Interventional Therapy Of Dendranthema Morifolium In Animal Model Of Common Heart Diseases

BACKGROUND:Arrhythmia is one of the major reasons of clinical symptom of many kinds of serious heart diseases, and ventricular fibrillation is the constant pathological cause of heart dyfunction. Recently, myocardial injury induced by ischemia/reperfusion or anoxia/reoxygenation has been of widespread concern because its pathological mechanism will explain a lot of clinical paradox which had not been elucidated. The mechanism of myocardial injury is tightly related with heart disease and death induced by coronary artery bypass grafting, thrombectomy, unstable angina and coronary artery spasm.The flower of Dendrantheira morifolium (ramat.) tzvel. (DM) has been used as a folk medicine for sudorific, antidote and clearing away pathogenic heat in the People's Republic of China. In the previous study of our laboratory, the water extract of DM increased coronary flow and contractility in the isolated rabbit heart, showing its effect out of the traditional frame of DM.The objectives of the present study were as follows: (1) To investigate the effect of Dendranthema morifoliiun (Ramat.) Tzvel (DM) on experimental arrhythmia induced by chloroform in mice and by aconitine in rats; (2) To investigate the effect of DM on isolated rat heart and ventricular myocytes during ischemia/ reperfusion and anoxia /reoxygenation.METHODS:1. Experimental arrhythmia model induced by aconitineMale Sprague-Dawley (SD) rats of 240~280 g were used. They were anaesthetized by an intraperitoneal injection of urethane (1.2 g/kg) and recorded by a electrocardiogram machine. Arrhythmias were induced by injecting aconitine (30 u.g/kg), in which the number or duration of ventricular premature beats (VPB), ventricular tachycardia (VT) and ventricular fibrillation (VF) were examined. Quantification of arrhythmias was achieved by using scoring systems: score = (logioVPBs) + 2(Iogi0number of episodes of VT) + 2[(logi0number of episodes of VF) + (logiototal duration of VF)].2. Experimental arrhythmia model induced by chloroformNIH mouse of 25~30g was put into a chamber with chloroform (2 ml). After the mouse halted its breath for 5 second, the electrocardiogram was recorded and the incident rate of VF and the conversion rate were examined.3. Isolated perfused rat heart preparationMale SD rat heart was retrogradely perfused with a 95%C>2+5%CO2 equilibrated Krebs-Henseleit (K-H) solution via the aorta at a constant pressure of 74 mmHg.Electrocardiogram and left ventricular contraction parameters were examined by MedLab data acquisition system.4. Preparation of isolated ventricular myocytesThe hearts were perfused with Ca2+-free Tyrode's solution with Langendorff apparatus. Then the perfusion solution was switched to a low Ca2+ (50 jaM) Tyrode's solution containing 0.03% collagenase and 1% bovine serum albumin (BSA) for 10 min. The ventricles were cut, minced, and gently triturated with a pipette in the low Ca2+Tyrode's solution containing BSA at 37'C for 10 min. The cells were resuspended in the Tyrode's solution in which the Ca2+ concentration was gradually increased to 1.25 mM over 40 min.5. Measurement of myocyte contractionMyocytes were placed in a chamber mounted on an inverted microscope. The chamber was perfused continuously at 2 ml/min with KH buffer, with 1% BSA and a gas phase of 95% O2 / 5% CO2 at room temperature. Myocyte contractions were elicited by field stimulation at a frequency of 0.5 Hz, and intensity twice the contraction threshold. A video-tracking system was used to measure the peak velocity of cell shortening (+dL/dtmax), the peak velocity of cell relengthening (-dL/dtmax), the amplitude of contraction (dL) and the end-diastolic length of the isolated myocyte.6. Intracellular calcium recordingMonitoring of intracellular calcium was carried out using the fluorescent dye fura-2. Fluorescence was measured on an Olympus inverted microscope equipped with a fluorometer system (T.I.L.L., Germany). The Ca2+-dependent signal of fura-2 was obtained byilluminating at 340 and 380 nm a...