阅读说明：本技术 一种慢性心衰急性加重型动物模型的制备方法 (Preparation method of acute aggravated animal model for chronic heart failure ) 是由 王维亭 郝春华 张珊 孙双勇 张蕊 徐向伟 赵专友 汤立达 于 2018-06-19 设计创作，主要内容包括：本发明属于医药领域,具体涉及一种慢性心衰急性加重型模型的制备方法。本发明构建的慢性心衰急性加重型模型,根据临床病因、诱因双重因素构建,一方面具备慢性心衰的病理疾病基础心肌梗死的病因,另一方面具备病因基础上体液负荷的诱因。该模型制备成功后,表现出心衰加重的两大特征,即在收缩功能已经减弱基础上的进一步下降、与临床症状相似的呼吸功能指标改变。该模型可用于新药有效性及新药作用机理等研究。(The invention belongs to the field of medicines, and particularly relates to a preparation method of an acute aggravation model of chronic heart failure. The acute aggravation model of the chronic heart failure is constructed according to dual factors of clinical etiology and inducement, so that the acute aggravation model of the chronic heart failure has the etiology of myocardial infarction based on pathological diseases of the chronic heart failure on one hand, and has the inducement of body fluid load based on the etiology on the other hand. After the model is successfully prepared, the model shows two characteristics of aggravation of heart failure, namely further decline on the basis of weakened contraction function and change of respiratory function indexes similar to clinical symptoms. The model can be used for the research on the effectiveness and the action mechanism of a new medicine and the like.)

1. A preparation method of an acute aggravated animal model of chronic heart failure has dual factors of clinical etiology and inducement.

2. The method of claim 1, wherein the model has a myocardial infarction etiology.

After SD rats are anesthetized, the left ventricular shortening Fraction (FS) before operation is determined, anterior descending coronary artery is ligated (only threading is performed in a sham operation group, ligation is not performed, and a Normal group is used), after 8 weeks, the FS after operation is determined, and rats with the FS reduced by more than 50% than that before operation are selected for later use.

4. The method of claim 1, wherein the model is predisposed to fluid loading.

5. Physiological saline is used as a volume loading mode, and the volume loading degree is divided into three loading modes of mild (1 ml/kg, namely HF + mild group), moderate (2 ml/kg, namely HF + moderate group) and severe (3 ml/kg, namely HF + severe group), and the infusion speed is 1 ml/min.

The Normal group used the same saline loading pattern as the heavy loading pattern.

7. The method of claim 1, wherein the model, when successful, exhibits further decline in contractile function based on the decrease in contractile function.

8. The method of claim 1, wherein the model is successful in demonstrating respiratory function index changes with similar clinical symptoms.

9. The method of claim 1, wherein the animal used in the model is a SD rat.

Technical Field

The invention belongs to the field of medicines, and particularly relates to a preparation method of an acute aggravation type animal model of chronic heart failure.

Background

Acute Heart Failure (AHF) refers to symptoms and signs that rapidly develop or worsen secondary to cardiac dysfunction, and includes both newly-developed AHF and acute decompensation of chronic heart failure or acute exacerbation (ADHF or AEHF), with AEHF being more common, accounting for about 70% to 80% of AHF. AHF prognosis is poor with a 3% hospitalization mortality, about 50% hospitalization at 6 months, and up to 60% 5-year mortality. AEHF itself has a pathophysiological basis for disease etiology such as structural or functional impairment of the heart, predisposing factors including infection, arrhythmia, hypertension, improper adjustment or cessation of medication (poor compliance with therapy), excessive intake of sodium salts, excessive or too rapid infusion of fluids, and the like. The construction of animal models of diseases according to clinical etiology, inducement and pathophysiological characteristics is an important means and basis for the research and development of new drugs in pharmacology and pharmacodynamics.

The animal model construction method of the new AHF mainly adopts drug poisoning induction, including verapamil method, propranolol, sodium pentobarbital, propafenone, adriamycin method, ethanol, imipramine and the like. Coronary ligation, balloon occlusion of the coronary artery leading to acute myocardial necrosis and injury have also been used by researchers. These methods generally do not have the pathological basis of long-term, terminal disease in AEHF and cannot be studied as models of AEHF. The mice AEHF can be induced by adopting the adriamycin and the lipopolysaccharide, but the research on signs and symptoms of the AEHF, and indexes related to the hemodynamics and the dyspnea is not seen.

The Zhongwei adopts tail vein injection of adriamycin, 5 mg/kg is carried out every week for 6 weeks continuously, the accumulated total dose is 30 mg/kg, and the tail vein injection of bacterial lipopolysaccharide is 5 mg/kg after 7 weeks, so that the ADHF animal model can be established. The ADHF model mouse has increased death rate, obvious myocardial cell damage, interstitial edema and inflammatory cell infiltration, and compared with the chronic heart failure group, the ADHF model mouse has obviously increased expression of brain natriuretic peptide, interleukin 6 and Bax protein, thereby proving that inflammatory cell factors are obviously increased after the bacterial lipopolysaccharide is injected, the myocardial cell apoptosis is obviously increased, and the cardiac function is acutely reduced. The model is characterized in that the myocardial toxicity injury lesion is the cause of disease and the infection is the cause of infection. The defect is that the adriamycin has wide toxicity, not only has cardiotoxicity, but also has bone marrow suppression, hepatotoxicity, gastrointestinal tract, skin and other nutrition metabolic system injuries, and has some differences with the basic pathological changes of a plurality of clinical patients.

The model is constructed based on dual factors of clinical etiology and inducement, on one hand, the model has the etiology of myocardial infarction based on pathological diseases of chronic heart failure, on the other hand, the model has inducement of body fluid load based on the etiology, and can make up for the defects of the existing model.

Disclosure of Invention

In view of the above, the present invention aims to provide a novel method for preparing an acute aggravation animal model of chronic heart failure. The acute aggravation model of the chronic heart failure prepared by the preparation method has dual factors of clinical etiology and inducement, and has the etiology of myocardial infarction based on pathological diseases of the chronic heart failure on one hand and the inducement of body fluid load based on the etiology on the other hand. The model shows two major characteristics of aggravation of heart failure, namely further decline on the basis of already weakened contractile function, and change of respiratory function index similar to clinical symptoms.

In order to realize the purpose of the invention, the invention adopts the following technical scheme:

after anesthesia, SD rats were anesthetized with Vevo 770^TM120 Doppler ultrasound 17.5 MHz probe echocardiography, M-mode images are acquired and stored in left ventricular short axis anatomical sites, and preoperative Foreshortening Score (FS) is determined by off-line workstation software.

The rat chest anterolateral operation field is disinfected conventionally. The skin and the muscular layer are cut in the direction parallel to the sternum, the ring muscular layer is sutured in a purse-string way, the fourth rib on the left side is cut off, the thoracic cavity is expanded by hemostatic forceps, and the heart is pulled out by a ring-shaped hook. Threading the anterior descending branch of coronary artery at the position 3-4 mm below the left auricle with 6/0 atraumatic silk thread to ligate the anterior descending branch of coronary artery. The heart is placed back in the chest cavity, the air in the chest cavity is pumped out, and the chest cavity is closed. Sham (Normal) animals were threaded only at the corresponding coronary sites and no ligation was performed, the other procedures were the same as those for ligation.

After 8 weeks, measuring the postoperative FS, selecting rats with the FS reduced by more than 50% compared with the preoperative one, and randomly dividing the rats into 3 groups according to the heart failure degree of the FS, wherein each group comprises 10 rats.

Physiological saline (NS) is used as a volume loading mode, and the volume loading degree is divided into three loading modes of mild (1 ml/kg, namely HF + mild group), moderate (2 ml/kg, namely HF + moderate group) and severe (3 ml/kg, namely HF + severe group), and the infusion speed is 1 ml/min. The Normal group used the same saline loading pattern as the heavy loading pattern.

After loading with saline, the Left Ventricular End Diastolic Pressure (LVEDP), the maximum rate of increase in left ventricular pressure (+ LVdp/dtmax), the maximum rate of decrease in left ventricular pressure (-LVdp/dtmax), and the Left Ventricular End Diastolic Pressure (LVEDP) were measured by reverse cannulation to the left ventricle through the right carotid artery using an SPR-320NR (2F) Millar pressure catheter. Central Venous Pressure (CVP) was measured using a fluid pressure transducer, right external jugular vein cannulation. A respiratory pressure transducer is fixed on the chest, and the respiratory frequency (R) is measured through respiratory waves. Heart Rate (HR) was measured from ECG II leads and various signals were acquired and recorded with an MP150 multi-lead physiological signal system. The model preparation was successful.

The animal used for preparing the acute aggravation animal model of the chronic heart failure at present is mainly an SD rat. In some embodiments, the SD rat of the present invention can be replaced by other species of rats, dogs, cats, pigs, rabbits, and guinea pigs.

The current causes for preparing acute aggravated animal models of chronic heart failure are myocardial infarction mainly caused by ligation of anterior descending coronary artery branches. Among other causes, in some embodiments, the agents of the invention can be replaced by other causes such as coronary atherosclerosis, coronary thrombosis, chemical coronary injury, drug and poison induced.

The cause for preparing the acute aggravated animal model of the chronic heart failure at present is mainly physiological saline (NS) load. In some embodiments, the inducers of the present invention may be replaced by other liquids such as blood plasma, high molecular dextran, and other drugs with increased activity, increased oxygen consumption, hypoxia, pressure increase, and heart rate influence.

The invention has the advantages that the constructed acute aggravation model of the chronic heart failure is constructed according to dual factors of clinical etiology and inducement, on one hand, the acute aggravation model has the etiology of myocardial infarction based on pathological diseases of the chronic heart failure, and on the other hand, the acute aggravation model of the chronic heart failure has the inducement of body fluid load based on the etiology. After the model is successfully prepared, the model shows two characteristics of aggravation of heart failure, namely further decline on the basis of weakened contraction function and change of respiratory function indexes similar to clinical symptoms. The model can be used for the research on the effectiveness and the action mechanism of a new medicine and the like.

Drawings

FIG. 1 illustrates exemplary embodiment systolic function + LVd after volume loadingp/dt _maxChange, n =10, compared to the Normal group,^*P<0.05,^**P<0.01. the abscissa is the loadThe last 60min period, ordinate + LVdp/dt _maxPercent change. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

FIG. 2 illustrates embodiment systolic function-LVd after volume loadingp/dt _maxChange, n =10, compared to the Normal group,^*P<0.05,^**P<0.01,^***P<0.001. the abscissa is 60min after load, and the ordinate is-LVdp/dt _maxPercent change. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

Fig. 3 illustrates an exemplary embodiment R frequency change after capacity loading, n =10, compared to the Normal set,^*P<0.05,^**P<0.01. the abscissa is the period of time 60min after loading and the ordinate is the percentage change in R frequency. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

Fig. 4 illustrates an exemplary embodiment HR change after capacity loading, n =10, compared to the Normal group,^*P<0.05,^**P<0.01. the abscissa is the time period 60min after loading and the ordinate is the percentage change in HR. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

Fig. 5 illustrates the CVP change after capacity loading, n =10, compared to the Normal set,^*P<0.05,^**P<0.01. the abscissa is the period of time 60min after loading and the ordinate is the percentage change in CVP. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

Fig. 6 illustrates the LVEDP change after capacity loading, n =10, compared to the Normal panel,^*P<0.05,^**P<0.01. the abscissa is the period of time 60min after loading and the ordinate is the percentage change in LVEDP. O is normal, □ is HF + mild, ● is HF + moderate, and Δ is HF + severe.

Detailed Description

The present invention will be further described with reference to the following examples. The examples are merely illustrative and are in no way meant to limit the scope of the invention in any way.

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention. Unless otherwise specified, the experimental materials used in the present invention are all commercially available products and commercially available.