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...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.展开更多
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-BTH...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.展开更多
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 stan...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.展开更多
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 patc...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.展开更多
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 myo...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.展开更多
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 si...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.展开更多
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 fro...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展开更多
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 ...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.展开更多
OBJECTIVE:To investigate the efficacy of water fraction from Dioscorea cirrhosa(WF)on oxidative damage and apoptosis of cardiomyocytes induced by H2O2,and to study its mechanism.METHODS:Cell viability was measured by ...OBJECTIVE:To investigate the efficacy of water fraction from Dioscorea cirrhosa(WF)on oxidative damage and apoptosis of cardiomyocytes induced by H2O2,and to study its mechanism.METHODS:Cell viability was measured by the MST assay kit.The content of malondialdehyde(MDA),release of lactate dehydrogenase(LDH)and activity of catalase(CAT)and superoxide dismutase(SOD)were detected by biochemical kit.The content of reactive oxygen species(ROS)was assessed by nonfluorescent probe 2′,7′-dichlorofluorescin diacetate(DCFH-DA).JC-1 was used to analyze the mitochondrial membrane potential(mtΔΨ)and Annexin-V-FITC/PI staining was applied to assess apoptosis of H9c2 by flow cytometry.Moreover,the expression of B-cell lymphoma-2(Bcl-2),Bcl-2-associated X(Bax),caspase-3,caspase-9,cleaved-caspase-3 and cleaved-caspase-9 proteins was determined by western blot analysis.RESULTS:WF increased cell viability and decreased LDH leakage in H9c2 cells exposed to H2O2.WF treatment decreased ROS and MDA level,enhanced SOD and CAT activities,improved mtΔΨand inhibited apoptosis.Western blot analysis demonstrated that the ratio of Bcl-2/Bax was increased and the expression cleaved-caspase-3,caspase-3,cleaved-caspase-9 and caspase-9 were decreased in group treated with WF.CONCLUSION:WF protects H9c2 myocardial cells on H2O2-induced oxidative stress and apoptosis by scavenging ROS,improving antioxidant capacity,protecting mitochondrial and regulating the proteins expression related to apoptosis.展开更多
Ca2+ sparks are the elementary events of intracellular Ca2+ release from the sarcoplasmic reticulum in cardiac myocytes. In order to investigate whether spontaneous L-type Ca2+ channel activation contributes to the ge...Ca2+ sparks are the elementary events of intracellular Ca2+ release from the sarcoplasmic reticulum in cardiac myocytes. In order to investigate whether spontaneous L-type Ca2+ channel activation contributes to the genesis of spontaneous Ca2+ sparks, we used confocal laser scanning microscopy and fluo-4 to visualize local Ca2+ sparks in intact rat ventricular myocytes. In the presence of 0.2 mmol/L CdCl2 which inhibits spontaneous L-type Ca2+ channel activation, the rate of occurrence of spontaneous Ca2+ sparks was halved from 4.20 to 2.04 events/(100 μm·s), with temporal and spatial properties of individual Ca2+ sparks unchanged. Analysis of the Cd2+-sensitive spark production revealed an open probability of ~10-5 for L-type channels at the rest membrane potentials (-80 mV). Thus, infrequent and stochastic openings of sarcolemmal L-type Ca2+ channels in resting heart cells contribute significantly to the production of spontaneous Ca2+ sparks.展开更多
Background Programmed cell death has previously been observed in animal models of viral myocarditis and in patients with end stage dilated cardio myopathy (DCM). The purpose of the present study was to investigate if ...Background Programmed cell death has previously been observed in animal models of viral myocarditis and in patients with end stage dilated cardio myopathy (DCM). The purpose of the present study was to investigate if apoptosis is also detectable in endomyocardial biopsies of patients with clinical and immuno histological signs of myocarditis. Methods Right ventricular myocardial biopsies were obtained from 53 patients with clinically suspected myocarditis (left ventricular ejection fraction 30±14%). Cardiac tissues from 4 patients with end stage heart failure were obtained during heart transplantation. Diagnosis of myocarditis and DCM was based on clinical presentation and on histological and immunohistological findings. The presence of apoptosis was investigated in paraffin embedded tissue sections by in situ end labeling of nuclear DNA strand breaks. Results By histological and immunohistological analysis 19 patients (36%) were diagnosed as myocarditis and 34 patients (64%) as DCM. Apoptosis was detected in 9 out of 19 patients with myocarditis, in 12 out of 34 patients with DCM (47% versus 35%, p=NS), and in all patients with end stage heart failure. Conclusions A high frequency of apoptosis was detectable in endomyocardial biopsies of patients with immunohistologically documented myocarditis. The rate of apoptosis in patients with myocarditis, however, was not different to patients with DCM. These findings suggest that apoptosis may be one common pathway in the pathogenesis of cardiac contractile dysfunction in patients with myocarditis and DCM.展开更多
We have studied transmembrane La3+ movement in rat ventricular myocytes for the first time by using the whole-cell patch-clamp recording mode. La3+ (0.01-5.0 mmol/L) could not bring out inward currents through the L-t...We have studied transmembrane La3+ movement in rat ventricular myocytes for the first time by using the whole-cell patch-clamp recording mode. La3+ (0.01-5.0 mmol/L) could not bring out inward currents through the L-type calcium channel in rat ventricular myocytes, while it could enter the cells by the same way carried by 1μmo1/L ionomycin. When the outward Na+ concentration gradient is formed, La3+ can enter the cells via Na-Ca exchange, and the exchange currents increase with the increase of external La3+ concentrations. But compared with Na-Ca exchange currents in the same concentration, the former is only 14%-38% of the latter. The patch-clamp experiment indicates that La3+ normally can not enter ventricular myocytes through L-type calcium channel, but it can enter the cells via Na-Ca exchange.展开更多
The temperature-dependence of infraeeliular free caleimn (Ca) was investigated in mdo-1 loaded ventricular myocytes from the ral, a non-hibernator, and from the ground squirrel, a hibernator. The dissociation constant...The temperature-dependence of infraeeliular free caleimn (Ca) was investigated in mdo-1 loaded ventricular myocytes from the ral, a non-hibernator, and from the ground squirrel, a hibernator. The dissociation constant of indo-l at different temperatures was calibrated both al pll-tat and at @-stat . and the result demonstrated that the @-stat ralibration should be prettrred . Analysis of the fluoreseent image showed a striking increase of Ca2 as well as spontaneous caleiuni waves in ral cells, indicating an overloaded cakuum. In contrast, cardiac myocytes of the ground sqnirraf were found to keep a constant (Ca2+) without caleium overload regardless of temperature variation. It is be-lieved that understanding of the mechanisms underlying the interccllular caleima homeostasis of hibrernators may lead to solutions of some medical questions .展开更多
The properties of the inward-rectifying potassium current (IK1) were studied in the single myocytes isolated from adult mouse ventricles by the whole-cell patch-damp technique for the first time. Most of the propertie...The properties of the inward-rectifying potassium current (IK1) were studied in the single myocytes isolated from adult mouse ventricles by the whole-cell patch-damp technique for the first time. Most of the properties of IK1 including channel conductances, activation, inactivation, rectification and external K+ sensitivity in mouse ventricular myocyte were similar to those in other species, but the current-voltage (1-V) curve of mouse ventricular myocyte showed no negative slope, i.e the slope in the range of membrane potential 50 mV positive to the reversal potential (VRev) was virtually flat and remained at a low current level ((59±39) pA). Under the superfusion of Tyrode’s solution with 3mmol/L K+ and 3mmol/L Cs+, IK1 in the above region nearly decreased to zero, and then the early after-depolarization (EAD) occurred. The results suggest that this distinctive characteristic of IK1 in mouse ventricular myocyte may relate to the high susceptibility to EA0 in mouse myocardium. The inhibition of IK1展开更多
The mechanism of reperfusion injury of ischemic myocardium is not clear yet, and there has not been effective therapeutic methods up to now. Recently, growing evidence shows that the myocardial ischemia-reperfusion in...The mechanism of reperfusion injury of ischemic myocardium is not clear yet, and there has not been effective therapeutic methods up to now. Recently, growing evidence shows that the myocardial ischemia-reperfusion injury is similar in pathologic character and展开更多
AIM: To investigate the effect of gingerol on colonic motility and the action of L-type calcium channel currents in this process.METHODS: The distal colon was cut along the mesenteric border and cleaned with Ca^(2+)-f...AIM: To investigate the effect of gingerol on colonic motility and the action of L-type calcium channel currents in this process.METHODS: The distal colon was cut along the mesenteric border and cleaned with Ca^(2+)-free physiological saline solution. Muscle strips were removed and placed in Ca^(2+)-free physiological saline solution, which was oxygenated continuously. Longitudinal smooth muscle samples were prepared by cutting along the muscle strips and were then placed in a chamber. Mechanical contractile activities of isolated colonic segments in rats were recorded by a 4-channel physiograph. Colon smooth muscle cells were dissociated by enzymatic digestion. L-type calcium currents were recorded using the conventional whole-cell patch-clamp technique.RESULTS: Gingerol inhibited the spontaneous contraction of colonic longitudinal smooth muscle in a dose-dependent manner with inhibition percentages of 13.3% ± 4.1%, 43.4% ± 3.9%, 78.2% ± 3.6% and 80.5% ± 4.5% at 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L, respectively(P < 0.01). Nifedipine, an L-type calcium channel blocker, diminished the inhibition of colonic motility by gingerol. Gingerol inhibited L-type calcium channel currents in colonic longitudinal myocytes of rats. At a 75 μmol/L concentration of gingerol, the percentage of gingerolinduced inhibition was diminished by nifedipine from 77.1% ± 4.2% to 42.6% ± 3.6%(P < 0.01). Gingerol suppressed IBa in a dose-dependent manner, and the inhibition rates were 22.7% ± 2.38%, 35.77% ± 3.14%, 49.78% ± 3.48% and 53.78% ± 4.16% of control at 0 m V, respectively, at concentrations of 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L(P < 0.01). The steady-state activation curve was shifted to the right by treatment with gingerol. The value of half activation was-14.23 ± 1.12 m V in the control group and-10.56 ± 1.04 m V in the 75 μmol/L group(P < 0.05) with slope factors, Ks, of 7.16 ± 0.84 and 7.02 ± 0.93(P < 0.05) in the control and 75 μmol/L groups, respectively. However, the steady-state inactivation curve was not changed, with a half-inactivation voltage, 0.5 V, of-27.43 ± 1.26 m V in the control group and-26.56 ± 1.53 m V in the 75 μmol/L gingerol group(P > 0.05), and a slope factor, K, of 13.24 ± 1.62 in the control group and 13.45 ± 1.68(P > 0.05) in the 75 μmol/L gingerol group.CONCLUSION: Gingerol inhibits colonic motility by preventing Ca^(2+) influx through L-type calcium channels.展开更多
The current study was to investigate mRNA expression of cyclin D 2 and p16 during the transition from cardiac myocyte hyperplasia to hypertrophy. Cultured cardiac myocytes (CM) and fibroblasts (FC) obtained from 1 day...The current study was to investigate mRNA expression of cyclin D 2 and p16 during the transition from cardiac myocyte hyperplasia to hypertrophy. Cultured cardiac myocytes (CM) and fibroblasts (FC) obtained from 1 day old Sparague Dawley rats were used in this study. We have determined (1) hyperplasia by cell growth curve and fluorescence activated cell sorting (FACS); and (2) ultrastructure by electron microscope observation; and (3) expressions of cyclin D 2 mRNA and p16 mRNA by using in situ hybridization and image analysis. The results were shown (1) Results of cell growth curve and FACS analysis showed CM could proliferate in the first 3 cultured days (4 days in postnatal development). But the ability decreased quickly, concomitant with the differentiation. (2) The ultrastructure of CM showed the large amount of myofilaments and mitochondrion and FC showed moderate amount of rough endoplasmic reticulum. (3) The expression of cyclin D 2 mRNA in 3 , 4 , 5 day CM group was 0.89 times(p<0.05), 0.80 times (p<0.05)and 0.56 times (p<0.01)of that in 1 day group respectively. P16 mRNA in 2 , 3 , 4 , 5 day CM group were 1.63 times(p<0.01),1.72 times(p<0.01),1.99 times (p<0.01)and 2.84 times (p<0.01) of that in 1 day group respectively. It can be concluded that cultured neonatal rat cardiac myocytes could proliferate during the first 3 cultured days, but the ability of proliferation decreased, from the fourth day, concomitant with differentiation. Cyclin D 2 and p16 have the key roles during the transition from myocyte hyperplasia to hypertrophy.展开更多
基金supported by a grant from the Health Department Foundation of Zhejiang Province(2010KYA102)
文摘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.
文摘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.
基金This research was supported by the National Natural Science Foundation of China (No. 39870334No. 39970273).
文摘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.
文摘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.
文摘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.
基金supported by National Plan of High-Tech Research and Development(863-2-2-2-4).
文摘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.
文摘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
基金Supported by the National Natural Science Foundation of China(No.81273934 and No.81874410)。
文摘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.
基金Supported by National Natural Science Foundation of China Basic Research on Ethnopharmacology of Guizhou Karst Area(No.U1812403)Study on the Active Components of Anti-oxidative Stress Activities of Shengxiong Glucose Injection Based on Pi3k/akt Pathway(No.81760699)+8 种基金Investigation of The Active Components and Mechanism of Polygonum Orientale Ameliorating Myocardial Ischemic Injury Based on"Components-Network Targets-Effects"(No.81803700)Central Guided Local Science and Technology Special Project Capability Enhancement of Guizhou Provincial Key Laboratory of Pharmaceutics(No.20184006)Guizhou Provincial Natural Science Foundation Effects of Gerbera Piloselloides on Airway Inflammation in Bronchial Asthma and Its Medicinal Substance Basis(20181128)Study on the Molecular Mechanism and Active Ingredients of Polygonum Orientale in Treating Myocardial Ischemic Disease Based on Systemic Pharmacology Strategy(No.20201z068)Guizhou Provincial People’s Pharmacy Effective Material Basic Research Science and Technology Innovation Talent Team(No.20165613)Cultivation of High-level Innovative Talents in Guizhou Province(Hundred Levels)(No.20165677)Study on the Effective Component Group and Its Mechanism of Anti-myocardial Ischemia Injury of Polygonum Orientale Based on"Component-network Target"(No.20175718)Study on the Effective Material Basis of Shengxiong Glucose Injection for Its Anti-oxidative Stress Activities Mediated by Pi3k/akt(No.20191280)Doctoral Start Fund of Guizhou Medical University Study on the Effective Material Basis of Gerbera Piloselloides Against Bronchial Asthma Based on In Vivo Processes(No.YJ2017-12)。
文摘OBJECTIVE:To investigate the efficacy of water fraction from Dioscorea cirrhosa(WF)on oxidative damage and apoptosis of cardiomyocytes induced by H2O2,and to study its mechanism.METHODS:Cell viability was measured by the MST assay kit.The content of malondialdehyde(MDA),release of lactate dehydrogenase(LDH)and activity of catalase(CAT)and superoxide dismutase(SOD)were detected by biochemical kit.The content of reactive oxygen species(ROS)was assessed by nonfluorescent probe 2′,7′-dichlorofluorescin diacetate(DCFH-DA).JC-1 was used to analyze the mitochondrial membrane potential(mtΔΨ)and Annexin-V-FITC/PI staining was applied to assess apoptosis of H9c2 by flow cytometry.Moreover,the expression of B-cell lymphoma-2(Bcl-2),Bcl-2-associated X(Bax),caspase-3,caspase-9,cleaved-caspase-3 and cleaved-caspase-9 proteins was determined by western blot analysis.RESULTS:WF increased cell viability and decreased LDH leakage in H9c2 cells exposed to H2O2.WF treatment decreased ROS and MDA level,enhanced SOD and CAT activities,improved mtΔΨand inhibited apoptosis.Western blot analysis demonstrated that the ratio of Bcl-2/Bax was increased and the expression cleaved-caspase-3,caspase-3,cleaved-caspase-9 and caspase-9 were decreased in group treated with WF.CONCLUSION:WF protects H9c2 myocardial cells on H2O2-induced oxidative stress and apoptosis by scavenging ROS,improving antioxidant capacity,protecting mitochondrial and regulating the proteins expression related to apoptosis.
基金supported by the National Natural Science Foundation of China(Grant No.30200088 for A-type grant)Chinese Major State Basic Research Development Program(Grant No.G2000057002).
文摘Ca2+ sparks are the elementary events of intracellular Ca2+ release from the sarcoplasmic reticulum in cardiac myocytes. In order to investigate whether spontaneous L-type Ca2+ channel activation contributes to the genesis of spontaneous Ca2+ sparks, we used confocal laser scanning microscopy and fluo-4 to visualize local Ca2+ sparks in intact rat ventricular myocytes. In the presence of 0.2 mmol/L CdCl2 which inhibits spontaneous L-type Ca2+ channel activation, the rate of occurrence of spontaneous Ca2+ sparks was halved from 4.20 to 2.04 events/(100 μm·s), with temporal and spatial properties of individual Ca2+ sparks unchanged. Analysis of the Cd2+-sensitive spark production revealed an open probability of ~10-5 for L-type channels at the rest membrane potentials (-80 mV). Thus, infrequent and stochastic openings of sarcolemmal L-type Ca2+ channels in resting heart cells contribute significantly to the production of spontaneous Ca2+ sparks.
文摘Background Programmed cell death has previously been observed in animal models of viral myocarditis and in patients with end stage dilated cardio myopathy (DCM). The purpose of the present study was to investigate if apoptosis is also detectable in endomyocardial biopsies of patients with clinical and immuno histological signs of myocarditis. Methods Right ventricular myocardial biopsies were obtained from 53 patients with clinically suspected myocarditis (left ventricular ejection fraction 30±14%). Cardiac tissues from 4 patients with end stage heart failure were obtained during heart transplantation. Diagnosis of myocarditis and DCM was based on clinical presentation and on histological and immunohistological findings. The presence of apoptosis was investigated in paraffin embedded tissue sections by in situ end labeling of nuclear DNA strand breaks. Results By histological and immunohistological analysis 19 patients (36%) were diagnosed as myocarditis and 34 patients (64%) as DCM. Apoptosis was detected in 9 out of 19 patients with myocarditis, in 12 out of 34 patients with DCM (47% versus 35%, p=NS), and in all patients with end stage heart failure. Conclusions A high frequency of apoptosis was detectable in endomyocardial biopsies of patients with immunohistologically documented myocarditis. The rate of apoptosis in patients with myocarditis, however, was not different to patients with DCM. These findings suggest that apoptosis may be one common pathway in the pathogenesis of cardiac contractile dysfunction in patients with myocarditis and DCM.
基金This work was supported by the National Natural Science Foundation of China (Grant No. 29890280).
文摘We have studied transmembrane La3+ movement in rat ventricular myocytes for the first time by using the whole-cell patch-clamp recording mode. La3+ (0.01-5.0 mmol/L) could not bring out inward currents through the L-type calcium channel in rat ventricular myocytes, while it could enter the cells by the same way carried by 1μmo1/L ionomycin. When the outward Na+ concentration gradient is formed, La3+ can enter the cells via Na-Ca exchange, and the exchange currents increase with the increase of external La3+ concentrations. But compared with Na-Ca exchange currents in the same concentration, the former is only 14%-38% of the latter. The patch-clamp experiment indicates that La3+ normally can not enter ventricular myocytes through L-type calcium channel, but it can enter the cells via Na-Ca exchange.
基金Project supported by the National Natural Science Foundation of China(Grant No. 39670276).
文摘The temperature-dependence of infraeeliular free caleimn (Ca) was investigated in mdo-1 loaded ventricular myocytes from the ral, a non-hibernator, and from the ground squirrel, a hibernator. The dissociation constant of indo-l at different temperatures was calibrated both al pll-tat and at @-stat . and the result demonstrated that the @-stat ralibration should be prettrred . Analysis of the fluoreseent image showed a striking increase of Ca2 as well as spontaneous caleiuni waves in ral cells, indicating an overloaded cakuum. In contrast, cardiac myocytes of the ground sqnirraf were found to keep a constant (Ca2+) without caleium overload regardless of temperature variation. It is be-lieved that understanding of the mechanisms underlying the interccllular caleima homeostasis of hibrernators may lead to solutions of some medical questions .
文摘The properties of the inward-rectifying potassium current (IK1) were studied in the single myocytes isolated from adult mouse ventricles by the whole-cell patch-damp technique for the first time. Most of the properties of IK1 including channel conductances, activation, inactivation, rectification and external K+ sensitivity in mouse ventricular myocyte were similar to those in other species, but the current-voltage (1-V) curve of mouse ventricular myocyte showed no negative slope, i.e the slope in the range of membrane potential 50 mV positive to the reversal potential (VRev) was virtually flat and remained at a low current level ((59±39) pA). Under the superfusion of Tyrode’s solution with 3mmol/L K+ and 3mmol/L Cs+, IK1 in the above region nearly decreased to zero, and then the early after-depolarization (EAD) occurred. The results suggest that this distinctive characteristic of IK1 in mouse ventricular myocyte may relate to the high susceptibility to EA0 in mouse myocardium. The inhibition of IK1
文摘The mechanism of reperfusion injury of ischemic myocardium is not clear yet, and there has not been effective therapeutic methods up to now. Recently, growing evidence shows that the myocardial ischemia-reperfusion injury is similar in pathologic character and
基金Supported by National Basic Research Program of China(973 Program)No.2013CB531703+3 种基金National Natural Science Foundation of ChinaNo.81273919Natural Science Foundation of Liaoning ProvinceNo.2012225020 and No.2013023002
文摘AIM: To investigate the effect of gingerol on colonic motility and the action of L-type calcium channel currents in this process.METHODS: The distal colon was cut along the mesenteric border and cleaned with Ca^(2+)-free physiological saline solution. Muscle strips were removed and placed in Ca^(2+)-free physiological saline solution, which was oxygenated continuously. Longitudinal smooth muscle samples were prepared by cutting along the muscle strips and were then placed in a chamber. Mechanical contractile activities of isolated colonic segments in rats were recorded by a 4-channel physiograph. Colon smooth muscle cells were dissociated by enzymatic digestion. L-type calcium currents were recorded using the conventional whole-cell patch-clamp technique.RESULTS: Gingerol inhibited the spontaneous contraction of colonic longitudinal smooth muscle in a dose-dependent manner with inhibition percentages of 13.3% ± 4.1%, 43.4% ± 3.9%, 78.2% ± 3.6% and 80.5% ± 4.5% at 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L, respectively(P < 0.01). Nifedipine, an L-type calcium channel blocker, diminished the inhibition of colonic motility by gingerol. Gingerol inhibited L-type calcium channel currents in colonic longitudinal myocytes of rats. At a 75 μmol/L concentration of gingerol, the percentage of gingerolinduced inhibition was diminished by nifedipine from 77.1% ± 4.2% to 42.6% ± 3.6%(P < 0.01). Gingerol suppressed IBa in a dose-dependent manner, and the inhibition rates were 22.7% ± 2.38%, 35.77% ± 3.14%, 49.78% ± 3.48% and 53.78% ± 4.16% of control at 0 m V, respectively, at concentrations of 25 μmol/L, 50 μmol/L, 75 μmol/L and 100 μmol/L(P < 0.01). The steady-state activation curve was shifted to the right by treatment with gingerol. The value of half activation was-14.23 ± 1.12 m V in the control group and-10.56 ± 1.04 m V in the 75 μmol/L group(P < 0.05) with slope factors, Ks, of 7.16 ± 0.84 and 7.02 ± 0.93(P < 0.05) in the control and 75 μmol/L groups, respectively. However, the steady-state inactivation curve was not changed, with a half-inactivation voltage, 0.5 V, of-27.43 ± 1.26 m V in the control group and-26.56 ± 1.53 m V in the 75 μmol/L gingerol group(P > 0.05), and a slope factor, K, of 13.24 ± 1.62 in the control group and 13.45 ± 1.68(P > 0.05) in the 75 μmol/L gingerol group.CONCLUSION: Gingerol inhibits colonic motility by preventing Ca^(2+) influx through L-type calcium channels.
文摘The current study was to investigate mRNA expression of cyclin D 2 and p16 during the transition from cardiac myocyte hyperplasia to hypertrophy. Cultured cardiac myocytes (CM) and fibroblasts (FC) obtained from 1 day old Sparague Dawley rats were used in this study. We have determined (1) hyperplasia by cell growth curve and fluorescence activated cell sorting (FACS); and (2) ultrastructure by electron microscope observation; and (3) expressions of cyclin D 2 mRNA and p16 mRNA by using in situ hybridization and image analysis. The results were shown (1) Results of cell growth curve and FACS analysis showed CM could proliferate in the first 3 cultured days (4 days in postnatal development). But the ability decreased quickly, concomitant with the differentiation. (2) The ultrastructure of CM showed the large amount of myofilaments and mitochondrion and FC showed moderate amount of rough endoplasmic reticulum. (3) The expression of cyclin D 2 mRNA in 3 , 4 , 5 day CM group was 0.89 times(p<0.05), 0.80 times (p<0.05)and 0.56 times (p<0.01)of that in 1 day group respectively. P16 mRNA in 2 , 3 , 4 , 5 day CM group were 1.63 times(p<0.01),1.72 times(p<0.01),1.99 times (p<0.01)and 2.84 times (p<0.01) of that in 1 day group respectively. It can be concluded that cultured neonatal rat cardiac myocytes could proliferate during the first 3 cultured days, but the ability of proliferation decreased, from the fourth day, concomitant with differentiation. Cyclin D 2 and p16 have the key roles during the transition from myocyte hyperplasia to hypertrophy.