Structure-function relationships and modifications of cardiac sarcoplasmic reticulum Ca2+-transport.

Sarcoplasmic reticulum (SR) is a specialized tubular network, which not only maintains the intracellular concentration of Ca2+ at a low level but is also known to release and accumulate Ca2+ for the occurrence of cardiac contraction and relaxation, respectively. This subcellular organelle is composed of several phospholipids and different Ca2+-cycling, Ca2+-binding and regulatory proteins, which work in a coordinated manner to determine its function in cardiomyocytes. Some of the major proteins in the cardiac SR membrane include Ca2+-pump ATPase (SERCA2), Ca2+-release protein (ryanodine receptor), calsequestrin (Ca2+-binding protein) and phospholamban (regulatory protein). The phosphorylation of SR Ca2+-cycling proteins by protein kinase A or Ca2+-calmodulin kinase (directly or indirectly) has been demonstrated to augment SR Ca2+-release and Ca2+-uptake activities and promote cardiac contraction and relaxation functions. The activation of phospholipases and proteases as well as changes in different gene expressions under different pathological conditions have been shown to alter the SR composition and produce Ca2+-handling abnormalities in cardiomyocytes for the development of cardiac dysfunction. The post-translational modifications of SR Ca2+ cycling proteins by processes such as oxidation, nitrosylation, glycosylation, lipidation, acetylation, sumoylation, and O GlcNacylation have also been reported to affect the SR Ca2+ release and uptake activities as well as cardiac contractile activity. The SR function in the heart is also influenced in association with changes in cardiac performance by several hormones including thyroid hormones and adiponectin as well as by exercise-training. On the basis of such observations, it is suggested that both Ca2+-cycling and regulatory proteins in the SR membranes are intimately involved in determining the status of cardiac function and are thus excellent targets for drug development for the treatment of heart disease.

Cardioprotective effects of cysteine alone or in combination with taurine in diabetes.

This study was undertaken to examine the effects of dietary supplementation of cysteine and taurine in rats with diabetes induced with streptozotocin (STZ, 65 mg/kg body weight). Experimental animals were treated orally (by gavage) with cysteine (200 mg/kg) and taurine (400 mg/kg), alone or in combination, daily for 8 weeks. In one group, rats were also pretreated 3 weeks before the induction of diabetes (prevention arm) whereas in the other, the treatment was started 3 days after the induction of diabetes (reversal arm). Diabetes increased heart weight/body weight (HW/BW) ratio, plasma glucose, triglyceride and cholesterol levels as well as depressed heart rate (HR), blood pressure, left ventricular systolic pressure (LVSP), rate of contraction (+dP/dt), rate of relaxation (-dP/dt), fractional shortening (FS) and cardiac output (CO). The left ventricular internal diameter in systole (LViDs) was increased whereas that in diastole (LViDd) was decreased. In the prevention arm, treatment of the diabetic animals with cysteine or taurine decreased HW/BW ratio and improved HR, FS, +dP/dt and -dP/dt, as well as normalized LViDs, without altering the increase in glucose level. Cysteine decreased plasma triglyceride and cholesterol levels and improved LVSP whereas CO was improved by taurine. In the reversal arm, cysteine alone or with taurine did not correct the changes in hemodynamic parameters, FS and plasma triglycerides. Diabetes-induced cardiac dysfunction and increases in plasma triglycerides can be prevented, but not reversed, by dietary cysteine alone or in combination with taurine.

Anti-atherosclerotic molecules targeting oxidative stress and inflammation.

The accumulation of lipids within arteries remains to be the initial impulse for the pathogenesis of atherosclerosis; however, both inflammation and oxidative stress are considered to play a critical role in this process. Several lipid lowering drugs are used as the first line therapy in atherosclerosis; however, different agents have been found to exhibit beneficial effects which are independent of their lipid lowering activity. Both statins and fibrates have been reported to exert anti-inflammatory and anti-oxidative effects in addition to their anti-atherosclerotic actions. Furthermore, anti-hypertensive, anti-diabetic and anti-platelet drugs, which reduce oxidative stress and inflammation, have been shown to attenuate atherosclerosis. In addition, novel substances such as HDL-related agents, cyclopentenone prostaglandins, lipoprotein-associated phospholipase A(2) inhibitors, 5-lipoxygenase pathway inhibitors, acyl CoA: cholesterol acyltransferase inhibitors, analogues of probucol and lysophosphatidic acid antagonists have been developed for the treatment of atherosclerosis as a consequence of their actions on oxidative stress and inflammation. The present article reviews the involvement of inflammation and oxidative stress in the pathogenesis of atherosclerosis and focuses on the mechanisms of some clinically used as well as potential anti-atherosclerotic substances with anti-inflammatory and anti-oxidative properties.

Losartan attenuates phospholipase C isozyme gene expression in hypertrophied hearts due to volume overload.

Because the left ventricular (LV) hypertrophy due to volume overload induced by arteriovenous (AV) shunt was associated with an increase in phospholipase C (PLC) isozyme mRNA levels, PLC is considered to be involved in the development of cardiac hypertrophy. Since the renin-angiotensin system (RAS) is activated in cardiac hypertrophy, the role of RAS in the stimulation of PLC isozyme gene expression in hypertrophied heart was investigated by inducing AV shunt in Sprague-Dawley rats. The animals were treated with or without losartan (20 mg/kg, daily) for 3 days as well as 1, 2 and 4 weeks, and atria, right ventricle (RV) and LV were used for analysis. The increased muscle mass as well as the mRNA levels for PLC beta1 and beta3 in atria and RV, unlike PLC beta3 gene expression in LV, at 3 days of AVshunt were attenuated by losartan. The increased gene expression for PLC beta1 at 2 weeks in atria, at 1 and 4 weeks in RV, and at 2 and 4 weeks in LV was also depressed by losartan treatment. Likewise, the elevated mRNA levels for PLC beta3 in RV at 1 week and in LVat 4 weeks of cardiac hypertrophy were decreased by losartan. On the other hand, the increased levels of mRNA for PLC gamma1 in RV and LV at 2 and 4 weeks of inducing hypertrophy, unlike in atria at 4 weeks were not attenuated by losartan treatment. While the increased mRNA level for PLC delta1 in LV was reduced by losartan, gene expression for PLC delta1 was unaltered in atria and decreased in RV at 3 days of inducing AV shunt. These results suggest that changes in PLC isozyme gene expression were chamber specific and time-dependent upon inducing cardiac hypertrophy due to AV shunt. Furthermore, partial attenuation of the increased gene expression for some of the PLC isozymes and no effect of losartan on others indicate that both RAS dependent and independent mechanisms may be involved in hypertrophied hearts due to volume overload.

Differential gene expression in infarct scar and viable myocardium from rat heart following coronary ligation.

Post-myocardial infarction (MI) remodeling of cardiac myocytes and the myocardial interstitium results in alteration of gross ventricular geometry and ventricular dysfunction. To investigate the mechanisms of the remodeling process of the heart after large MI, the expression of various genes in viable left ventricle and infarct scar tissue were examined at 16 weeks post-MI. Steady-state expression of Na(+)-K+ ATPase alpha-1 and -2, phospholamban (PLB), alpha-myosin heavy chain (alpha-MHC), ryanodine receptor (Rya) and Ca2+ ATPase (Serca2) mRNAs were decreased in the infarct scar vs noninfarcted sham-operated controls (P < 0.05). On the other hand, Gialpha2 and beta-MHC mRNAs were upregulated (P < 0.05, respectively) in the infarct scar whereas Na(+)-K+ ATPase-beta, Na(+)-Ca2+ exchanger and Gs mRNAs were not altered vs control values. In viable left ventricle, the alpha-1 subunit of Na(+)-K+ ATPase, alpha-3, beta-isoforms, Rya, beta-MHC, Gialpha2, Gs and Na(+)-Ca2+ exchanger were significantly elevated while expression of the alpha-2 subunit of Na(+)-K+ ATPase, PLB and Serca2 were significantly decreased compared to controls. Expression of CK2alpha mRNA was elevated in noninfarcted heart (145 +/- 15%) and diminished in the infarct scar (66 +/- 13%) vs controls. Expression of beta-MHC mRNA was elevated in both viable and infarct scar tissues of experimental hearts (140 +/- 31% and 183 +/- 30% vs. controls, respectively). These results suggest that cardiac genes in the infarcted tissue and viable left ventricle following MI are differentially regulated.

Attenuation of changes in G(i)-proteins and adenylyl cyclase in heart failure by an ACE inhibitor, imidapril.

Cardiac dysfunction in animals with congestive heart failure due to myocardial infarction (MI) is known to be associated with a wide variety of defects in receptor and post-receptor mechanisms. Since the heart function have been shown to be improved by treatment with different angiotensin converting enzyme (ACE) inhibitors, we examined the effects of imidapril, an ACE inhibitor, on changes in post-receptor mechanisms involving adenylyl cyclase (AC) and G proteins in the failing heart. Heart failure in rats was induced by occluding the coronary artery and 3 weeks later the animals were treated daily with 1 mg/kg (orally) imidapril for 5 weeks. The animals were assessed for their left ventricular function and crude membranes were isolated from the viable left ventricle and examined for AC activities as well as G-protein activities and expression. Animals with heart failure exhibited depressions in ventricular function and AC activities in the absence or presence of forskolin, NaF and Gpp(NH)p. The AC activity in the presence of pertussis toxin was increased whereas that in the presence of cholera toxin was decreased in the failing heart. Protein contents and mRNA levels for G(i)-proteins were increased whereas those for G(s)-proteins were unaltered in the infarcted heart. All these changes due to MI were prevented by imidapril treatment. The results indicate that the depressed cardiac function in the failing heart may partly be due to the direct effects of changes in AC and G(i) proteins.

Neurohormones and oxidative stress in nonischemic cardiomyopathy: relationship to survival and the effect of treatment with amlodipine.

OBJECTIVES: The purpose of this study was to assess the effects of amlodipine on neurohormones and oxidative stress in nonischemic cardiomyopathy, and determine the relationship between baseline and posttreatment levels of these markers with survival. BACKGROUND: Neurohormones and oxidative stress are important in the pathophysiology of heart failure. Calcium-channel blockers are associated with poor outcomes in patients with heart failure, in part due to neurohormonal activation. In contrast, amlodipine, a second-generation dihydropyridine, has a more favorable clinical profile. METHODS: In the Prospective Randomized Amlodipine Survival Evaluation 2 (PRAISE-2) trial, a subset of 181 patients with nonischemic cardiomyopathy were randomized to amlodipine (10 mg/day) or placebo. Blood samples were evaluated at baseline, 2 weeks and 26 weeks for norepinephrine, epinephrine, angiotensin II, dopamine, N-terminal pro-atrial natriuretic peptide (Nt-pro-ANP), brain natriuretic peptide (BNP), adrenolutin and malondialdehyde. RESULTS: There was no difference in levels of neurohormones or oxidative stress markers between the amlodipine and placebo groups at the different times. Both Nt-pro-ANP and BNP decreased at 2 weeks and at 26 weeks. Baseline Nt-pro-ANP correlated with survival in multivariate analysis (P =.001). A strong relationship was found between a reduction in BNP at 26 weeks and survival, with a hazard ratio of 0.153 (95% CI 0.051-0.461, P =.017). No relationship was found between markers of oxidative stress and survival. CONCLUSIONS: We conclude that amlodipine does not affect circulating neurohormones and oxidative stress markers in patients with nonischemic cardiomyopathy treated with angiotensin-converting enzyme inhibitors, digoxin and diuretics. In addition, low circulating Nt-pro-ANP and a reduction in BNP over time confers a good prognosis.

Prognostic importance of the oxidized product of catecholamines, adrenolutin, in patients with severe heart failure.

OBJECTIVES: The purpose of this study was to assess whether adrenolutin, the inert product of the highly reactive molecules aminochromes, is increased in severe chronic heart failure and whether it is associated with a poor prognosis. BACKGROUND: Experimental evidence suggests that oxidative products of catecholamines, aminochromes, are more cardiotoxic than unoxidized catecholamines and may be increased in heart failure. METHODS: Adrenolutin was measured at baseline and at 1 and 3 months in 263 patients with chronic New York Heart Association class III or IV heart failure and a left ventricular ejection fraction of 22% +/- 7%. Adrenolutin levels were compared with normal levels, and their relation to prognosis was evaluated. RESULTS: Baseline adrenolutin was increased (55 +/- 90 pg/mL vs 8.4 +/- 9.1 pg/mL for control, P <.02) and remained increased at 1 month (49 +/- 65 pg/mL). During a mean follow-up of 309 +/- 148 days (22-609 days), 57 patients died. Baseline adrenolutin levels correlated with mortality rates by univariate and multivariate analyses (relative risk 1.06, 95% CI 1.01-1.10 for each 17.9-pg/mL rise, P =.032). Left ventricular ejection fraction (P =.013) and New York Heart Association class (P =.009) were the only other variables associated with survival. Age, sex, plasma creatinine, plasma N-terminal atrial natriuretic peptide, and plasma norepinephrine levels were not retained in our model. Adrenolutin levels 1 month after random assignment were not significantly correlated with total mortality rate (P =.061) but were correlated with mortality rate from low output (relative risk 1.14, 95% CI 1.06-1.22, P =.002). CONCLUSIONS: Plasma adrenolutin is increased in patients with heart failure and correlates with a poor prognosis independent of other important predictors of survival. This finding has potentially important pathophysiologic, prognostic, and therapeutic implications.

Sarpogrelate diminishes changes in energy stores and ultrastructure of the ischemic-reperfused rat heart.

Although the involvement of serotonin in exacerbating vascular abnormalities in ischemic heart disease has been established, its role in mediating changes in cardiac function due to ischemia reperfusion (IR) is poorly understood. The aim of this study was to investigate the effect of a serotonin blocker, sarpogrelate (5-HT2A antagonist), in preventing cardiac injury due to IR. Isolated rat hearts were subjected to 30 min of global ischemia followed by 1 h of reperfusion. Sarpogrelate (50 nM-0.9 microM) was infused 10 min before ischemia as well as during the reperfusion period. The IR-induced changes in left ventricular developed pressure, left ventricular end diastolic pressure, rate of pressure development, and rate of pressure decay were attenuated (P < 0.05) with sarpogrelate treatment. Sarpogrelate also decreased the ultrastructural damage and improved the high energy phosphate level in the IR hearts (P < 0.05). This study provides evidence for the attenuation of IR-induced cardiac injury by 5-HT2A receptor blockade and supports the view that serotonin may contribute to the deleterious effects of IR in the heart.

Modulation of cardiac sarcoplasmic reticulum gene expression by lack of oxygen and glucose.

Although ischemia reperfusion has been shown to depress gene expression of the sarcoplasmic reticulum (SR) proteins, such as the ryanodine receptor, Ca2+-pump ATPase, phospholamban, and calsequestrin in the heart, the mechanisms of these changes are not understood. Given the occurrence of hypoxia and the lack of glucose during the ischemic phase, we investigated the effects of these factors on the cardiac SR gene expression. Isolated rat hearts perfused in the absence of oxygen and/or glucose for 30 min showed an increase in the expression of SR genes. However, perfusion of hearts for 60 min with normal oxygenated medium after 30 min of lack of both oxygen and glucose depressed the transcript levels for the SR proteins; these changes did not occur when hearts were deprived of either oxygen or glucose. The effect of intracellular Ca2+-overload, which occurs during reperfusion, was studied by using hearts perfused for 5 min with Ca2+-free medium and then reperfused for 30 min. Ca2+-depletion/repletion induced a dramatic decrease in the transcript levels of the SR genes. These results suggest that the lack of both oxygen and glucose during ischemia are necessary for reperfusion-induced depression in SR gene expression, possibly due to the occurrence of intracellular Ca2+-overload.

Depressed levels of Ca2+-cycling proteins may underlie sarcoplasmic reticulum dysfunction in the diabetic heart.

In view of the depressed sarcoplasmic reticulum (SR) Ca2+-pump and Ca2+-release activities in the diabetic heart and the critical role of phosphorylation in regulating the SR function, we examined the status of Ca2+-calmodulin-dependent protein kinase (CaMK) and cAMP-dependent protein kinase (PKA)-mediated phosphorylations in the diabetic heart. Diabetes was induced in male Sprague-Dawley rats by an injection of streptozotocin (65 mg/kg i.v.), and the animals were killed 6 weeks later for assessment of the ventricular SR function. Depressed cardiac performance and SR Ca2+-uptake and -release activities in diabetic animals were accompanied by a significant decrease in the level of SR Ca2+-cycling proteins, such as ryanodine receptor, Ca2+-pump ATPase, and phospholamban. On the other hand, the CaMK- and PKA-mediated phosphorylations of these Ca2+-cycling proteins, the endogenous SR CaMK and PKA activities, and the endogenous SR and cytosolic phosphatase activities were increased in the diabetic heart. Treatment of 3-week diabetic animals with insulin partially or fully prevented the diabetes-induced changes in cardiac performance, SR Ca2+-uptake and -release activites, and SR protein content, whereas the diabetes-induced changes in SR CaMK- and PKA-mediated phosphorylations and activities, as well as phosphatase activities, were not significantly affected. These results suggest that the reduced content of the Ca2+-cycling proteins, unlike alterations in PKA and phosphatase activities, appear to be the major defect underlying SR dysfunction in the diabetic heart.

Ca2+-antagonists inhibit the N-methyltransferase-dependent synthesis of phosphatidylcholine in the heart.

Evidence indicates that, in addition to the L-type Ca2+ channel blockade, Ca2+-antagonists target other functions including the Ca2+-pumps. This study was conducted to test the possibility that the reported inhibition of heart sarcolemmal (SL) and sarcoplasmic reticular (SR) Ca2+-pumps by verapamil and diltiazem could be due to drug-induced depression of phosphatidylethanolamine (PE) N-methylation which modulates these Ca2+-transport systems. Three catalytic sites individually responsible for the synthesis of PE monomethyl (site I), dimethyl (site II) and trimethyl (phosphatidylcholine (PC), site III) derivates were examined in SL and SR membranes by employing different concentrations of S-adenosyl-L-methionine (AdoMet). Total methyl group incorporation into SL PE, in vitro, was significantly depressed by 10(-6)-10(-3) M verapamil or diltiazem at site III. The catalytic activity of site I was inhibited by 10(-3) M verapamil only, whereas the site II activity was not affected by these drugs. The inhibition induced by verapamil or diltiazem (10(-5) M) was associated with a depression of the Vmax value without any change in the apparent affinity for AdoMet. Both drugs decreased the SR as well as mitochondrial PE N-methylation at site III. A selective depression of site III activity was also observed in SL isolated from hearts of rats treated with verapamil in vivo. Furthermore, administration of [3H-methyl]-methionine following the treatment of animals with verapamil, reduced the synthesis of PC by N-methyltransferase. Verapamil also depressed the N-methylation-dependent positive inotropic effect induced by methionine in the isolated Langendorff heart. Both agents depressed the SL Ca2+-pump and although diltiazem also inhibited the SR Ca2+-pump, verapamil exerted a stimulatory effect. In addition, verapamil decreased SR Ca2+-release. These results suggest that verapamil and diltiazem alter the cardiac PE N-methyltransferase system. This action is apparently additional to the drugs' effect on L-type Ca2+ channels and may serve as a biochemical mechanism for the drugs' inhibition of the cardiac Ca2+-pumps and altered cardiac function.

Sarpogrelate inhibits serotonin-induced proliferation of porcine coronary artery smooth muscle cells: implications for long-term graft patency.

BACKGROUND: Serotonin can induce proliferation of vascular smooth muscle cells. We assessed the ability of a specific serotonin receptor antagonist, sarpogrelate, to inhibit proliferation of cultured porcine coronary artery smooth muscle cells. METHODS: Cell proliferation and mitotic activity were measured using 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide. To determine the effect of sarpogrelate on DNA (deoxyribonucleic acid), RNA (ribonucleic acid), and protein synthesis, radioactive incorporation of 3H-thymidine, 3H-uridine, and 3H-phenylalanine, respectively, was used. Synthesis of DNA was also assessed by flow cytometry with propidium iodide as a fluorochrome. RESULTS: Serotonin, platelet-derived growth factor, endothelin, and angiotensin II all induced proliferation of porcine coronary artery smooth muscle cells. Sarpogrelate specifically inhibited the serotonin-induced cytokine trigger but did not influence platelet-derived growth factor-, endothelin-, or angiotensin II-induced cell proliferation. Sarpogrelate inhibited the serotonin-induced increase in intracellular free ionized calcium concentration, prevented mitogen-activated protein kinase activation, and down-regulated expression of the protooncogenes c-fos and c-jun. Sarpogrelate acted at the G1 phase of the cell cycle. CONCLUSIONS: These data suggest that sarpogrelate could be used as a therapeutic agent to inhibit serotonin-induced neointimal hyperplasia and improve patency of coronary artery bypass grafts.

Role of oxidative stress in catecholamine-induced changes in cardiac sarcolemmal Ca2+ transport.

Although an excessive amount of circulating catecholamines is known to induce cardiomyopathy, the mechanisms are poorly understood. This study was undertaken to investigate the role of oxidative stress in catecholamine-induced heart dysfunction. Treatment of rats for 24 h with a high dose (40 mg/kg) of a synthetic catecholamine, isoproterenol, resulted in increased left ventricular end diastolic pressure, depressed rates of pressure development, and pressure decay as well as increased myocardial Ca2+ content. The increased malondialdehyde content, as well as increased formation of conjugated dienes and low glutathione redox ratio were also observed in hearts from animals injected with isoproterenol. Furthermore, depressed cardiac sarcolemmal (SL) ATP-dependent Ca2+ uptake, Ca2+-stimulated ATPase activity, and Na+-dependent Ca2+ accumulation were detected in experimental hearts. All these catecholamine-induced changes in the heart were attenuated by pretreatment of animals with vitamin E, a well-known antioxidant (25 mg/kg/day for 2 days). Depressed cardiac performance, increased myocardial Ca2+ content, and decreased SL ATP-dependent, and Na+-dependent Ca2+ uptake activities were also seen in the isolated rat hearts perfused with adrenochrome, a catecholamine oxidation product (10 to 25 microg/ml). Incubation of SL membrane with different concentrations of adrenochrome also decreased the ATP-dependent and Na+-dependent Ca2+ uptake activities. These findings suggest the occurrence of oxidative stress, which may depress the SL Ca2+ transport and result in the development intracellular Ca2+ overload and heart dysfunction in catecholamine-induced cardiomyopathy.

Depressed responsiveness of phospholipase C isoenzymes to phosphatidic acid in congestive heart failure.

The cardiac sarcolemmal membrane cis -unsaturated fatty acid-sensitive phospholipase D hydrolyzes phosphatidylcholine to form phosphatidic acid. The functional significance of phosphatidic acid is indicated by its ability to increase [Ca(2+)](i)and augment cardiac contractile performance via the activation of phospholipase C. Accordingly, we tested the hypothesis that a defect occurs in the membrane level of phosphatidic acid and/or the responsiveness of cardiomyocytes to phosphatidic acid in congestive heart failure due to myocardial infarction. Myocardial infarction was produced in rats by ligation of the left coronary artery while sham-operated animals served as control. At 8 weeks after surgery, the experimental animals were at a stage of moderate congestive heart failure. Compared to sham controls, phosphatidic acid-mediated increase in [Ca(2+)](i), as determined by the fura 2-AM technique, was significantly reduced in failing cardiomyocytes. Immunoprecipitation of sarcolemmal phospholipase C isoenzymes using specific monoclonal antibodies revealed that the stimulation of phospholipase C gamma(1)and delta(1)phosphatidylinositol 4,5-bisphosphate hydrolyzing activities by phosphatidic acid was decreased in the failing heart. Although the activity of phospholipase C beta(1)in the failing heart was higher than the control, phosphatidic acid did not stimulate this isoform in control sarcolemma, and produced an inhibitory action in the failing heart preparation. Furthermore, the specific binding of phosphatidic acid to phospholipase C gamma(1)and delta(1)isoenzymes was decreased, whereas binding to phospholipase beta(1)was absent in the failing heart. A reduction in the intramembranal level of phosphatidic acid derived via cis -unsaturated fatty acid-sensitive phospholipase D was also seen in the failing heart. These findings suggest that a defect in phosphatidic acid-mediated signal pathway in sarcolemma may represent a novel mechanism of heart dysfunction in congestive heart failure.

Low level of sarcolemmal phosphatidylinositol 4,5-bisphosphate in cardiomyopathic hamster (UM-X7.1) heart.

OBJECTIVE: Phosphatidylinositol 4,5-bisphosphate (PtdIns 4,5-P(2)) is not only a precursor to inositol 1,4,5-trisphosphate (Ins 1,4, 5-P(3)) and sn-1,2 diacylglycerol, but also essential for the function of several membrane proteins. The aim of this study was to evaluate the changes in the level of this phospholipid in the cell plasma membrane (sarcolemma, SL) of cardiomyopathic hamster (CMPH) heart. METHODS: We examined the cardiac SL PtdIns 4,5-P(2) mass and the activities of the enzymes responsible for its synthesis and hydrolysis in 250-day-old UM-X7.1 CMPH at a severe stage of congestive heart failure (CHF) and in age-matched controls (Syrian Golden hamsters). RESULTS: The SL PtdIns 4,5-P(2) mass in CMPH was reduced by 72% of the control value. The activities of PtdIns 4 kinase and PtdIns 4-P 5 kinase were depressed by 69 and 50% of control values, respectively. Although, the total phospholipase C (PLC) activity was moderately, although significantly, decreased (by 18% of control), PLCdelta(1) isoenzyme activity in the SL membrane was elevated, with a concomitant increase in its protein content, whereas PLCbeta(1) and gamma(1) isoenzyme activities were depressed despite the increase in their protein levels. A 2-fold increase in the Ins 1,4,5-P(3) concentration in the cytosol of the failing heart of CMPH was also observed. CONCLUSIONS: Reduced SL level of PtdIns 4, 5-P(2) may severely jeopardize cardiac cell function in this hamster model of CHF. In addition, the profound changes in the profile of heart SL PLC isoenzyme could alter the complex second messenger responses of these isoenzymes, and elevated Ins 1,4,5-P(3) levels may contribute to intracellular Ca(2+) overload in the failing cardiomyocyte.

Sarcoplasmic reticulum and cardiac oxidative stress: an emerging target for heart disease.

The sarcoplasmic reticulum (SR) is a major player in maintaining cardiac function, as it is intimately involved in the regulation of Ca2+-movements on a beat-to-beat basis. SR dysfunction due to abnormalities in SR protein content has been reported in different cardiac diseases such as ischaemic heart disease, myocardial infarction, congestive heart failure and various cardiomyopathies; thus the genes expressing the SR Ca2+-pump, Ca2+-channels, calsequestrin, phospholamban and other regulatory proteins are considered important targets for drug development. In our experience, ischaemic preconditioning (IP) and pharmacological therapies, such as anti-oxidants, beta-adrenergic receptor blockers, angiotensin receptor (AT-1) blockers, angiotensin converting enzyme inhibitors (ACE-I) and angiotensin receptor blockers are effective therapies that improve cardiac performance in the failing heart by improving SR function. Accordingly, this paper is intended to shed light on the knowledge in the field of cardiac therapy targeted to improve and protect SR function.

Attenuation of changes in sarcoplasmic reticular gene expression in cardiac hypertrophy by propranolol and verapamil.

The effects of propranolol and verapamil on contractile dysfunction, subcellular remodeling and changes in gene expression in cardiac hypertrophy due to pressure overload were examined. Rats were subjected to banding of the abdominal aorta and then treated with either propranolol (10 mg/kg daily), verapamil (5 mg/kg daily) or vehicle for 8 weeks after the surgery. Depression of the left ventricular function in the hypertrophied heart was associated with decreases in myofibrillar and myosin Ca2+ ATPase activities as well as Ca2+-pump and Ca2+-release activities of the sarcoplasmic reticulum (SR). The level of alpha-myosin heavy chain (alpha-MHC) mRNA was decreased while that of beta-MHC mRNA was increased in the pressure-overloaded heart. The level of SR Ca2+-pump ATPase (SERCA2) mRNA and protein content for SERCA2 were decreased in the pressure overloaded heart. Treatment of the hypertrophied animals with propranolol or verapamil resulted in preservation of the left ventricular function and prevention of the subcellular alterations. Shift in the alpha- and beta-MHC mRNA levels and changes in the expression in SERCA2 mRNA level and protein content were also attenuated by these treatments. The results suggest that blockade of beta-adrenoceptors or voltage-dependent calcium channels normalizes the cardiac gene expression, prevents subcellular remodeling and thus attenuates heart dysfunction in rats with cardiac hypertrophy. Furthermore, both cardiac beta-adrenoceptors and L-type Ca2+-channels may be involved in the genesis of cardiac hypertrophy due to pressure overload.

Role of cardiac renin-angiotensin system in sarcoplasmic reticulum function and gene expression in the ischemic-reperfused heart.

The aim of this study was to explore the possible participation of cardiac renin-angiotensin system (RAS) in the ischemia-reperfusion induced changes in heart function as well as Ca2+-handling activities and gene expression of cardiac sarcoplasmic reticulum (SR) proteins. The isolated rat hearts, treated for 10 min without and with 30 microM captopril or 100 microM losartan, were subjected to 30 min ischemia followed by reperfusion for 60 min and processed for the measurement of SR function and gene expression. Attenuated recovery of the left ventricular developed pressure (LVDP) upon reperfusion of the ischemic heart was accompanied by a marked reduction in SR Ca2+-pump ATPase, Ca2+-uptake and Ca2+-release activities. Northern blot analysis revealed that mRNA levels for SR Ca2+-handling proteins such as Ca2+-pump ATPase (SERCA2a), ryanodine receptor, calsequestrin and phospholamban were decreased in the ischemia-reperfused heart as compared with the non-ischemic control. Treatment with captopril improved the recovery of LVDP as well as SR Ca2+-pump ATPase and Ca2+-uptake activities in the postischemic hearts but had no effect on changes in Ca2+-release activity due to ischemic-reperfusion. Losartan neither affected the changes in contractile function nor modified alterations in SR Ca2+-handling activities. The ischemia-reperfusion induced decrease in mRNA levels for SR Ca2+-handling proteins were not affected by treatment with captopril or losartan. The results suggest that the improvement of cardiac function in the ischemic-reperfused heart by captopril is associated with the preservation of SR Ca2+-pump activities; however, it is unlikely that this action of captopril is mediated through the modification of cardiac RAS. Furthermore, cardiac RAS does not appear to contribute towards the ischemia-reperfusion induced changes in gene expression for SR Ca2+-handling proteins.