Effect of phosphocreatine on transient outward potassium current in ischemic ventricular mid-myocardial cells of rats

AIM: To determine the effect of exogenous phosphocreatine (PCr) at different concentrations on transient outward potassium (I_to) current in rat ischemic ventricular mid-myocardial (M) cells and to explore the antiarrhythmia mechanism in the treatment of ischemic heart disease. METHODS: M cells were isolated enzymatically from left ventricular mid-myocardium of rats. Peak I_to current was recorded by patch-clamp technique in the whole-cell configuration when M cells were superfused with normal Tyrode solution, simple ischemic solution,and simulated ischemic solution containing PCr at concentrations of 5, 10, 20 and 30 mmol/L for 10 min. RESULTS: Peak I_to current density of M cells superfused with simple simulated ischemic solution was significantly reduced by (76.1?6.3)% (P<0.05) compared with M cells superfused with Tyrode solution. Ischemic solution containing 5, 10, 20 and 30 mmol/L PCr reduced peak I_to current density by (57.1?9.6)% (P<0.05), (40.3?10.3)% (P<0.05), (34.3?9.6)% (P<0.05) and (32.1?10.6)% (P<0.05),respectively. There was statistical difference among ischemic solution without PCr and containing PCr at concentrations of 5 and 10 mmol/L groups (P<0.05). No statistical difference among groups of 10, 20 and 30 mmol/L PCr was observed (P>0.05). CONCLUSION: PCr reverses the inhibition of I_to current under ischemic condition in M cells, which may be the mechanism responsible for arrhythmia prevention in ischemic heart disease. PCr at concentrations of 0~10 mmol/L exerts significant dose-effect relationship.     

Alteration of Na+ currents in ventricular myocytes from 1-week infarcted rabbit heart

AIM:To study the current density and function of Na⁺ channel in cells from the epicardial border zone of the 1-week infarcted rabbit heart.METHODS:Rabbits were infarcted by ligation of the left anterior descending coronary artery.1 week later, INa was recorded using whole cel patch-clamp techniques in ventricular my ocytes from infarcted heart(IZs)and compared with the INa from noninfarcted heart(NZs).RESULTS:Peak I_Na current density(at -30 mV) was significantly reduced in IZs(22.48±4.62 PA/PF, n=14) compared with NZs(45.50±5.33 PA/PF, n=12), P＜0.01;V_0.5 of the availability curve(I/Imax curve)was shifted significantly in the hyperpolarizing direction in IZs(-89.1±5.6 mV, n=6) compared with NZs (-76.2±5.3 mV, n=5), P＜0.05.Recovery of I_Na from inactivation was significantly slower in IZs.CONCLUSIONS:I_Nais reduced, and its kinetics are altered in IZs.These changes may underlie the altered excitability and postrepolarization refractoriness of ventricular fibers of the IZs, thus contributing to reentrant arrhymias in the infarcted heart.     

Effect of hypoxia on persistent sodium current in ventricular myocytes from 3-week infarcted rat heart

AIM: To investigate the effect of hypoxia on persistent sodium current (INap) in single ventricular myocyte isolated from acute myocardial infarction (AMI) heart of rats and to study the mechanisms of cardiac arrhythmias that occur after AMI. METHODS: AMI model was induced by ligating the left anterior descending coronary artery in rats. The whole-cell patch clamp technique was used to record the current in epicardial myocytes in infarcted region from rats at 3 week after AMI. RESULTS: In normoxic conditions, the current density of INap in cardiomyocytes of fake operation (FO) and AMI hearts was 0.144±0.022 pA/pF (n=9), 0.121±0.013 pA/pF (n=9，P<0.01)， respectively, which was blocked by tetrodotoxin (TTX). The amplitude of INap was gradually increased with the prolongation of hypoxia time, but the increase in extent of INap in FO cells was significant bigger than that in AMI cells. The INap was blocked by 1 mmol/L glutathione. CONCLUSIONS: After AMI, the amplitude of INaP in infarcted and noninfarcted myocardium showed differences both in normoxic and hypoxic conditions, which increased dispersion of repolarization. This may be one of the reasons of reentrant ventricular arrhythmias that occur after AMI.     

Effects of stretch on transient outward potassium and inward rectifier potassium current in cultured neonatal rat atrial myocytes

AIM：To investigate the effects of mechanical stretch on transient outward potassium current (Ito), inward rectifier potassium current (IK1) and action potential duration(APD) of cultured neonatal rat atrial myocytes. METHODS：Neonatal rat atrial myocytes were isolated and cultured on silicone sheeting with or without stretch for 24 h. The silicone membrane area was increased by 12% in stretched group. The cells without stretch served as control. Ito, IK1 and APD were recorded by the technique of whole-cell patch clamp. RESULTS：Compared with control group, Ito density in stretched myocytes was significantly reduced ［(16±04) pA/pF vs (121±29) pA/pF, P<001］, whereas IK1 density was increased ［(-108±08) pA/pF vs (-88±09) pA/pF, P<001］. The APDs at 50% and 90% levels of repolarization (APD50 and APD90) in the stretched cells were obviously decreased than those in non-stretched cells ［(105±14) ms vs (155±24) ms, (300±28) ms vs (563±36) ms, P<001］. CONCLUSION：Stretch stimulation leads to the reduction of Ito density, the increase in IK1 density and the shortness of APD in cultured rat atrial neonatal myocytes, which may contribute to atrial electrical remodeling induced by pressure overload.     

Characteristics of transient outward potassium current in repolarization 1 phase from the canine right ventricular mid-myocardial cells

AIM: To examine the electrophysiological characteristics of transient outward potassium current(Ito₁) in repolarization 1 phase from the canine right ventricular M cells. METHODS: By use of whole cell patch-clamp technique, we quantitatively researched the ionic intensity, density of Ito₁ and the notch magnitude of action potential in repolarization 1 phase. RESULTS: (1) The activating process of Ito₁ of canine right ventricular M cells presented evident voltage-dependency. Under the condition of 37℃, 5 000 ms, 0 mV and +70 mV, the average peak Ito₁ intensity of right ventricular M cell were (690±380) pA and (3 130±1 910) pA, respectively (P＜0.01). (2) The Ito₁ intensity of canine right ventricular M cell possessed obvious frequent dependency. Under the condition of 37℃,+70 mV, 500 ms and 5 000 ms, the average peak Ito₁ intensity were (1 690±830) pA,(3 130±1 910) pA, respectively(P＜0.01), corresponding to the increase of action potential "spike-dome" magnitude in repolarization 1 phase. CONCLUSION: Potent Ito₁ as well as the "spike-dome"-like action potential figure mediated by Ito₁ is one of the prominent electrophysiological characteristics of the canine right ventricular M cells.     

新书推荐：《分子克隆实验指南》（第三版）

AIM: To study the effect of panax notoginseng sponins (PNS) on L-type Ca²⁺ current in isolated right ventricle myocytes from chronic hypoxic rats. METHODS: Using whole cell patch clamp recording technique,we measured I_Ca-L in isolated right ventricle myocytes which were divided into three group:control group, chronic hy-poxic group and chronic hypoxic group with PNS(100 mg·kg^-1·d^-1). RESULTS: The result showed I_Ca-L of cells from chronic hypoxic group were significantly larger than the other two groups(P＜0.05). CONCLUSION: PNS decreases L-type Ca²⁺ current of the right ventricle myocytes from chronic hypoxic rats.     

Effects of simvastatin on transient outward potassium current in ventricular myocytes of normocholesterolemic rabbits suffering from ischemia and reperfusion

AIM: To investigate the effects of simvastatin on transient outward potassium current (Ito) in left ventricular myocytes of rabbit heart undergoing ischemia-reperfusion, so as to explore the cellular (ionic) mechanism of statin treatment for antiarrhythmia. METHODS: Forty-five rabbits were randomly divided into three groups: ischemic-reperfusion group (I-R), simvastatin intervention group (statin) and sham-operation control group (sham). Anesthetized rabbits were subjected to 30 min ischemia by ligation of the left anterior descending coronary artery and 60 min reperfusion after oral administration of simvastatin at dose of 5 mg·kg-1·d-1(statin group) or placebo (I-R group) for 3 d. Single ventricular myocytes were isolated enzymatically from the epicardial zone of the infracted region derived from the hearts in I-R, statin group and the same anatomy region in sham. Whole cell patch clamp technique was used to record Ito. Simultaneously, the level of serum cholesterol was examined. RESULTS: No significant difference in serum cholesterol concentration among three groups was observed. The Ito current density (at +60 mV) was significantly decreased in I-R ［(9.49 ±1.91) pA/pF, n=11］ compared with sham ［(17.41± 3.13) pA/pF, n=15, P＜0.01］ and statin ［(15.24 ± 2.41) pA/pF, n=11, P＜0.01］, although there was slight reduction in statin group compared with sham (P＜0.05). CONCLUSION: Ischemia-reperfusion induces significant down-regulation of Ito, which may underlie the altered electrical activity and prolong abnormal transmembrane action potential duration of the surviving ventricular myocytes. Pretreatment with simvastatin attenuates these changes without lowering the serum cholesterol level, suggesting that simvastatin may reverse this electrical remodeling, thus contributing to the ionic mechanism of statin treatment for antiarrhythmia.     

Effects of piperine on abnormalities of inward rectifier potassium current and ultra rapid delayed rectifier potassium current induced by hydrogen peroxide in rabbit atrial myocytes

AIM: To study the protective effect of piperine on abnormalityies of inward rectifier potassium current (I_K1) and ultra rapid delayed rectifier potassium current (I_KUr) induced by hydrogen peroxide (H₂O₂) in single rabbit atrial myocytes. METHODS: The technique of whole-cell patch-clamp was used to study the effect of H₂O₂ at concentration of 50 μmol/L on I_K1 and I_KUr in single rabbit atrial myocytes. The protective effect of pretreatment with piperine (7 μmol/L) was also observed. RESULTS: The piperine at concentration of 7 μmol/L had no significant effect on I_K1 and I_KUr and their channel dynamics. In the presence of H₂O₂ at concentration of 50 μmol/L, the peak currents of I_K1 and I_KUr reduced significantly (P<0.05).The steady-state activation curve of I_KUr was shifted right, the steady-state inactivation curve of I_KUr was shifted left, and the recovery from inactivation of I_KUr was shifted downward. The I_KUr showed frequency-dependent characteristics. Piperine at concentration of 7 μmol/L significantly alleviated the inhibitory effect of H₂O₂ on I_K1 and I_KUr (P<0.01). In addition, piperine protected against the changes of I_KUr dynamics induced by H₂O₂. CONCLUSION: Piperine alleviates the abnormalities of I_K1 and I_KUr induced by oxidative stress in atrial myocytes.     

Effects of cardiomyopeptidin on sodium current in ventricular myocytes of guinea pigs

AIM: To determine the effect of cardiomyopeptidin on sodium current (I_Na) in ventricular myocytes of guinea pigs and to explore the mechanism of cardiomyopeptidin action at the ionic channel level. METHODS: Single ventricular myocytes of guinea pigs were obtained by enzymatic dissociation method. The whole-cell patch-clamp recording technique was used to record the change of I_Na. RESULTS: Cardiomyopeptidin (1, 5, 10, 50, 100 and 500 mg/L) decreased I_Na in a dose-dependent manner. The inhibition rates were (0±1)%, (6±2)%, (10±2)%, (15±1)%, (22±9)% and (30±6)%, respectively. The time to peak (TTP) was delayed from (2.8±0.7) ms to (3.0±0.8) ms (P<0.05) by cardiomyopeptidin (50 mg/L). In the presence of cardiomyopeptidin (50 mg/L), the current density-voltage curve of I_Na was shifted and without change of its active potential, peak potential, reversal potential, and the shape of the curve. The steady activation curve, the steady inactivation curve and the steady inactivation recovery curve of I_Na were not affected. CONCLUSION: Cardiomyopeptidin inhibits the I_Na in guinea pig ventricular myocytes, which may be one of the mechanisms of its antiarrhythmic effect.     

Effect of hypercholesterolemia on the ionic currents in cardiac ventricular myocytes of rats

AIM: To determine whether chronic hypercholesterolemia affects ionic currents on cardiac ventricular myocytes of rats. METHODS: Whole-cell patch-clamp technique was used to record the ionic currents in single cardiac myocytes isolated from normal cholesterolemia and hypercholesterolemia rats. RESULTS: In the hypercholesterol group (group Ⅱ), serum total-cholesterol level was significantly higher than that of normal group (group Ⅰ) ［(3.10±0.62)mmol·L-1 vs (1.18±0.37)mmol·L-1, P<0.01, n=20］. The serum triglyceride content of group II was remarkably higher than that of group Ⅰ ［(1.51±0.30)mmol·L-1 vs (0.43±0.15)mmol·L-1, P<0.01, n=20］. In ventricular myocytes of rats, 50% repolarization of action potential duration (APD50) prolonged from (70.86±8.12)ms (group Ⅰ) to (116.16±6.90)ms (group Ⅱ) (n=10 in each group, P<0.01); APD90 prolonged from (95.10±7.27)ms (group Ⅰ) to (144.04±7.39)ms (group Ⅱ) (n=10 in each group, P<0.01); at the test potential of -120 mV, Ik1 increased from (-16.98±4.54) pA/pF(group Ⅰ) to (-19.92±4.08) pA/pF (group Ⅱ) (n=12 in each group, P<0.05); at the test potential of 0 mV, ICa-L decreased from (-8.56±1.29) pA/pF (group Ⅰ) to (-5.24±0.90) pA/pF (group Ⅱ) (n=10 in each group, P<0.01); at the test potential of +60 mV, Ito decreased from (13.20±1.97) pA/pF (group Ⅰ) to (10.30±1.97) pA/pF (group Ⅱ) (n=8 in each group, P<0.05). CONCLUSION: Hypercholesterolemia affects the ionic currents on cardiomyocytes of rats greatly, which may be the ionic mechanism of cardiac toxicity induced by hypercholesterolemia.     

Effect of platelet activating factor on the action potential and potassium currents in guinea-pig ventricular myocytes

AIM: To investigate the effects of platelet activating factor (PAF) on the action potential and potassium currents in guinea-pig ventricular myocytes. METHODS: By using whole-cell patch clamp technique, the effects of PAF on APD₉₀, I_K1 and I_K were investigated in enzymatically dispersed single guinea-pig ventricular myocytes. RESULTS: With 5 mmol/L ATP in the pipette electrode, 1 μmol/L PAF increased APD₉₀ from (225.8±23.3) ms to (352.8±29.8) ms (n=5, P<0.05), decreased I_K1 and I_K tail currents from (-6.1±1.3) nA to (-5.6±1.1) nA (n=5, P<0.05) at -120 mV and from (173.5±16.7) pA to (152.1±11.5) pA (P<0.05, n=4) at +30 mV, respectively. In contract, without ATP in the pipette electrode, 1 μmol/L PAF shortened APD₉₀ from (153.0±24.6) ms to (88.2±19.4) ms (n=5, P<0.01). Incubation of myocytes with 1 μmol/L glibenclamide, a blocker of I_KATP restored prolongation of APD induced by PAF. CONCLUSION: In guinea-pig ventricular myocytes, with 5 mmol/L ATP in the pipette, PAF prolonged APD partly due to the inhibition of I_K and I_K1, while with 0 mmol/L ATP in the pipette, PAF induced an activation of I_KATP, hence a decrease in APD was observed. Therefore, PAF might amplify the heterogeneity between ischemia and normal cardiac myocytes during ischemic reperfusion, which might play a vital role in the pathogenesis of the arrhythmias induced by ischemia/reperfusion.     

Changes of potassium currents in rabbit ventricle with healed myocardial infarction

AIM: To elucidate the mechanism of arrhythmia in healed myocardial infarction (HMI), and to investigate the changes of action potential duration (APD),transient outward potassium current (I_to), delayed rectifier potassium current (I_K) and inward rectifier potassium current (I_K1) of left ventricular myocytes in noninfarcted zone of HMI. METHODS: 12 rabbits were randomly assigned in two groups: HMI group (thoracotomy and ligation of the circumflex coronary); sham-operated group (thoracotomy but no conorary ligation). 3 months after operation, whole cell patch clamp technique was used to record APD, I_to, I_K and I_K1 of ventricular myocytes in non-infarcted zone. RESULTS: Membrane capacitance was larger in HMI group than that in sham-operated group. Action potential duration was lengthened significantly in HMI group and early after depolarization (EAD) appeared in HMI group. The densities of I_to, I_K,tail and I_K1 were reduced significantly in HMI group (P<0.01), from (6.72±0.42) pA/pF, (1.54±0.13) pA/pF and (25.6±2.6) pA/pF in Sham-operated group to (4.03±0.33) pA/pF, (1.14±0.11) pA/pF and (17.6±2.3) pA/pF, respectively. CONCLUSION: The reduced densities of I_to, I_K,tail and I_K1 in ventricular myocytes of non-infarcted zone in HMI are responsible for the prolongation of APD and the presentation of EAD, which play important roles in the malignant arrhythmia of HMI.     

Downregulated transient outward potassium channel protein Kv4.2 and Kv4.3 expression in PVN contributes to sympathoexcitation in rats with chronic heart failure

AIM: To investigate the transient outward potassium channel protein expression in paraventricular nucleus(PVN) and its contribution to renal sympathetic nerve activity(RSNA) in rats with chronic heart failure(CHF).METHODS: A rat model of CHF was prepared by acute myocardial infarction that was induced by ligation of the left anterior descending coronary artery. Four weeks after heart failure, echocardiogram was applied to identify the CHF model and plasma norepinephrine(NE), serum NH₂-terminal pro-brain natriuretic peptide(NT-proBNP) were detected by ELISA. The expression of ransient outward potassium channel proteins Kv4.2 and Kv4.3 at mRNA and protein levels was determined by real-time PCR and Western blot. The mean arterial pressure(MAP), heart rate(HR) and RSNA were measured in anesthetized rats with PVN microinjection of potassium channel blockers 4-AP. RESULTS: In CHF group, the rat cardiac function and Kv4.2 and Kv4.3 expression in PVN were obviously lower while plasma NE and serum NT-proBNP were obviously higher than those in sham group. Microinjection of 4-AP into PVN induced an increase in MAP, HR and RSNA in both sham and CHF rats, while the CHF rats exhibited smaller responses to 4-AP than sham-operated rats.CONCLUSION: Downregulation of Kv4.2 and Kv4.3 expression in the PVN may be a potential mechanism for sympathoexciation in the rats with chronic heart failure.     

Knockdown of IK1 potassium channel inhibits expression and function of ClC-3 chloride channel

AIM: To investigate whether the ClC-3 chloride channel is an acting target of the IK1 potassium channel, and to study the action of IK1 potassium channel on the functional activities and expression of ClC-3 chloride channels. METHODS: IK1 gene was silenced by IK1 siRNA in poorly-differentiated nasopharyngeal carcinoma cells (CNE-2Z). Real-time PCR and Western blot were used to detect the expression of ClC-3 at mRNA and protein levels. The distribution of ClC-3 protein in the cells was observed under confocal immunofluorescence microscope. The chloride current was recorded by the patch-clamp technique. RESULTS: IK1 siRNA was successfully transfected into the CNE-2Z cells and knocked down the expression of IK1 potassium. The mRNA expression of ClC-3 was increased by the IK1 siRNA. IK1 siRNA inhibited the expression of ClC-3 protein. A chloride current was activated by hypotonic challenges, and the hypotonicity-induced current was reduced in the cells which successfully transfected with IK1 siRNA. CONCLUSION: The knockdown of IK1 potassium channels inhibits the expression and function of ClC-3 chloride channel.     

Effects of hydrogen peroxide on sodium current in acutely isolated rat hippocampal CA1 neurons

AIM and METHODS: The effects of hydrogen peroxide on Na⁺ currents were studied in freshly dissociated rat hippocampal CA1 neurons using the whole-cell patch-clamp techinique. RESULTS: ①H₂O₂ caused a dose-dependent and voltage-dependent increase in the voltage-activated Na⁺ currents. The amplitudes of Na⁺ currents were increased (48.0±4.2)% and (88. 2±5. 1)% (n=10) by H₂O₂ at 10 μmol/L and 100 μmol/L, respectively. ②H₂O₂ (10 μmol/L) did not affect the activation process, but changed the inactivation process significantly. Before and after application of 10 μmol/L of H₂O₂, the half-inactivation voltage was (-64.58±1.22)mV and (-53.55±0.94)mV (n=10, P<0.01), but the slope factor was not changed. CONCLUSION: As a product of oxidation metabolism, H₂O₂ is related to some diseases in the central nervous system.     

Blunted response of L-type calcium current to simulated ischemia and reperfusion in ventricular myocytes from diabetic rabbits

AIM: To evaluate the effects of simulated acute ischemia and reperfusion on L-type calcium current (I_{Ca, L}) in ventricular myocytes from diabetic and non-diabetic rabbits.METHODS: Using whole-cell patch clamp techniques, I_{Ca, L} was measured in left ventricular myocytes isolated from 6-week alloxan-induced diabetic rabbits and age-matched control ones at baseline, 5 min of simulated ischemia, and 5 min of reperfusion.RESULTS: There were no significant differences on baseline maximum I_{Ca, L} densities between diabetic and control ventricular myocytes. In control cells (n=11), maximal I_{Ca, L} densities of baseline, ischemia and reperfusion were (-8.36±1.63)pA/pF, (-5.90±1.75)pA/pF and (-4.22±1.02)pA/pF, respectively. The I_{Ca, L} of ischemia was less than that of baseline (P<0.01), while the I_{Ca, L} of reperfusion was less than those of baseline (P<0.01) and ischemia (P<0.05). In diabetic cells (n=9), the I_{Ca, L} of baseline, ischemia and reperfusion were (-7.55±1.62)pA/pF, (-6.05±1.58)pA/pF and (-5.12±1.13)pA/pF, respectively. Only I_{Ca, L} of reperfusion was less than that of baseline (P<0.01), while I_{Ca, L} of ischemia was not significantly different from that of baseline (P>0.05) or reperfusion (P>0.05).CONCLUSION: I_{Ca, L} in diabetic ventricular myocytes represents blunted response to acute ischemic injury, being decreased more slowly than that in control cells. Post-ischemic reperfusion is still a potent inhibitor against I_{Ca, L} in both diabetic and non-diabetic cells. This study may be indicative of the mechanism about ischemia-reperfusion injury to diabetic myocardium and the therapy for diabetic patients with ischemic heart disease.     

Effects of angiotensionⅡon metalloproteinases and matrix in hypertrophied myocardium from infarcted rats

AIM: To investigate the roles of angiotensionⅡ (AngⅡ) receptors (AT₁, AT₂) antagonists on matrix metalloproteinases (MMPs) and extracellular matrix (ECM) system in septal myocardium from infarcted rats.METHODS: The model of rat myocardium infarction (MI) was established by permanent ligation of the left coronary artery. The treatments of the AT₁ receptor antagonist valsartan (10 mg·kg^-1·d^-1) or AT₂ receptor antagonist PD123319 (30 mg·kg^-1·d^-1) were started 7 days prior to surgery. On day 14 after MI, protein levels of MMP-2, 3, 9, fibronectin (FN), tenascin-C (TN-C) in interventricular septum (IS) were determined. The distributions of FN and TN-C were also determined by immunofluorescence.RESULTS: Pathological changes of IS on day 14 after MI showed typical myocardial hypertrophy. Protein expressions of MMP-2, 3, 9 and TN-C of IS in banding group were higher than those in sham-operation group (P<0.01). The expressions of TIMP-1 and FN were lower than those in sham-operation group (P<0.01). Protein expressions of MMP-2, 3, 9 and TN-C in valsartan group were obviously lower than those in banding and PD123319 groups (P<0.01). TIMP-1 and FN protein expressions in valsartan group were higher than those in banding and PD123319 groups (P<0.01). No difference between banding and PD123319 groups was observed (P>0.05).CONCLUSION: AngⅡis involved in myocardium remodeling in infarcted rats, which is mediated via AT₁ receptor to degrade matrix by MMPs. The heart protection of AT₁ receptor antagonists may relate to inhibition of MMPs.     

Effects of endothelin and endothelin receptor antagonist on funny current and its gene expression in neonatal rat ventricular myocytes

AIM: To analyze the effect of endothelin-1 (ET-1) and ET-1 receptor antagonist on funny current (If) and its gene (hyperpolarization-activated cation channel, HCN) expression in neonatal rat ventricular myocytes. METHODS: Fresh ventricular myocytes were isolated from 1-3 d rats. The expression of If gene was measured by real-time quantitative polymerase chain reaction (real-time PCR). If was recorded and studied through whole-cell patch clamp. The cardiomyocytes were stimulated by ET-1 (0, 1, 10, 100 nmol/L) for 3 hours, or ET-1 (100 nmol/L) for 0, 0.5, 1, 3, 6 hours or ET-1 plus ETA- or ETB-receptor antagonist (BE-18257B or IRL-1038), respectively. RESULTS: HCN1, HCN2, HCN3, HCN4 represented (0.23±0.01)%, (83.58±0.04)%, (0.79±0.01)%, (15.44±0.01)% of total HCN mRNA, respectively. If was recorded. When cells were stimulated by ET-1 (10, 100 nmol/L), HCN2 was significantly increased by 0.1, 2 times and HCN4 was increased by 0.1, 0.5 times. When cells were stimulated for 0.5, 1, 3 and 6 hours, HCN2 was significantly increased by 0.3, 1, 5, 5.1 times and HCN4 was increased by 0.1, 0.6, 2, 2.1 times. The treatment of the combination of ET-1 plus BE-18257B significantly decreased HCN2 and HCN4 level. However, HCN1 and HCN3 had no statistically significant change. If would be increased by ET -1 and this effect was reverted by BE-18257B. IRL-1038 had no effects on If and HCN. CONCLUSION: (1) HCN2 and HCN4 represent a large amount of total HCN mRNA in neonatal rat ventricular myocytes. (2) If and the expression of HCN2 and HCN4 are increased by ET-1, this effect is reverted by BE-18257B.     

Transplantation of bcl-2 gene-modified bone marrow mesenchymal stem cells improves cardiac function and angiogenesis in rabbit ischemic car-diac insufficiency model

AIM: To investigate the effects of transplantation of bone marrow mesenchymal stem cells (BMSCs) modified by bcl-2 gene on myocardial cell apoptosis, angiogenesis and cardiac function in the rabbit after acute myocardial infarction (MI).METHODS: The rabbit BMSCs were isolated, cultured and purified in vitro. The BMSCs were transfected with adenovirus or adenovirus-Bcl-2. The rabbit model of MI was established by ligation of left anterior descending branch. The rabbits were injected with Ad-Bcl-2-BMSCs (MI+Bcl-2-BMSCs group), Ad-BMSCs (MI+BMSCs group) and DMEM (MI group) in infarction marginal zone 2 weeks after ligation. The cardiac function was evaluated by echocardiography.The apoptosis of myocardial cells was measured by TUNEL. The mRNA expression of VEGF was detected by real-time PCR. The expression of CD31 was examined by immunohistochemical staining, and new blood capillaries were counted at 4 weeks after BMSCs transplantation. The correlation of the above values with cardiac function was analyzed.RESULTS: The cardiac function was better, the apoptotic rate was lower, the mRNA expression of VEGF and the capillary density were higher in both MI+Bcl-2-BMSCs group and the MI+BMSCs group than those in MI group, and those in MI+Bcl-2-BMSCs group increased more obviously.The left ventricular ejection fraction (LVEF) had a negative correlation with the myocardial cell apoptosis rate. A positive correlation with the mRNA expression level of VEGF and the capillary density was also observed.CONCLUSION: The transplantation of BMSCs modified by bcl-2 gene significantly reduces the myocardial cell apoptosis, promotes angiogenesis, improves heart function of the rabbits with MI.