Boris Z. Simkhovich

Air Pollution and Cardiovascular Injury: Epidemiology, Toxicology, and Mechanisms

Recent epidemiologic studies show that increased levels of air pollutants are positively associated with cardiovascular morbidity and mortality. Inhalation of air pollutants affects heart rate, heart rate variability, blood pressure, vascular tone, blood coagulability, and the progression of atherosclerosis. Several categories within the general population (i.e., people with pre-existing cardiovascular disease and diabetic and elderly individuals) are considered to be more susceptible to air pollution-mediated cardiovascular effects. Major mechanisms of inhalation-mediated cardiovascular toxicity include activation of pro-inflammatory pathways and generation of reactive oxygen species. Although most studies focus on the influence of systemic effects, recent studies indicate that ultrafine particles may be translocated into the circulation and directly transported to the vasculature and heart where they can induce cardiac arrhythmias and decrease cardiac contractility and coronary flow.

Transplantation of neonatal cardiomyocytes after permanent coronary artery occlusion increases regional blood flow of infarcted myocardium.

BACKGROUND Cellular cardiomyoplasty is a promising approach for rebuilding scar tissue after acute myocardial infarction. However, the angiogenic potential of transplanted immature cardiomyocytes and their effect on regional myocardial blood flow (RMBF) after coronary artery occlusion remain to be evaluated. METHODS AND RESULTS Intramyocardial injection of cultured neonatal cardiomyocytes (4 x 10(6) cells/50-70 microliter) into the scar 1 week after permanent coronary occlusion in rats resulted in improved RMBF in the infarct 4 weeks after transplantation (radioactive microspheres, 0.97 +/- 0.18 ml/min/g) in comparison to medium-injected hearts (0.61 +/- 0.11 ml/min/g, P < 0.047). The macroscopic perfusion defect after in vivo staining with the blue dye 50% Uniperse blue was significantly smaller in the cell transplantation group (1.5 +/- 0.3% of the heart) compared to the medium group (3.0 +/- 0.6%, P < 0.017). Clusters of engrafted cells within the scar demonstrated a high capillary density (1217 +/- 114 perfused (blue) capillaries/mm(2)); however, in the scar tissue itself capillary density in the cell group (156 +/- 62/mm(2)) did not significantly differ from the medium group (125 +/- 10/mm(2)), suggesting that neo-angiogenesis was confined to regions of successful engraftment (non-infarcted tissue: 1924 +/- 114 perfused capillaries/mm(2)). The transplantation group was characterized by smaller diastolic and systolic left ventricular volumes, as assessed by intravenous ventriculography, along with thickened infarcts (0.93 +/- 0.07 vs. 0.75 +/- 0.04 mm, P < 0.020) and lower infarct expansion indices (0.64 +/- 0.07 vs. 0.83 +/- 0.06, P < 0.023), as determined by post-mortem morphometry of histologic slides. CONCLUSIONS Transplantation of neonatal cardiomyocytes induced neo-angiogenesis in zones of successful cell engraftment within the scar, which effectively enhanced tissue perfusion.

Brief episode of ischemia activates protective genetic program in rat heart: a gene chip study

OBJECTIVE Brief episodes of ischemia of 20 min or less have the potential to protect the heart. Such episodes are associated primarily with reversible ischemic injury yet they induce changes in gene expression. The purpose of the study was to determine whether activation of protective genes takes place within 4 h following a brief episode of ischemia that would mimic angina pectoris. METHODS Three groups of rats were studied. In the control (Ctrl) group, hearts were immediately excised following anesthesia; in the sham-operated (SO) group, opened-chest rats received 4 h and 20 min of no intervention; and in the group subjected to ischemia (SI) hearts received 20 min of proximal coronary occlusion followed by 4 h of reperfusion. Hearts from the SI group were divided into nonischemic (NI) and ischemic (Isc) areas. Changes in gene expression pattern were analyzed by using Affymetrix Gene Chips. RESULTS Ischemia led to strong upregulation of mRNA transcripts for heat shock proteins 70, 27, 105, 86 and 40 kDa, vascular endothelial growth factor, brain-derived neurotrophic factor, plasminogen activator inhibitor-1, activating transcription factor 3, B-cell translocation gene 2, and growth arrest and DNA damage inducible 45 alpha protein compared to the NI tissue. The majority of mRNAs whose levels increased following brief ischemia were of a protective nature. CONCLUSION Genetic reprogramming emerging during or following brief episodes of ischemia that simulate angina, can be characterized as protective in nature. Developing new therapeutic strategies aimed to promote this protective response represents a legitimate target for future research.

Particulate air pollution and coronary heart disease.

Purpose of review Air pollution poses a significant health risk. The article focuses on the adverse effects of air pollution on the cardiovascular system. Recent findings Short-term and long-term studies clearly indicate that relatively modest exposures to particulate matter in the ambient air are associated with increased morbidity and mortality due to coronary heart disease. In humans, inhalational exposure to particulate air pollutants decreases heart rate variability, causes ST-segment depression and endothelial dysfunction, increases blood pressure and blood coagulability, and accelerates the progression of atherosclerosis. Mechanisms of air pollution-induced cardiotoxicity include increased generation of reactive oxygen species followed by activation of proinflammatory and prothrombotic pathways. In experimental settings, ultrafine air pollutants instilled directly into the cardiac vasculature depress cardiac contractility and decrease coronary flow. Both effects are attenuated by the use of a free radical scavenger. Summary Reactive oxygen species-related mechanisms of air pollution cardiotoxicity might become a valid target in developing new pharmacological strategies aimed at decreasing adverse effects of air pollution during extreme episodes (fires, earthquakes, industrial accidents, acts of terrorism). Educating patients and the general population on the negative cardiovascular effects of air pollution might be helpful in decreasing the risk of developing air pollution-related coronary heart disease.

In vivo and in vitro models to test the hypothesis of particle-induced effects on cardiac function and arrhythmias.

Exposure to ultrafine particles (UFPs) by inhalation increases the number and severity of cardiac events. The specific mechanism(s), of action are unknown. This study was designed to examine whether UFPs could exert a direct effect on the cardiovascular system without dependence upon lung-mediated responses. The direct effects of UFPs were determined in normal rats (infused intravenously with UFPs), and in the isolated Langendorff perfused rat heart. UFPs from either ambient air (UFAAs) or diesel engine exhaust (UFDGs) were studied. Infusion of UFDGs prepared in our laboratory caused ventricular premature beats (VPBs) in 2 of 3 rats in vivo. Ejection fraction in creased slightly (∼4.5%) in rats receiving UFPAA and was unchanged in the UFDG and saline groups in vivo. In the isolated rat heart, perfused according to Langendorff, UFDGs caused a marked ncrease in left-ventricular end-diastolic pressure (LVEDP; from 12.0±4.6 mmHg to 24.8±11.2 mmHg, p<0.05) after 30 min of exposure. UFPs isolated from industrial diesel particulate matter (UFIDs), obtained from the National Institute of Standards and Technology, caused a significant decrease in left-ventricular systolic pressure (LVSP; from 85.7±4.0 mmHg to 37.9±20.3 mmHg, p<0.05) and ±dp/dt (from 2365±158 mmHg/s to 1188±858 mmHg/s, p<0.05) at 30 min after the start of infusion. This effect was absent when the soluble fraction (containing no particles) isolated from the UFIDs was studied. These findings indicate that UFPs can have direct effects on the cardiovascular system that are independent of effects of particles on the lungs.

Critical Role of Nuclear Calcium/Calmodulin-dependent Protein Kinase IIδB in Cardiomyocyte Survival in Cardiomyopathy

Calcium/calmodulin-dependent protein kinase II (CaMKII) plays a central role in cardiac contractility and heart disease. However, the specific role of alternatively spliced variants of CaMKII in cardiac disease and apoptosis remains poorly explored. Here we report that the δB subunit of CaMKII (CaMKIIδB), which is the predominant nuclear isoform of calcium/calmodulin-dependent protein kinases in heart muscle, acts as an anti-apoptotic factor and is a novel target of the antineoplastic and cardiomyopathic drug doxorubicin (Dox (adriamycin)). Hearts of rats that develop cardiomyopathy following chronic treatment with Dox also show down-regulation of CaMKIIδB mRNA, which correlates with decreased cardiac function in vivo, reduced expression of sarcomeric proteins, and increased tissue damage associated with Dox cardiotoxicity. Overexpression of CaMKIIδB in primary cardiac cells inhibits Dox-mediated apoptosis and prevents the loss of the anti-apoptotic protein Bcl-2. Specific silencing of CaMKIIδB by small interfering RNA prevents the formation of organized sarcomeres and decreases the expression of Bcl-2, which all mimic the effect of Dox. CaMKIIδB is required for GATA-4-mediated co-activation and binding to the Bcl-2 promoter. These results reveal that CaMKIIδB plays an essential role in cardiomyocyte survival and provide a mechanism for the protective role of CaMKIIδB. These results suggest that selective targeting of CaMKII in the nuclear compartment might represent a strategy to regulate cardiac apoptosis and to reduce Dox-mediated cardiotoxicity.

Metabolic mechanism by which mild regional hypothermia preserves ischemic tissue.

Background: Our laboratory demonstrated that mild regional hypothermia reduced myocardial infarct size by an average of 65% in the rabbit model of regional ischemia. The exact mechanism for this benefit has not been explored. We hypothesized that a moderate reduction in regional myocardial temperature could preserve cardiac energy metabolism and thus protect the myocardium from sustained ischemic insult. Methods and Results: Anesthetized open-chest rabbits were randomized to normothermic sham-operated (NS, n = 6), hypothermic sham-operated (HS, n = 6), normothermic ischemic (NI, n = 10), and hypothermic ischemic (HI, n = 10) groups. Both sham-operated groups received no occlusions, and both ischemic groups were subjected to 20 minutes of coronary occlusion. To achieve regional cooling of the hearts in the hypothermic groups, a bag of ice water was placed directly on the risk area 15 minutes prior to coronary artery occlusion/no intervention and maintained for the duration of the subsequent 20 minutes of ischemia/no intervention (in the HI and HS groups respectively). Hypothermia preserved adenosine triphosphate (ATP) and glycogen stores in the ischemic area by 42.9% and 84.2%, respectively (1.20 ± 0.11 µmoles ATP/g wet tissue vs 0.84 ± 0.06 µmoles ATP/g wet tissue and 8.16 ± 0.95 µmoles of glucosyl unit/g wet tissue vs 4.43 ± 0.44 µmoles of glucosyl unit/g wet tissue in the HI and the NI groups, respectively). In addition, hypothermia resulted in a trend toward creatine phosphate preservation in the nonischemic area. Conclusions: This is the first demonstration that local therapy with mild reductions in myocardial temperature preserves energy metabolism both in the ischemic and the nonischemic areas as well. The preservation in ATP is the likely mechanism by which regional hypothermia is preserving ischemic myocardium.

Ranolazine as a Cardioplegia Additive Improves Recovery of Diastolic Function in Isolated Rat Hearts

Background— Ranolazine (Ran), an antianginal agent, inhibits late Na+ current. The purpose of this study was to determine whether there was an added benefit of adding Ran to cardioplegia (CP) in a model of global ischemia/reperfusion. Methods and Results— Isolated rat hearts were Langendorff-perfused and exposed to 40-minute normothermic, cardioplegic global ischemia and 30 minutes of reperfusion. Before ischemia and during reperfusion, hearts were treated with no drug (control) or with the late Na+ current inhibitors Ran (5 &mgr;mol/L) or tetrodotoxin (1 &mgr;mol/L). Ischemic cardioplegic arrest led to an increase of left ventricular end-diastolic pressure (LVEDP) by ≥20 mm Hg (ie, cardiac contracture). Ten out of 11 hearts treated with CP alone developed contracture, whereas 6 out of 11 hearts treated with CP plus Ran developed contracture. Ran added to CP reduced LVEDP at the end of ischemia from 41±5 mm Hg in CP alone to 26±3 mm Hg in CP plus Ran (P=0.024). Area under the curve for LVEDP during the entire ischemic period was also smaller in CP plus Ran versus CP alone. The percent increase (from baseline) of LVEDP measured at the end of 30-minute reperfusion was smaller for CP plus Ran (66±18%) versus CP alone (287±90%; P=0.035). The area under the curve for LVEDP during reperfusion was smaller in CP plus Ran versus CP alone. Tetrodotoxin (1 &mgr;mol/L) also reduced cardiac contracture during ischemia/reperfusion, compared to CP alone. Conclusions— Our results suggest that Ran may have therapeutic potential as an adjunct to CP and further support a protective role of Na+ current inhibition during ischemia/reperfusion.

Cellular Mechanisms of Infarct Size Reduction with Ischemic Preconditioning: Role of Calcium?

Abstract: Brief episodes of ischemia protect or “precondition” the heart and reduce infarct size caused by a subsequent sustained ischemic insult. Despite a decade of intensive investigation, the cellular mechanism(s) responsible for this paradoxical protection remain poorly understood. In this review, we focus on the emerging concept that alterations in intracellular calcium homeostasis may participate in either triggering and/or mediating infarct size reduction with preconditioning.

Direct evidence that ischemic preconditioning does not cause protein kinase C translocation in rabbit heart

OBJECTIVE Indirect pharmacological evidence suggests that myocardial protection conferred by ischemic preconditioning in rabbit myocardium is mediated through the translocation of protein kinase C (PKC). To test this hypothesis, we performed direct biochemical measurements of subcellular distribution of PKC in rabbit hearts. METHODS Two protocols were utilized. In Protocol I the preconditioned group (PC) underwent two 5-min episodes of brief coronary artery occlusion each followed by 5 min of reperfusion, while the control group consisted of time-matched, sham-operated (non-ischemic) animals (SO). Tissue samples were homogenized and cytosolic and particulate fractions were obtained by ultracentrifugation. In Protocol II one group of rabbits received ischemic preconditioning as in Protocol I followed by 10 min of sustained ischemia (PC + IS); a second control group was subjected to 10 min of sustained ischemia (IS); and the third group was time-matched, sham-operated (non-ischemic) animals (SO). Homogenized tissue samples were separated into cytosolic, nuclear and membrane fractions. RESULTS In Protocol I, no differences in the subcellular distribution of PKC between the SO and PC groups were observed. In Protocol II, when samples were obtained at 10 min of sustained ischemia, no changes in the subcellular distribution of PKC were observed between SO, IS and PC + IS groups. CONCLUSION Our results indicate that translocation of protein kinase C is not an important mediator of ischemic preconditioning in the rabbit ischemia model.