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Method for establishing human arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model

A technology for arrhythmia and disease models, applied in the field of biomedicine, can solve problems such as application limitations, difficulty in obtaining, and harsh in vitro culture conditions

Inactive Publication Date: 2019-03-01
ZHEJIANG UNIV
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Problems solved by technology

For the study of cardiovascular diseases, primary human cardiomyocytes are the most ideal cell resources, but they are difficult to obtain and the conditions of in vitro culture are harsh, so their application is greatly limited

Method used

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  • Method for establishing human arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model
  • Method for establishing human arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model
  • Method for establishing human arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model

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Embodiment Construction

[0023] 1) Isolation and primary culture of skin fibroblasts: After obtaining the skin, excise the fat tissue and cut the skin tissue into small pieces, rinse with PBS+1% penicillin and streptomycin once, absorb the PBS and add collagen The enzyme was digested at 37°C for 6 hours. After the digestion, DMEM+10% FBS+1% penicillin and streptomycin medium was added and cultured in an incubator. Wait for the skin group to adhere to the wall and change the medium every 4 days. Never blow the skin tissue, fibroblasts can be observed after 1 week.

[0024] 2) Establishment of ARVC patient-specific iPS cell line: fibroblasts were divided into 1×10 4 ,2×10 4 ,3×10 4 ,4×10 4 , 6×10 4 ,8×10 4 Gradient inoculated in a 48-well plate, and after observation on the second day, select cells that were just overgrown in 1 well to infect with Sendai virus. After 24 hours, replace with DMEM+10%FBS+1% penicillin and streptomycin medium, and change the medium every other day On day 7, the cells ...

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Abstract

The invention discloses a method for establishing a human arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model. The method comprises the steps that primary culture is carried out on the skin of a patient with the ARVC to obtain fibroblasts, and specific induced pluripotent stem cells (iPSC) of the patient with the ARVC are obtained by non-integrated Sendai virus induction, and thendirectionally differentiated into specific cardiomyocytes of the patient with ARVC so that a stable in-vitro cardiomyocyte disease model can be established. The method can establish the stable humanARVC model based on an iPSC technology; the morphology and function of the model are both in accordance with the pathological characteristics of the ARVC, and the model is suitable for the research ofARVC related pathological mechanisms and individualized drug screening.

Description

technical field [0001] The invention belongs to the field of biomedical technology, and discloses a method for establishing a "human-derived" arrhythmogenic right ventricular cardiomyopathy (ARVC) disease model, which can be used for the research of related pathological mechanisms of ARVC and the research and development of therapeutic and preventive drugs . Background technique [0002] Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a cardiomyopathy that is more common in young and middle-aged men, with a male to female ratio of about 2.7:1. The pathological features of the disease are myocardial atrophy and fibrofatty tissue replacement in the right ventricle, and the clinical manifestations are often right ventricle enlargement, arrhythmia, sudden death and heart failure, and it is a common cause of death in exercise-induced sudden death. 30% to 50% of ARVC cases are familial, mainly manifested as autosomal dominant inheritance, with variable penetrance. Curr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077C12N5/074
CPCC12N5/0657C12N2500/90C12N2501/33C12N2506/45C12N2533/50
Inventor 梁平沈嘉希蒋晨阳孙雅逊苏俊宫庭钰
Owner ZHEJIANG UNIV
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