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Diagnosis and treatment for rheumatoid arthritis and osteoarthritis based on SPAST gene

A rheumatoid and osteoarthritis technology, applied in disease diagnosis, anti-inflammatory agents, bone diseases, etc., can solve problems such as difficult diagnosis and difficult identification of clinical features

Active Publication Date: 2018-10-12
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In primary care, the diagnosis of early rheumatoid arthritis is as difficult as other musculoskeletal problems: clinical features are poorly discernible, inflammatory markers such as erythrocyte sedimentation rate and C-reactive protein are not characteristic, and specific markers such as rheumatoid factor ( Seronegative in 31% of patients) and anti-cyclic citrullinated peptide antibodies (anti-CCP, seronegative in 33% of patients) were also commonly seronegative
There are no molecular markers for the differential diagnosis of rheumatoid arthritis and osteoarthritis, and the applicant carried out this study under such a background

Method used

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  • Diagnosis and treatment for rheumatoid arthritis and osteoarthritis based on SPAST gene
  • Diagnosis and treatment for rheumatoid arthritis and osteoarthritis based on SPAST gene
  • Diagnosis and treatment for rheumatoid arthritis and osteoarthritis based on SPAST gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Example 1 Screening of differentially expressed genes in osteoarthritis synovial tissue, rheumatoid arthritis synovial tissue and normal synovial tissue

[0075] The synovial tissue of 5 patients with rheumatoid arthritis came from patients with rheumatoid arthritis who underwent knee replacement or synovectomy in the orthopedics department of the hospital. The diagnosis of all cases met the classification criteria of rheumatoid arthritis revised by the American Rheumatology Association in 1987 , including 3 females and 2 males, with an average age of (55±8) years and an average disease duration of (12±7) years. The synovial tissues of 5 patients with osteoarthritis were obtained from osteoarthritis patients undergoing knee joint replacement or synovectomy in the Department of Orthopedics of the hospital. Age (63±5) years old, mean disease duration (12±8) years. 6 cases of normal synovial tissues came from joint synovial tissues of patients undergoing trauma surgery. ...

Embodiment 2

[0122] Example 2 Large sample verification of differentially expressed genes

[0123] Based on the results of previous high-throughput sequencing and according to the size of the P value, we selected the SPAST gene for verification. The up-regulation was even greater in patients with osteoarthritis.

[0124] 1. Detection of differential expression of SPAST genes at the transcriptional level

[0125] 1. According to the method of Example 1, 100 cases of rheumatoid arthritis synovial tissue, 100 cases of osteoarthritis synovial tissue, and 100 cases of normal synovial tissue were collected.

[0126] 2. Carry out the extraction and quality detection of RNA according to the method of Example 1

[0127] 3. Reverse transcription

[0128] 1 μg of total RNA was reverse-transcribed to synthesize cDNA using reverse transcription buffer. Use 25μl reaction system, take 1μg total RNA for each sample as template RNA, add the following components to the PCR tube respectively: DEPC water,...

Embodiment 3

[0149] Embodiment 3 SPAST gene expression plasmid construction

[0150] 1. Design and synthesis of interference RNA

[0151] According to the SPAST gene sequence, siRNA was designed and synthesized by Shanghai Gemma Pharmaceutical Technology Co., Ltd. Shanghai Gemma Pharmaceutical Technology Co., Ltd. also provides a negative control siRNA (siRNA-NC) that has no sequence homology with the SPAST gene.

[0152] siRNA-SPAST:

[0153] The sense strand is 5'-UCAAUACCUUUCUUAUACCAU-3' (SEQ ID NO.5);

[0154] The antisense strand is 5'-GGUAUAAGAAAGGUAUUGAAG-3'(SEQ ID NO.6),

[0155] 2. In vitro culture of synovial tissue cells

[0156] After the aseptically obtained synovial tissues of rheumatoid arthritis and osteoarthritis were washed with PBS, they were repeatedly cut into tissue pieces of about 1mm x 1mm x 1mm with sterile surgical scissors, and collagenase II (0.5mg / ml) After digestion at 37°C for 2 hours, filter through a 200-mesh gauze, centrifuge to remove the supernatan...

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Abstract

The invention discloses a diagnostic marker for rheumatoid arthritis and / or osteoarthritis. The diagnostic marker is an SPAST gene or SPAST protein, and the purpose of diagnosis is achieved by detecting the expression level of the SPAST gene. The research proves that the mRNA expression level of the SPAST gene of patients with rheumatoid arthritis and / or osteoarthritis is obviously improved compared with a normal person. According to the correlation between the SPAST gene and the rheumatoid arthritis and / or osteoarthritis, a kit for diagnosing rheumatoid arthritis and / or osteoarthritis can beprepared, and the kit can be widely applied to clinic. In addition, the invention further discloses application of the SPAST gene or the SPAST protein in the aspects of treating rheumatoid arthritis and / or osteoarthritis.

Description

technical field [0001] The present invention relates to the fields of molecular diagnosis and treatment, more specifically, the present invention relates to a diagnostic method by means of detecting SPAST abnormality; and a therapeutic agent for inhibiting SPAST gene or protein. Background technique [0002] Rheumatoid arthritis is an autoimmune disease characterized by polyarticular involvement that can progress to joint destruction and deformity, often involving peripheral joints. The etiology is unclear, but extraarticular organ involvement such as pulmonary interstitial disease and Sjogren's syndrome are also common. Appropriate early treatment can improve joint symptoms and function, reduce mortality, and reduce complications. [0003] A 2010 systematic review found that the prevalence of rheumatoid arthritis in North America and Northern Europe was estimated to range from 0.5% to 1.1%. The incidence in developing countries is relatively lower (0.1% to 0.5%). Rheumat...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883G01N33/68A61K45/00A61P29/00A61P19/02
CPCA61K45/00A61P19/02A61P29/00C12Q1/6883C12Q2600/158G01N33/6893G01N2800/102
Inventor 肖枫李曙光孙耀兰常鹏
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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