CN-118985533-B - Method for establishing antigen-specific T cell-induced type 1 diabetes animal model

Abstract

The invention provides a preparation method of a type 1 diabetes non-human mammal model. Specifically, the invention induces islet beta cell destruction in a non-immunodeficiency mouse by using the T cells of the specific targeting GAD, constructs T cell autoimmune type 1 diabetes which accords with the actual pathogenesis, and has application prospect in the field of diabetes research.

Inventors

• ZHOU ZIKAI
• HU DENGHUI

Assignees

• 中国科学院上海药物研究所

Dates

Publication Date

20260505

Application Date

20230516

Claims (6)

1. 1. A method of making a non-human mammalian model of type 1 diabetes mellitus, wherein said animal is a non-immunodeficient animal, and said method comprises administering to said animal a specific GAD-targeted CD8 + T cell and streptozotocin; the T cells of the specific targeting GAD specifically bind to an epitope peptide shown as AFLHATDLL (SEQ ID NO. 1); And the animal is a mouse; Wherein, adoptive transfer 200 μl containing 5×10ζ6T cells is injected into mice via tail vein, and 40 mg/kg streptozotocin solution is injected intraperitoneally at day 2 and 3, respectively.
2. 2. The method of claim 1, wherein the animal is a C57BL/6 mouse.
3. 3. The method of claim 1, wherein the T cells are prepared by: providing a population of cells immunostimulatory by a GAD antigenic peptide, isolating T cells from the population of cells; the immunostimulation includes stimulation with a GAD antigen peptide having an amino acid sequence shown as AFLHATDLL (SEQ ID NO. 1).
4. 4. Use of an animal model prepared by the method of claim 1 for studying the pathogenesis of T1 DM.
5. 5. Use of an animal model prepared by the method of claim 1 for screening or identifying substances that prevent or treat T1 DM.
6. 6. A composition for preparing a T1DM non-human mammalian model according to the preparation method of claim 1, wherein the composition comprises: m1) CD8 + T cells specifically targeted to GAD, and M2) streptozotocin; wherein the T cells specifically targeting GAD specifically bind to an epitope peptide as shown in AFLHATDLL (SEQ ID NO. 1); and the animal is a mouse.

Description

Method for establishing antigen-specific T cell-induced type 1 diabetes animal model Technical Field The present invention relates to the field of biological models. In particular, the invention relates to a method for establishing an antigen-specific T cell-induced type 1 diabetes animal model. Background Type 1diabetes (Type 1diabetes mellitus,T1DM) is an organ-specific autoimmune disease characterized by inflammatory infiltrates of the islets and by the constant destruction of autoreactive immune cells, especially T cells, to islet beta cells, which results in the inability to produce sufficient insulin to maintain homeostasis of blood glucose metabolism, hyperglycemia, etc. T1DM animal models have greatly driven research on T1DM pathogenesis, drug development, and the like. Among the commonly used research models for T1DM are NOD mouse models and chemical substances, such as Streptozotocin (STZ) or tetraoxypyrimidine (alloxan), injection-induced T1DM models. The T1DM phenotype of NOD mice is caused by genetic defects, and the mice exhibit multiple immunophenotype abnormalities, including defects in antigen presenting cell immunoregulatory function, defects in T-strangulation Baku, NK cell function, and the like, and thus cannot truly mimic the pathology of the T1DM population. The injection of STZ and other medicines induces T1DM model mainly because the chemical substances directly destroy beta cells, but not autoimmune causes T1DM. Therefore, to better advance related research and drug development efforts, there remains a need in the art to develop methods for constructing T1DM animal models that can mimic the pathological processes of T1DM populations in non-immunodeficient wild animals. Disclosure of Invention The invention aims to provide a method for constructing a wild type T1DM animal model. In a first aspect of the invention, there is provided a method of preparing a model of a non-human mammal having type 1 diabetes mellitus, said animal being a non-immunodeficient animal, and said method comprising administering to said animal T cells specifically targeted to GAD. In another preferred embodiment, the animal is a rodent or primate, preferably comprising a mouse, rat, rabbit, monkey. In another preferred embodiment, the animal is a C57BL/6 mouse. In another preferred embodiment, the T cells are prepared by the following method: Providing a population of cells that are immunostimulatory with a GAD antigenic peptide, and isolating T cells from the population of cells. In another preferred embodiment, the immunostimulation comprises in vivo immunostimulation, or in vitro immunostimulation. In another preferred embodiment, the immunostimulation comprises stimulation with a GAD antigen peptide having an amino acid sequence as shown in AFLHATDLL (SEQ ID NO. 1) or YAFLHATDL (SEQ ID NO. 2). In another preferred embodiment, the GAD-specific T cells specifically bind an epitope peptide as shown in AFLHATDLL (SEQ ID NO. 1). In another preferred embodiment, the T cell is a CD4 + T cell, a CD8 + T cell, or a combination thereof. In another preferred embodiment, the T cell is a CD8 + T cell. In another preferred embodiment, the T cells are used in an amount of no more than 5X 10. Sup.6 cells/animal, preferably no more than 3X 10. Sup.6 cells/animal, more preferably no more than 1X 10. Sup.6 cells/animal. In another preferred embodiment, the method further comprises administering a compound selected from the group consisting of: STZ, tetraoxypyrimidine, or a combination thereof. In another preferred embodiment, the total amount of STZ administered is no more than 300mg/kg body weight, preferably no more than 200mg/kg body weight, more preferably no more than 150mg/kg body weight, more preferably no more than 80mg/kg body weight. In another preferred embodiment, the STZ is administered at a single dose of no more than 80mg/kg body weight, preferably no more than 60mg/kg body weight, more preferably no more than 40mg/kg body weight. In another preferred embodiment, the number of islet beta cells is reduced, preferably by more than 50%, more preferably by more than 80% in said animal. In another preferred embodiment, the decrease in islet beta cell number is caused by T cell specific killing. In a second aspect of the invention there is provided the use of an animal model prepared by a method according to the first aspect of the invention for studying T1DM pathogenesis. In a third aspect of the invention there is provided the use of an animal model prepared by a method according to the first aspect of the invention for screening or identifying substances which prevent or treat T1 DM. In a fourth aspect of the invention, there is provided a method of making a GAD-specific T cell, the method comprising the steps of: 1) Providing a population of GAD immunostimulatory cells, isolating a population of T cells from the population of cells; 2) T cells that specifically bind to the antigenic peptide shown in SEQ ID NO.1