A cerebrospinal fluid low-abundance protein biomarker for opticospinal multiple sclerosis diagnosis and application thereof

Abstract

The application discloses a cerebrospinal fluid low-abundance protein biomarker for opticospinal myelitis diagnosis and application thereof, and belongs to the field of biomedical technology. The application identifies and screens protein markers capable of having significant difference between NMO patients and healthy people through proteomics technology, and verifies the markers, thereby further improving the accuracy and reliability of diagnosis. Finally, the RAPGEF2 protein which can be used as a biomarker is screened, thereby providing a simple and effective clinical diagnosis means, and NMO patients can be accurately identified in an early stage, and doctors can make more accurate diagnosis and treatment decisions in clinic.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a low-abundance protein biomarker in cerebrospinal fluid for the diagnosis of neuromyelitis optica and its application. Background Technology

[0002] Neuromyelitis optica (NMO) is a chronic, immune-mediated disease of the central nervous system, the etiology of which remains incompletely understood. Early diagnosis of NMO is crucial for intervention and treatment, but currently, highly specific and sensitive diagnostic methods are lacking. Traditional diagnostic methods rely primarily on clinical symptoms, imaging examinations, and cerebrospinal fluid (CSF) analysis, but these methods all have limitations. For example, imaging examinations cannot accurately detect NMO lesions in the early stages, and while CSF analysis can provide some information, it cannot provide a definitive diagnostic basis for all patients. Existing biomarker research lacks unified standards, and the diagnostic value of many biomarkers has not been fully validated.

[0003] In recent years, the application of low-abundance proteomics technology has provided new insights for the early diagnosis of non-moxicillin-related diseases (NMO). Differential proteomics analysis of cerebrospinal fluid from patients and healthy individuals can uncover novel potential biomarkers. Low-abundance proteins refer to proteins with low concentrations in samples that are difficult to detect using traditional methods; these proteins may play a crucial role in the early stages of disease. However, current research based on low-abundance proteomics still faces challenges such as imprecise biomarker screening methods and a lack of validation tools. Existing technologies mostly limit research to screening differentially expressed proteins using techniques such as mass spectrometry, with limited in-depth exploration of the functional validation and clinical applications of these proteins.

[0004] For example, some studies have attempted to analyze the differential abundance of CSF low-abundance proteins between NMO patients and healthy individuals using mass spectrometry, and have identified some potential biomarkers. However, the clinical validation of these biomarkers lacks sufficient data support, and their diagnostic efficacy has not been widely validated. Therefore, there is an urgent need for a novel biomarker detection method that can both identify differentially expressed proteins using low-abundance proteomics technology and be validated through reliable methods. Summary of the Invention

[0005] Technical problem solved: To address the above-mentioned technical problem, this invention provides a cerebrospinal fluid low-abundance protein biomarker for the diagnosis of neuromyelitis optica and its application, which can accurately identify NMO patients at an early stage, provide a basis for clinical diagnosis and treatment, and improve diagnostic efficiency.

[0006] Technical solution: A low-abundance protein biomarker in cerebrospinal fluid for the diagnosis of neuromyelitis optica, wherein the biomarker is RAPGEF2 protein.

[0007] Preferably, the RAPGEF2 protein is derived from a cerebrospinal fluid sample.

[0008] Preferably, the amino acid sequence of the RAPGEF2 protein is as shown in SEQ ID NO: 1.

[0009] SEQ ID NO: 1: MKPLAIPANHGVMGQQEKHSLPADFTKLHLTDSLHPQVTHVSSSHSGCSITSDSGSSSLS DIYQATESEAGDMDLSGLPETAVDSEDDDDEEDIERASDPLMSRDIVRDCLEKDPIDRTD DDIEQLLEFMHQLPAFANMTMSVRRELCAVMVFAVVERAGTIVLNDGEELDSWSVILNGS VEVTYPDGKAEILCMGNSFGVSPTMDKEYMKGVMRTKVDDCQFVCIAQQDYCRILNQVEK NMQKVEEEGEIVMVKEHRELDRTGTRKGHIVIKGTSERLTMHLVEEHSVVDPTFIEDFLL TYRTFLSSPMEVGKKLLEWFNDPSLRDKVTRVVLLWVNNHFNDFEGDPAMTRFLEEFENN LEREKMGGHLRLLNIACAAKAKRRLMTLTKPSREAPLPFILLGGSEKGFGIFVDSVDSGS KATEAGLKRGDQILEVNGQNFENIQLSKAMEILRNNTHLSITVKTNLFVFKELLTRLSEE KRNGAPHLPKIGDIKKASRYSIPDLAVDVEQVIGLEKVNKKSKANTVGGRNKLKKILDKT RISILPQKPYNDIGIGQSQDDSIVGLRQTKHIPTALPVSGTLSSSNPDLLQSHHRILDFS ATPDLPDQVLRVFKADQQSRYIMISKDTTAKEVVIQAIREFAVTATPDQYSLCEVSVTPE GVIKQRRLPDQLSKLADRIQLSGRYYLKNNMETETLCSDEDAQELLRESQISLLQLSTVE VATQLSMRNFELFRNIEPTEYIDDLFKLRSKTSCANLKRFEEVINQETFWVASEILRETN QLKRMKIIKHFIKIALHCRECKNFNSMFAIISGLNLAPVARLRTTWEKLPNKYEKLFQDL QDLFDPSRNMAKYRNVLNSQNLQPPIIPLFPVIKKDLTFLHEGNDSKVDGLVNFEKLRMI AKEIRHVGRMASVNMDPALMFRTRKKKWRSLGSLSQGSTNATVLDVAQTGGHKKRVRRSS FLNAKKLYEDAQMARKVKQYLSNLELEMDEESLQTLSLQCEPATNTLPKNPGDKKPVKSE TSPVAPRAGSQQKAQSLPQPQQQPPPAHKINQGLQVPAVSLYPSRKKVPVKDLPPFGINS PQALKKILSLSEEGSLERHKKQAEDTISNASSQLSSPPTSPQSSPRKGYTLAPSGTVDNF SDSGHSEISSRSSIVSNSSFDSVPVSLHDERRQRHSVSIVETNLGMGRMERRTMIEPDQY SLGSYAPMSEGRGLYATATVISSPSTEELSQDQGDRASLDAADSGRGSWTSCSSGSHDNI QTIQHQRSWETLPFGHTHFDYSGDPAGLWASSSHMDQIMFSDHSTKYNRQNQSRESLEQA QSRASWASSTGYWGEDSEGDTGTIKRRGGKDVSIEAESSSLTSVTTEETKPVPMPAHIAV ASSTTKGLIARKEGRYREPPPTPPGYIGIPITDFPEGHSHPARKPPDYNVALQRSRMVAR SSDTAGPSSVQQPHGHPTSSRPVNKPQWHKPNESDPRLAPYQSQGFSTEEDEDEQVSAV.

[0010] Application of the biomarker RAPGEF2 protein in the preparation of diagnostic reagents for neuromyelitis optica.

[0011] Preferably, the RAPGEF2 protein is upregulated in the cerebrospinal fluid of patients with neuromyelitis optica.

[0012] Preferably, the diagnostic reagent is used for the early diagnosis of neuromyelitis optica and to assess the patient's disease activity.

[0013] Beneficial effects: This invention can improve the accuracy of diagnosis of neuromyelitis optica: Through differential analysis using proteomics technology, the biomarker RAPGEF2 with clinical application value was successfully screened, indicating that it has good diagnostic performance.

[0014] This invention can reduce the misdiagnosis rate: by using reliable protein biomarkers, accurate diagnosis can be made in the early stages of NMO, thereby reducing the misdiagnosis rate, enabling early intervention, and improving patient prognosis.

[0015] This invention has broad clinical application prospects: the cerebrospinal fluid protein biomarkers and their detection methods of this invention can provide clinicians with a simple and effective auxiliary diagnostic tool, which is widely applicable to the diagnosis of NMO.

[0016] In summary, this invention can significantly improve the diagnostic accuracy of neuromyelitis optica, and its feasibility and effectiveness in practical applications have been demonstrated through clinical validation data. Attached Figure Description

[0017] Figure 1 Volcano plot of differential analysis of low-abundance proteomics in cerebrospinal fluid between patients with neuromyelitis optica (NMO) and healthy controls (HC); Figure 2 Heatmap of differential analysis of low-abundance proteomics in cerebrospinal fluid between patients with neuromyelitis optica (NMO) and healthy controls (HC); Figure 3 GO pathway enrichment analysis of upregulated proteins in cerebrospinal fluid low-abundance proteomics differential analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC). Figure 4 GO pathway enrichment analysis of downregulated proteins in cerebrospinal fluid low-abundance proteomics differential analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC). Figure 5The violin figure illustrates the difference in RAPGEF2 protein levels in the cerebrospinal fluid between patients with neuromyelitis optica (NMO) and healthy controls (HC). Detailed Implementation

[0018] The present invention will be further described below with reference to the accompanying drawings and specific embodiments.

[0019] Example 1: Differential analysis of cerebrospinal fluid proteomics Experimental materials: NMO patients: Four patients who met the diagnostic criteria for neuromyelitis optica (NMO) were selected. All patients were newly diagnosed and untreated, and other neurological diseases were excluded.

[0020] Healthy control group (HC): Six healthy volunteers were selected, matched for sex and age with the NMO group, and had no neurological diseases.

[0021] Cerebrospinal fluid sample collection: After obtaining informed consent, all participants underwent lumbar puncture to collect cerebrospinal fluid samples, and all samples were stored at -80°C.

[0022] Low abundance enrichment: Low-abundance proteins in cerebrospinal fluid (CSF) from multiple sclerosis (MS) patients and healthy individuals (HC) were enriched using magnetic column enrichment method.

[0023] Proteomics analysis: High-throughput qualitative and quantitative analysis of proteins in cerebrospinal fluid samples was performed using liquid chromatography-mass spectrometry (LC-MS / MS). Each sample was analyzed in triplicate to ensure data reliability and reproducibility.

[0024] Data processing and differential analysis were performed using DIANN software. Proteins with a fold change greater than 1.5 and a p-value less than 0.05 were selected as differentially expressed proteins.

[0025] Screening and validation of differentially expressed proteins: The above analysis identified differentially expressed proteins associated with neuromyelitis optica (NMO), such as RAPGEF2. These proteins were significantly upregulated in the cerebrospinal fluid of NMO patients.

[0026] Example 2: Enrichment analysis of GO and KEGG pathways GO pathway enrichment analysis: Gene ontology (GO) pathway enrichment analysis was performed on differentially upregulated proteins to analyze their relationship with biological processes such as immune response and cell adhesion.

[0027] The results showed that the upregulated proteins in NMO patients were closely related to immune-related biological processes such as immune cell activation and antigen presentation.

[0028] These analytical results provide theoretical support for RAPGEF2 as a biomarker for neuromyelitis optica.

[0029] Comparative experiments and data analysis Comparative experiment: The method of the present invention was compared with traditional NMO diagnostic methods (such as imaging examinations and routine CSF analysis). The results showed that the proteomics-based diagnostic method of the present invention has higher sensitivity and specificity in early diagnosis.

[0030] Data Analysis: This invention successfully screened RAPGEF2 as a potential biomarker for NMO through differential analysis and pathway enrichment analysis, and the data supports its feasibility in clinical diagnosis.

Claims

1. A low-abundance protein biomarker in cerebrospinal fluid for the diagnosis of neuromyelitis optica, characterized in that, The biomarker is the RAPGEF2 protein.

2. The cerebrospinal fluid low-abundance protein biomarker for the diagnosis of neuromyelitis optica according to claim 1, characterized in that, The RAPGEF2 protein was derived from a cerebrospinal fluid sample.

3. A cerebrospinal fluid low-abundance protein biomarker for the diagnosis of neuromyelitis optica according to claim 1, characterized in that, The amino acid sequence of the RAPGEF2 protein is shown in SEQ ID NO:

1.

4. The use of the biomarker RAPGEF2 protein of claim 1 in the preparation of a diagnostic reagent for neuromyelitis optica.

5. The application according to claim 4, characterized in that, The RAPGEF2 protein was upregulated in the cerebrospinal fluid of patients with neuromyelitis optica.

6. The application according to claim 4, characterized in that, The diagnostic reagent is used for the early diagnosis of neuromyelitis optica and to assess the patient's disease activity.

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