Biomedical subjects

P Athias

Publications and source records attributed to P Athias.

At least 19 recordsLinked to original sources

PubMed2006

[Development of cardiac physiopathological models from cultured cardiomyocytes].

The cultures of neonatal rat cardiomyocytes represent a very useful tool for the observation and the understanding of the cellular aspects of the electrophysiological, contractile, morphological, metabolic and molecular properties of the myocardium. This model is characterized by a homogeneous population of cardiac muscular cells and by vast possibilities of control of the chemical and physical environment of the cells, allowing the in vitro mimicry of a wide range of cardiac pathological situations. The cardiomyocyte cultures are thus suited to very varied experimental protocols, allowing multiparametric analysis of the cardiocellular effects of different stress such as hypoxia-reoxygenation, of ischemia-reperfusion, of the free radical attack and of thermal shock. These investigations can be combined with the study of the effects and of the cytotoxicity of pharmacological agents, not limited to the putatively cardioactive drugs. The present review proposes an outline of the procedures for the isolation, the culture and the use of neonatal cardiomyocytes. To illustrate the potentialities of this preparation, we describe more specifically the protocols and the various consequences at the cellular scale of an in vitro model of myocardial ischemia reperfusion.

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PubMed2003

Calcitonin gene-related peptide partly protects cultured smooth muscle cells from apoptosis induced by an oxidative stress via activation of ERK1/2 MAPK.

Oxidative stress induced by a glucose/glucose oxidase (G/GO) generator system dose-dependently decreased the viability of cultured vascular smooth muscle cells (VSMC) as estimated by MTT assay. Cell death was induced in 40% of cells exposed to 0.2 IU/ml of the free radical generating mixture. Annexin-V labeling, Hoechst staining together with DNA laddering demonstrated that apoptosis was responsible for this cell loss. Pretreatment of the cells with 10(-8) M calcitonin gene-related peptide (CGRP) significantly attenuated the damaging effect of the oxidative stress. Indeed, cell viability was estimated to be 80% in CGRP-treated group, instead of 60% in absence of CGRP treatment. This protective effect of CGRP was antagonized by 8-37 CGRP, an antagonist of CGRP-1 receptors, whereas it was not reproduced by amylin, a CGRP analogue. As indicated by the reduction in Hoechst staining and in DNA laddering, CGRP prevented the onset of apoptosis. We also demonstrated that the peptide significantly up-regulated the activation of ERK1/2 and P38 kinases. Inhibitors of the kinases prevented the protective effect of CGRP. We conclude that CGRP antagonizes oxidative stress-induced apoptosis by up-regulating MAP kinase activation and that activation of these kinases was necessary to protection.

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PubMed2001

Influence of deep hypothermia on the tolerance of the isolated cardiomyocyte to ischemia-reperfusion.

The influence of deep hypothermia (4 degrees C) during a substrate-free, hypoxia-reoxygenation treatment was investigated on cardiomyocytes (CM) prepared from newborn rat heart in culture in an in vitro, substrate-free model of ischemia-reperfusion. The transmembranous potentials were recorded with standard microelectrodes. The contractions were monitored photometrically. The RNA messenger (mRNA) and protein expression for protein (HSP70) were analysed by RT-PCR (reverse transcriptase-polymerase chain reaction) and Western blotting, respectively. Simulated ischemia (SI) caused a gradual decrease and then a cessation of the spontaneous electromechanical activity. During the reoxygenation, the CM recovered normal function, provided that SI did not exceed 2.5 h. When SI duration was increased up to 4 h, reoxygenation failed to restore the spontaneous electromechanical activity. Conversely, the exposure of the CM to SI together with deep hypothermia decreased the functional alterations observed, and provided a complete electromechanical recovery after 2.5 h as well as after 4 h of SI. Deep hypothermia alone failed to induce HSP70 mRNA and protein production. On the contrary, HSP70 mRNA production increased after 2.5 and 4 h of deep hypothermia followed by 1 h of rewarming, proportionally to the duration of the cooling period. This augmentation in mRNA was associated with a rise in HSP70 protein content. In summary, it appeared that deep hypothermia exerts a strong cytoprotective action during SI only, whereas cooling CM before SI has no beneficial effect on subsequent SI. Moreover, these results suggested the persistence of a signaling system and/or transduction in deeply cooled, functionally depressed cells. Finally, CM in culture appeared to be a model of interest for studying heart graft protection against ischemia-reperfusion and contributed to clarifying the molecular and cellular mechanisms of deep hypothermia on myocardium.

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PubMed2001

Changes in HSP70 and P53 expression are related to the pattern of electromechanical alterations in rat cardiomyocytes during simulated ischemia.

The objective was to relate the response of the HSP70 and P53 genes to the cessation and the recovery of cardiac muscle cell functions when submitted to ischemia-reperfusion. We have measured the electromechanical activity, the released enzymes and HSP70 RNA and protein levels in cultured neonatal rat cardiomyocytes (CM) in a substrate-free, hypoxia-reoxygenation model of ischemia-reperfusion. In parallel the expression of the two genes P53 (the key apoptosis regulator gene) and P21/Waf1 (the P53 target gene) has been evaluated. The functional recovery during post-'ischemic' reoxygenation was associated with an overexpression of HSP70 and P53 lasting until the functional parameters reverted back to the normal, prehypoxic values. In contrast, extending the substrate-free hypoxic treatment worsens the dysfunction of the cardiac muscle cell and, in these conditions, reoxygenation failed to restore cell functions and to activate HSP70. Finally, in the conditions of reversible 'ischemic' cell injury, an early and transitory activation of P53 was associated with the functional recovering process of the CM submitted to simulated ischemia. These observations are suggestive of a contributive role of both HSP70 and P53 to a cytoprotective program activated by reoxygenation in post-'ischemic' CM.

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PubMed2000

Oxidative injury of isolated cardiomyocytes: dependence on free radical species.

The contribution of lipid peroxidation to myocardial injury by free radicals (FR) is still unclear. Consequently, we examined the functional damages inflicted on cultured rat cardiomyocytes (CM) during FR stress provoked by the xanthine/xanthine oxidase system (X/XO) or by a hydroperoxidized fatty acid ((9 Z, 11 E, 13 (S), 15 Z)-13-hydroperoxyocta-decatrienoic acid; 13-HpOTrE), in order to simulate in vitro the initial phase and the propagation phase of the FR attack, respectively. Transmembrane potentials were recorded with glass microelectrodes and contractions were monitored photometrically. The EPR spectroscopy showed that X/XO produced superoxide and hydroxyl radicals during 10 min. The X/XO system altered sharply and irreversibly the spontaneous electrical and mechanical activities of the CM. However, the gas chromatographic analysis showed that these drastic functional damages were associated with comparatively moderate membrane PUFA degradation. Moreover, the EPR analysis did not reveal the production of lipid-derived FR. 13-HpOTrE induced a moderate and reversible decrease in electrical parameters, with no change in CM contractions. These results indicate that the functional consequences of FR attack are dependent on the radical species present and do not support the idea that the membrane lipid breakdown is a major factor of myocardial oxidant dysfunction.

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PubMed1999

Hypoxia-reoxygenation and polyunsaturated fatty acids modulate adrenergic functions in cultured cardiomyocytes.

The polyunsaturated fatty acids (PUFAs) of the omega 3 series are known to modulate adrenergic functions in ventricular myocytes. This study evaluated the influence of hypoxia duration and PUFA composition on the ability of cultured rat cardiomyocytes in producing alpha- and beta-adrenergic messengers (IPs and cAMP). After hypoxia (1.5, 2.5 or 3.5 h) followed by reoxygenation (1h). IP and cAMP production was induced by phenylephrine or isoproterenol stimulation, respectively. Hypoxia did not affect the basal level of messenger production in unstimulated cells, but decreased the cAMP production elicited by isoproterenol stimulation (up to 50%). The decrease in IP production after phenylephrine stimulation was observed only after long-term hypoxia duration close to irreversible cellular damages. The use of modified culture media supplemented with either arachidonic acid (AA) or docosahexaenoic acid (DHA) induced cardiomyocytes displaying either an arachidonic acid membrane profile (35% AA and 2% DHA in the phospholipids) or a docosahexaenoic acid membrane profile (15% AA and 20% DHA). These modifications did not alter the basal level of either messenger production in unstimulated cells nor the IP released after alpha-adrenergic stimulation. Conversely, the decrease in cAMP production was significantly more pronounced in docosahexaenoic acid-enriched cells than in arachidonic acid-enriched cells. This study suggests that hypoxia alters the beta-adrenergic messenger production, and that the alpha-system may balance the depression of the beta-system. The depression of the beta-adrenergic function induced by the incorporation of docosahexaenoic acid in membrane phospholipids may contribute to the beneficial effect of this fatty acid in the reperfused heart.

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PubMed1999

Cross-influence of membrane polyunsaturated fatty acids and hypoxia-reoxygenation on alpha- and beta-adrenergic function of rat cardiomyocytes.

The purpose of the present investigation was to determine whether the beneficial effects of polyunsaturated fatty acids (PUFA) may influence ischemia-reperfusion-induced alterations of myocardial alpha- and beta-adrenoceptor (alpha-AR, beta-AR) responsiveness. This study was carried out using monolayer cultures of neonatal rat ventricular myocytes in a substrate-free, hypoxia-reoxygenation model of ischemia. The cardiomyocytes (CM) were incubated during 4 days in media enriched either with n-6 PUFA (arachidonic acid, AA) or with n-3 PUFA (eicosapentaenoic acid, EPA, and docosahexaenoic acid, DHA). The n-6/n-3 ratio in n-3 CM was close to 1.2, compared to 20.1 in n-6 CM. The contractile parameters of n-6 CM and n-3 CM were similar in basal conditions as well as during hypoxia and reoxygenation. In basal conditions, the phospholipid (PL) enrichment with long chain n-3 PUFA resulted in an increased chronotropic response to isoproterenol (ISO) and to phenylephrine (PHE). After posthypoxic reoxygenation, the chronotropic response to beta-AR activation in n-6 CM was significantly enhanced as compared with the control response in normoxia. In opposition, the ISO-induced rise in frequency in n-3 CM in control normoxia and after reoxygenation was similar. In these n-3 CM, the changes in contractile parameters, which accompanied the chronotropic response, were also similar in reoxygenation and in normoxic periods, although the rise in shortening velocity was slightly increased after reoxygenation. In response to PHE addition, only the chronotropic effect of n-6 CM appeared significantly enhanced after hypoxic treatment. These results suggested that increasing n-3 PUFA in PL reduced the increase in alpha- and beta-AR functional responses observed after hypoxia-reoxygenation. This effect may partly account for the assumed cardiac protective effect of n-3 PUFA, through the attenuation of the functional response to catecholamines in the ischemic myocardium.

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PubMed1998

Correlation between direct ESR spectroscopic measurements and electromechanical and biochemical assessments of exogenous free radical injury in isolated rat cardiac myocytes.

Reactive free radical species appear to be involved in the ischemic injury of cardiac muscle, although the mechanisms by which oxygen-derived free radicals affect the heart cell function are not known. In the present study, cultured ventricular myocytes were exposed to an exogenous oxygen radical generating system. The myocyte-enriched, primary cultures were prepared from ventricles of new-born rat heart and exposed to a xanthine/xanthine oxidase (X+XO) system. The transmembrane potentials were recorded with glass microelectrodes. Cell contractions were monitored photometrically. The release of lactate dehydrogenase (LDH) in the medium was analysed. Quantitative measurement and the time course of the radical generation were performed by the electron paramagnetic resonance (EPR) spin trapping technique with the spin trap 5,5-dimethyl-1-pyroline-N-oxide (DMPO). We verified that X and XO alone had no significant functional and biochemical effects. The X+XO system produced a rapid decrease in the action potential amplitude. This effect was accompanied by a strong decrease in contractility and spontaneous rate. The time course of these functional defects were correlated with a progressive efflux of LDH from the cardiomyocytes. Prolonging the exposure to the X+XO system provoked the cessation of the spontaneous beatings and the progressive loss of the resting diastolic potential, together with a near total release of the cellular LDH. The LDH release and the functional depression were both efficiently prevented by catalase. On the contrary, superoxide dismutase (SOD) slowed down but did not protect against the functional and biochemical effects of the free radicals. In comparison, the EPR spectra obtained indicated that the X+XO system was associated with an important generation of superoxide anions but also with a small hydroxyl production. SOD scavenged the superoxide but a small .OH production persisted. Catalase (CAT) did not modify the superoxide generation but decreased the hydroxyl adduct formation. These results suggest that, although the generation of superoxide anions by the X+XO system was higher than the hydroxyl production, the functional injury and enzyme leakage seemed mainly mediated through a hydrogen peroxide-hydroxyl radical pathway. Cultured ventricular myocytes can be thus used as a valuable model to investigate the cellular mechanism of oxidant-induced damage in the heart.

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PubMed1997

Influence of phospholipid long chain polyunsaturated fatty acid composition on neonatal rat cardiomyocyte function in physiological conditions and during glucose-free hypoxia-reoxygenation.

There is evidence that dietary polyunsaturated fatty acids (PUFA) may protect against cardiovascular diseases, but the involvement of the cardiac muscle cell in this beneficial action remain largely unknown. The present study compared the respective influence of n-3 and n-6 PUFA on the function of cultured neonatal rat cardiomyocytes (CM). Cells were grown for 4 days in media enriched either n-3 (eicosapentaenoic acid, EPA and docosahexaenoic acid, DHA) or n-6 (arachidonic acid, AA) PUFA. The PUFA n-6/n-3 ratio in the phospholipids was close to 1 and 20 in the n-3 and n-6 cells, respectively. The transmembrane potentials were recorded using microelectrodes and the contractions were monitored with a photoelectric device. In physiological conditions, the increase of n-6 PUFA level in the phospholipids resulted in a significant decrease in the maximal rate of initial depolarization (-16%). In opposition, the action potential amplitude and duration were not altered, and the cell contraction outline was not affected. Ischemia was simulated in vitro using a substrate-free, hypoxia-reoxygenation procedure in a specially designed gas-flow chamber. The progressive loss of electrical activity induced by the substrate-free, hypoxic treatment was affected by the n-6/n-3 ratio, since the n-6 rich CM displayed a slower depression of the AP amplitude and duration parameters. Conversely, the recovery of the resting potential (MDP) during reoxygenation was faster in n-3 CM, whereas the recovery of the contraction parameters was unaffected by the fatty acid composition of the cells. These results suggested that, in physiological conditions, the modification of long chain PUFA balance in the phospholipids of cardiac muscle cells may modulate the initial AP upstroke, which is governed by sodium channels. Moreover, the presence of n-3 PUFA appeared to accelerate the electrical depression during substrate-free hypoxia but in turn to allow a faster recovery upon reoxygenation.

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PubMed1997

Protective effects of trimetazidine on hypoxic cardiac myocytes from the rat.

The electrophysiological effects of the antianginal drug trimetazidine (TMZ) were investigated in cultured rat ventricular myocytes using a substrate-free hypoxia model of ischemia. The transmembrane potentials were recorded with glass microelectrodes and the contractions were simultaneously monitored with a video motion detector. The cardiomyocytes were treated with TMZ (1-5.10(-4) M final concentration) in the bath. The untreated and the drug-treated cells were submitted either to 150 min normoxia or to 150 min hypoxia followed by 90 min reoxygenation in the absence of oxidizable substrate. In normoxic conditions, TMZ did not affect the maximal diastolic potential (MDP) but significantly lowered the plateau potential level (OS) and decreased the upstroke velocity (Vmax) and the spontaneous action potential rate (APR). Conversely, TMZ significantly increased action potential duration at 80% repolarization (APD80). Under substrate-free hypoxia, the untreated cells displayed a progressive contractile failure and an important decrease in OS and APD. In parallel, early postdepolarizations triggering high rate spikes were observed. Prolonging oxygen depletion led to the cessation of the spontaneous electrical activity and thereafter to a gradual decrease in MDP. Near normal rhythmic action potentials and contractions resumed after reoxygenation. Comparatively, the treatment by 5.10(-4) M TMZ almost completely prevented the decrease in plateau amplitude, resting membrane potential, Vmax, APD80, and rate caused by substrate-free hypoxia. Moreover, the hypoxia-induced arrhythmias and the cessation of spontaneous electromechanical activities did not occur in the presence of TMZ (5.10(-4) M). After reoxygenation, the TMZ-treated cells exhibited a higher action potential amplitude than that of the untreated cells, although the TMZ-induced depressive effects on the spontaneous frequency and the Vmax persisted. In conclusion, this study shows that TMZ (5.10(-4) M) is efficient in protecting the isolated cardiac myocytes against the functional alterations induced by substrate-free hypoxia and led thus to a better recovery upon reoxygenation. The cytoprotective action may be linked, at least in part, to apparent ion channel blocking effects of the drug, which appeared in basal conditions at concentrations used in this study.

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PubMed1996

Membrane docosahexaenoic acid vs. eicosapentaenoic acid and the beating function of the cardiomyocyte and its regulation through the adrenergic receptors.

The beta-adrenergic system in cardiac muscle cells is influenced by the n-3 polyunsaturated fatty acid (PUFA) content in cell membranes. This study deals with the specific effect of docosahexaenoic acid (DHA) as compared to other n-3 PUFA, without modification of the arachidonic acid content. Increasing the DHA content in the phospholipids of isolated cardiomyocytes did not provoke electrical or contraction modifications, except for a slightly lower plateau phase (-2 mV). Conversely, the beta-adrenergic function was affected at several levels: (i) the receptor affinity for dihydroalprenolol tended to decrease (Kd) without alteration of the number of beta-binding sites (beta max); (ii) the isoproterenol-induced (10(-7) M and 10(-6) M) cAMP production was significantly decreased (-20%); and (iii) the positive chronotropic response to beta-adrenergic stimulation (isoproterenol, 10(-7) M) was significantly enhanced (+80%). In order to further investigate the relationship between the decreased cAMP and the increased chronotropic response, the cells were treated with dibutyryl-cAMP, a permeant analogue of cAMP, which elicited a significantly higher chronotropic response in DHA-enriched cells than in EPA-enriched cells +50%). The increase in DHA content in cardiac cell membranes phospholipids may thus affect the beta-adrenergic system through an increase of cAMP efficiency. Although the membrane phosphatidylinositols were largely involved in the PUFA alterations, none of the parameters related to alpha-adrenergic system (chronotropic response, receptor density, affinity for prasozin, and inositide phosphate production) were influenced by the membrane DHA content.

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PubMed1995

Trimetazidine: in vitro influence on heart mitochondrial function.

The mechanism of action of the antianginal trimetazidine (TMZ) remains largely unknown. In cultured rat ventricular myocytes in physiologic conditions, TMZ (5 x 10(-4) M) reduced the plateau potential level, the upstroke velocity, and the spontaneous action potential rate. When the cardiomyocytes were submitted to hypoxia (150 or 240 minutes) in a glucose-free medium, treatment with TMZ largely prevented the hypoxia-induced electromechanical alterations, i.e., the decrease in plateau amplitude, in resting membrane potential, in action potential duration, in rate, and in contractility. No hypoxia-induced arrhythmia was observed in the TMZ-treated cells. Moreover, the lactate dehydrogenase leakage was significantly reduced in the TMZ-treated cardiomyocytes (-58% and -36%, after 150 and 240 minutes of hypoxia, respectively). The drug was not efficient in reducing the hypoxia-induced decrease in adenosine triphosphate (ATP) content. The cellular ATP content was slightly lower in the TMZ-treated cells in normoxic conditions and in hypoxic conditions, but only in the glucose-free medium. To investigate further the relation between TMZ and energy metabolism, the respiration parameters were measured in heart mitochondria isolated from control and TMZ-treated rats (6 mg/kg/day, 7 days) with different substrates. This treatment resulted in a slight alteration of pyruvate oxidation, which was observed in the absence and in the presence of TMZ (10(-4) M) in the respiration medium. Conversely, a potent inhibition of palmitoylcarnitine oxidation was measured when TMZ was added to the respiration medium. Neither pretreatment of the rats, nor addition of TMZ to the medium affected the oxidation of glutamate or citrate.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗

PubMed1995

Effect of docosahexaenoic acid and eicosapentaenoic acid in the phospholipids of rat heart muscle cells on adrenoceptor responsiveness and mechanism.

The specific effect of docosahexaenoic (DHA; C22:6 n-3), as compared to eicosapentaenoic acid (EPA; C20:5 n-3), on adrenoceptor function was investigated in cultured rat myocardial cells. The cardiomyocytes were grown for 24 h in a conventional seric medium, and then incubated for 96 h in a medium enriched with either DHA or EPA. After this treatment, the phospholipids of the DHA-treated cells contained approximately 20% of the total fatty acids as C22:6 n-3, and those of EPA-treated cells displayed a high content in C20:5 n-3 and its elongation product C22:5 n-3 (30% of total fatty acids). Additionally, the n-3/n-6 polyunsaturated fatty acid ratio was the same in the two groups of cells. These modifications were roughly similar in all the phospholipid classes. The contractions were monitored photometrically and no significant difference in basal frequency and contraction parameters could be detected. The stimulation of the beta-adrenergic receptors (isoproterenol 10(-7) M) resulted in a positive chronotropic effect, which was significantly higher in the DHA-rich cells. Conversely, the higher DHA content in the phospholipids appeared to induce a decrease in the affinity of the beta-receptors for the ligand (dihydroalprenolol) without alteration of the number of beta-receptor binding sites and provoked a significant decrease in isoproterenol-stimulated cAMP production (-19%). To investigate further these controversial data, the cardiomyocytes were treated with dibutyryl-cAMP, which elicited a positive chronotropic response significantly higher in the DHA-rich cells. The alpha-adrenergic stimulation by phenylephrine (3 x 10(-6) M) increased the spontaneous rate, but in a similar manner in the DHA- and EPA-enriched cells. Similarly, neither the alpha-adrenergic receptor binding characteristics nor the production of phosphoinositides was modulated by the membrane DNA content, although the phosphatidylinositol PUFAs were significantly altered. In conclusion, increasing the DHA content in membrane phospholipids did not affect the alpha-adrenergic system, but exerted a specific positive influence on the beta-adrenergic transduction mechanism, essentially through an increase of cAMP efficiency.

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PubMed1995

Influence of the phospholipid content in docosahexaenoic acid on electrophysiology and contraction of rat heart muscle cells.

The cardiovascular beneficial effects of fish oils are currently attributed to docosahexaenoic (C22:6 n-3) and eicosapentaenoic (C20:5 n-3) acids, although most investigations have focused on eicosapentaenoic acid. This study was devoted to the specific effect of docosahexaenoic acid, as compared to eicosapentaenoic acid, on the basal electrophysiological and mechanical characteristics of cultured rat myocardial cells. The myocyte cultures were prepared from newborn rat heart ventricles. The cells were grown for 24 hours in a conventional seric medium, and then incubated in a medium enriched with either docosahexaenoic acid or eicosapentaenoic acid for 96 hours. This treatment resulted in docosahexaenoic acid-rich cells (16% of the phospholipid fatty acids) and docosahexaenoic acid-poor cells (1.5%), both displaying the same phospholipid n-6/n-3 polyunsaturated fatty acid ratio. The transmembrane potentials were recorded with glass microelectrodes. Contractions were monitored photometrically. The action potential amplitude was slightly smaller in docosahexaenoic acid-rich cells (-4 mv), due to a lower plateau phase. There was no difference in action potential duration and spontaneous rate. The contraction measurements were not significantly different between the two groups of cells. We conclude that increasing the docosahexaenoic acid content in cardiomyocyte membrane phospholipids may have modulated the calcium ionic channels governing the plateau phase of the action potential, whereas the other physiological activities remained unaffected.

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PubMed1994

Some biochemical aspects of the protective effect of trimetazidine on rat cardiomyocytes during hypoxia and reoxygenation.

This study was undertaken to evaluate the direct cardioprotective effect of trimetazidine (TMZ), an anti-anginal drug devoid of haemodynamic action, on isolated myocytes. Cultured rat ventricular myocytes were treated with the drug 16 h and 1 h before the experiments. The drug-treated cells and control cells were placed in a substrate free medium and submitted in a specially designed device to either normoxia (N4), or hypoxia (150 min, H2.5, or 240 min, H4), or 150 min hypoxia followed by 90 min reoxygenation (HR). The treatment of the cells with TMZ (5 x 10(-4) M) resulted in a significant decrease of lactate dehydrogenase (LDH) leakage (-58% in H2.5, -36% in H4 and -37% in HR). The LDH release provoked by oxidizing agents. H2O2 and 13-s-HpOTrE (13(S)-hydroperoxyoctadecatrienoic acid) during post-hypoxic reoxygenation was also lowered by TMZ. However, this effect reflected the beneficial action of TMZ during hypoxia since the drug was not efficient in altering the LDH leakage induced by the oxidizing agents in normal conditions. Moreover, the hypoxia-induced decrease of ATP content was not affected by TMZ, and resynthesis of ATP during substrate-free reoxygenation was similar in TMZ-treated and control cells. The respiration parameters have been studied in rat heart mitochondria isolated from control and TMZ-treated rats, in the presence or absence of TMZ in the respiration medium (10(-4) M). The main result was a rapid and potent inhibition of palmitoylcarnitine oxidation, when TMZ was added to the respiration medium. The chronic treatment only resulted in a slight alteration of pyruvate oxidation. In conclusion, a pre-treatment of ventricular myocytes with TMZ resulted in an increased cell resistance to hypoxic stress, as evidenced by LDH leakage. This cytoprotective effect of TMZ should not be mediated through an antioxidant activity, but could be related to a modification of lipid metabolism.

Adenine Nucleotides↗

PubMed1993

Anthracycline toxicity is potentiated by iron and inhibited by deferoxamine: studies in rat heart cells in culture.

The interrelation between iron, iron chelation, and anthracycline toxicity was investigated in a heart cell culture system. Two indicators of cellular damage have been used, lactate dehydrogenase (LDH) release and cell contractility. Both of these indicators have shown a marked increase in doxorubicin toxicity by prior iron loading. This was not a simple additive effect, because at the concentrations used, iron had only a minimal effect on LDH release and no effect at all on contractility, whereas doxorubicin had only a minor effect on contractility. Deferoxamine (DF) treatment of iron-loaded heart cells resulted in a marked decrease in anthracycline toxicity as judged both by LDH leakage and cell contractility. However, DF treatment of normal heart cells had no measurable protective effect against doxorubicin toxicity, whether DF was administered before or simultaneously with doxorubicin. Doxorubicin treatment did not alter cellular malondialdehyde (MDA) concentrations in either normal or iron-loaded cells. Conversely, the protective effect of DF in iron-loaded cells and its failure to prevent anthracycline toxicity in normal cells were both associated with a significant decrease in MDA measurements. Our data indicate that iron overload aggravates anthracycline toxicity and that this interaction may be prevented by effective iron chelating therapy. Because patients requiring anthracycline therapy often have increased tissue iron stores caused by multiple blood transfusions and bone marrow suppression, our observations may have important implications for the prevention of anthracycline toxicity.

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PubMed1992

Eicosapentaenoic and docosahexaenoic acids in cultured rat ventricular myocytes and hypoxia-induced alterations of phospholipase-A activity.

Hypoxia was reported to induce a decrease in phosphatidylcholine-hydrolyzing phospholipase activity (PC-PLA) in cultured rat cardiomyocytes. This work was intended to compare the influence of the presence of either eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) in the phospholipids on the PC-PLA activity in normoxic and hypoxic conditions. The enrichment of the medium with EPA or DHA resulted in cell phospholipids containing about 2% or 22% DHA, respectively. These cells were then submitted for 3.5 h to either normoxia or hypoxia and the PC-PLA activities were assayed using [1-14C] dioleoyl-PC (pH 8.4 for PC-PLA2 and 4.9 for PC-PLA1). The results show that both enzymic activities are significantly higher in DHA-rich cardiomyocytes. Hypoxia induced a significant decrease in PC-PLA2 (about 25%) which was not statistically different between the two groups of cells. The hypoxia-induced decrease in PC-PLA1 was not found significant. In conclusion, the nature of the long chain n-3 polyunsaturated fatty acids in the phospholipids appears to contribute to the regulation of PC-PLA activity but not to influence its decrease during hypoxia.

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PubMed1992

Incorporation of cyclic fatty acid monomers in lipids of rat heart cell cultures.

Primary cultures of newborn rat cardiomyocytes were grown in medium supplemented with cyclic fatty acid monomers (CFAM) which had been isolated from heated linseed oil. The cells were harvested, and lipids were extracted and fractionated using silica cartridges and high-performance liquid chromatography. The CFAM structures isolated from cellular lipids were determined and compared to those that had been supplemented to the medium, using gas-liquid chromatography coupled with mass spectrometry (GC/MS). We found that CFAM were incorporated into phospholipids and neutral lipids of cardiomyocytes. Furthermore, CFAM with a cyclopentyl ring structure were more abundant in cardiomyocytes than were the cyclohexyl ring isomers. Our data suggest that CFAM of the 5-carbon and 6-carbon ring series are metabolized differently in newborn rat cardiomyocytes.

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