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In vitro amplification adjustable T cell method

An in vitro amplification and regulatory technology, applied in the field of biomedicine, can solve the problems of inability to achieve tolerance, prolong the survival of allografts, lack of immunosuppressive function, etc., and achieve the effect of sufficient quantity and great clinical application value

Inactive Publication Date: 2009-12-16
吕凌
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Problems solved by technology

Such as: (1) Blocking co-stimulatory pathways: Although in transplanted animal models, the survival time of allografts can be significantly prolonged by blocking co-stimulatory pathways, but this method cannot achieve sustained and stable tolerance
(2) Immature allogeneic dendritic cells (DC): At present, immature DC induced in vitro can induce antigen-specific T cell anergy in vivo and in vitro in some cases, and in vivo infusion can significantly prolong allogeneic transplantation. survival of the drug, but only short-term tolerance
Regulatory T cells induced by TGF-β in iTreg are currently the most valued, but human Foxp+ T cells induced in vitro have the phenotype of regulatory cells and lack immunosuppressive function [18]

Method used

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  • In vitro amplification adjustable T cell method
  • In vitro amplification adjustable T cell method

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Embodiment 1

[0030] Materials: CD4+ T cell isolation kit (purchased from Miltenyi Biotec, USA), anti-human CD3 (BD ​​Com., USA) coated culture plate (purchased from VWR, USA), systemic immunodeficiency (SCID) mice (purchased from Jackson Laboratory, USA).

[0031] Instruments: magnetic bead separator (Auto MACS from Miltenyi, Germany), flow cytometer (BD company, model: Vantage SE).

[0032] Reagent: Inducer: anti-CD3CD28-beads (the ratio of cells to cells is 1:10, Invitrogen), rhIL-2 (final concentration after addition is 100IU / mL), rhTGF-β (final concentration after addition is 5ng / mL) Purchased from BD Company of the United States and Rapamycin (final concentration after adding was 10nM) was purchased from Wyeth Pharmaceuticals. Amplifier: rhIL-2 (20IU / mL), anti-humanCD28 (1ng / ml), Rapamycin (10nM, Wyeth Pharmaceuticals); culture medium was supplemented with 100U / ml penicillin in complete RPMI-1640 medium, 100μg / ml chain Mycin, 2mM L-glutamic acid, 10mM 4-hydroxyethylpiperazineethanes...

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Abstract

The invention relates to a method for amplifying cells, in particular to an in vitro amplification adjustable T cell method, belonging to the technical field of biomedical science. The invention uses Rapamycin combined with TGF-beta to induce human T cell in vitro to become adjustable T cell having immune suppression function, having the following advantages in : 1. enough quantity; and 2. capability of resisting differentiation towards Th 17 cell. The invention overcomes plural defects of natural adjustable T cell, and has larger curing advantages in curing inflammation diseases, autoimmune diseases and prevention of immunity rejection of organ implantation.

Description

technical field [0001] The invention relates to a method for expanding cells, in particular to a method for expanding regulatory T cells in vitro, and belongs to the technical field of biomedicine. Background technique [0002] For nearly half a century, while immunosuppressive drugs have significantly prolonged the survival time of allografts, they also have a series of serious problems due to their own toxic side effects, high prices, and clinical complications caused by drug-induced non-specific immunosuppression. , greatly restricting the further development of clinical organ transplantation. It has become one of the basic topics in the field of transplantation immunology to reduce or even eliminate immunosuppressants by inducing graft-specific immune tolerance or achieving a "clinically almost tolerated" state similar to the application of immunosuppressants. Although many investigators have successfully induced a state of immune tolerance in allografts in different an...

Claims

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Application Information

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IPC IPC(8): C12N5/08
Inventor 吕凌张峰王学浩
Owner 吕凌
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