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Method for realizing efficient differentiation of induced pluripotent stem cells towards mature endothelial cells

A technology of pluripotent stem cells and endothelial cells, applied in the field of efficient differentiation of induced pluripotent stem cells to mature endothelial cells, can solve the problems of imperfect endothelial function, lack of tube-forming ability, long induction period, etc., and achieve significant in vitro tube-forming ability , Improve the efficiency of endothelial transformation and reduce the difficulty of operation

Active Publication Date: 2018-08-10
ALLIFE MEDICAL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these induction methods also have certain limitations, such as long induction period (generally 9-20 days), low conversion efficiency (20%-60%), cumbersome purification process (screening by MACS or flow cytometry), Insufficient endothelial function (lack of tube-forming ability in vitro or in vivo), etc.

Method used

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  • Method for realizing efficient differentiation of induced pluripotent stem cells towards mature endothelial cells
  • Method for realizing efficient differentiation of induced pluripotent stem cells towards mature endothelial cells
  • Method for realizing efficient differentiation of induced pluripotent stem cells towards mature endothelial cells

Examples

Experimental program
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Effect test

Embodiment 1

[0097] The method for highly efficient differentiation of induced pluripotent stem cells into mature endothelial cells in this embodiment, the specific process is as follows figure 1 shown, including the following steps:

[0098] Step 1: Pretreat the induced pluripotent stem cells to obtain a cell suspension. The specific method is:

[0099] In the cell culture plate, the induced pluripotent stem cells were revived and subcultured, washed twice with PBS, and then washed every 10cm 2 The ratio of adding liquid to the bottom area of ​​the cell culture plate is 0.5mL, add the cell digestion solution preheated at 37°C for enzymatic digestion, and obtain the enzymatic digestion solution; when 70-80% of the induced pluripotent stem cells start to fall off , adding 5 times the volume of DMEM / F12 medium of the above enzymatic digestion solution for neutralization, and then transferring the induced pluripotent stem cells to a centrifuge tube. Wherein, the above-mentioned DMEM / F12 me...

Embodiment 2

[0117] The method for highly efficient differentiation of induced pluripotent stem cells into mature endothelial cells in this embodiment, the specific process is as follows figure 1 shown, including the following steps:

[0118] Step 1: Pretreat the induced pluripotent stem cells to obtain a cell suspension. The specific method is:

[0119] In the cell culture plate, the induced pluripotent stem cells were revived and subcultured, washed twice with PBS, and then washed every 10cm 2 The ratio of adding liquid to the bottom area of ​​the cell culture plate is 0.5mL, add the cell digestion solution preheated at 37°C for enzymatic digestion, and obtain the enzymatic digestion solution; when 70% of the induced pluripotent stem cells start to fall off, add The DMEM / F12 medium with 5 times the volume of the above enzymatic digestion solution was neutralized, and then the induced pluripotent stem cells were transferred to a centrifuge tube. Wherein, the above-mentioned DMEM / F12 me...

Embodiment 3

[0136] The method for highly efficient differentiation of induced pluripotent stem cells into mature endothelial cells in this embodiment, the specific process is as follows figure 1 shown, including the following steps:

[0137] Step 1: Pretreat the induced pluripotent stem cells to obtain a cell suspension. The specific method is:

[0138] In the cell culture plate, the induced pluripotent stem cells were revived and subcultured, washed twice with PBS, and then washed every 10cm 2 The ratio of adding liquid to the bottom area of ​​the cell culture plate is 0.5mL, add the cell digestion solution preheated at 37°C for enzymatic digestion, and obtain the enzymatic digestion solution; when 80% of the induced pluripotent stem cells start to fall off, add The DMEM / F12 medium with 5 times the volume of the above enzymatic digestion solution was neutralized, and then the induced pluripotent stem cells were transferred to a centrifuge tube. Wherein, the above-mentioned DMEM / F12 me...

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Abstract

The invention discloses a method for realizing efficient differentiation of induced pluripotent stem cells towards mature endothelial cells, belonging to the technical field of cell engineering. The method comprises the following steps: carrying out pretreatment culturing on the induced pluripotent stem cells; carrying out induction to form side plate mesoblastic cells; carrying out inducing to form endothelial progenitor cells tending to the endothelial cells; and carrying out inducing to form the mature endothelial cells. According to the method, various cell inducing factors are matched through increase and decrease, meanwhile, the reasonable upgrading is carried out on the inducing method, thus the directional differentiation of the induced pluripotent stem cells towards the endothelial cells can be efficiently induced, under the condition that a small number of induced pluripotent stem cells are adopted and the purifying operation is not needed, a large number of mature endothelial cells with high purity and complete functions are obtained.

Description

technical field [0001] The invention relates to a method for efficiently differentiating induced pluripotent stem cells into mature endothelial cells, belonging to the technical field of cell engineering. Background technique [0002] Vascular Endothelial Cells (vEC) is a layer of specialized squamous epithelial cells that make up the inner wall of blood vessels, and is an important guarantee for maintaining the stability of the blood circulation system throughout the body. Diabetes, hypertension and other chronic pathologies are often accompanied by endothelial cell dysfunction or damage, which can easily induce cardiovascular diseases such as atherosclerosis and even myocardial infarction, seriously threatening the lives and health of patients. Therefore, regulating the repair or replacement of damaged vascular endothelium has important clinical value for the prevention and treatment of cardiovascular diseases. Studies have pointed out that mature vascular endothelial cel...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N5/10
CPCC12N5/069C12N2501/727C12N2506/45
Inventor 顾雨春张会远谭声江
Owner ALLIFE MEDICAL SCI & TECH CO LTD
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