Method for establishing a mouse model of heart failure with preserved ejection fraction

Abstract

This invention belongs to the field of animal model construction technology, specifically relating to a method for establishing a mouse model of heart failure with abnormal ejection fraction. The method involves feeding mice a diet containing adenine for 25–30 days. C57BL / 6J mice were fed a diet containing adenine for 25–30 days, and cardiac function was assessed using a Vevo 2100 small animal ultrasound imaging system. The results showed increased ejection fraction, decreased cardiac output, reduced left ventricular diameter and volume during systole and diastole, and mild cardiac fibrosis. This demonstrates that adenine diet can induce heart failure with abnormal left ventricular ejection fraction in mice, providing a practical direction for the study of heart failure.

Description

Technical Field

[0001] This invention relates to a method for establishing a mouse model of heart failure with abnormal ejection fraction, belonging to the field of animal model construction technology. Background Technology

[0002] Heart failure (HF) is a global problem, and the number of HF patients has been rapidly increasing in recent years. HF is defined as a clinical syndrome whose symptoms and / or signs are caused by structural and / or functional cardiac abnormalities and confirmed by objective evidence of elevated natriuretic peptide levels and / or pulmonary or systemic congestion. In 2021, a universal definition and classification of ejection fraction (EF) was proposed. Ejection fraction refers to the percentage of stroke volume relative to the end-diastolic volume of the ventricle (i.e., cardiac preload), and is one of the important indicators for determining the type of heart failure. The calculation formula is: EF = (EDV - ES) × 100% / EDV, where EF is the ejection fraction; EDV is the end-diastolic volume of the ventricle; and ES is the end-systolic volume of the ventricle. In routine practice, heart failure is usually classified according to the left ventricular ejection fraction (LVEF). The definition of normal LVEF varies depending on guidelines or medical / scientific societies. For example, the British Society of Echocardiography defines normal LVEF as ≥55%. The American Society of Echocardiography and the European Society for Cardiovascular Imaging define normal LVEF as 52%–72% for men and 54%–74% for women.

[0003] In recent years, the terminology for the definition of heart failure has been revised from "HF with reduced ejection fraction, preserved ejection fraction, and intermediate (or borderline) ejection fraction" to "HF with reduced ejection fraction, preserved ejection fraction, and mildly reduced ejection fraction", and "HF with improved ejection fraction" has been introduced. The definition of this classification is as follows: (1) Heart failure with reduced ejection fraction (HFrEF): Heart failure with LVEF ≤ 40%; (2) Heart failure with mildly reduced ejection fraction (HFmrEF): LVEF of 41% to 49%; (3) Heart failure with preserved ejection fraction (HFpEF): Heart failure with LVEF ≥ 50%; (4) Heart failure with improved ejection fraction: Heart failure with baseline LVEF ≤ 40%, LVEF increased by ≥ 10 points from baseline, and LVEF > 40% on the second measurement.

[0004] A new type of heart failure, heart failure with supra-normal ejection fraction (HFsnEF), has recently been proposed, characterized by elevated left ventricular ejection fraction (LVEF) (LVEF > 65%). Some studies have reported that patients with elevated LVEF may have an increased risk of long-term major adverse cardiovascular events and a U-shaped death pattern. Currently, the prognosis of HFsnEF is not fully established, but it appears to be associated with an increased risk of long-term major adverse cardiovascular events. Patients with LVEF > 60% have reportedly smaller ventricular diameters (relatively increased wall thickness, smaller left ventricular end-diastolic volume, and smaller left ventricular end-systolic volume). Patients with higher LVEF also have lower stroke volume index (stroke volume index is the ratio of stroke volume (SV) to body surface area (BSA), measured in mL / m²). 2 These structural changes are typically associated with smaller cardiac chambers and left ventricular concentricity. This is due to the shortening and thickening of crossing fibers, which increases the contraction rate of cardiomyocytes at the endocardial level, thereby increasing the ejection fraction. These structural changes are associated with abnormal and impaired left ventricular diastolic function.

[0005] Heart failure with an abnormal ejection fraction is a newly discovered type of heart failure, and there are currently no animal models available for basic research. Therefore, how to establish an animal model of heart failure with an abnormal ejection fraction is a technical problem that urgently needs to be solved by those skilled in the art. Summary of the Invention

[0006] This invention addresses the lack of existing animal models for studying heart failure with abnormal ejection fraction by providing a method for establishing a mouse model of heart failure with abnormal ejection fraction.

[0007] The technical solution of the present invention to solve the above-mentioned technical problems is as follows: A method for establishing a mouse model of heart failure with abnormal ejection fraction, wherein the mice are fed a diet containing adenine for 25 to 30 days.

[0008] Based on the above technical solution, the present invention can also be improved as follows:

[0009] Furthermore, the adenine-containing diet is animal feed containing adenine.

[0010] Furthermore, in the animal feed containing adenine, the mass fraction of adenine is 0.15%-0.20%.

[0011] Furthermore, the adenine-containing animal feed contains 0.20% adenine by mass.

[0012] Furthermore, feed the animals 5g-10g of adenine-containing feed daily.

[0013] Furthermore, the rearing period is 28 days.

[0014] Furthermore, the animal feed is rodent feed.

[0015] Furthermore, the rodent feed includes:

[0016] 20% casein;

[0017] 39.75% corn starch;

[0018] 13.2% corn dextrin;

[0019] 10% sucrose;

[0020] 5% cellulose;

[0021] 7% soybean oil;

[0022] 3.5% complex minerals;

[0023] 1% multivitamins.

[0024] Furthermore, the mice were C57BL / 6J wild-type male mice.

[0025] The beneficial effects of this invention are as follows: C57BL / 6J mice were continuously fed an adenine-containing diet for 25-30 days. The cardiac function of the mice was detected using a Vevo 2100 small animal ultrasound imaging system. Compared with the control group, the adenine-fed mice showed an increased ejection fraction (from 45.6702±8.90 to 71.94±8.56), a decreased cardiac output (from 13307.5±9109.55 to 2994.34±1188.75), and a decreased systolic left ventricular diameter (from 2.97±0). The decrease in left ventricular ejection fraction (LVEF) from 0.22 to 1.68±0.33, the decrease in diastolic left ventricular diameter (from 3.87±2.77 to 0.17±0.24), the decrease in systolic left ventricular volume (from 34.04±6.41 to 5.79±2.96), the decrease in diastolic left ventricular volume (from 62.73±28.02 to 7.52±5.48), and the presence of mild cardiac fibrosis demonstrate that an adenine diet can induce heart failure with an abnormal left ventricular ejection fraction in mice, providing a practical direction for the study of heart failure. Attached Figure Description

[0026] Figure 1 M-mode echocardiogram images of the heart obtained from mice in the examples and comparative examples were obtained using the vevo2100 small animal ultrasound imaging system.

[0027] Figure 2 Ejection fraction calculated based on the results of the vevo2100 small animal ultrasound imaging system;

[0028] Figure 3The cardiac output was calculated based on the results of the vevo2100 small animal ultrasound imaging system.

[0029] Figure 4 The changes in interventricular septal thickness, left ventricular diameter, and left ventricular volume during systole and diastole during the experimental stage were recorded in mice after 29 days of feeding.

[0030] Figure 5 Images of mouse heart tissue after masone staining for both examples and comparative studies;

[0031] Figure 6 The example shows the changes in mouse body weight. Detailed Implementation

[0032] The principles and features of the present invention are described below with reference to the accompanying drawings. The examples given are only for explaining the present invention and are not intended to limit the scope of the present invention.

[0033] Examples and comparative examples:

[0034] (1) Animal preparation:

[0035] Eight-week-old wild-type male C57BL / 6J mice, weighing 18g–20g, were selected from Vital River Pharmaceuticals and housed in the animal facility of Beijing Century Altar Hospital under sterile, free-range diet. The ambient temperature was 22℃–25℃, and the humidity was 40%–60%.

[0036] (2) Group modeling:

[0037] The animals were divided into a control group (comparative sample) and a model group (example sample), with 8 animals in each group.

[0038] The control group mice were fed a normal rodent diet for 28 days.

[0039] The model group mice were fed a diet containing 0.2% adenine synthesized by Medison for 28 days. Adenine was purchased from Sigma-Aldrich in the United States.

[0040] (3) Indicator detection:

[0041] The cardiac function of mice was assessed using the Vevo 2100 small animal ultrasound imaging system, and cardiac images were acquired. (See [link to documentation]). Figure 1This figure mainly shows the ventricular wave groups of the hearts of mice in each group (which can show the motion curves of the ventricular wall and interventricular septum). By measuring parameters such as the amplitude and slope of the curves, cardiac function indicators such as ventricular wall thickness, ventricular cavity size, EF, and FS can be accurately calculated, which is an important basis for quantitative assessment of cardiac function. Based on the test results, ejection fraction, cardiac output, systolic interventricular septal thickness (IVS; s), diastolic interventricular septal thickness (IVS; d), systolic left ventricular diameter (LVID; s), diastolic left ventricular diameter (LVID; d), systolic left ventricular volume (Lv vol; s), and diastolic left ventricular volume (Lvvol; d) can be calculated.

[0042] (4) Observation of fibrosis in mouse heart tissue by masson staining:

[0043] Paraffin sections were dewaxed and hydrated, then stained with mordant at 37°C for 1 hour, and rinsed with tap water until the yellow color disappeared; stained with azurite blue for 5 minutes, rinsed briefly with tap water, stained with fuchsin for 10 minutes, and rinsed briefly with tap water; then quickly rinsed in 70%, 80%, 90%, and 95% alcohol for 30 seconds in sequence, and finally settled in 100% alcohol to stop the color separation; dehydrated by applying 100% alcohol three times for 5 minutes each time; cleared by applying xylene twice for 5 minutes each time; and then mounted with neutral resin and observed and photographed under an optical microscope.

[0044] Test Result Analysis:

[0045] On day 29, the body weight of the mice was measured, and it was found that the body weight of the model group mice was significantly reduced (see...). Figure 6 ).

[0046] Cardiac function tests revealed that the model group mice exhibited significant myocardial hypertrophy and cardiac chamber stenosis, increased ejection fraction, and decreased cardiac output (see [link to relevant documentation]). Figure 1-3 Increased interventricular septal thickness during systole and diastole, decreased left ventricular diameter during systole and diastole, and decreased left ventricular volume during systole and diastole (see [link to relevant documentation]). Figure 4 ),in, Figures 2-4Results are expressed as mean ± standard deviation. * indicates P < 0.05, ** indicates P < 0.01, *** indicates P < 0.001, and **** indicates P < 0.0001.

[0047] Masson staining revealed mild cardiac fibrosis in the model group mice (see...). Figure 5 This invention demonstrates the successful construction of a heart failure model with abnormal ejection fraction. This model provides a practical research method for studying the pathogenesis of human heart failure with abnormal ejection fraction and exploring new treatment methods.

[0048] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for establishing a mouse model of heart failure with abnormal ejection fraction, characterized in that, Mice were fed an adenine-containing diet for 28 days, with 5-10g of adenine-containing animal feed per day. The adenine-containing diet is an animal feed containing adenine; the mass fraction of adenine in the adenine-containing animal feed is 0.20%. The animal feed is rodent feed; by mass fraction, the rodent feed comprises: 20% casein; 39.75% corn starch; 13.2% corn dextrin; 10% sucrose; 5% cellulose; 7% soybean oil; 3.5% compound minerals; and 1% compound vitamins. The mice were wild-type male C57BL / 6J mice.

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