Inducing agent and medium for inducing directed differentiation of embryonic stem cells into cardiomyocytes

A technology of embryonic stem cells and differentiation medium, which is applied in the field of induction differentiation medium, can solve the problems of too much expression and too little can not achieve the expected effect, and achieve good curative effect

Active Publication Date: 2016-01-20
罗国安
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0017] The safety of transgenes is one of the first considerations. To do this, the expression of transgenes in specific cells must be precisely controlled. Too much or too little expression will not achieve the desired effect.

Method used

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  • Inducing agent and medium for inducing directed differentiation of embryonic stem cells into cardiomyocytes
  • Inducing agent and medium for inducing directed differentiation of embryonic stem cells into cardiomyocytes
  • Inducing agent and medium for inducing directed differentiation of embryonic stem cells into cardiomyocytes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1: Salvianolic acid B induces directed differentiation of embryonic stem cells CGR8 into cardiomyocytes

[0055] 1. Salvianolic acid B affects the proliferation activity of embryonic stem cells

[0056] Cell line: mouse embryonic stem cell line CGR8 (129 / Ola-derivedwild-type ES cells, ATCC).

[0057] The main experimental reagents: GMEM medium; special grade fetal bovine serum (FBS); mouse leukemia inhibitory factor (LeukaemiaInhibitoryFactor, LIF).

[0058] After digestion and counting of normal cultured CGR8, adjust the concentration of cell suspension to 3×10 3 cells / mL. The cell suspension was added to a 96-well culture plate at 200 μL per well, and incubated in an incubator for 2 hours. After the ES cells adhered to the wall, a complete medium containing salvianolic acid B (see Table 1 for the composition) was added for culture. The complete medium The concentration of salvianolic acid B in the range of 10 -3 ~10 -6 M, the complete medium without salvi...

Embodiment 2

[0080] Example 2: Salvianolic acid B and ginsenoside Rg1 induce embryonic stem cell CGR8 to differentiate into cardiomyocytes

[0081] 1. Salvianolic acid B and ginsenoside Rg1 affect the proliferation activity of embryonic stem cells

[0082] The composition of cell lines, main experimental reagents and complete medium is the same as that described in Example 1.

[0083] Adjust the concentration of CGR8 cell suspension to 3×10 3 cells / mL. The cell suspension was added to a 96-well culture plate, 200 μL per well. Incubate in the incubator for 2 hours. After the ES cells adhere to the wall, add a complete medium containing salvianolic acid B and ginsenoside Rg1 (same as Example 1) for culture. The concentration of salvianolic acid B and ginsenoside Rg1 is: 10 -3 +10 -3 M, 10 -4 +10 -4 M, 5×10 -5 +5×10 -5 M, 10 -5 +10 -5 M, 5×10 -6 +5×10 -6 M, the complete medium without salvianolic acid B and ginsenoside was used as the control group. After 3 days of culture, the p...

Embodiment 3

[0088] Example 3: Salvianolic acid B and total ginsenosides induce embryonic stem cell CGR8 to differentiate into cardiomyocytes

[0089] 1. Salvianolic acid B and total ginsenosides affect the proliferation activity of embryonic stem cells

[0090] The composition of cell lines, main experimental reagents and complete medium is the same as that described in Example 1.

[0091] Adjust the concentration of CGR8 cell suspension to 3×10 3 cells / mL. The cell suspension was added to a 96-well culture plate, 200 μL per well. Incubate in the incubator for 2 hours. After the ES cells adhere to the wall, add complete medium containing salvianolic acid B and total ginsenosides for culture. The concentration of salvianolic acid B and total ginsenosides is: 10 -3 M+1.5g / L (highest), 10 -4 M+0.15g / L (high), 5×10 -5 M+0.078g / L (medium), 10 -5 M+0.015g / L (low) and 5×10 -6 M+0.0078g / L (minimum). The complete medium without drugs was used as the control group. After 3 days of culture,...

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Abstract

The invention discloses an inductive agent for inducing embryonic stem cells to directionally differentiate into myocardial cells and a culture medium. The inductive agent contains salvianolic acid B. The culture medium is prepared through the following method: the inductive agent is added into a differential medium for culturing the embryonic stem cells, wherein the concentration of the salvianolic acid B of the inductive agent in the induced differential medium is 10<-6>-10<-3> moles. The invention further discloses applications of the inductive agent and the culture medium in inducing the embryonic stem cells to directionally differentiate into the myocardial cells. The inductive agent is toxic-free and has a function of inducing the embryonic stem cells to directionally differentiate, the functional myocardial cells after directional differentiation can provide suitable cell donors for myocardial cell transplantation therapy.

Description

technical field [0001] The invention relates to an inducer for inducing embryonic stem cells to differentiate into cardiomyocytes, a medium for inducing differentiation and applications thereof. Background technique [0002] Heart failure caused by cardiomyocyte dysfunction is a major disease worldwide, and the final cause of death in heart failure patients is mainly loss of cardiac pumping function or arrhythmia. The one-year mortality rate of patients with severe heart failure exceeds 50%. Although heart transplantation is possible for terminally ill patients, about 20% of patients die while waiting for organ transplantation due to lack of donor organs. Due to the high morbidity and mortality of heart failure, the death of transplanted hearts, complications such as immune rejection, and the late failure of transplanted hearts, it is urgent to develop new therapeutic methods to improve the function of cardiomyocytes and prevent the occurrence of heart failure. Current the...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0735C12N5/071C12N5/02
Inventor 罗国安王义明
Owner 罗国安
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