Method for establishing a rat model of acute heart failure

Abstract

The present research aims at successfully establishing a rat acute heart failure model, observing the heart pathological changes of the animal and detecting the heart and blood related indexes to evaluate the stability of the model, so as to make the rat acute heart failure model with short model making period, stable heart failure characteristics, good repeatability and closeness to clinic, facilitate the further research on the pathogenesis and pathophysiological changes of the acute heart failure model and provide certain basis for how to implement the heart protection intervention measures. The rat acute heart failure model construction method of the present application is to inject acetone and isopropyl alcohol into the vein of the rat, stimulate with electromagnetic waves, frighten with sound, stimulate with formaldehyde and stimulate with potassium permanganate, so as to induce the rat acute heart failure model with short model making period, stable heart failure characteristics, good repeatability and closeness to clinic. The rat acute heart failure model constructed by the present application only needs 10 days, the heart failure characteristics are stable, the model is closer to the clinical symptoms and has high repeatability.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of disease animal model construction, and particularly relates to a method for establishing a rat acute heart failure model. BACKGROUND

[0002] Heart failure (HF) is a kind of pathophysiological syndrome caused by myocardial structure and function changes and activation of neuroendocrine hormones due to pressure overload, volume overload, primary myocardial disease and other causes, resulting in low systolic or diastolic function of the heart, reduced contractility of myocardial cells, reduced cardiac ejection, and inability to meet the needs of tissue cell function and metabolism. Acute heart failure (AHF) is a kind of syndrome caused by rapid occurrence or aggravation of heart failure symptoms and signs, and is most common in acute left heart failure. It is caused by acute onset or aggravation of heart function abnormalities, resulting in significant and rapid decrease of cardiac output, sudden increase of pulmonary circulation pressure, and increase of peripheral circulation resistance, thereby leading to insufficient perfusion of tissues and organs and acute congestion syndrome. When AHF occurs, the effective blood perfusion of all tissues and organs in the body will decrease rapidly, which will lead to severe ischemia and hypoxia of the body, thereby reducing the generation of adenosine triphosphate (ATP) and making it difficult to maintain the energy consumption required for physiological activities. The brain is greatly affected, and the consequences are quite serious. Heart failure patients often have different degrees of myocardial structure and function remodeling, and the pathological changes of cardiac remodeling include myocardial cell enlargement, increased extracellular matrix deposition, thickening of the media of intramyocardial coronary arteries, and accumulation of collagen fibers in myocardial interstitium and perivascular, which can cause abnormal fibrosis.

[0003] Heart failure is a group of syndromes caused by various structural or functional diseases of the heart, resulting in impaired ventricular filling and(or) ejection function, and the cardiac output cannot meet the metabolic needs of the body tissues, with congestion in the pulmonary and(or) systemic circulation, and insufficient blood supply to organs and tissues. It is a serious stage and end-stage of various cardiovascular diseases, with high morbidity and mortality, and is a major threat to human health. Heart failure has a complex pathogenesis, poor treatment effect, and poor clinical prognosis. Current research on its treatment cannot meet the clinical requirements. Further exploration of the pathogenesis of heart failure, finding more effective drugs and treatment methods, and improving the prognosis of heart failure are urgent problems to be solved. Heart failure animal models are developed by various means to simulate the pathophysiological development of human heart failure, providing a scientific platform for the study of its occurrence, development, treatment, and prognosis. In recent years, animal experiment research has been widely used in the etiology, pathogenesis, prevention, and treatment of heart failure. At present, there are various methods to replicate heart failure animal models, and the changes in cardiac function and characteristics of different heart failure animal models are different. In the replication of heart failure animal models, the research target should be considered comprehensively to meet the requirements of experimental research as much as possible. Successful replication of heart failure models suitable for experimental requirements is an important prerequisite for the study of heart failure. However, research on the pathogenesis of heart failure and other related fields has not yet reached the clinical requirements, and clinical human experiments are strictly limited due to medical ethics. Mature heart failure animal models can be used to systematically study the pathogenesis of heart failure, verify the clinical efficacy and adverse reactions of anti-heart failure drugs, and can also be applied to experimental research on stem cell transplantation for myocardial injury.

[0004] In the prior art, the methods for establishing a heart failure model include: 1) pressure overload method: aortic constriction method, renal artery constriction method, and pulmonary artery constriction method. 2) volume overload method: peritoneal venous fistula arteriovenous shunt method, high salt method after kidney removal, and aortic valve insufficiency method. 3) myocardial ischemia / myocardial infarction method. 4) drug method. Surgical modeling methods for establishing in vivo models include abdominal aortic constriction surgery and coronary artery ligation surgery, and drug modeling methods include adriamycin and isopropyl epinephrine. Methods for establishing in vitro models include pentobarbital sodium-induced heart failure models. The establishment of transgenic heart failure models involves using transgenic technology to overexpress some factors in the myocardium of animals related to heart failure, or knocking out some base pairs related to myocardial contraction using gene technology, resulting in heart failure.

[0005] With the rapid development of experimental zoology, although many reliable heart failure models have appeared in recent years and have made great contributions to clinical trials, there are still some drawbacks, such as long model preparation period, unstable heart failure characteristics, poor repeatability, and failure to accurately reflect the changes in clinical pathology. In order to more efficiently and systematically study the pathogenesis of acute heart failure disease, more accurately evaluate the effects of related drugs and study the mechanism of drug action, an acute heart failure model with a short model preparation period, stable heart failure characteristics, good repeatability, and closer to clinical symptoms is urgently needed. SUMMARY

[0006] The present study aims to successfully establish a rat acute heart failure model and observe the pathological changes of the heart of the animal and detect the heart and blood related indexes to evaluate the stability of the model, so as to prepare a rat acute heart failure model with a short model preparation period, stable heart failure characteristics, good repeatability, and closer to clinical symptoms, which is convenient for further studying the pathogenesis and pathophysiological changes of the acute heart failure model and providing a certain basis for how to implement heart protection interventions.

[0007] The present application aims to develop an animal model that can focus on the classic symptoms of acute heart failure and can well evaluate the efficacy of drugs for acute heart failure. The rat acute heart failure model constructed in the present application shows a high success rate and stability in inducing classic symptoms of acute heart failure (dyspnea, impaired cardiac ejection capacity, myocardial fibrosis, myocardial tissue inflammatory cell infiltration, etc.) and myocardial tissue collagen deposition, pulmonary edema, and other symptoms accompanying acute heart failure. The present study can well simulate the clinical onset of acute heart failure and lay a good foundation for rapid screening of therapeutic drugs and interventions. The modeling method is stable and reliable and close to clinical practice.

[0008] The present application is implemented by the following technical solutions:

[0009] A method for establishing a rat acute heart failure model, characterized in that it comprises the following steps:

[0010] (1) Adaptive feeding: select Wistar male rats with normal development, good activity, 6-10 months of age, and a body weight of 260±10g, place the rats in a feeding cage, and adaptively feed them for one week; the temperature is 22-25℃, the relative humidity is 50%-55%, the light and dark cycle is 12h alternately once, and the rats can freely obtain feed and water.

[0011] (2) Intravenous injection: the rats are adaptively fed for one week before modeling; on the 1st, 3rd, and 7th days of modeling, the rats are intravenously injected with acetone 50mg / kg (calculated according to the body weight of the rats), and on the 2nd, 4th, and 8th days of modeling, the rats are intravenously injected with isopropyl alcohol 80mg / kg (calculated according to the body weight of the rats).

[0012] (3), electromagnetic wave stimulation: on the first to the third day of modeling, the rats are irradiated with electromagnetic waves using a specific wave pop instrument, twice a day, each time for 1 hour; the electromagnetic waves emitted by the specific wave pop instrument have a wavelength of 70-80 μm and an intensity of 150-200 mw / cm2.

[0013] (4), sound fright: on the fourth to the seventh day of modeling, animal calls at about 80 decibels are played in the feeding cage, three times a day, each time for 30 minutes.

[0014] (5), formaldehyde stimulation: on the eighth to the tenth day of modeling, a bottle of formalin is placed in the feeding cage, and the bottle opening is opened to allow it to volatilize.

[0015] (6), potassium permanganate stimulation: during the ten days of modeling, the rats are fed with granular feed containing potassium permanganate, and the content of potassium permanganate in the feed is 6.5 g / kg; after ten days of modeling, the rat acute heart failure model can be obtained.

[0016] After modeling, the model is evaluated by observing the life state of the rats, the open field experiment, the dynamic observation of heart ultrasound, the detection of plasma BNP level, the HE staining of myocardium, the HE staining of lung tissue, the detection of serum inflammatory factors, the detection of myocardial apoptosis rate, the determination of heart index, the detection of myocardial mRNA content by q-PCR method, the detection of myocardial infarction area, and the detection of myocardial protein expression by Western Blot method. The results show that the rats have typical symptoms of acute heart failure, which meets the reliability and effectiveness of the animal model of acute heart failure, that is, the pathogenic cause, symptom manifestation, and pathophysiology of the disease can be successfully prepared into a rat acute heart failure model according to the construction method of the application.

[0017] The technical scheme of the application has the following advantages:

[0018] 1. The currently commonly used acute heart failure model has the problems of long model preparation period, unstable heart failure characteristics, poor repeatability, and failure to accurately reflect the changes in clinical pathology. However, the rat acute heart failure model construction method of the application can induce a rat acute heart failure model with a short model preparation period, stable heart failure characteristics, good repeatability, and closer to clinical conditions through intravenous injection of acetone and isopropyl alcohol + electromagnetic wave stimulation + sound fright + formaldehyde stimulation + potassium permanganate stimulation. Such a multi-factor induced acute heart failure model can better simulate the characteristics of human acute heart failure. The model preparation period of the prior art acute heart failure model is usually about 6 weeks, while the rat acute heart failure model constructed by the application only needs 10 days, and the heart failure characteristics are stable, closer to clinical symptoms, and have high repeatability.

[0019] 2、The non-single factor induced acute heart failure animal model of the present application is a relatively ideal animal model from the aspects of pathophysiological theory, simulation of typical clinical symptoms and treatment prediction, and will provide support for the pathogenesis exploration of acute heart failure and provide a theoretical basis for the development of new solutions for the treatment of acute heart failure. The acute heart failure animal model constructed by the present application has high efficiency, stable effect and small individual difference; the multiple factors are synergistic, and the constructed disease model is more accurate. The model shows high success rate and stability in inducing classic symptoms of acute heart failure (dyspnea, impaired cardiac ejection capacity, myocardial fibrosis, myocardial tissue inflammatory cell infiltration, etc.) and symptoms accompanying acute heart failure such as myocardial collagen deposition and pulmonary edema.

[0020] 3、In the present application, acetone 50mg / kg is intravenously injected to the rats in the morning of the 1st, 3rd and 7th day of modeling, and isopropyl alcohol 80mg / kg is intravenously injected to the rats in the morning of the 2nd, 4th and 8th day of modeling. Acetone can stimulate the renin-angiotensin-aldosterone system (RASS system) of the rats, accelerate the synthesis of myocardial collagen and the proliferation of fibroblasts, cause excessive deposition of collagen and hypertrophy of fibroblasts. Isopropyl alcohol can regulate the expression of MMPs, destroy the balance of the interaction of the MMPs / TIMPs system, cause increased degradation of fibrous collagen, extracellular matrix remodeling and ventricular dilation, and promote the occurrence of myocardial fibrosis. Multiple intravenous injections of acetone and isopropyl alcohol to the rats in succession can greatly induce myocardial fibrosis and myocardial cell apoptosis of the rats, thereby inducing the occurrence of acute heart failure.

[0021] The irradiation of electromagnetic waves with specific wavelength and intensity can damage the Na + -K + -ATPase of the rats, destroy the Na + 、K + balance inside and outside the cell membrane, make a large amount of Na + enter the cell, a large amount of K + flow out of the cell, and a large amount of Na + in the cell exchange with Ca2 2+ , causing the aggregation of Ca 2+ in the cell, thereby damaging the myocardial cells. After intravenous injection of acetone and isopropyl alcohol, the irradiation of specific electromagnetic waves to the rats not only consolidates and strengthens the induction effect of intravenous injection, but also further induces myocardial cell damage, which is greatly beneficial to the induction of acute heart failure in rats.

[0022] A certain degree of sound fright can accelerate the heart rate of rats, induce myocardial cell hypertrophy to some extent, make the structure disorder, even muscle fiber rupture, and further increase myocardial oxygen consumption, aggravate the cardiac load, and ultimately lead to heart failure. Three days of sound fright after intravenous injection and specific electromagnetic wave irradiation can further induce and aggravate myocardial injury.

[0023] Formaldehyde can stimulate the neuroendocrine system of rats to some extent, and the secreted substances can induce myocardial cell apoptosis, reduce myocardial cells, cause myocardial contractility to decrease, and further aggravate heart failure. On the basis of intravenous injection of acetone and isopropyl alcohol, specific electromagnetic wave irradiation and sound fright in the early stage of modeling, formaldehyde stimulation in the later stage of modeling can play a role in connecting the past and the future, and further aggravate myocardial cell apoptosis.

[0024] Potassium permanganate in feed can induce the formation of rat myofibroblasts through the TGF-β1 / Smad3 signaling pathway, cause excessive deposition of extracellular matrix, and inhibit the expression of metalloproteinases, thereby inducing myocardial fibrosis. Adding potassium permanganate to the feed of rats can cooperate with other steps to maximize the induction and aggravation of myocardial fibrosis and inflammatory response in rats, thereby ensuring the stability of the rat model of acute heart failure with heart failure characteristics.

[0025] 4、The present study can provide a good theoretical support and model reference for further analyzing the immune mechanism of humans after acute heart failure and evaluating new treatment methods. The research team has repeatedly verified the model, and the results are reliable. The model has been used to evaluate the efficacy of various drugs, and the application value of the model has been verified. The animal model shows typical characteristics of acute heart failure: difficulty breathing, impaired cardiac ejection capacity, myocardial fibrosis, and inflammatory cell infiltration in myocardial tissue. The model can be used for basic research in the field of acute heart failure, and lays a foundation for exploring the pathogenesis of acute heart failure. The present study can provide a reliable animal model for the evaluation of the efficacy of drugs for acute heart failure, which has the advantages of short model preparation period, stable heart failure characteristics, good repeatability, and closer clinical symptoms. DETAILED DESCRIPTION

[0026] A method for establishing a rat model of acute heart failure, characterized in that it comprises the following steps:

[0027] (1) Adaptive feeding: select Wistar male rats with normal development, good activity, 6-10 months old, and body weight of 260±10 g, place the rats in a feeding cage, and adaptively feed them for one week; the temperature is 22-25℃, the relative humidity is 50%-55%, the light-dark cycle is 12h alternately once, and the rats can freely obtain feed and water.

[0028] (2), intravenous injection: rats were adaptively fed for one week before modeling; on the 1st, 3rd, 7th morning of modeling, rats were intravenously injected with acetone 50 mg / kg (based on the weight of rats), and on the 2nd, 4th, 8th morning of modeling, rats were intravenously injected with isopropyl alcohol 80 mg / kg (based on the weight of rats).

[0029] (3), electromagnetic wave stimulation: on the 1st-3rd day of modeling, rats were irradiated with electromagnetic waves using a specific wave pop instrument, twice a day, each time for 1 hour; the electromagnetic waves emitted by the specific wave pop instrument have a wavelength of 70-80 μm and an intensity of 150-200 mw / cm2.

[0030] (4), sound fright: on the 4th-7th day of modeling, animal calls at about 80 decibels were played in the feeding cage, three times a day, each time for 30 minutes.

[0031] (5), formaldehyde stimulation: during the 8th-10th day of modeling, a bottle of formalin was placed in the feeding cage and the bottle opening was opened to allow it to volatilize.

[0032] (6), potassium permanganate stimulation: during the modeling period, rats were fed with granular feed containing potassium permanganate, and the content of potassium permanganate in the feed was 6.5 g / kg; after modeling for ten days, the rat model of acute heart failure was obtained.

[0033] After modeling, the model was evaluated by observing the life state of rats, open field experiment, dynamic observation of heart ultrasound, plasma BNP level detection, myocardial HE staining, lung tissue HE staining, serum inflammatory factor detection, myocardial tissue apoptosis rate detection, heart index determination, q-PCR method for detecting myocardial tissue mRNA content, myocardial infarction area detection, Western Blot method for detecting myocardial protein expression, etc. (the control group was injected with normal saline and routinely fed), and the results were as follows:

[0034] 1. Life state observation

[0035] After modeling, the rats' food intake, water intake, and body weight changes were observed and recorded, and the rats' mental state, activity, urine volume changes, hair color changes, and general conditions such as limb edema, and cyanosis of the lips were observed.

[0036] The results showed that the control group rats were active and quick in response, healthy in food and water intake, and had obvious weight gain. The model group rats showed low mental state, reduced movement, fatigue, poor appetite, cyanosis of the lips, yellowish hair color, severe hair loss, diarrhea, and symptoms such as subcutaneous edema and weight loss.

[0037] 2. Open field experiment

[0038] The experimental rats were placed in a 50 cm x 50 cm x 40 cm black bottom blue side wall of the open field reaction box, the video recorder was vertically shot, and the black box bottom in the picture was kept uniform square, moderate size. Under the quiet condition, the experimental rats were placed in the center of the box bottom, and the 5 min spontaneous activity of the rats was recorded. The residual and odor in the box were eliminated before the experiment of the lower rats. The animal behavior analysis system was used to analyze the video, and the differences of the open field distance and the open field speed between the groups were compared.

[0039] Compared with the control group, the open field distance and the open field speed of the model group decreased significantly.

[0040] 3. Dynamic observation of heart ultrasound

[0041] The rats were weighed and anesthetized, and the heart function of the rats was detected by ultrasonic Doppler. The left ventricular short axis fractional shortening (LVFS), left ventricular ejection fraction (LVEF), left ventricular end systolic size (LVESD), and left ventricular end diastolic size (LVEDD) were measured. All data were detected for 3 cardiac cycles and the average value of the measurement was calculated.

[0042] Compared with the control group, the heart of the model rats was enlarged and the systolic function decreased, which was manifested as significant increase of LVESD and LVEDD, and significant decrease of LVFS and LVEF, and the differences were statistically significant.

[0043] 4. Detection of plasma BNP level

[0044] The abdominal aortic blood of the rats was collected in a test tube containing anticoagulant, centrifuged at 3000 r / min for 3 min, and the plasma was stored at-80℃. The average level of plasma BNP of the rats in each group was detected by ELISA.

[0045] The average level of plasma BNP of the model rats was (120.46±9.82) ng·L -1 , which was significantly higher than that of the control group (66.56+7.02) ng·L -1 .

[0046] 5. Myocardial HE staining

[0047] The rats were anesthetized and sacrificed, and the heart specimens were quickly taken. The apex part of the tissue was fixed with 4% formaldehyde solution, paraffin-embedded, and sectioned. The myocardial tissue morphology was observed under a light microscope after routine HE staining.

[0048] The myocardial tissue was stained with hematoxylin-eosin (HE staining), which showed that the myocardial tissue of the model group was damaged, showing myocardial disease-like changes: severe myocardial cell proliferation, hypertrophy and disordered arrangement, irregular nuclear arrangement, disordered muscle fiber arrangement, different degrees of inflammatory cell infiltration and interstitial edema. The morphology of myocardial cells, cytoplasm, interstitium and transverse (longitudinal) stripes were normal in the normal group.

[0049] 6. Lung tissue HE staining

[0050] The lung tissue of the rat was taken, washed with ice double distilled water to remove residual blood, then soaked in 10% formaldehyde solution for 24 h fixation, dehydrated, paraffin-embedded, 3-5 μm continuous sectioning, HE staining, conventional ethanol gradient dehydration, transparency, neutral resin sealing, and the pathological morphological changes of the lung tissue were observed under an optical microscope.

[0051] The alveolar results of the control group were complete, there was no effusion in the alveolar cavity, and there was no congestion of the interstitial blood vessels and no inflammatory cell infiltration. The lung tissue of the model group: the alveolar wall capillary was dilated and congested, the alveolar wall fibrous connective tissue was significantly proliferated, a small amount of edema fluid, part of the inflammatory cells and red blood cells leaked out, local hemosiderin deposition was visible, and the interstitial blood vessels were dilated and congested.

[0052] 7. Serum inflammatory factor detection

[0053] ELISA was used to detect the levels of IFN-γ, IL-2, IL-4, IL-6 and IL-10 in the serum of the rat. The abdominal aortic blood of the rat was centrifuged at 3000 r / min for 10 min at 4℃, the serum was taken, aliquoted, and stored at -20℃ for standby, and the index detection was performed according to the steps of the corresponding kit instruction.

[0054] Compared with the control group, the levels of inflammatory factors IFN-γ and IL-2 in the myocardial tissue of the model group were significantly increased, and the levels of IL-4, IL-6 and IL-10 were significantly decreased.

[0055] 8. Myocardial tissue apoptosis rate detection

[0056] The heart of the rat was taken out, and part of the myocardial tissue was fixed in 10% formaldehyde solution for TUNEL staining to observe the myocardial cell apoptosis and calculate the apoptosis rate.

[0057] Compared with the control group, the myocardial cell apoptosis index of the model group was significantly increased.

[0058] 9. Heart index determination

[0059] After removing the ascites in the abdominal cavity of the rat, the weight was determined again, the ascites weight was calculated as the body weight minus the weight after removing the ascites, the heart was taken out, the non-myocardial tissue was removed, the blood was washed, the moisture was absorbed with filter paper, and the whole heart weight was determined. The heart index (HW / BW) was calculated as the whole heart weight / body weight.

[0060] Compared with the control group, the body weight and the heart weight of the model group were significantly decreased, and the ascites weight and the heart index had an increasing trend.

[0061] 10. Detection of the mRNA contents of TGF-β1, Smad3 and Caspase-3 in the myocardial tissue by quantitative polymerase chain reaction (q-PCR)

[0062] The myocardial tissue of rats was cut and put into liquid nitrogen for homogenate. Steps: total RNA was obtained by Trizol method, the RNA concentration was determined by ultraviolet spectrophotometer, cDNA was obtained by transcription kit, reaction was carried out by PCR kit, and the reaction results were analyzed by PCR instrument.

[0063] Compared with the control group, the content of TGFβ1, Smad3 and Caspase-3 mRNA in the myocardial tissue of the model group increased.

[0064] 11. Detection of myocardial infarction area (IS)

[0065] After paraffin-embedded myocardial section, Masson staining was carried out, and under optical microscope, myocardial collagen fibers were blue, muscle fibers, cytoplasm and red blood cells were red, and cell nuclei were blue-brown. Five sections were taken from each heart, observed under a stereoscope and photographed, and the relative IS of myocardium was calculated by Image-ProPlus6.0 software, IS (%) = infarcted myocardial arc length / [(left ventricular inner circumference + left ventricular outer circumference) / 2] x 100%.

[0066] The myocardial IS detection results showed that the myocardial infarction of the model group was significantly higher than that of the control group.

[0067] 12. Western Blot method for detecting expression of NF-κBp65 and TLR4 protein in myocardium of rats

[0068] 0.5-0.6 g of heart tissue was taken, pre-cooled protein lysis solution was added for grinding and homogenate, then placed on ice for lysis for 30 min, high-speed centrifugation for 15 min to obtain supernatant (i.e. total protein solution), 5x protein loading buffer and protein sample were added for denaturation, gel electrophoresis, membrane transfer, antibody incubation and exposure to obtain protein bands. The film was scanned and archived, and the optical density value of the target band was processed and analyzed by ImageJ software.

[0069] Compared with the control group of rats, the expression of NF-κB protein and TLR4 protein in the myocardium of the model group of rats increased significantly, proving that the modeling was successful.

Claims

1. A method for establishing a rat model of acute heart failure, characterized by, Comprise the following steps: (1) adaptive feeding: select normal development, good activity, 6-10 months old, body weight 260±10 g Wistar male rats, rats are placed in the cage, adaptive feeding for one week; temperature is 22-25℃, relative humidity 50%-55%, light and dark cycle for 12h alternately 1 times, rats can freely obtain feed and water; (2) intravenous injection: rats adaptive feeding for one week start modeling; in the first, 3, 7 days in the morning respectively to rats intravenous injection of acetone 50mg / kg, in the second, 4, 8 days in the morning respectively to rats intravenous injection of isopropyl alcohol 80mg / kg; (3), electromagnetic wave stimulation: on the 1st-3rd day of modeling, electromagnetic wave irradiation was performed on the rats using a specific wave pop instrument, twice a day, each time for 1 hour; the electromagnetic wave emitted by the specific wave pop instrument had a wavelength of 70-80 pm and an intensity of 150-200 mw / cm 2 ; (4) sound fright: in the modeling of the 4-7 days, in the cage play 80 decibel or so animal call, play three times a day, each play 30 minutes; (5) formaldehyde stimulation: in the modeling of the 8-10 days, in the cage put a bottle of formalin, and open the bottle opening to volatilize; (6) potassium permanganate stimulation: during the modeling of ten days, to rats feed containing potassium permanganate granular feed, the potassium permanganate content in feed is 6.5g / kg; modeling ten days, namely can obtain rat acute heart failure model.