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Application of CXCL8-CXCR1 of tree shrew in treatment of rheumatoid arthritis

A technology for arthritis and rheumatoid arthritis, applied in the application field of tree shrew CXCL8-CXCR1 in the treatment of rheumatoid arthritis, which can solve the problems of patients with adverse reactions, intolerance, and incomplete understanding of the pathological mechanism of rheumatoid arthritis

Inactive Publication Date: 2015-11-18
YUNNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Many studies have shown that abnormal cellular immunity and humoral immunity mediated by a variety of inflammatory cells (lymphocytes, neutrophils, macrophages, synoviocytes, etc.) can lead to local and systemic immune dysfunction in joints (Grespanetal.2008 ), but the pathological mechanism of rheumatoid arthritis is still not completely clear, and there is a lack of effective clinical treatment
At present, the commonly used drugs for clinical treatment of rheumatoid arthritis are divided into four categories, namely, nonsteroidal anti-inflammatory drugs (nonsteroidal anti-inflammatory drugs, NSAIDs), immunosuppressants, glucocorticoids and herbal medicines (Chinese Medical Association Rheumatology Society. 2004), such as methotrexate (MTX), azathioprine, cyclophosphamide, etc., but long-term use of these drugs will cause adverse reactions in many patients, resulting in intolerance and discontinuation

Method used

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  • Application of CXCL8-CXCR1 of tree shrew in treatment of rheumatoid arthritis
  • Application of CXCL8-CXCR1 of tree shrew in treatment of rheumatoid arthritis
  • Application of CXCL8-CXCR1 of tree shrew in treatment of rheumatoid arthritis

Examples

Experimental program
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Effect test

Embodiment 1

[0024] Example 1: Cloning and analysis of tree shrew CXCL8 and CXCR1 genes.

[0025] 1. The normally raised healthy tree shrews were anesthetized with ether until lethargic, and their necks were cut to death, and 15 kinds of tissues (stomach, lung, kidney, liver, thymus, spleen, brain, heart, adrenal gland, bladder) were collected in a sterile ultra-clean bench. , thyroid, pancreas, lymph node, bone, whole blood) were aliquoted into sterile 1.5ml centrifuge tubes, RNA was extracted from the tissue, and the extracted RNA was reverse-transcribed into cDNA using Takara's PrimeScriptRTreagentKitwithcDNAEraser (PerfectRealTime). Select the predicted tree shrew CXCL8 (XM_006142918.1) and CXCR1 (XM_006157465.1) genes to design primers as follows:

[0026] tsCXCL8:Forward:5'-TCTCGTGTAAACATGACTTCCAAGC-3';

[0027] Reverse: 5'-TTATGCATCTTGCTTCTCAGCTTTC-3';

[0028] tsCXCR1:Forward:5'-ATGTGGAACATCACTGATCTGTGGA-3';

[0029] Reverse: 5'-TCAAAGGTTTGAAGAGGTATTGATA-3'.

[0030] The comple...

Embodiment 2

[0040] Example 2: Functional analysis of tree shrew chemokines CXCL8 and CXCR1.

[0041] 1. Get 4 ml of heparin anticoagulated peripheral blood from 4 healthy male tree shrews (body weight 130 ± 20 g, 4-6 months old) in a 15 ml centrifuge tube respectively, and use mouse lymphocyte separation medium Ficoll-Hypaque (P8620, Beijing Suolaibao Biotechnology Company) isolated tree shrew whole blood total lymphocytes. CD3+T cells, CD19+B cells and CD11b+ neutrophils (MiltenyiBiotec, Germany) were sorted by immunomagnetic bead sorting, and RNA was extracted, cDNA was synthesized by reverse transcription, and CXCR1 was analyzed in lymphocytes, CD3+T cells , CD19+ B cells and CD11b+ neutrophils ( Figure 4 A).

[0042] 2. Take 700ul of peripheral blood from 3 tree shrews and put them into 1.5ml clean centrifuge tubes, centrifuge at 12000rpm at 4°C for 5min, separate into two layers (the upper layer is serum, the lower layer is red blood cells), and transfer the supernatant to another...

Embodiment 3

[0047]Example 3: Establishment of tree shrew rheumatoid arthritis model.

[0048] 1. Select a total of 20 healthy male tree shrews with a body weight of (130±20) g, 4-6 months old, and divide them into equal groups according to the principle of weight equalization, that is, 10 control groups and 10 collagen-induced arthritis groups. Then the lyophilized II collagen was dissolved in 0.1 mol / L glacial acetic acid solution with a concentration of 2 mg / ml, and stirred overnight at 4°C. Put II collagen solution and an equal volume of complete Freund's adjuvant CFA in an ice bath until completely emulsified, and vortex with a shaker until homogenized.

[0049] 2. From the first day (marked as day0) to the tree shrews in the collagen-induced arthritis group, each tree shrew was injected with 0.25ml of CII / CFA emulsifier from the base of the tail with a concentration of 1mg / ml, and the control group was injected with the same volume of normal saline. In addition, on the seventh day (...

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Abstract

The invention identifies a chemotactic factor CXCL8 gene and a chemokine receptor CXCR1 gene of a tree shrew and proteins encoded by the two genes, and determines that a signal path formed by CXCL8 and CXCR1 participates in adjustment of peripheral blood lymphocyte migration functionally. In a tree shrew CIA (collagen induced arthritis) animal model, reduction of the concentration of CXCL8 in peripheral blood or inhibition of the function of the receptor CXCR1 can effectively reduce migration of immune lymphocyte cells to peripheral bones and joints, and the effect of treatment of rheumatoid arthritis is achieved. A therapeutic molecular target is provided for a patient suffering from clinic rheumatoid arthritis.

Description

technical field [0001] The present invention relates to the expression level, distribution and physiological function of tree shrew (Treeshrew) CXCL8 and CXCR1 proteins in the tree shrew immune system; relates to the regulation of the tree shrew CXCL8-CXCR1 signal pathway on the migration of peripheral lymphocytes and its role in tree shrew collagen-induced joints CIA model and its application in the treatment of rheumatoid arthritis (RA). Background technique [0002] The tree shrew (Treeshrew) belongs to the tree shrew family of the order Climbing shrews, and is a small mammal similar in size to a squirrel. It is mainly distributed in South Asia, Southeast Asia and southern my country (Fane et al. 2013). There are 1 genera, 1 species and 6 subspecies of tree shrews in my country, and 1 species is Burmese tree shrew ( Tupaiabelangeri ), the tree shrew has a body length of 14-23cm and a body weight of 80-200g. The taxonomic status of tree shrews has been controversial, and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/54C12N15/24A61K38/20A61P19/02A61P29/00
CPCC07K14/5421A61K38/00
Inventor 李梅章丁磊蒋宗敏
Owner YUNNAN UNIV
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