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Polypeptide for inducing CD4CD25 regulatory T cells and application

A regulatory and cellular technology, applied in the field of immunology, to achieve the effect of inhibiting inflammatory pathological reactions and broad application prospects

Inactive Publication Date: 2009-11-18
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report on pathogenic polypeptides that can induce Tregs and have immunosuppressive properties in China

Method used

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  • Polypeptide for inducing CD4CD25 regulatory T cells and application
  • Polypeptide for inducing CD4CD25 regulatory T cells and application
  • Polypeptide for inducing CD4CD25 regulatory T cells and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1: After in vivo immunization with peptides or pretreatment of mouse spleen and lymph node cells in vitro, CD4 + CD25 + Foxp3 + Changes in the number of T cells;

[0048] 1) Synthesize Schistosoma japonicum-T cell epitope peptide SJMHE1, its amino acid sequence (SEQ.ID.NO.1) is VPGGGTALLRCIPVLDTLSTKNED(Val Pro Gly Gly Gly Thr Ala Leu Leu Arg Cys Ile ProVal Leu Asp Thr Leu Ser Thr Lys Asn Glu Asp). And synthesize the control peptide OVA 323-339 , its sequence is (SEQ.ID.NO.2): ISQAVHAAEINEAGRY (Ile Ser Gln Ala Val His Ala Ala Glu Ile Asn GluAla Gly Arg Tyr); the synthetic purity of the peptide and the control peptide are both ≥99%. The above-mentioned peptides were all produced by Shanghai Shenggong Company, and the purity of the peptides was greater than 99%, which was identified by HPLC.

[0049] 2) The peptide (SJMHE1), control peptide (OVA 323-339 ), PBS and IFA to make an emulsifier. IFA was purchased from Sigma Company. Each 100 μl emulsifier containe...

Embodiment 2

[0053] Example 2: Peptide immunization in vivo or pretreatment of mouse CD4 in vitro + CD25 + After Tregs cells, the effect on the suppressive function of Tregs;

[0054] 1) Immunize the mice according to the method in the above-mentioned Example 1, separate the spleen and lymph nodes of the immunized mice in each group, make a single cell suspension, lyse the erythrocytes with the erythrocyte lysate, and use Mouse CD4 + CD25 + Regulatory T cell IsolationKit (purchased from Miltenyi Biotec Company, item number: 130-091-041) to isolate mouse CD4 + CD25 + , CD4 + CD25 - T cells; non-CD4 + Cells were treated with 50 μg / ml mitomycin at 37°C, 5% CO 2 After 30 minutes of treatment in the incubator, it is APC. Take 5×10 4 / well purified CD4 + CD25 + T cells, 1×10 5 / well CD4 + CD25 -T cells, 1×10 5 / well APC, add 0.1 μg / ml SJMHE1 or OVA 323-339 , under anti-CD3 stimulation, 37 °C, 5% CO 2 After culturing in the incubator for 3 days, the 3 H incorporation assay for d...

Embodiment 3

[0057] Example 3: Peptide immunization in vivo or pretreatment of mouse CD4 in vitro + CD25 + After Tregs cells, the peptide on CD4 + CD25 + The mechanism of action of Tregs inhibitory effect;

[0058] 1) Immunize the mice according to the method in the above-mentioned Example 1, separate the spleen and lymph nodes of the immunized mice in each group, make a single cell suspension, and after lysing the erythrocytes with the erythrocyte lysate, separate the small cells according to the method in the above-mentioned Example 2. mouse CD4 + CD25 + , CD4 + CD25 - T cells; non-CD4 + Cells were treated with 50 μg / ml mitomycin at 37°C, 5% CO 2 After 30 minutes of treatment in the incubator, it is APC. Add 0.4 μm Millicell (Millipore Company, semi-permeable membrane separator, which can separate cells and only allow soluble molecules to pass through) in the 24-well culture plate, and take 2.5×10 4 / well purified CD4 + CD25 + Add T cells to the upper part of Millicell, 5×10 ...

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Abstract

The invention belongs to the field of immunology, and in particular relates to a polypeptide for inducing CD4CD25 regulatory T cells (Tregs) and with immune-suppression function on schistosome source and application. The amino acid sequence of the polypeptide capable of inducing the CD4CD25 Tregs is VPGGGTALLRCIPVLDTLSTKNED. After in vivo immunization or in vitro pretreatment of the polypeptide on spleen and lymphocyte of a mouse, the number of CD4CD25Foxp3 T cells is remarkably increased; and after the in vivo immunization or the in vitro pretreatment of the polypeptide on the CD4CD25 Tregs cells of the mouse, the suppression function of the CD4CD25 Tregs is remarkably enhanced. The polypeptide provided by the invention can effectively suppress inflammation pathologic reaction, and has wide application prospect.

Description

technical field [0001] The invention belongs to the field of immunology, and in particular relates to a method for inducing CD4 + CD25 + Regulatory T cells, schistosome-derived polypeptides with immunosuppressive effects and applications. Background technique [0002] In recent years, the "hygiene hypothesis" believes that the incidence of allergic diseases and autoimmune diseases in western developed countries has increased significantly, which is due to the reduced probability of infection in childhood. In the long history of co-evolution between humans and pathogens, pathogens have evolved multiple mechanisms to regulate the host's immune regulatory network. One of the most important mechanisms is that pathogen infection can induce the production of regulatory T cells--professional immune cells with immunosuppressive functions, mainly including Trl cells, Th3 cells, CD4 cells induced by peripheral antigen activation. + CD25 + T regulatory cells (Tregs), etc., play an ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/00A61K38/16A61P37/06A61P29/00
Inventor 苏川汪雪峰周莎贺蕾刘丰
Owner NANJING MEDICAL UNIV
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