Search

CN-121970720-A - Construction method and application of spontaneous sustained ventricular tachycardia animal model

CN121970720ACN 121970720 ACN121970720 ACN 121970720ACN-121970720-A

Abstract

The invention relates to a construction method and application of a spontaneous sustained ventricular tachycardia animal model, in particular to a spontaneous sustained ventricular tachycardia rat model based on MYL4 gene defect combined myocardial infarction, which utilizes the synergistic effect of MYL4 gene conservation gene defect and dual pathological factors of acquired myocardial infarction to construct the model, wherein the induction rate is more than or equal to 90 percent, and the spontaneous duration time is 1-3 min, which is obviously superior to the existing construction method of the spontaneous sustained ventricular tachycardia model. The model can simulate the pathophysiological process of ventricular tachycardia after the cardiovascular diseases related to the MYL4 genes are combined with myocardial infarction, provides a general technical support for ventricular arrhythmia pathogenesis research, drug screening and medical instrument research and development, and has wide scientific research and clinical conversion values.

Inventors

  • LI HAILING
  • SONG PEIRAN
  • ZHONG YUAN
  • PENG WENHUI

Assignees

  • 上海市第十人民医院

Dates

Publication Date
20260505
Application Date
20251230

Claims (10)

  1. 1. A method for constructing an animal model of arrhythmia, comprising the steps of: Step A, selecting a conserved arrhythmia related gene based on a target species, and constructing and selecting a gene defect homozygote animal model; B, constructing a myocardial damage animal model by adopting the homozygous gene defect animal model obtained in the step A; step C, screening arrhythmia animal models meeting the standard through phenotype monitoring; wherein the arrhythmia animal model comprises a atrial flutter arrhythmia animal model, a ventricular flutter arrhythmia animal model, a spontaneous sustained ventricular tachycardia animal model, a slow arrhythmia animal model and a atrial reentry tachycardia animal model.
  2. 2. The construction method according to claim 1, wherein the conserved arrhythmia-related genes comprise SCN5A, KCNQ1, KCNH2, CACNA1C, GJA1, RYR2, CASQ2, MYL4, and/or, The construction of the myocardial injury animal model is selected from one of permanent coronary artery ligation model construction, drug/chemical induction model construction, electric stimulation induction model construction, ischemia/reperfusion injury model construction, physical/mechanical injury model construction and genetic engineering model construction.
  3. 3. The method of constructing according to claim 1, wherein the step of constructing the spontaneous sustained ventricular tachycardia animal model includes: S1, constructing an MYL4 gene defect animal model, breeding and feeding, and selecting an MYL4 gene defect homozygote animal model; S2, constructing a myocardial infarction animal model by adopting the MYL4 gene defect homozygote animal model obtained in the step S1 and implementing left anterior descending coronary artery ligation; And S3, after the myocardial infarction animal model in the step S2 is constructed, carrying out long-term electrocardiographic monitoring on the animal, and screening out a spontaneous sustained ventricular tachycardia animal model induced by MYL4 gene defects combined with myocardial infarction according to monitoring results.
  4. 4. A method of construction according to claim 3, wherein the animal comprises a rodent, a lagomorpha, a canine, a porcine.
  5. 5. The method according to claim 3, wherein the animal is a rat, and wherein in step S1, the MYL4 gene-deficient animal model is a MYL4 gene-deficient rat model comprising MYL4 gene-knockout rats and MYL4 functional-deficient rats resulting from gene editing.
  6. 6. The construction method according to claim 5, wherein in step S1, the MYL4 gene-deficient homozygous rat model is a homozygous MYL4p.E11K point mutant rat model, and/or, In the step S1, the expression level of MYL4 genes of the MYL4 gene defective homozygote rat model is reduced by more than 90% compared with the wild type, and the induction condition of the ventricular tachycardia is satisfied.
  7. 7. The construction method according to claim 5, wherein in the step S2, the construction method of the myocardial infarction rat model comprises the steps of fixing the rat on an operating table after anesthesia, separating the trachea and intubation, connecting a small animal breathing machine, separating muscles layer by layer from the 4 th-5 th intercostal incision on the left chest, opening the chest cavity, exposing the heart, finding the left anterior descending branch of the coronary artery between the left atrial appendage and the pulmonary artery cone, ligating the left anterior descending branch near the trunk of the left coronary artery by using a non-invasive cotton suture, observing that the color of the anterior wall of the left ventricle becomes pale and the myocardial movement is weakened, namely ligating is successful, closing the chest cavity layer by layer, removing the trachea after spontaneous respiration recovery of the rat, and preventing infection after operation.
  8. 8. The method according to claim 5, wherein in the step S3, the screening is judged by monitoring for 3 minutes under anesthesia at 1-4 weeks after the operation, continuously 3 or more ventricular premature beats and continuously 30S or more, judging as spontaneous continuous ventricular tachycardia, and/or, The construction method further comprises a step S4 of carrying out verification analysis on the self-sustained ventricular tachycardia animal model obtained by construction, wherein the verification comprises functional verification and/or pathological verification, the functional verification comprises ultrasonic cardiography detection of left ventricular ejection fraction, left ventricular end diastole inner diameter and left ventricular end systole inner diameter, and the pathological verification comprises HE staining observation of myocardial necrosis and Masson staining detection of fibrosis.
  9. 9. The construction method according to claim 5, wherein the construction success rate of the spontaneous sustained ventricular tachycardia rat model is equal to or more than 80%, the arrhythmia induction rate is equal to or more than 90%, and the ventricular tachycardia is 80%.
  10. 10. The construction method according to any one of claims 1 to 9 or the application of an arrhythmia animal model obtained by construction thereof, wherein the application is at least one selected from the group consisting of application of the arrhythmia animal model in determining a molecular mechanism for synergistically inducing arrhythmia by gene defect and myocardial injury, application of the arrhythmia animal model in screening anti-arrhythmia drugs, and application of the arrhythmia animal model in preparing an arrhythmia therapeutic medical device.

Description

Construction method and application of spontaneous sustained ventricular tachycardia animal model Technical Field The invention relates to the technical field of medical animal models, in particular to a method for constructing a spontaneous sustained ventricular tachycardia animal model and application thereof, and specifically relates to a method for constructing a spontaneous sustained ventricular tachycardia animal model induced by combining MYL4 gene defects and myocardial infarction and application thereof. Background Ventricular tachycardia is a serious arrhythmia, especially sustained ventricular tachycardia, which can often lead to heart failure, cardiogenic shock and even sudden death, and seriously threatens human life and health. Clinically, myocardial infarction is one of the important causes of inducing ventricular tachycardia, and a gene defect may also increase the risk of developing ventricular tachycardia. However, there are few studies on myocardial infarction-induced spontaneously sustained ventricular tachycardia at present, mainly due to the lack of ideal animal models capable of simulating the pathological process. In the prior art, the construction method of the ventricular tachycardia animal model mainly comprises a drug induction method, an electric stimulation method, a myocardial infarction model method and the like. The drug induction method generally uses aconitine, ouabain and other drugs, and can rapidly induce ventricular tachycardia, but has poor model stability and can not simulate related pathological mechanisms, the electric stimulation method induces ventricular tachycardia by in vitro electric stimulation heart, has short model duration and is difficult to meet long-term research requirements, and the simple myocardial infarction model can simulate arrhythmia after myocardial infarction, but only has 10%（Oknińska M, Paterek A, Bierła J, Czarnowska E, Mączewski M, Mackiewicz U. Effect of age and sex on the incidence of ventricular arrhythmia in a rat model of acute ischemia. Biomed Pharmacother. 2021 Oct;142:111983.）, induction rate which can be increased to 50% after electric stimulation is generally required to be overlapped, and has a duration <10 s（Jiao KL, Li YG, Zhang PP, Chen RH, Yu Y. Effects of valsartan on ventricular arrhythmia induced by programmed electrical stimulation in rats with myocardial infarction. J Cell Mol Med. 2012 Jun;16(6):1342-51.）. The MYL4 gene is located on human chromosome 17, and the encoded protein is "atrial myosin light chain-1", which is critical for heart development and contractile function, and is highly conserved in mammalian myocardial tissue. In recent years, it has been found that mutation of a gene encoding a myocardial structural protein (such as MYL 4) can cause severe arrhythmia by changing the electrophysiological properties of the heart, but whether the gene is related to spontaneous sustained ventricular tachycardia is unknown, and no report of constructing an animal model by combining MYL4 gene mutation with myocardial injury modeling is currently involved. Therefore, the method for constructing the spontaneous sustained ventricular tachycardia animal model with high induction rate and high stability of dual pathological factors of congenital gene defects combined with acquired myocardial infarction attack is significant for deeply researching pathogenesis of related arrhythmia and developing effective therapeutic drugs and medical appliances. Disclosure of Invention In order to overcome at least one defect in the prior art, such as the defect that a ventricular tachycardia animal model cannot stably simulate continuous spontaneous ventricular tachycardia in the prior art, the invention provides a spontaneous continuous ventricular tachycardia animal model induced by combining MYL4 gene defects and myocardial infarction, and a construction method and application thereof, and the stability and success rate of continuous ventricular tachycardia animal model construction are greatly improved. In order to achieve the above purpose, the invention adopts the following technical scheme: the first aspect of the present invention provides a method for constructing an animal model of arrhythmia, comprising the steps of: Step A, selecting a conserved arrhythmia related gene based on a target species, and constructing and selecting a gene defect homozygote animal model; B, constructing a myocardial damage animal model by adopting the homozygous gene defect animal model obtained in the step A; step C, screening arrhythmia animal models meeting the standard through phenotype monitoring; Wherein the arrhythmia animal model comprises a atrial flutter arrhythmia animal model, a ventricular fibrillation animal model, a spontaneous sustained ventricular tachycardia animal model, a slow arrhythmia animal model and a atrial reentry tachycardia animal model. Further, the conserved arrhythmia-related genes include SCN5A (sodium channel): long