动物模型 z-bo 2020-12-07 726

　　Introduction: In the field of life sciences including pharmacological research, pigs are expected to become another model besides dogs or monkeys. Since pigs have biological characteristics similar to the cardiovascular, respiratory, metabolic, and gastrointestinal systems in addition to human skin, the use of experimental animals and/or monkeys may cause intense concern among animal lovers. Since miniature pigs weigh between 34 and 68 kg in adulthood, very small miniature pigs weigh 7 kg at 6 months of age. However, there is still a lack of background knowledge about the cardiovascular response caused by typical myocardial ion channel modulators. This is essential for applying this new animal to the field of cardiac safety pharmacology. In this study, we assessed the cardiovascular effects of pisineamide, verapamil and E-4031, and divided miniature pigs into in vivo experimental models, which can preferentially inhibit cardiac Na+, Ca2+ and K+ channels, respectively. Sober animals usually have more repolarization reserves than healthy people, making the animals less sensitive to drug-induced QT prolongation. Make animals insensitive to drug-induced QT prolongation. Halothane anesthesia inhibits cardiac IKS and IKR and autonomic tension to a certain extent, resulting in a decrease in repolarization reserve. The degree of repolarization reserve of health-conscious and halothane anesthetized dogs has been considered to be similar. Therefore, the halothane anesthetized dog drug induced QT interval extension well mimics the QT interval extension of healthy people. With halothane inhalation anesthesia, the cardiovascular response induced by the drug was directly compared with previous reports under the same anesthesia condition.

　　 Cardiovascular variables: Miniature pigs are 8±2 months old and weigh 8.6±1.1 kg (n=13). Intramuscular injection of ketamine (16 mg/kg)/xylazine (1.6 mg/kg) was pre-anesthetized. A 24G cannula was inserted into the superficial auricular vein, and propofol was injected intravenously 20 mg/d to induce anesthesia. After tracheal intubation, anesthesia was maintained with 1% halothane and 100% oxygen. The tidal volume and respiratory rate were set at 10 mL/kg and 15 breaths/minute, respectively. Two groups of clinically available 6F catheter sheaths were inserted into the right femoral artery and vein. Heparin calcium 100 IU/kg was injected intravenously through the right femoral vein catheter to prevent blood clotting. The left ventricular pressure at the peak time of the R wave on the ECG is defined as the left ventricular end-diastolic pressure. In sinus rhythm, the maximum left ventricular pressure (LVDP/DTMax) and left ventricular end-diastolic pressure are obtained to estimate the left ventricular contractility and preload. The electrocardiogram is obtained from A-B leads. The QT interval is corrected by the Van de Water formula: QTc=QT-0.087×(RR-1000), because this formula was originally developed for dogs.

　　Experimental operation: monitor aortic and left ventricular pressure and electrocardiogram, and use real-time automatic data analysis system for analysis. The three complexes recorded continuously are used to calculate the average value of cardiovascular variables. After the basic evaluation, 1 mg/kg of pisineamide was injected intravenously within 10 minutes, and each variable was evaluated at 5, 10, 15, 20, and 30 minutes after the start of the administration (n=4). The dose of verapamil was 0.1 mg/kg, and pircinamide was evaluated in the same way (n=4). E-4031 was injected intravenously at a low dose of 0.01 mg/kg within 10 minutes, and each parameter was evaluated at 5, 10, 15, 20 and 30 minutes after the start of the administration (n=5). Additional E-4031 was infused at a high dose of 0.1 mg/kg for more than 10 minutes, and each variable was evaluated at 5, 10, 15, 20, 30, 45, and 60 minutes after the start of administration. Based on previous reports, the doses of pircinamide, verapamil and E-4031 were determined. Plasma drug concentration: Use a heparinized syringe to draw 2 ml of blood from the femoral artery, 10, 15 and 30 minutes after the start of pircinamide and verapamil, and 5, 10, 15 and 30 minutes after the start of low-dose E-4031 Measure the plasma drug concentration at 5, 10, 15, 30 and 60 minutes after the start of the high-dose E-4031; centrifuge the blood sample at 1500°C for 30 min, prepare the plasma at 4°C, and then store it at -80°C until measured Drug concentration. HPLC method was used to determine the plasma concentration of pisineamide, verapamil and E-4031.

　　Result: Describes the effects of pisineamide, verapamil and E-4031 on electrocardiogram, aortic pressure and left ventricular pressure. The time course of plasma concentration, heart rate, mean blood pressure, left ventricular end diastolic pressure and LVD/DTMax, PR interval, QRS width, QT interval and QTc.

　　 The pharmacokinetic and pharmacodynamic study of pisineamide: The peak plasma concentration of pisineamide was observed 10 min after the start of the infusion, which was 2.37±0.22μg/ml. The pre-drug control values (C) of heart rate, mean blood pressure, left ventricular end-diastolic pressure and LVDP/DTMAX are 83±6 BPM, 58±3 mmHg, 11±2 mmHg, and 612±43 mmHg, respectively /second. Picinamide significantly reduced heart rate, mean blood pressure and LVDP/DTMax 10-30 minutes after the start of administration, but left ventricular end-diastolic pressure did not change significantly. The prodrug control values (C) of PR interval, QRS width, QT interval and QTc were 128±9 ms, 88±9 ms, 418±37 ms and 441±39, respectively. Pisinamide significantly prolongs the PR interval, QRS width, QT interval and QTc in 10-30 minutes.

　　 Pharmacokinetic and pharmacodynamic study of verapamil: The peak plasma concentration of verapamil was observed 10 minutes after the start of the infusion, which was 627±77 ng/ml. The pre-drug control values (C) of heart rate, mean blood pressure, left ventricular end diastolic pressure and LVD/DTMax were 72±4 BPM, 48±6 mmHg, 12±1 mmHg and 456±39 mmHg, respectively second. Verapamil reduces heart rate and average blood pressure 5-30 minutes after the start of administration. The LVDP/DTMAX was reduced 15-30 minutes after administration, and the left ventricular end diastolic pressure did not change significantly. The prodrug control values (C) of PR interval, QRS width, QT interval and QTc were 132±7 ms, 80±5 ms, 444±19 ms and 458±16, respectively. Verapamil prolonged the PR interval in 10 to 30 minutes, shortened the QRS width in 10 to 20 minutes, shortened the QTc in 10 to 30 minutes, and had no significant change in the QT interval.

　　 The pharmacokinetic and pharmacodynamic study of E-4031: Low-dose and high-dose E-4031 reached a peak value 10 minutes after the infusion started, which were 29±12 and 594±309 ng/ml, respectively. The pre-drug control values (C) of heart rate, mean blood pressure, left ventricular end-diastolic pressure and LVDP/DTMax were 82±19 BPM, 66±7 mmHg, 14±1 mmHg, and 734±56 mmHg/ second. The low dose 15-20 minutes after the start of the infusion reduced the heart rate, while other cardiac hemodynamic variables did not change significantly. The high dose decreased the heart rate 5-30 minutes after the infusion started, the average blood pressure was lowered in 5-60 minutes, and the LVDP/DTMAX was lowered in 10-30 minutes, but the left ventricular end diastolic pressure did not change significantly. The prodrug control values (C) of PR interval, QRS width, QT interval and QTc were 132±14 ms, 73±5 ms, 416±47 ms and 426±31, respectively. Low-dose prolonged QT interval, 5-30 minutes QTc prolonged, 5-60 minutes high-dose QTc prolonged, PR interval or QRS width did not change significantly.

　　 Conclusion: This study is characterized by miniature pigs as an in vivo experimental model. By evaluating the cardiovascular effects of piscinamide, verapamil and E-4031, the cardiovascular effects of humans and dogs were compared for the first time. The peak plasma concentrations of pisineamide, verapamil, and E-4031 in the plasma of mini-pigs are 1.7 to 4.8 times higher than that of dogs. This may be due to the effective volume of drugs in mini-pigs and less fat content in the body than dogs. The degree of drug-induced cardiovascular response is usually greater in minipigs than dogs. It can be explained by the following possibilities: the unique pharmacokinetic curve of miniature pigs, fewer large reflexes mediate sympathetic nerve enhancement and smaller repolarization reserves. This makes minipigs a new animal model for evaluating the safety of the heart of chemical drugs.