced relaxations as has been reported for EDHF. These findings provide further evidence that EETs are EDHFs.     

Hyperglycemia reduces coronary collateral blood flow through a nitric oxide-mediated mechanism

We tested the hypothesis that hyperglycemia alters retrograde coronary collateral blood flow by a nitric oxide-mediated mechanism in a canine Ameriod constrictor model of enhanced collateral development. Administration of 15% dextrose to increase blood glucose concentration to 400 or 600 mg/dl decreased retrograde blood flow through the left anterior descending coronary artery to 78 +/- 9 and 82 +/- 8% of baseline values, respectively. In contrast, saline or L-arginine (400 mg x kg(-1) x h(-1)) had no effect on retrograde flow. Coronary hypoperfusion and 1 h of reperfusion decreased retrograde blood flow similarly in saline- or L-arginine-treated dogs (76 +/- 11 and 89 +/- 4% of baseline, respectively), but these decreases were more pronounced in hyperglycemic dogs (47 +/- 10%). L-arginine prevented decreases in retrograde coronary collateral blood flow during hyperglycemia (100 +/- 5 and 95 +/- 6% of baseline at blood glucose concentrations of 400 and 600 mg/dl, respectively) and after coronary hypoperfusion and reperfusion (84 +/- 14%). The results suggest that hyperglycemia decreases retrograde coronary collateral blood flow by adversely affecting nitric oxide availability.     

Influence of low-flow infusion and magnesium on tissue necrosis during regional ischemia in the canine myocardium.

Passive intracoronary perfusion of therapeutic agents has been used in the clinical setting to attenuate the effects of brief episodes of myocardial ischemia. The objective of this study was to assess the effects of low-flow coronary infusion with or without Mg2+ on tissue necrosis and cardiac hemodynamics after prolonged regional ischemia. In 33 anesthetized dogs (5 excluded during study), the left anterior descending coronary artery was occluded for 6 h. Dogs were assigned to three groups: the first group (n = 8) was subjected to 6 h coronary occlusion without low-flow perfusion (controls), the second group (n = 10) received a low-flow coronary infusion of Ringer's lactate (Mg(2+)-free), and the third group (n = 10) received a low-flow coronary infusion of Ringer's lactate plus Mg2+ sulfate (15 mM). Tissue necrosis was evaluated using tetrazolium staining and was normalized to the principal baseline predictors of infarct size including anatomic risk zone (microsphere autoradiography) and coronary collateral flow. In control hearts, infarct size comprised 51.1 +/- 4.1% of the risk zone (40.8 +/- 5.1% left ventricular cross-sectional area (LV)). In the Mg(2+)-free and Mg2+ groups, risk zone size was 17.3 +/- 2.2 and 16.8 +/- 1.8% LV (p < 0.05 vs. controls), while infarct size was 23.1 +/- 3.1 and 24.9 +/- 8.1% (p < 0.05 vs. controls), respectively. Coronary collateral flow in the endocardium was similar for all of the experimental groups; however, hearts subjected to ischemia with low-flow perfusion of Ringer's lactate demonstrated significantly higher epicardial coronary collateral flow levels compared with controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of thromboxane A2 in reperfusion injury

Thromboxane A2 (TXA2) receptor antagonists can limit infarct size in models of coronary occlusion and reperfusion, but it was unknown if these compounds can mitigate reperfusion injury. Anesthetized open chest dogs were subjected to left circumflex coronary (LCX) occlusion for 90 min. Two minutes before reperfusion, the dogs were given iv saline (0.9% NaCl) or the TXA2 antagonist SQ 29,548 (0.2 mg/kg + 0.2 mg/kg/hr). Reperfusion was instituted for 5 hr at which time infarct size was determined. Regional myocardial blood flow was determined before, during, and after occlusion. SQ 29,548 treatment resulted in a significant reduction in infarct size (57 +/- 7 and 34 +/- 8% of the left ventricular area at risk infarcted in the saline and SQ 29,548 groups, respectively). No differences in collateral flow during occlusion were observed between groups, but SQ 29,548 treatment resulted in a significantly higher subendocardial reperfusion flow (54 +/- 10 and 93 +/- 14 ml/min/100g for the saline and SQ 29,548 groups, respectively). Thus, TXA2 seems to play a role in exacerbating reperfusion injury and TXA2 receptor blockade may have potential as a mode of therapy for ischemia-reperfusion damage.     

Effects of coronary ischemia on lung fluid balance in conscious sheep

It has been suggested that coronary ischemia increases extravascular lung water. To determine whether pulmonary microvascular permeability is increased by coronary ischemia, we measured pulmonary hemodynamics, lung lymph flow (QL), and lymph-to-plasma protein concentration ratio (L/P) in 12 sheep with chronic lung lymph fistulas. Studies were done in 3 groups: in group 1 (n = 7) a marginal branch of the left circumflex artery (Lcx) was occluded, in group 2 (n = 5) left atrial pressure (Pla) was mechanically raised by 10 mmHg, and in group 3 (n = 5) Lcx was occluded and Pla was raised by 10 mmHg. In group 1, coronary occlusion increased QL (4.6 +/- 0.4 to 8.3 +/- 2.6 ml/h) without changes in L/P. In group 2, elevated Pla increased QL (5.1 +/- 1.2 to 10.1 +/- 3.0 ml/h) with decreases in L/P (0.71 +/- 0.02 to 0.61 +/- 0.02). In group 3, coronary occlusion with elevated Pla caused a further increase in QL (5.0 +/- 1.5 to 16.9 +/- 4.6 ml/h) without significant decreases in L/P (0.71 +/- 0.01 to 0.65 +/- 0.06). Lung lymph concentrations of 6-keto-prostaglandin F1 alpha (a degradation product of prostacyclin) increased transiently after coronary occlusion. These results indicate that coronary occlusion can increase transcapillary protein transport in lungs of conscious sheep and simultaneously increase prostacyclin production in the lung.     

Early treatment with deferoxamine limits myocardial ischemic/reperfusion injury

Oxygen-derived free radicals (the superoxide anion O2- and hydroxyl radical.OH) have been implicated in myocardial injury associated with coronary artery occlusion followed by reperfusion. Transition metals (such as iron or copper) are needed to catalyze the formation of the .OH radical and subsequent .OH-mediated lipid peroxidation, yet the role of these transition metals in the pathogenesis of myocyte necrosis remains undefined. To address this issue, 21 dogs underwent 2 h of coronary artery occlusion and 4 h of reperfusion. Each animal was randomly assigned into 1 of 3 treatment groups: 7 received the iron chelator deferoxamine beginning 30 min preocclusion, 7 received deferoxamine beginning 5 min prior to reperfusion, while 7 dogs served as saline controls. Deferoxamine effectively chelated free iron in both treatment groups (total urine iron content averaged 42 +/- 16, 662 +/- 177 and 803 +/- 2.5 micrograms in control, pretreated, and deferoxamine at reperfusion groups respectively; p less than 0.05), but had no significant effect on in vivo area at risk (AR), hemodynamic parameters, collateral blood flow during occlusion, or myocardial blood flow following reperfusion. Area of necrosis (AN) in dogs pretreated with deferoxamine (34.6 +/- 3.7% of the AR; p less than 0.05) was significantly smaller than that observed in the saline control group (55.4 +/- 4.7% of the AR). Deferoxamine administered at the time of reperfusion, however, had no significant effect on infarct size (AN/AR = 54.3 +/- 8.7%, p = NS vs. controls). Thus, early treatment with the iron chelator deferoxamine acutely reduced the extent of myocyte necrosis produced by 2 h of transient coronary artery occlusion in the canine model.(ABSTRACT TRUNCATED AT 250 WORDS)     

A Device for Performing Automated Balloon Catheter Inflation Ischemia Studies

Coronary collateral growth (arteriogenesis) is a physiological adaptive response to transient and repetitive occlusion of major coronary arteries in which small arterioles (native collaterals) with minimal to no blood flow remodel into larger conduit arteries capable of supplying adequate perfusion to tissue distal to the site of occlusion. The ability to reliably and reproducibly mimic transient, repetitive coronary artery occlusion (ischemia) in animal models is critical to the development of therapies to restore coronary collateral development in type II diabetes and the metabolic syndrome. Current animal models for repetitive coronary artery occlusion implement a pneumatic occluder (balloon) that is secured onto the surface of the heart with the suture, which is inflated manually, via a catheter connected to syringe, to effect occlusion of the left anterior descending coronary artery (LAD). This method, although effective, presents complications in terms of reproducibility and practicality. To address these limitations, we have designed a device for automated, transient inflation of balloon catheters in coronary artery occlusion models. This device allows repeated, consistent inflation (to either specified pressure or volume) and the capability for implementing very complex, month-long protocols. This system has significantly increased the reproducibility of coronary collateral growth studies in our laboratory, resulting in a significant decrease in the numbers of animals needed to complete each study while relieving laboratory personnel from the burden of extra working hours and enabling us to continue studies over periods when we previously could not. In this paper, we present all details necessary for construction and operation of the inflator. In addition, all of the components for this device are commercially available and economical (Table S1). It is our hope that the adoption of automated balloon catheter inflation protocols will improve the experimental reliability of transient ischemia studies at many research institutions.     

Serum Cystatin C Reflects Angiographic Coronary Collateralization in Stable Coronary Artery Disease Patients with Chronic Total Occlusion

Objective

We investigated whether and to what extent cystatin C was associated with angiographic coronary collateralization in patients with stable coronary artery disease and chronic total occlusion.

Methods

Serum levels of cystatin C and high-sensitive C-reactive protein (hsCRP) and glomerular filtration rate (GFR) were determined in 866 patients with stable angina and angiographic total occlusion of at least one major coronary artery. The degree of collaterals supplying the distal aspect of a total occlusion from the contra-lateral vessel was graded as poor (Rentrop score of 0 or 1) or good coronary collateralization (Rentrop score of 2 or 3).

Results

In total, serum cystatin C was higher in patients with poor collateralization than in those with good collateralization (1.08 ± 0.32 mg/L vs. 0.90 ± 0.34 mg/L, P < 0.001), and correlated inversely with Rentrop score (adjusted Spearmen’s r = -0.145, P < 0.001). The prevalence of poor coronary collateralization increased stepwise with increasing cystatin C quartiles (P for trend < 0.001). After adjusting for age, gender, risk factors for coronary artery disease, GFR and hsCRP, serum cystatin C ≥ 0.97 mg/L remained independently associated with poor collateralization (OR 2.374, 95% CI 1.660 ~ 3.396, P < 0.001). The diagnostic value of cystatin C levels for detecting poor coronary collateralization persisted regardless of age, gender, presence or absence of diabetes, hypertension or renal dysfunction.

Conclusions

Serum cystatin C reflects angiographic coronary collateralization in patients with stable coronary artery disease, and cystatin C ≥ 0.97 mg/L indicates a great risk of poor coronary collaterals.     

Progression of myocardial injury during coronary occlusion in the collateral-deficient heart: a non-wavefront phenomenon

It is widely accepted that, during acute coronary occlusion, ischemic cell death progresses from the subendocardium to the subepicardium in a wavefront fashion. This concept, which implies that the subendocardium is the most susceptible myocardial region to ischemic injury, was established using a canine model with an extensive system of subepicardial coronary collaterals. In humans, particularly in those with coronary artery disease, there is a wide range in the distribution and functional capacity of the collateral circulation, which may affect the pattern of infarct evolution. Using an ovine model with a limited system of preformed subendocardial coronary collaterals, we characterized the effect of increasing lengths of ischemia on regional blood flow and infarct size in three regions of the ventricular wall: subendocardium, midmyocardium, and subepicardium. Our results demonstrate that the myocardium and microvasculature in these three regions are equally susceptible to injury after 45 min of ischemia. When ischemic time is increased to 1 h, infarct size in the midmyocardium (90 +/- 2%) is greater than in the subendocardium (76 +/- 4%, P = 0.004) and subepicardium (84 +/- 3%, P = 0.13). Microvascular dysfunction as assessed as a percentage of baseline flow is also greater in the midmyocardium (14 +/- 5%) compared with the subendocardium (20 +/- 3%, P = 0.23) and subepicardium (51 +/- 9%, P = 0.007). These findings suggest that, in subjects with a limited system of coronary collateral circulation, the midmyocardium is the most susceptible myocardial region to ischemia and the subendocardium is the most resistant. Myocardial viability during coronary occlusion appears to be primarily determined by the distribution and functional capacity of the collateral circulation.     

A3 adenosine receptor agonist IB-MECA reduces myocardial ischemia-reperfusion injury in dogs

We examined the effect of the A3 adenosine receptor (AR) agonist IB-MECA on infarct size in an open-chest anesthetized dog model of myocardial ischemia-reperfusion injury. Dogs were subjected to 60 min of left anterior descending (LAD) coronary artery occlusion and 3 h of reperfusion. Infarct size and regional myocardial blood flow were assessed by macrohistochemical staining with triphenyltetrazolium chloride and radioactive microspheres, respectively. Four experimental groups were studied: vehicle control (50% DMSO in normal saline), IB-MECA (100 microg/kg iv bolus) given 10 min before the coronary occlusion, IB-MECA (100 microg/kg iv bolus) given 5 min before initiation of reperfusion, and IB-MECA (100 microg/kg iv bolus) given 10 min before coronary occlusion in dogs pretreated 15 min earlier with the ATP-dependent potassium channel antagonist glibenclamide (0.3 mg/kg iv bolus). Administration of IB-MECA had no effect on any hemodynamic parameter measured including heart rate, first derivative of left ventricular pressure, aortic pressure, LAD coronary blood flow, or coronary collateral blood flow. Nevertheless, pretreatment with IB-MECA before coronary occlusion produced a marked reduction in infarct size ( approximately 40% reduction) compared with the control group (13.0 +/- 3.2% vs. 25.2 +/- 3.7% of the area at risk, respectively). This effect of IB-MECA was blocked completely in dogs pretreated with glibenclamide. An equivalent reduction in infarct size was observed when IB-MECA was administered immediately before reperfusion (13.1 +/- 3.9%). These results are the first to demonstrate efficacy of an A3AR agonist in a large animal model of myocardial infarction by mechanisms that are unrelated to changes in hemodynamic parameters and coronary blood flow. These data also demonstrate in an in vivo model that IB-MECA is effective as a cardioprotective agent when administered at the time of reperfusion.     

Diabetes and hyperglycemia impair activation of mitochondrial K(ATP) channels

Hyperglycemia is an important predictor of cardiovascular mortality in patients with diabetes. We investigated the hypothesis that diabetes or acute hyperglycemia attenuates the reduction of myocardial infarct size produced by activation of mitochondrial ATP-regulated potassium (K(ATP)) channels. Acutely instrumented barbiturate-anesthetized dogs were subjected to a 60-min period of coronary artery occlusion and 3 h of reperfusion. Myocardial infarct size (triphenyltetrazolium chloride staining) was 25 +/- 1, 28 +/- 3, and 25 +/- 1% of the area at risk (AAR) for infarction in control, diabetic (3 wk after streptozotocin-alloxan), and hyperglycemic (15% intravenous dextrose) dogs, respectively. Diazoxide (2.5 mg/kg iv) significantly decreased infarct size (10 +/- 1% of AAR, P < 0.05) but did not produce protection in the presence of diabetes (28 +/- 5%) or moderate hyperglycemia (blood glucose 310 +/- 10 mg/dl; 23 +/- 2%). The dose of diazoxide and the degree of hyperglycemia were interactive. Profound (blood glucose 574 +/- 23 mg/dl) but not moderate hyperglycemia blocked the effects of high-dose (5.0 mg/kg) diazoxide [26 +/- 3, 15 +/- 3 (P < 0.05), and 11 +/- 2% (P < 0.05), respectively]. There were no differences in systemic hemodynamics, AAR, or coronary collateral blood flow (by radioactive microspheres) between groups. The results indicate that diabetes or hyperglycemia impairs activation of mitochondrial K(ATP) channels.