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Method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro

A technology for hematopoietic stem cells and cell differentiation, which is applied in the field of inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro, and can solve problems such as signal stimulation of thymic basal cells

Pending Publication Date: 2021-05-28
CELL VALLEY (NANJING) BIOTECHNOLOGY CO
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the T cells obtained above cannot be further differentiated into T helper cells (Tregs, Th2, etc.)

Method used

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  • Method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro
  • Method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro
  • Method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Material: Human peripheral blood.

[0054] Instrument: flow cytometer, flow cytometer (BD company, model: Vantage SE).

[0055] Reagents: Flow cytometric antibodies (CD3, CD4, CD34, CD38, CD39, Foxp3, etc.) were purchased from eBioscience, rhIL-2 (after adding the final concentration was 100IU / ml), rhIL-15 (10ng / ml), atRA (after adding The final concentration is 10 nM) was purchased from Wyeth Pharmaceuticals. Medium 1 was prepared by adding penicillin 100U / ml, streptomycin 100μg / ml and 50mM dihydroxyethanol to α-MEM medium. Medium 2 is composed of complete RPMI-1640 medium supplemented with 100U / ml penicillin, 100u g / ml streptomycin, 2mM L-glutanoic acid, 10mM 4-hydroxyethylpiperazineethanesulfonic acid, 0.1mM non-essential amino acids, 1mM Sodium pyruvate (purchased from BioSource International) and 50 uM dihydroxyethanol (purchased from Sigma Aldrich) was prepared.

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Abstract

The invention provides a method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro. The method comprises the following specific steps: 1, collecting blood; 2, separating and sorting the blood to obtain CD34+, CD38+ and CD39+; 3, selecting the remaining cells in the step 2 to prepare DC cells; 4, carrying out first induction on the cells obtained by sorting in the step 2; 5, performing induction for the second time; and 6, carrying out third induction to obtain CD4+, CD25+ and Foxp3+ regulatory T cells. The method has the beneficial effects that CD34+, CD38+ and CD39+ cells are separated from human peripheral blood, thymosin [alpha]1, IL-2, IL-3, IL-15, TGF-[beta], atRA, TRAF6 and irradiated DC cells are added, and finally human hematopoietic stem cells are induced into regulatory T cells with an immunosuppression function.

Description

technical field [0001] The invention relates to the field of biotechnology, and more specifically relates to a method for inducing hematopoietic stem cells to differentiate into regulatory T cells in vitro. Background technique [0002] Organ transplantation has become the only effective way to treat end-stage organ failure. After organ transplantation, patients must take immunosuppressive drugs for life to prevent transplant rejection and graft-versus-host disease (GVHD) and other transplant immune diseases. Long-term use of immunosuppressive drugs can induce infection, tumor recurrence, and chronic graft dysfunction. In order to reduce the above side effects, organ transplantation centers in various countries have tried to stop using immunosuppressive drugs, but the stopping of drugs may also lead to diseases such as abnormal liver and kidney function, metabolic disorders and even chronic rejection, and those who successfully stopped using drugs are all reported in indivi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C12N5/0784C12N13/00
CPCC12N5/0637C12N5/0639C12N13/00C12N2506/11C12N2502/1121C12N2501/30C12N2501/22C12N2501/15C12N2501/2302C12N2501/2303C12N2501/2304C12N2500/44C12N2501/2315C12N2500/38C12N2500/32C12N2500/30C12N2501/70
Inventor 吕凌古鉴
Owner CELL VALLEY (NANJING) BIOTECHNOLOGY CO
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