Application of mitochondrial transplantation in treatment of primary dilated heart disease

A mitochondrial, primary technology, applied in cardiovascular system diseases, medical preparations containing active ingredients, unknown raw materials, etc. Cardiac cavity dilation and improvement of cardiac dysfunction

Pending Publication Date: 2020-07-17
HEFEI KNATURE BIO PHARM CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But so far, there is no relevant record on the treatment of primary dilated heart disease by mitochondrial transplantation

Method used

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  • Application of mitochondrial transplantation in treatment of primary dilated heart disease
  • Application of mitochondrial transplantation in treatment of primary dilated heart disease
  • Application of mitochondrial transplantation in treatment of primary dilated heart disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] According to the experimental road map of the present invention, see figure 1 , follow the steps below:

[0040] Matrigel Coated Petri Dishes

[0041] 1.1 Matrigel (BD, bioscience) is aliquoted and stored at -20°C / -80°C, and the aliquoted Matrigel is thawed at 4°C overnight before use.

[0042] 1.2 Pre-cool Matrigel and DMEM+F12 medium at 4°C, dilute according to the ratio of 800μl / 50ml and mix immediately. This process should be performed quickly to prevent the gel from solidifying. Add 1ml of medium to each well of the 6-well plate.

[0043] 1.3 Seal the culture dish with a parafilm to prevent evaporation, and put it in a 37°C cell incubator for at least 2 hours before using it for subsequent experiments. If it is not used temporarily, store it at 4°C.

[0044] 1.4 Before using the Matrigel-coated 6-well plate, remove the medium in the well plate and add the medium to be used.

[0045] 2. Culture and passage of IPS cells

[0046] 2.1 When the cell density reaches ...

Embodiment 2

[0061] The animal tissue and cell mitochondria preparation methods were operated on ice according to the requirements of the Beyontian animal tissue and cell mitochondria separation kit used, the lysis conditions of the lysate were kept the same, and the number of grinding rods was 40-50 times.

[0062] After the mitochondria were prepared, they were dissolved in PBS and stored on ice. A part of the mitochondrial solution was drawn, diluted and counted to adjust the concentration of the mitochondrial working suspension. Mitochondria were used immediately after preparation and transplanted into animal myocardium within 1 hour. The longer the mitochondria are kept in vitro, the poorer their viability.

Embodiment 3

[0064] Three groups were set up in the experiment: Sham group, DCM group, DCM+mitochondria transplantation group. Adult C57BL / 6 mice were depilated in the chest in advance, anesthetized with 2% isoflurane gas inhalation, and left thoracotomy was performed to separate the pectoralis major from the pectoralis minor, and the pleura and pericardium were opened with hemostats. Extrude the heart and transplant 100 μl at multiple points on the surface of the left ventricle of the myocardium (each 100 μl contains 10 5 mitochondria) normal saline suspension, and intraperitoneal injection of 15 mg / kg doxorubicin at the same time; the same volume of PBS was injected intraperitoneally in the DCM group, and 15 mg / kg doxorubicin was injected intraperitoneally; in the Sham group, only the heart was extruded by thoracotomy, and the same volume of doxorubicin was injected intramuscularly. PBS, no intraperitoneal injection of doxorubicin.

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Abstract

The invention belongs to the field of primary dilated heart disease treatment, and particularly relates to application of mitochondrial transplantation in treatment of primary dilated heart disease. Humanized induced multifunctional stem cells are obtained and differentiated and induced into beating myocardial cells in vitro, human myocardial cell mitochondria are obtained through separation, mouse myocardial mitochondria are separated at the same time, and human mitochondria and mouse mitochondria are separately obtained for later use. A mouse dilated cardiomyopathy model is constructed by intraperitoneal injection of doxorubicin, the human and mouse mitochondria are transplanted at multiple points on the left ventricular myocardial surface layer of a mouse, the cardiac function of the mouse is detected by an echocardiogram after 6 days, and the change of the myocardial structure of the mouse is detected by histopathology. Research results show that in the construction process of a doxorubicin-induced mouse dilated cardiomyopathy model, the transplanted human or mouse myocardial cell mitochondria can significantly enhance myocardial contraction and relaxation functions and inhibitheart double-chamber cardiac cavity dilation.

Description

technical field [0001] The invention belongs to the field of treating primary dilated heart disease, and in particular relates to the application of mitochondrial transplantation in the treatment of primary dilated heart disease. Background technique [0002] Dilated cardiomyopathy (dilated cardiomyopathy, DCM) is a primary myocardial disease, mainly manifested as congestive heart failure caused by enlarged ventricular cavity and decreased systolic function. According to statistics, the mortality rate of dilated cardiomyopathy within one year accounts for 20%, and the mortality rate within four years accounts for 56%. Due to the complex pathogenic factors of dilated cardiomyopathy, in most cases, there is no clear treatment measure, and the clinical use of drugs such as ACE inhibitors, diuretics, β-blockers, digitalis and anticoagulants can only relieve the disease. Delay symptoms of heart failure, prolong the patient's life, 40% of the incidence of heart failure within two...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/34A61P9/04A61P9/00
CPCA61K35/34A61P9/04A61P9/00
Inventor 葛均波孙爱军孙晓垒
Owner HEFEI KNATURE BIO PHARM CO LTD
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