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Biomedical subjects

V Panagia

Publications and source records attributed to V Panagia.

At least 91 records · Page 5Linked to original sources
PubMed1984

Characterization of heart sarcolemmal phospholipid methylation.

The transmethylation of phosphatidylethanolamine (PE) to phosphatidylcholine (PC) was studied in rat heart sarcolemmal membrane. Kinetically, three apparent Km values for S-adenosyl-L-methionine (AdoMet) were obtained when the total [3H]methyl groups incorporation into the phospholipids was examined in the presence of 0.01-250 microM AdoMet. A first methyltransferase active site having a very low Km (0.1 microM) for AdoMet showed a partial requirement for Mg2+ and an optimum pH of 8.0 with a major formation of phosphatidyl-N-monomethylethanolamine (PMME). Both Ca2+ and K+ were inhibitory to this site. A second active site with a Km of 3.6 microM showed an optimum pH of 7.0 with predominant formation of phosphatidyl-N,N-dimethylethanolamine (PDME) and no Mg2+ requirement; in addition, transmethylation activity was also observed over a broad alkaline pH range (9-11) with an optimum at pH 10.5. This site was insensitive to Ca2+ but was stimulated by Na+, while K+ had an inhibitory effect. A third active site with a Km of 119 microM showed an optimum pH of 10.5 with major formation of PC and no Mg2+ requirement. This site was also insensitive to Ca2+ but markedly inhibited by both K+ and Na+. Under optimal conditions, the activities of all three methyltransferase sites were linear for at least 30 min of incubation and the sensitivity to the inhibitory effect of S-adenosyl-L-homocysteine was different for each site. Addition of exogenous PMME and PDME as substrates enhanced the synthesis of the corresponding methylated products by 3-5-fold and 3-8-fold, respectively. In contrast, exogenous PE failed to increase methyltransferase activity. These results provide evidence for the existence of three distinct methyltransferase active sites in rat heart sarcolemma.

Animals↗
PubMed1984

Sarcolemmal alterations during the development of genetically determined cardiomyopathy.

The purpose of this study was to identify alterations in specific enzyme and Ca2+ binding activities in cardiac sarcolemmal fractions from UM-X7.1 myopathic Syrian hamsters during the development of cardiomyopathy. Experimental and healthy control animals were examined from 25 to 200 days of age. Sarcolemmal Na+, K+-ATPase activity was depressed in the myopathic hamsters throughout the time course of this study. Sarcolemmal ATP-independent Ca2+ binding was found to be depressed in experimental animals as early as 55 days of age. Ca2+ -stimulated, Mg2+ -dependent ATPase activity was depressed in the experimental animals by 90 days of age and this decrease in enzyme activity was accompanied by a decrease in ATP-dependent Ca2+ binding capacity of the sarcolemmal membranes. Mg2+ -ATPase and Ca2+ -ATPase activities were only affected in the latter stages of the disease (155 to 200 days old). NaF, epinephrine and Gpp(NH)p stimulation of the sarcolemmal adenylate cyclase activity was also observed to be attenuated during the latter stages of the disease. These defects in adenylate cyclase system of the sarcolemmal fraction appeared specific since basal adenylate cyclase activity was not altered at any age studied. The results demonstrate that the earliest lesions in sarcolemmal activity in myopathic hamster heart occur in Na+, K+-ATPase and ATP-independent Ca2+ binding capacity. These defects correspond temporally to the initial stages of cardiac necrotic development in this strain of myopathic hamster.

Adenosine Triphosphatases↗
PubMed1984

Sarcoplasmic reticular Ca2+-pump adaptation in cardiac hypertrophy due to pressure overload in pigs.

Ca2+-transport activities of the sarcoplasmic reticulum isolated from the non-failing hypertrophied left ventricle due to supravalvular banding of aorta for 4 and 8 weeks in pigs were compared with those from the sham operated controls. The rate of Ca2+-uptake, but not the capacity, in 4 weeks hypertrophied heart preparations was increased. On the other hand, both the rate and capacity of sarcoplasmic reticulum to accumulate Ca2+ were decreased at 8 weeks of heart hypertrophy. The Ca2+-binding and Ca2+-stimulated Mg2+ dependent ATPase activities of the reticular fractions from 4 and 8 weeks hypertrophied hearts were not significantly different from the control values. No changes in the sarcoplasmic reticular phospholipid contents were evident except that sphingomyelin was significantly decreased in 4 weeks and diphosphatidylglycerol was increased in both 4 and 8 weeks hypertrophied preparations. These results suggest that adaptation of the sarcoplasmic reticulum function in physiologic and pathologic hypertrophied hearts may occur by changing the efficiency of Ca2+-pump system during the development of cardiac hypertrophy.

Animals↗
PubMed1984

Progress and problems in understanding the involvement of calcium in heart function.

In this article we have briefly reviewed the role of Ca2+ in the excitation contraction coupling in the myocardium and have indicated that cardiac contraction and relaxation are initiated upon raising and lowering the intracellular concentration of free Ca2+, respectively. Different mechanisms for the entry of Ca2+ through sarcolemma as well as release of Ca2+ from sarcoplasmic reticulum and possibly mitochondria have been outlined for initiating cardiac contraction. Relaxation of the cardiac muscle appears to be intimately dependent upon efflux of Ca2+ through sarcolemma as well as sequestration of Ca2+ by the intracellular storage sites, particularly sarcoplasmic reticulum and possibly mitochondria. The actions of some pharmacological and pathophysiological interventions have been explained on the basis of changes in subcellular Ca2+ movements in myocardium. Quinidine, which produced an initial positive inotropic action on rat heart was also found to increase sarcolemmal Ca2+-ATPase activity without any changes in the Na+-K+ ATPase. Other antiarrhythmic agents, procainamide and lidocaine, also increased sarcolemmal Ca2+-ATPase activity without affecting the Na+-K+ ATPase. On the other hand, both Ca2+-ATPase and Na+-K+ ATPase activities were increased in heart sarcolemma obtained from cardiomyopathic hamsters. In this model the increased Ca2+-ATPase activity may promote the occurrence of intracellular Ca2+ overload in the cardiac cell whereas the increased Na+-K+ ATPase activity may increase Ca2+ efflux through Na+-Ca2+ exchange systems as an adaptive mechanism. It has been suggested that some caution should be exercised while interpreting the data from in vitro experiments in terms of functional changes in the myocardium. Furthermore, it has been proposed that the pathophysiology and pharmacology of Ca2+ movements at different membrane sites be understood fully in normal and diseased myocardium in order to improve the therapy of heart disease.

Adenosine Triphosphate↗
PubMed1984

Sarcolemmal phosphatidylethanolamine N-methylation in diabetic cardiomyopathy.

Phosphatidylethanolamine N-methylation was studied in cardiac sarcolemma 8 weeks after the induction of chronic experimental diabetes in rats by a streptozotocin injection (65 mg/kg, iv). Incorporation of radiolabeled methyl groups from S-adenosyl-L-methionine into intramembranal phosphatidylethanolamine, assayed under optimal conditions, confirmed the existence of three catalytic sites involved in the sequential methyl transfer reactions. Total methyl group incorporation at all three sites was significantly depressed in diabetic myocardium, but this change was reversible by a 14-day insulin therapy to the diabetic animals. Measurements of phospholipid N-methylation activity at different times indicated that the depression was evident at 6 weeks after the induction of diabetes. This defect was also seen for the individual methylated lipid products (monomethyl-, dimethylphosphatidylethanolamine, and phosphatidylcholine) specifically formed at each catalytic site. Experiments with different concentrations of S-adenosyl-L-methionine revealed that, for all three sites, the apparent affinity for the methyl donor did not change, whereas the apparent Vmax values were significantly lowered in diabetes. The results of this study identify a defect in the sarcolemmal phosphatidylethanolamine N-methylation in diabetic cardiomyopathy.

Animals↗
PubMed1984

Direct effects of adriamycin on the rat heart sarcolemma.

Effects of adriamycin on rat heart sarcolemma were investigated by determining several membrane enzyme activities in the absence or presence of different concentrations of the drug. The adenylate cyclase activity was found to be stimulated by 0.001 and 0.01 microM of adriamycin whereas this enzyme activity was inhibited at 100 and 1000 microM concentrations of the drug. Magnesium dependent Na+, K+ ATPase was not effected by adriamycin, however, in presence of 2 mM ouabain, adriamycin (100 microM) had a depressant effect on the Na+, K+ ATPase activity. Ca2+ ATPase activity was found to be stimulated by adriamycin. The drug had no effect on the Mg2+ ATPase and 5'-nucleotidase activities. These data suggest direct as well as specific effects of adriamycin which may be important in drug-induced cardiotoxicity.

5'-Nucleotidase↗
PubMed1983

Association of adenylate cyclase inhibition by NaF with loss of a factor in rat heart sarcolemma.

Heart sarcolemma from different animals was isolated for studying the effect of NaF on membrane-bound adenylate cyclase activity. Unlike dog and rabbit heart, a depression of adenylate cyclase by NaF was observed in sarcolemma from rat heart. There was a progressive attenuation of the NaF ability to stimulate the enzyme at different steps of the sarcolemmal isolation procedure. The activation by epinephrine in the presence of Gpp(NH)p also decreased progressively but unlike NaF, this agent did not show an inhibition of the enzyme. The inhibitory action of NaF was not reversed upon the treatment of heart membranes with deoxycholate or by Ca2+. Lubrol extract (supernatant) of a particulate fraction from rat heart, which showed NaF activation, returned the stimulatory response of the sarcolemmal adenylate cyclase to NaF. These results suggest that some regulatory factor is required for the stimulation of adenylate cyclase by NaF in myocardium and rat heart is susceptible for the loss of such a factor during the sarcolemmal isolation by the hypotonic shock-LiBr treatment method.

Adenylyl Cyclase Inhibitors↗
PubMed1983

Properties of 5'-nucleotidase in rat heart sarcolemma.

The activity of 5'-nucleotidase (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) was examined in membrane fractions isolated by hypotonic shock-LiBr treatment (fraction HL) and sucrose gradient separation (fraction S) of rat ventricle homogenate. The enzyme activity in these two fractions differed significantly in several respects. In fraction HL, 5'-nucleotidase had a high affinity for AMP (Km 35 microM), and ATP was a potent competitive inhibitor. In contrast, the 5'-nucleotidase displayed by fraction S showed a low substrate affinity (Km 130 microM) and less sensitivity to ATP. Treatment of membranes with trypsin and neuraminidase markedly stimulated 5'-nucleotidase in fraction HL, whereas only a modest effect was observed in fraction S. Exposure of the membranes to Triton X-100 resulted in a 60% and 10% increase in the enzyme activity in fractions HL and S, respectively. The characteristic activity ratios of 5'-nucleotidase at 200 microM relative to 50 microM AMP in fractions HL and S were modified by alamethicin in an opposite way and became identical. Although concanavalin A almost completely inhibited the 5'-nucleotidase activity in both membrane preparations at a concentration of 2 microM, Hill plots of the data on concanavalin A inhibition revealed a coefficient of 2.2 for fraction S and 1.1 for fraction HL. The differences in 5'-nucleotidase activity of the two membrane fractions are considered to be due to differences in the orientation of the vesicles of the sarcolemmal preparations. These results suggest that two distinct catalytic sites for 5'-nucleotidase are present at the intra- and extracellular surface of the rat heart sarcolemma.

Alamethicin↗
PubMed1982

Role of membrane integrity and cation interaction for heart sarcolemmal adenylate cyclase and Na+-K+ ATPase.

The adenylate cyclase and Na+ -K+ ATPase activities decreased on storage at 4 degrees C as well as on freezing and thawing of the rat heart sarcolemma. Treatment of the sarcolemmal fraction with phospholipase C and trypsin also depressed the adenylate cyclase and Na+ -K+ ATPase activities; the Na+ -K+ ATPase was more sensitive to these treatments than the adenylate cyclase. When the sarcolemmal enzyme activities were determined in the presence of different concentrations of some cations the adenylate cyclase activity was enhanced and the Na+ -K+ ATPase activity was depressed by monovalent cations (Na+, K+, Rb+, Cs+, Li+, and NH+4). Divalent cations such as Sr2+, Ba2+, Co2+, and Mn2+ had biphasic or no effects on the adenylate cyclase activity but inhibited the Na+ -K+ ATPase activity. Although Ca2+, Ni2+, Cd2+, Cu2+, Hg2+, and Zn2+ depressed both Na+ -K+ ATPase and adenylate cyclase activities, the degree of inhibition of these enzymes was different. These results reveal the role of membrane integrity for full expression of the adenylate cyclase and Na+ -K+ ATPase activities, whereas both monovalent and divalent cations appear to regulate sarcolemma-bound enzyme activities.

Adenylyl Cyclases↗
PubMed1982

Properties of Ca2+- or Mg2+-dependent ATPase in rat heart sarcolemma.

Rat heart sarcolemma was shown to hydrolyze ATP in the presence of Ca2+ or Mg2+; Ka values for Ca2+ and Mg2+ were in the range of 0.58-0.67 and 0.72-0.83 mM, whereas Vmax values were 33-38 and 21-28 mumol Pi/mg per hr, respectively. Both Ca2+ ATPase and Mg2+ ATPase showed low- and high-affinity sites for ATP; the Km value for the low-affinity sites for both enzyme activities was 300-325 microM, whereas Km values for high-affinity sites were 75-85 and 100-108 microM, respectively. The pattern of nucleotide hydrolysis in the presence of Ca2+ was found to be different from that with Mg2+. Although both high concentrations of ADP and Pi inhibited the enzyme activities, Mg2+ ATPase was more sensitive to ADP and less sensitive to Pi in comparison to Ca2+ ATPase. Storage of sarcolemma at about 0 degrees C showed a greater increase in ATP hydrolysis with Ca2+ than with Mg2+. The inhibitory effect of Mg2+ on Ca2+ ATPase, unlike that of Ni2+, Co2+, and Cu2+, was more than that on Mg2+ ATPase. Treatment of membranes with sodium dodecylsulfate or deoxycholate produced a greater reduction in Mg2+ ATPase than in Ca2+ ATPase. These results further support the view that Ca2+ ATPase and Mg2+ ATPase may be two separate enzymes in heart sarcolemma. It is suggested that Ca2+-dependent ATPase may be involved in opening calcium channels for the entry of calcium, whereas Mg2+ ATPase may serve as a Mg2+ pump mechanism for the efflux of magnesium from the cardiac cell.

Adenosine Triphosphatases↗
PubMed1982

Ca2+- and Mg2+-dependent ATPase activities in the deoxycholate-treated rat heart sarcolemma.

1. Isolated rat heart sarcolemma was treated with different concentrations of an ionic detergent, deoxycholate (DOC) and ATP hydrolysis in the presence of Ca2+ or Mg2+ was determined. 2. Both Ca2+-dependent ATPase and Mg2+-dependent ATPase activities were decreased in the DOC-treated membranes; however, the depression of Mg2+-dependent ATPase activity was greater than that of Ca2+-dependent ATPase. 3. The differential changes in Ca2+-dependent ATPase and Mg2+-dependent ATPase activities were apparent when incubations with DOC were carried out for different time intervals and at different temperatures. 4. In DOC-treated preparations, the Km value for Ca2+-dependent ATPase was decreased whereas that for Mg2+-dependent ATPase was increased. The half maximal velocities of the Ca2+-dependent ATPase and Mg2+-dependent ATPase enzyme reactions in the treated preparations were obtained at a DOC: membrane protein ratio of 3.0 and 0.6, respectively. 5. In the DOC-treated membranes exhibiting the half maximal velocities of enzyme reactions, the Ka value for Ca2+-dependent ATPase was drastically reduced but remained unchanged for Mg2+-dependent ATPase. 6. The DOC treatment was associated with a loss of protein as well phospholipids and resulted in changes in the ultrastructural integrity of the membrane. 7. Varying degrees of decreases in the activities of sarcolemmal adenylate cyclase. (Na+-K+)-ATPase, 5'-nucleotidase and calcium binding were seen upon DOC treatment. 8. The extent of reduction in Ca2+-dependent ATPase and Mg2+-dependent ATPase activities were also different when the membrane was treated with a non-ionic detergent, Lubrol PX. 9. These data suggest that Ca2+-dependent ATPase in heart sarcolemma is more resistant than Mg2+-dependent ATPase to detergent treatments and further indicate some differences in the properties of these enzymes.

Adenosine Triphosphatases↗
PubMed1981

Role of phosphatidylinositol in basal adenylate cyclase activity of rat heart sarcolemma.

The adenylate cyclase activity and phospholipid composition were determined in rat heart sarcolemma after treating the membranes with a phosphatidylinositol-specific phospholipase C. Complete hydrolysis of phosphatidylinositol in sarcolemma was associated with a marked loss of the basal adenylate cyclase activity. The recombination of the supernatant with the phosphatidylinositol-depleted membranes was found to reactivate the adenylate cyclase activity. The soluble component(s) in the supernatant, which restored the adenylate cyclase activity, was thermolabile and precipitated by ammonium sulfate. Extensive hydrolysis of phosphatidylcholine, phosphatidylethanolamine and sphingomyelin in sarcolemma with a Clostridium welchii phospholipase C treatment did not affect the basal adenylate cyclase activity. These results suggest that phosphatidylinositol anchors component(s) essential for the expression of basal adenylate cyclase activity to the myocardial cell membrane.

Adenylyl Cyclases↗
PubMed1981

Solubilization of rat heart sarcolemma 5'-nucleotidase by phosphatidylinositol-specific phospholipase C.

The influence of a phosphatidylinositol-specific phospholipase C treatment on rat heart sarcolemmal 5'-nucleotidase was investigated. Upon complete hydrolysis of all phosphatidylinositol in the sarcolemma, 75% of 5'-nucleotidase activity was found in the solubilized form. The insolubilized enzyme after this treatment has the same Km for AMP as the untreated, sarcolemmal-bound enzyme (0.04 mM), whereas the solubilized enzyme has a 40-fold increase in Km for AMP (0.16 mM). Other sarcolemmal-bound enzymes were not affected by the same treatment. Hence, the specific involvement of phosphatidylinositol in the binding of 5'-nucleotidase to the sarcolemma of the rat heart is clearly demonstrated.

5'-Nucleotidase↗
PubMed1981

Effect of cyclic AMP phosphodiesterase inhibitors on cardiac sarcolemmal 5'-nucleotidase.

Two different membrane preparations (fraction HL and fraction S) were used to study the effect of caffeine, theophylline, dipyridamole and papaverine on cardiac sarcolemmal 5'-nucleotidase. Caffeine and theophylline inhibited the 5'-nucleotidase in both fractions, but theophylline appeared to be a more potent agent. The inhibitory effect of caffeine seemed to be the same in both membrane preparations, but the enzyme in fraction S appeared to be more sensitive to theophylline than that present in fraction HL. The inhibition was of the competitive type in both preparations. The inhibitory effect of caffeine and theophylline on 5'-nucleotidase may not be related to their actions on phosphodiesterase since dipyridamole and papaverine, which are also known to inhibit phosphodiesterase, did not affect the 5'-nucleotidase activity. It is suggested that by inhibiting 5'-nucleotidase, caffeine and theophylline may influence the production of adenosine in the myocardium and thereby may interfere with an important mechanism for the regulation of coronary blood flow.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1980

Subcellular calcium transport during contractile failure and recovery in heart perfused with Na-free medium.

Perfusion of isolated rat hearts with Na-free medium resulted in an immediate increase in contractile force followed by a decline and complete loss of contractile force within 25 s. The recovery of the contractile force upon reperfusion was only partial if the duration of Na-free perfusions was 10 min or longer. Ca binding and uptake activities of mitochondria obtained from hearts perfused with Na-free medium did not change significantly. However, Ca binding and uptake activities of microsomes were depressed after 5 min of perfusion. The critical concentration of Na in the perfusion medium for inducing these changes was found to be less than 35 mM. The microsomal Ca-ATPase activity was decreased after 10 min of Na-free perfusion. Only partial recovery of microsomal Ca uptake was observed upon reperfusion of hearts preperfused with Na-free medium for 20 min or longer whereas Ca-ATPase activity in these hearts did not recover at all. These results suggest that the defect in the microsomal Ca transport may be secondary to the development of contractile failure and may partially be associated with the inability of Na-depleted hearts to recover fully their contractile force.

Adenosine Triphosphatases↗
PubMed1980

Species-related difference in the heart sarcolemmal enzyme activities.

The sarcolemmal membranes were isolated by the hypotonic shock-LiBr treatment method from rat, guinea pig, rabbit, and dog hearts and their various biochemical activities were measured. The Mg2+-ATPase, Ca2+-ATPase, and 5'-nucleotidase were most active in rat heart homogenates as well as sarcolemmal membranes, whereas the adenylate cyclase and Na+, K+-ATPase were most active in guinea pig heart preparations. ATP-independent calcium binding activity of rat heart sarcolemma was the same as that of guinea pig heart membrane, whereas ATP-dependent calcium binding of rat heart preparation was negligible. Treatment of sarcolemma with 0.6 M KCl increased the adenylate cyclase and Na+, K+-ATPase activity in rat heart, but these activities were unaltered or decreased in other species. The gel electrophoretic pattern of protein bands and phospholipid composition of rat heart sarcolemma were different from those of guinea pig heart. Sarcolemma isolated from hearts by sucrose gradient method also showed species-dependent difference in the membrane-bound enzyme activities. These results have been interpreted to suggest species-related difference in calcium movements across sarcolemma and this may contribute to determining species-dependent difference in the regulation of myocardial calcium metabolism.

Adenosine Triphosphatases↗