Robert T. Rowland

Preconditioning and Hypothermic Cardioplegia Protect Human Heart Equally Against Ischemia

BACKGROUND The purpose of this study was to determine whether transient ischemic preconditioning protects human myocardium against normothermic ischemic injury. METHODS Isolated human right atrial trabeculae were suspended in an organ bath with oxygenated Tyrodes solution at 37 degrees C and field stimulated at 1 Hz. Developed force was recorded. Trabeculae (Warm I/R) received normoxic perfusion before 45 minutes of normothermic simulated ischemia (hypoxic, substrate-free buffer with pacing at 3 Hz) and 120 minutes of reperfusion. Preconditioned trabeculae (Warm IPC) were subjected to 5 minutes of normothermic simulated ischemia and 10 minutes of perfusion before normothermic simulated ischemia-reperfusion injury. Trabeculae (Cold I/R) were subjected to hypothermic (4 degrees C) ischemia (hypoxic buffer) for 4 hours and 60 minutes of reperfusion (37 degrees C). Preconditioned trabeculae (Cold IPC) were pretreated with 5 minutes of normothermic simulated ischemia before hypothermic ischemia and 60 minutes of reperfusion. At the end of reperfusion, trabeculae were frozen at -70 degrees C and assayed for tissue creatine kinase activity. RESULTS At the end of reperfusion, warm preconditioned trabeculae (Warm IPC) recovered 51% +/- 5% of baseline developed force, whereas warm I/R trabeculae recovered 24% +/- 3% (p < 0.05). Tissue creatine kinase levels reflecting preserved tissue viability were sustained in Warm IPC trabeculae (1,183 +/- 204 U/g), whereas nonpreconditioned control trabeculae (Warm I/R) exhibited lower levels of enzymatic activity (403 +/- 32 U/g) (p < 0.05). In contrast, Cold IPC trabeculae recovered 47% +/- 5% and Cold I/R, 56% +/- 8% of baseline developed force at the end of reperfusion (p > 0.05). CONCLUSIONS We conclude that transient ischemic preconditioning protects human myocardium against normothermic ischemic injury.

Optimal myocardial preservation: cooling, cardioplegia, and conditioning.

Myocardial preservation techniques have evolved in conjunction with cardiac surgery and currently offer substantial protection against myocardial injury. We propose that cardiac preconditioning, a robust, endogenous mechanism of cardioprotection, is emerging as an important adjunct to current cardioplegic techniques. By reviewing the physiologic basis for current cardioplegic strategies, and understanding the cardioprotective benefits of preconditioning, we postulate that cardiac preconditioning may represent an important, clinically accessible component of myocardial protection.

Myocardial gene reprogramming associated with a cardiac cross-resistant state induced by LPS preconditioning

Lipopolysaccharide (LPS) preconditioning induces cardiac resistance to subsequent LPS or ischemia. This study tested the hypothesis that resistance to LPS and resistance to ischemia are two manifestations of cardiac cross-resistance which may involve reprogramming of cardiac gene expression. Rats were preconditioned with a single dose of LPS (0.5 mg/kg ip). Cardiac resistance to LPS was examined with a subsequent LPS challenge. Cardiac resistance to ischemia was determined by subjecting hearts to ischemia-reperfusion. Total RNA was extracted from myocardium for Northern analysis of mRNAs encoding protooncoproteins, antioxidant enzymes, and contractile protein isoforms. Rats preconditioned with LPS 1-7 days earlier acquired cardiac resistance to endotoxemic depression. This resistance temporally correlated with resistance to ischemia. Pretreatment with cycloheximide (0.5 mg/kg ip) abolished resistance to both LPS and ischemia. LPS preconditioning induced the expression of c- jun and c- fos mRNAs. LPS also transiently increased mRNAs encoding catalase and Mn-containing superoxide dismutase. The expression of both α- and β-myosin heavy chain mRNAs was upregulated, whereas the expression of cardiac α-actin mRNA was suppressed. We conclude that 1) LPS induces sustained cardiac resistance to both LPS and ischemia, 2) resistance to ischemia and resistance to LPS seem to be two mechanistically indistinct components of cardiac cross-resistance, and 3) the cardiac cross-resistance is associated with reprogramming of myocardial gene expression.

The obligate role of protein kinase C in mediating clinically accessible cardiac preconditioning

BACKGROUND Cardiac preconditioning is an adaptation of cardiomyocytes that promotes tolerance to a subsequent ischemic insult. Adenosine receptor signaling is proposed as a mediator of preconditioning, but its mechanism of protection remains unknown. We hypothesized that protection against hypoxia-reoxygenation (H/R) injury could be conferred in a rat ventricle by adenosine-mediated protein kinase C (PKC) activation and that adenosine-mediated cardioprotection could be extended to human ventricular muscle. METHODS Isolated rat and human ventricular muscle (VM) strips were subjected to 30 minutes of hypoxia and 60 minutes of reoxygenation (H/R control). The VM was pretreated with 125 mumol/L adenosine, an adenosine antagonist ((p-Sulfophenyl) theophylline [SPT] 50 mumol/L) and adenosine (adenosine + SPT), or with a PKC inhibitor (chelerythrine, 10 mumol/L) and adenosine (adenosine + chelerythrine) before H/R Developed force (DF) and tissue creatine kinase (CK) activity were assessed at end reoxygenation. Human trabeculae were obtained from diseased explanted hearts at cardiac transplantation and were also subjected to H/R injury. Human VM was pretreated with adenosine (125 mumol/L) before H/R injury. Results are expressed as mean +/- standard error of mean. RESULTS In the rat, adenosine pretreatment conferred protection of DF against H/R injury (adenosine, 62% +/- 6%; H/R control, 27% +/- 2%, p < 0.05). Adenosine + SPT or adenosine + chelerythrine eliminated the functional recovery conferred by adenosine. This recovery of contractile function was associated with greater tissue CK activity (adenosine, 415 +/- 40 units/gm; H/R control, 78 +/- 13 units/gm, p < 0.05). The protective effects of adenosine against H/R were present in the human ventricle and with recovery of DF in adenosine (66% +/- 5%) and H/R control (24% +/- 4%), p < 0.05. CONCLUSIONS Adenosine, a clinically accessible agonist, induces protection against H/R injury through a PKC-mediated mechanism in the rat ventricle. Further, the protection conferred by adenosine against H/R extends to the human ventricle.

Constructive Priming Of Myocardium Against Ischemia-reperfusion Injury

ABSTRACT Ischemia and ischemic stress hormones induce endogenous cardiac protection against ischemia-reperfusion (I/R) injury. Although ischemia and ischemic stress hormones are accompanied by increased [Ca2+], it is unknown whether either opening of the sarcoplasmic reticular ryanodine Ca2+ channel (SR RyR) or inhibition of Ca2+ uptake by the sarcoendoplasmic reticular Ca2+-ATPase (SERCA) prior to I/R can similarly induce post-l/R functional protection. To study this, isolated, crystalloid perfused Sprague-Dawley rat hearts were used to assess the effects of inducing a pre-ischemic [Ca2+], load by either priming the SR RyR with ryanodine (Ry, 5 nM/2 min) or by transient blockade of the SERCA 10 min prior to global I/R (20 min). A pre-ischemic Ca2+ load by either SR RyR activation or SERCA blockade improved post-ischemic myocardial functional recovery (developed pressure, end diastolic pressure, coronary flow, heart rate, and left ventricular creatine kinase activity). We conclude that 1) Ca2+-induced myocardial functional protection involves the SR Ca2+ source, 2) a pre-ischemic Ca2+ load induced with either Ry or thapsigargin constructively primes against myocardial I/R injury, and 3) Ca2+-induced cardioadaptation to I/R injury may have important therapeutic implications prior to planned ischemic events such as cardiac allograft preservation and cardiac bypass surgery.

Potential gene therapy strategies in the treatment of cardiovascular disease

Gene therapy is the introduction of new genetic material into somatic cells to synthesize missing or defective proteins. Efficient methods for the introduction of genetic material into cells are available, both in vitro and in vivo. These strategies involve chemical, physical, and viral-mediated mechanisms of gene transfer. Application of these gene transfer techniques has led to the development of potential gene-based treatment strategies that could combat vascular and myocardial disease. Gene therapy in the treatment of cardiovascular disease promises to alter atherosclerotic risk factors, prevent vascular thrombotic disease, retard progression of disease in the peripheral vasculature, provide drug delivery systems, and prevent myocardial infarction in patients with coronary artery disease. This exciting technology will eventually become the ultimate intervention in the treatment of cardiovascular disease.

Mechanisms of immature myocardial tolerance to ischemia: Phenotypic differences in antioxidants, stress proteins, and oxidases

BACKGROUND Previous work has suggested tolerance to ischemic injury in newborn myocardium. Although various mechanisms for this protection have been proposed, a link between oxidant-antioxidant factors, stress protein expression, and protection from cardiac ischemia/reperfusion (I/R) injury has not been made in newborn myocardium. We hypothesized total newborn myocardial resistance to I/R is related to decreased oxygen radical producing potential, increased free radical scavenging capacity and augmented stress protein expression. The purposes of the study were to examine in newborn and adult rat hearts (1) functional recovery from I/R, (2) catalase and xanthine oxidase (XO) activities, and (3) heat shock protein 72 (HSP 72) expression. METHODS Isolated rat hearts (7 to 10 days versus 60 days) were perfused on a nonworking Langendorff apparatus at 60 mm Hg (Krebs-Henseleit buffer, pH 7.4, 37 degrees C) and subjected to 20 minutes of global ischemia and 40 minutes of reperfusion. Left ventricular developed pressure was recorded by using a left ventricular catheter. Catalase and XO were measured by means of standard assays, and HSP 72 was assessed with in situ immunohistochemistry. RESULTS Newborn rat hearts had greater percentage functional recovery of left ventricular developed pressure after I/R (66.0% +/- 4.2% versus 44.3% +/- 3.5%; p < 0.05). The newborn myocardium also had increased catalase activity (1027.9 +/- 20.6 units/gm versus 707.3 +/- 38.7 units/gm; p < 0.05), whereas the activity of XO was decreased relative to the adult (0.23 +/- 0.01 mU/gm versus 7.6 +/- 1.4 mU/gm; p < 0.05). Furthermore, the expression of HSP 72 was greater in the newborn than the adult control. CONCLUSIONS Relative to adult hearts, newborn rat hearts are more tolerant to a global ischemic insult followed by reperfusion. This improved functional recovery is associated with decreased oxidant production potential (XO), increased scavenging capacity (catalase), and augmented stress protein expression (HSP 72).