Characteristics of the Nonselective Cation Current (NSCCs) in Rabbit Left Ventricular Epicardial, Midmyocardial and Endocardial Myocytes

Objectives Recent studies have described regional differences in the electrophysiology and pharmacology of ventric- ular myocardium in canine, feline, rat, guinea pig, and human hearts. This has been shown to be due to a smaller IKs and a lager sodium-calcium exchange current （INa-Ca） and late INa in M region （ deep subepicardial to midmyocardial）. Studies from our laboratory have found a new repolarization current-nonselective cation current （NSCCs） existing in rabbit fight ventricular myocytes. Methods We examined the characteristics of NSCCs in epicardial, M region, and endocardial cells isolated from the rabbit left ventricle with standard microelectrode and whole-cell patch-clamp tech- niques. The permeability to Na^＋ , K^＋ , Li^＋ , Cs^＋ but not to Cl^- indicating that it was a nonselective cation current. Gd^＋3 （0. 1 mmol/1） and La^3＋ （0. 1 retool/1 ） can block the current markedly. Results Further characterization of NSCCs was significantly smaller in M cells than in epicardial and endocardial cells. NSCCs current density was significantly smaller in M cells than in epicardial and endocardial cells. With repolarization to - 80 mV, INa current density was （ -0. 44 ±0. 05） PA/PF in endocardial cells, （ -0. 12 ±0. 05） PA/PF in M cells and （ - 0. 28 ±0. 07） PA/PF in epicardial cells ; and with repolarization to ＋ 30 mV, INa, current density was （ 1.09 ± 0. 29） PA/PF in endocardial cells, （0. 38±0. 09） PA/PF in M cells and （0. 91 ± 0. 32） PA/PF in epicardial cells. Conclusions Transmural dispersion of repolarization was due to the heterogeneity of NSCCs in rabbit left ventricle epicardial, endocardial myocytes and M cells. These findings may advance our understanding of the ionic basis for our understanding of factors contributing to the development of cardiac arrhythmias.

Protective function of tocilizumab in human cardiac myocytes ischemia reperfusion injury

Objective:To investigate the protective function of tocilizumab in human cardiac myocytes ischemia-reperfusion injury.Methods:The human cardiac myocytes were treated by tocilizumab with different concentrations(1.0 mg/mL,3.0 mg/mL,5.0 mg/mL) for 24 h.then cells were cultured in ischemia environment for 24 h and reperfusion environment for 1 h.The MTT and flow cytometry were used to detect the proliferation and apoptosis of human cardiac myocytes,respectively.The mRNA and protein expressions of Bcl-2 and Bax were measured by qRT-PCR and western blot,respectively.Results:Compared to the negative group,pretreated by tocilizumab could significantly enhance the proliferation viability and suppress apoptosis of human cardiac myocytes after suffering ischemia reperfusion injury(P<0.05).The expression of Bcl-2 in tocilizumab treated group were higher than NC group(P<0.05).while the Bax expression were lower(P<0.05).Conclusions:Tocilizumab could significantly inhibit apoptosis and keep the proliferation viability of human cardiac myocytes after suffering ischemia reperfusion injury.Tocilizumab may obtain a widely application in the protection of ischemia reperfusion injury.

Effect of Cholic Acid on Fetal Cardiac Myocytes in Intrahepatic Choliestasis of Pregnancy

This study examined the effect of cholic acid （CA） on cultured cardiac myoeytes （CMs） from neonatal rats with an attempt to explore the possible mechanism of sudden fetal death in intra- hepatic cholestasis of pregnancy （ICP）. Inverted microscopy was performed to detect the impact of CA on the beating rates of rat CMs. MTT method was used to study the effect of CA on the viability of CMs. CMs cultured in vitro were incubated with 10 ~maol/L Ca2＋-sensitive fluorescence indicator fluo-3/AM. The fluorescence signals of free calcium induced by CA were measured under a laser scanning confocal microscope. The results showed that CA decreased the beating rates of the CMs in a dose-dependent manner. CA could suppress the activities of CMs in a time- and dose-dependent manner. CA increased the concentration of intracellular free calcium in a dose-dependent manner. Our study suggested that CA could inhibit the activity of CMs by causing calcium overload, thereby leading to the sudden fetal death in ICP.

Voltage- and Use-dependent Effect of 7-chlor-benzylte-trahydropalmatine on Sodium Currents in Guinea Pig Ventricular Myocytes

The whole-cell patch-clamp technique was employed to obtain information about the voltage-dependence and kinetics of interaction of 7-chlor-benzylte-trahydropalmatine (7-Cl-BTHP) with cardiac sodium channels. 7-Cl-BTHP (30 mol/L) significantly decreased the peak sodium current (from 7. 8±1. 8 nA to 5. 3±1. 4 nA, P<0. 01, n=5), without producing a shift of the current-voltage curve. It shifted the inactivation curves of sodium current to hyperpolarized potentials, and the V(0.5) was shifted from - (82. 5±2. 5) mV to - (95±2.4) mV (P <0. 05, n=4). 7-Cl-BTHP produced a significant use-dependent effect that was proportional to the duration of the voltage step. In addition, 7-Cl-BTHP slowed the recovery of sodium channel from inactivation, which could explain its use-dependent effects on sodium current. The characteristics of 7-Cl-BTHP blockage suggest that this agent binds preferentially to inactivated sodium channels.

EFFECTS OF ADENOSINE IN SIMULATED ISCHEMIA ANDREPERFUSION ON ISOLATED GUINEA-PIG VENTRICULAR MYOCYTES

Objective To investigate the effects of adenosine （Ado） on myocardiac electrophysiology in simu- lated ischemla and reperfusion in guinea-pig ventricular myocytes. Methods Electrical activity was recorded using standard intracellular microelectrode technique. Right ventricle was superfused with simulated ischemic Tyrode’s so- lution for 15 min, and reperfued with normal Tyrode’s solution for 30 min. Results The results showed Ado had no measurable effects on guinea-pig ventricular myocytes in normal Tyrode’s solution. In the presence of Ado, maximal diastolic potential tended to be more depolarized during ischemia, and action potential （AP） parameters were abbrevi- ated greatly in a concentration-dependent manner. Especially, the concentration of Ado 100 μmol·L-1 had significant effects on AP parameters in ischemic phase [APD30, APD50, and APD90 reduced by （86±8）% versus （65±6）%, （70 ±7）% versus （50±6）%, and （60±6）% versus （42±4）% for control after 15 min, P<0.O5]. During reperfu- sion, AP parameters did not completely return to initial values in presence of Ado. This study illustrated that Ado significantly decreased incidence of arrhythmias induced by ischemia and reperfusion (in presence of Ado 100 μmol· L-1, the incidence of DAD decreased by 17% versus 82% for control during reperfusion). Conclusion Ado has no significant effects on guinea-pig ventricle in normal conditions, abbreviates greatly AP parameters during ischemia with a concentration-dependent manner, and has marked antiarrhythmic effects in ischemia and reperfusion.

In Vitro Study on Mesenchymal Stem Cells:Anti-apoptotic Effects on Cardiac Myocytes

Objectives To investigate the anti-apoptotic effects of mesenchymal stem cells （MSCs） on hypoxic injured cardiac myocytes in vitro. Methods MSCs were isolated from bone marrow of Sprague-Dawley （SD） rats, and cardiac myocytes from neonatal rats. The rat cardiac myocytes were co-cultured with MSCs or MSC-conditioned media in anoxia （95% N2 ±5% CO2） for 72 hours. Cell apoptosis was measured by Hoechst 33258 staining. The expression of Bcl-2 and Bax in cardiac myocytes was tested by Western Blot. Results The apoptotic rate was 51.6% ± 2.4% when cardiac myocytes were cultured in continuous hypoxia and was significantly decreased when cardiac myocytes were cocultured with MSCs or MSC-conditioned media （ 15.1% ± 5.4% and 24. 0% ± 4.2% respectively, P 〈 0. 001 ）. The decreased expression of Bax in the cardiac myocytes was greatly related to the decreasing of apoptosis, but there was no difference in Bcl-2 expression among these groups. Conclusions Co-cultured MSCs showed significant anti-apoptotic effects on cardiac myocytes in continuous hypoxia. The mechanism may be the interact of cell to cell and paracrine of cytokines which effected the expression of Bax in the cardiac myocytes.

Antioxidant Effect of Human Selenium-containing Single-chain Fv in Rat Cardiac Myocytes

Reactive oxygen species（ROS） plays a key role in human heart diseases. Glutathione peroxidase（GPX） functions as an antioxidant as it catalyzes the reduction of hydroperoxide. In order to investigate the antioxidant effect of human selenium-containing single-chain Fv（Se-scFv-B3）, a new mimic of GPX, a model system of hydrogen peroxide（H202）-induced rat cardiac myocyte damage was established. The cardiac myocyte damage was characte- rized in terms of cell viability, lipid peroxidation, cell membrane integrity, and intracellular H202 level. The Se-scFv-B3 significantly reduced H2O2-induced cell damage as shown by the increase of cell viability, the decline of malondialdehyde（MDA） production, lactate dehydrogenase（LDH） release, and intracellular H2O2 level. So Se-scFv-B3 may have a great potential in the treatment of human heart diseases induced by ROS.

Characterization and Differentiation into Adipocytes and Myocytes of Porcine Bone Marrow Mesenchymal Stem Cells

Bone marrow mesenchymal stem cells （BMSCs） could differentiate into various cell types including adipocytes and myocytes, which had important scientific significance not only in the field of tissue regeneration, but also in the field of agricultural science. In an attempt to exhibit the characterization and differentiation into adipocytes and myocytes of porcine BMSCs, we isolated and purified porcine BMSCs by red blood cell lysis method and percoll gradient centrifugation. The purified cells presented a stretched fibroblast-like phenotype when adhered to the culture plate. The results of flow cytometry analysis and immunofluorescence staining demonstrated that the isolated cells were positive for mesenchymal surface markers CD29, CD44 and negative for hematopoietic markers CD45 and the adhesion molecules CD31. Cells were induced to differentiate into adipocytes with adipogenic medium containing insulin, dexamethasone, oleate and octanoate. Oil Red O staining demonstrated that the porcine BMSCs successfully differentiated to adipocytes. Moreover, the findings of real-time PCR and Western blotting indicated that the induced cells expressed adipogenic marker genes （PPAR-y, C/EBP-c~, perilipin, aP2） mRNA or proteins （PPAR-3,, perilipin, aP2）. On the other hand, porcine BMSCs were induced into myoctyes with myogenic medium supplemented with 5-azacytidine, basic fibroblast growth factor, chick embryo extract and horse serum. Morphological observation by hochest 33342 staining showed that the induced cells presented as multi-nucleus muscular tube structure. And myogenic marker genes （Myf5, desmin） mRNA or proteins （MyfS, MyoD, myogenin, desmin） were found in the induced cells. In addition, the results of immunofluorescence staining revealed that myogenic marker （Myf5, MyoD, myogenin, desmin, S-MyHC） proteins was positive in the induced cells. Above all, these results suggested that the isolated porcine BMSCs were not only consistent with the characterization of mesenchymal stem cells, but also exhibited the multipotential capacity to form adipocytes and myocytes, which provided the basis to investigate the regulation mechanism involved in the selective differentiation of porcine BMSCs.

Antioxidant Effect of Selenium-containing Glutathione S-Transferase in Rat Cardiomyocytes

As one of the most important antioxidant enzymes, glutathione peroxidase（GPX） protects cells and tissues from oxidative damage, and plays an important role in cardiovascular and cerebrovascular injuries induced by oxida- tive stress. The antioxidant effect of selenium-containing glutathione S-transferase（Se-GST）, a mimic of GPX was investigated on rat cardiomyocytes. To explore the protection function of Se-GST in hydrogen peroxide（H202） chal- lenged rat cardiomyocytes, we examined malondialdehyde（MDA）, lactate dehydrogenase（LDH）, superoxide dismu- tase（SOD） and cell apoptosis. The results demonstrate exposure of rat cardiomyocytes to H202 for 6 and 12 h induced the significant increases of MDA, LDH and apoptosis rate of cardiomyocytes, but pretreatment of rat cardiomyocytes with Se-GST at 0.0005 or 0.001 unit/mL prevents oxidative stress induced by H202 with the decreases of cell apopto- sis. All the results hint Se-GST has antioxidant activity for oxidative stress challenged rat cardiomyocvtes.

Effects ofβ_2-Adrenergic Antagonist on Cytosolic Ca^(2+) in Ventricular Myocytes from Infarcted Rat Heart

Objectives To investigate the effects of β2-adrenergic antagonist on cytosolic Ca^2 ＋ （[Ca^2＋ ]i） in ventricular myocytes from infarcted rat heart. Methods A ligature was placed around left anterior descending coronary artery of rat hearts. Rats in the control group were sham-operated. Cardiomyocytes were dissociated at two, four, eight weeks after myocardial infarction （MI） and [Ca^2＋]i was measured via fura-2 fluorescence. The response of cardiomyocytes to isoproterenol in presence or absence of betal-adrenergic antagonist atenolol, beta2-adrenergic antagonist ICI118, 551 or non-selective β1, 2- adrenergic antagonists propranolol was examined. Results The followings were found that ICI 118, 551 had no significant effects on the rise of [Ca^2＋]i induced by isoproterenol in normal ventricular myocytes （P 〉 0.05）, ICI118, 551 only significantly attenuated the rise of [Ca^2＋]i induced by isoproterenol at four weeks and eight weeks after MI （24.5%±5.7% vs 57.8% ± 13.2%, P〈 0.01; 12.2%±7.9% vs 44.6%±11.3%, P〈 0.01）. Atenolol had suppressive effects only in the control group and the post-MI group of two weeks （P 〈 0.05）, and propranolol had suppressive effects in the control and all the three post-MI groups （P 〈 0.01）. Conclusions Beta2-adrenergic antagonist ICI118, 551 may exert negative effects on Ca^2＋ overload initiated by sympathetic stimulation after MI.

Effects of simvastatin on hypertrophy and PTEN expression of rat cardiac myocytes

目的将在心脏的 myocytes 在信号小径由浆液和在染色体十上删除的磷酸酶和 tensin 相当或相同的事物(PTEN ) 的角色导致了的有教养的老鼠的肥大上学习 simvastatin 的效果。方法有教养的新生的 Sprague-Dawley (SD ) 老鼠心脏的 myocytes 与 15% 胎儿的牛的浆液被对待，或没有 simvastatin 的浆液，或不同 consentrations。图象分析系统被用来测量心脏的 myocytes 表面区域。myocytes 的蛋白质合成被测量经由[3H ] 白氨酸加入方法。atrial natriuretic 的表示水平在 myocytes 的肽(ANP ) mRNA 与反向的抄写聚合酶链反应(RT-PCR ) 被决定。在心脏的 myocytes 的 PTEN 的 mRNA 和蛋白质表示层次分别地与 RT-PCR 和西方的污点被调查。在 24 个小时的结果，心脏的 myocytes 表面区域在 15% 浆液组是显著地更高的(1611.

The Effect of Levobunolol Hydrochloride on the Calcium andPotassium Channels in Isolated Ventricular Myocytes of Guinea Pig

The effects of levobunolol hydrochlorid (Bun) on the type L calciumchannel currents (Ica) and delayed rectifier potassium channel currents (Ik) in isolated ventricular myocytes of guinea pig were studied by using patch clamp wholecell recording techniques. The results were showed that: 1) Bun caused a dosedependent decrease in Ica and a dose-dependent increase in Ik of the ventricular myocytes.The threshold concentrations of Bun for Ica and Ik were 10-8 mol/L and10-7 mol/L respectively. The maximum effective concentration of Bun for both Ica and Ik was 3 × 10-5 mol/L, and half-maximal concentration was 3 × 10-6 mol/L;2 ) Ik was blocked by 2× 10-6mol/L tetraethylammonium (TEA). A concentration of 3 × 10-6 mol/L Bun showed a decreasing effect on the Ica as revealed by the current-voltage relationship curve, i. e., Bun caused an elevation of the curve; 3)When Ica was blocked by 2 × 10-6 mol/L Isoptin (Verapamil), at a concentrationof 3 × 106- mol/L Bun showed an increasing effect on Ik and the effect could be blocked by TEA. The above-mentioned results indicated that Bun had an inhibito-ry effect on Ica and a fascilitatory effect on Ik The results suggested that themolecular mechanisms of antihypertensive, heart rate slowing and β-receptorblocking effects of Bun might be due to decrease of Ica and increase of Ik.

Protein Kinase C Enhances the Swelling-Induced Chloride Current in Human Atrial Myocytes

Swelling-activated chloride currents（ICl.swell） are thought to play a role in several physiologic and pathophysiologic processes and thus represent a target for therapeutic approaches. However, the mechanism of ICl.swell regulation remains unclear. In this study, we used the whole-cell patch-clamp technique to examine the role of protein kinase C（PKC） in the regulation of ICl.swell in human atrial myocytes. Atrial myocytes were isolated from the right atrial appendages of patients undergoing coronary artery bypass and enzymatically dissociated. ICl.swell was evoked in hypotonic solution and recorded using the whole-cell patch-clamp technique. The PKC agonist phorbol dibutyrate（PDBu） enhanced ICl.swellin a concentration-dependent manner, which was reversed in isotonic solution and by a chloride current inhibitor, 9-anthracenecarboxylicacid. Furthermore, the PKC inhibitor bis-indolylmaleimide attenuated the effect and 4α-PDBu, an inactive PDBu analog, had no effect on ICl.swell. These results, obtained using the whole-cell patch-clamp technique, demonstrate the ability of PKC to activate ICl,swell in human atrial myocytes. This observation was consistent with a previous study using a single-channel patch-clamp technique, but differed from some findings in other species.

Effect of post-burn serum on free cytosolic calcium in isolated cardiac myocytes of adult rats

The ohjective of this study was to determine whether the free intracellular calcium concentration ([Ca2+] ) of isolatedcardiac myocytes increased with the stimulation of post-burn serum(PBS) in adult rats. Cardiac myocytes were isolated by collage-nase using Langendorff’s perfusion apparatus, and [Ca2+], was measured using the fluorescent indicator Fain-2. The normal[Ca2+], was 101. 3 ± 21. 3 nmol/L in cardic myocytes. PBS at various postburn home could very significantly increase the[Ca2+]i (P< 0. 01 ) and, 6 h PBS had the strongest effect. However, no significant difference was found between the effects of2 h PBS and 4 h PBS (P >0. 05 ). Both calcium channel antagonist verapamil(30 umol/L) and the inhibitor of ryanodine receptoron sarcoplasmic reticulum procaine (2 mmol/L), very significantly inhibited the action of 6 h PBS, with the inhibition rate of47. 7% and 67. 6% respectively. The inhibiting rate of procaine was significantly greater than that of verapamil (P < 0. 01 ). Theresults suggested that PBS could stimulate the increase of [Ca2+], in isolated cardiac myocytes of adult rats, in which calcium release from intracellular stores might play greater roles. Agents modulating the calcium release from intracellular stores are expectedto have great significance in preventing the organic injuries due to the increases of [ Ca2+]i.

Effects of simulated microgravity on nitric oxide level in cardiac myocytes and its mechanism

The depression of cardiac contractility induced by space microgravity is an important issue of aerospace medicine research, while its precise mechanism is still unknown. In the present study, we explored effects of simulated microgravity on nitric oxide (NO) level, inducible nitric oxide synthase (iNOS) expression and related regulative mechanism using electron spin resonance (ESR) spectroscopy, immunocytochemistry and in situ hybridization. We found a remarkable in-crease of NO level and up-regulation of iNOS and iNOS mRNA expression in rat cardiac myocytes under simulated microgravity. Staurosporine (a nonselective protein kinase inhibitor), calphostin C (a selective protein kinase C inhibitor), partially inhibited the effect of simulated microgravity. Thus regulative effect of simulated microgravity on iNOS expression is mediated at least partially via activation of protein kinase C. These results indicate that NO system in cardiac myocytes is sensi-tive to simulated microgravity and may play an important role in the depression of cardiac contrac-tility induced by simulated microgravity.

Differences of promethazine and terfenadine on ion channels in guinea pig ventricular myocytes

Promethazine, a first generation antihistamine,has an antiarrhythmic effect on ischemia-reperfusion inducing arrhythmias^1 and experimental arrhythmias.^2 However, terfenadine as a second generation of antihistamine, has been reported to elicit hypotension, bradycardia, prolongation of the QTc interval and torsades de pointes （TdP） like ventricular arrhythmia.^3 This may be due to the blockage on rectifier postassium current （Ik） of terfenadine, resulting in the prolongation of the action potential duration （APD） and dispersion of the repolarization duration, which might provoke a specific form of polymorphic ventricular tachydysrhythmia, i.e. TdP.^4 In clinical practice,however, the class Ⅲ antiarrhythmic agents, which target on the lk and prolong the action potential duration and QTc interval, rarely lead to arrhythmias.Other actions must be considered to underlie the arrhythmogenic tendency of terfenadine besides its inhibition on Ik. Though both promethazine and terfenadine block the H1 receptor, there must be a different pharmacology profile between the two compounds on ion channels of cardiac myocytes.Whole-cell patch clamp technique was used to investigate the effects of these two antagonists of the H1 receptor on the main ion currents in cardiac electrical activities.

Effects of adiponectin on oxidative stress and apoptosis in human cardiac myocytes cultured with high glucose

Background Diabetic cardiomyopathy is the major cause of morbidity and mortality in diabetic patients. Oxidative stress plays an important role in diabetic cardiomyopathy. This study aimed to investigate the effects of adiponectin on oxidative stress and apoptosis in human cardiac myocytes （HCM） cultured with high glucose. Methods The cells were assigned to three group： control group, high glucose group and high glucose plus adiponectin group. After culture for 24, 48, 72 hours, oxidative stress was evaluated by detecting levels of malondialdehyde （MDA） and superoxide dismutase （SOD） in the supernatant of culture media. The expression of p66Shc and Heme oxygenase-1 （HO-1） was detected by real-time polymerase chain reaction （PCR）. Flow cytometry was designed to observe and detect cellular apoptosis. Results Our findings showed significant increase in MDA levels and decrease in SOD activity in the high glucose group compared with the control group （P 〈0.05）. However, MDA levels were significantly decreased and SOD activity was significantly increased in the adiponectin group compared with those in the high-glucose group （P 〈0.05）. The mRNA expression of HO-1 in the high glucose group was significantly increased in a time-dependent manner compared with that in the control group （P 〈0.05）. Adiponectin further increased the mRNA expression of HO-1 induced by high glucose in a time-dependent manner （P 〈0.05）.The expression of p66Shc was significantly increased in high glucose group compared with that in the control group （P 〈0.05）. Adiponectin significantly suppressed the upregulation of p66Shc induced by high glucose （P 〈0.05）. The apoptotic rate of cardiomyocytes was significantly increased in the high glucose group compared with that in the control group while the apoptotic rate in the adiponectin group was remarkably declined in comparison with that in the high glucose group. Conclusion Adiponectin reduces high glucose-induced oxidative stress and apoptosis and plays a protective role in myocardial cells by upregulating the HO-1 expression and downregulating p66Shc expression.

Blocking effect of Puerarin on calciu m channel in isolated guineapig ventricular myocytes

To identify whether Puerarin (Puer) has blocking effects on L type calcium channel Methods We used whole cell recording of patch clamp techniques to analyse calcium channel current in ventricular myocytes isolated from Langendorff guinea pig hearts by collagenase Results In control group, peak calcium channel current was decreased by 10 6% and 29 8% at 5, 10?min of recording time, respectively For the same period, 0 1?mmol/L Puer reduced calcium current by 23 9% and 72 4% 0 1 and 1?mmol/L Puer inhibited the amplitude of peak Calcium channel current by 40 6% and 63 2% after perfusing for 10?min Compared with control group in which “rundown' phenomenon was observed, Puer also demonstrated its blocking effect on L type channel While washout of Puer with perfusing solution, calcium channel current went down further instead of recovery Current voltage (Ⅰ Ⅴ) curves showed that the levels of calcium channel current were obviously decreased by Puer (both 0 1?mmo/L and 1?mmo/L) form -40?mV to+10?mV Conclusion Puer has blocking effect on L type calcium channel in a concentration dependent manner

Panax quinquefolium saponin Optimizes Energy Homeostasis by Modulating AMPK-Activated Metabolic Pathways in Hypoxia-Reperfusion Induced Cardiomyocytes

Objective To investigate the effects and underlying mechanisms of Panax quinquefolium saponin(PQS)on energy deficiency in hypoxia-reperfusion(H/R)induced cardiomyocytes.Methods The H/R injury involved hypoxia for 3 h and then reperfusion for 2 h.Cardiomyocytes recruited from neonatal rat ventricular myocytes(NRVMs)were randomly divided into control,H/R,H/R+compound C(C.C),H/R+PQS,and H/R+C.C+PQS groups.BrdU assay,lactase dehydrogenase(LDH)leakage and early apoptosis rate were evaluated to assess cell damages.Contents of high energy phosphate compounds were conducted to detect the energy production.Protein expression levels of adenosine monophosphate-activated protein kinase a(AMPKα),glucose transporter 4(GLUT4),phosphate fructose kinase 2(PFK2),fatty acid translocase/cluster of differentiation 36(FAT/CD36),and acetyl CoA carboxylase 2(ACC2)in the regulatory pathways were measured by Western blotting.Immunofluorescence staining of GLUT4 and FAT/CD36 was used to observe the mobilization of metabolic transporters.Results PQS(50 mg/L)pretreatment significantly alleviated H/R-induced inhibition of NRVMs viability,up-regulation of LDH leakage,acceleration of early apoptosis,and reduction of energy production(P<0.05).Compared with the H/R group,up-regulated expression of AMPKα,GLUT4,PFK2,FAT/CD36 and ACC2 were observed,and more GLUT4 and FAT/CD36 expressions were detected on the membrane in the H/R+PQS group(P<0.05).These effects of PQS on H/R-induced NRVMs were eliminated in the H/R+C.C+PQS group(P<0.05).Conclusion PQS has prominent advantages in protecting NRVMs from H/R-induced cell damages and energy metabolic disorders,by activation of AMPKα-mediated GLUT4-PFK2 and FAT/CD36-ACC2 pathways.

Blocking Effect of Salvianolic Acid A on Calcium Channels in Isolated Rat Ventricular Myocytes

Objective： To study the effect of salvianolic acid A （SAA） on L-type calcium current （I-CaL） in isolated ventdcular myocytes of Sprague-Dawley rats. Methods： SPA powder was dissolved in normal Tyrode＇s solution to reach the concentrations of 1, 10, 100, and 1000 μmol/L. The traditional whole-cell patch-clamp recording technique was employed to evaluate the effects of SAA on I-CaL in single ventricular myocytes which were prepared by Langendorff perfusion apparatus from Sprague-Dawley rats. Results： SPA （1, 10, 100, and 1000 μmol/L） inhibited I-CaL peak value by 16.23%± 1.3% （n=6, P〈0.05）, 22.9% ± 3.6% （n=6, P〈0.05）, 53.4% ± 3.0% （n=8, P〈0.01）, and 62.26% ± 2.9% （n=8, P〈0.01）, respectively. SAA reversibly inhibited I-CaL in a dose-dependent manner and with a half-blocking concentration （IC50） of 38.3 μmol/L. SAA at 100 μmol/L elevated the I-V curve obviously, and shifted the half-active voltage （V0.5） from （-15.78± 0.86） mV to （-11.24 ± 0.77） mV （n=6, P〈0.05） and the slope （K） from 5.33 ±0.74 to 4.35±0.74 （n=6, P〉0.05）. However, it did not alter the shapes of I-V curve, steady-state inactivation curve, or recovery from inactivation curve. Conclusions： SAA inhibited I-CaL in a dose-dependent manner. It shifted the steady-state activation curve to a more positive voltage, which indicated that the drug affected the activated state of calcium channels, and suggested that the Ca2. antagonistic effect of SPA be beneficial in the treatment of myocardial ischemia reperfusion injury.