A biomarker for opticospinal myelitis diagnosis and application thereof

Abstract

The application discloses a 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 by using ELISA and other methods, so that the accuracy and reliability of diagnosis are further improved. Finally, the CD14 protein which can be used as a biomarker is screened, so that a simple and effective clinical diagnosis means is provided, and NMO patients can be accurately identified in the early stage, so that doctors can make more accurate diagnosis and treatment decisions in the clinic.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a biomarker 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 proteomics technology has provided new insights for the early diagnosis of non-moxicillin-related diseases (NMOs). Differential proteomics analysis of cerebrospinal fluid from patients and healthy individuals can uncover new potential biomarkers. However, current proteomics-based research still faces challenges such as imprecise biomarker screening methods and a lack of validation tools. Existing technologies largely limit research to screening differentially expressed proteins using techniques like 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 CSF protein 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 through proteomics technology and be validated through reliable methods. Summary of the Invention

[0005] Technical problem solved: To address the above-mentioned technical problems, this invention provides a 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 biomarker for the diagnosis of neuromyelitis optica, wherein the biomarker is CD14 protein.

[0007] Preferably, the CD14 protein is derived from a cerebrospinal fluid sample.

[0008] Preferably, the amino acid sequence of the CD14 protein is shown in SEQ ID NO: 1.

[0009] SEQ ID NO: 1 is as follows: MERASCLLLLLLPLVHVSATTPEPCELDDEDFRCVCNFSEPQPDWSEAFQCVSAVEVEIH AGGLNLEPFLKRVDADADPRQYADTVKALRVRRLTVGAAQVPAQLLVGALRVLAYSRLKE LTLEDLKITGTMPPLPLEATGLALSSLRLRNVSWATGRSWLAELQQWLKPGLKVLSIAQA HSPAFSCEQVRAFPALTSLDLSDNPGLGERGLMAALCPHKFPAIQNLALRNTGMETPTGV CAALAAAGVQPHSLDLSHNSLRATVNPSAPRCMWSSALNSLNLSFAGLEQVPKGLPAKLR VLDLSCNRLNRAPQPDELPEVDNLTLDGNPFLVPGTALPHEGSMNSGVVPACARSTLSVG VSGTLVLLQGARGFA Application of the biomarker CD14 protein in the preparation of diagnostic reagents for neuromyelitis optica.

[0010] Preferably, the CD14 protein is upregulated in the cerebrospinal fluid of patients with neuromyelitis optica.

[0011] Preferably, the diagnostic reagent is used for the early diagnosis of neuromyelitis optica and to assess the patient's disease activity.

[0012] Beneficial effects: This invention can improve the accuracy of diagnosis of neuromyelitis optica: Through differential analysis using proteomics technology, the biomarker CD14 with clinical application value was successfully screened out. The AUC value of CD14 is 0.7214, indicating that it has good diagnostic performance.

[0013] This invention simplifies the diagnostic process: it utilizes simple and rapid detection methods such as ELISA to verify biomarkers, reducing the complexity of the experimental process and improving diagnostic efficiency.

[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 cerebrospinal fluid proteomics analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC); Figure 2 Protein abundance order map of cerebrospinal fluid proteomics between patients with neuromyelitis optica (NMO) and healthy controls (HC); Figure 3 GO pathway enrichment analysis of upregulated proteins in cerebrospinal fluid proteomics differential analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC). Figure 4 KEGG pathway enrichment analysis of upregulated proteins in cerebrospinal fluid proteomics differential analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC). Figure 5 PPI protein interaction network diagram of upregulated proteins in cerebrospinal fluid proteomics differential analysis between patients with neuromyelitis optica (NMO) and healthy controls (HC); Figure 6 The violin figure illustrates the difference in CD14 protein levels in the cerebrospinal fluid of patients with neuromyelitis optica (NMO) and healthy controls (HC) as verified by ELISA. Figure 7 : Receiver operating characteristic (ROC) curve of CD14 protein expression level in neuromyelitis optica (NMO). Detailed Implementation

[0018] The present invention will be further described below with reference to the accompanying drawings and specific embodiments. The English abbreviations and corresponding Chinese names involved in the present invention are as follows: PPI (Protein-Protein Interaction): Protein-protein interaction GO (Gene Ontology): Gene Ontology KEGG (Kyoto Encyclopedia of Genes and Genomes): The Kyoto Encyclopedia of Genes and Genomes ROC (Receiver Operating Characteristic): Receiver Operating Characteristic Curve AUC (Area Under Curve): Area under the curve ELISA (Enzyme-Linked Immunosorbent Assay): Enzyme-linked immunosorbent assay 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.

[0019] Healthy control group (HC): Six healthy volunteers were selected, matched for sex and age with the NMO group, and had no neurological diseases.

[0020] 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.

[0021] Proteomics analysis: High-throughput qualitative and quantitative analysis of proteins in cerebrospinal fluid samples was performed using liquid chromatography-mass spectrometry (LC-MS / MS). The results are as follows: Figure 1 and 2 As shown. Each sample was repeated three times to ensure the reliability and reproducibility of the data.

[0022] Data processing and differential analysis were performed using DIANN software. Proteins with a fold change greater than 1.3 and a p-value less than 0.05 were selected as differentially expressed proteins.

[0023] Screening and validation of differentially expressed proteins: The above analysis identified differentially expressed proteins, such as CD14, associated with neuromyelitis optica (NMO). These proteins were significantly upregulated in the cerebrospinal fluid of NMO patients.

[0024] The expression levels of these differentially expressed proteins were verified using an enzyme-linked immunosorbent assay (ELISA). The ELISA results are as follows: Figure 6 As shown, the mean concentration of CD74 in the cerebrospinal fluid of MS patients was (80.21±43.35) ng / mL, while that in the healthy control group was (47.82±21.99) ng / mL, and the difference between the two groups was statistically significant (t=0.0048, p<0.001). The results indicate that the CD74 protein level in the NMO group was significantly higher than that in the healthy control group (HC).

[0025] 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.

[0026] The results are as follows Figure 3 As shown, the results indicate that the upregulated proteins in NMO patients are closely related to immune-related biological processes such as immune cell activation and antigen presentation.

[0027] KEGG pathway enrichment analysis: Further KEGG pathway analysis was performed, and the results were as follows: Figure 4 As shown, the results revealed that the differentially upregulated proteins are involved in important cell signaling pathways such as immune response pathways.

[0028] These analytical results provide theoretical support for CD14 as a biomarker for neuromyelitis optica.

[0029] Example 3: PPI Network Analysis PPI network construction: A protein-protein interaction (PPI) network of differentially upregulated proteins was constructed using the STRING database. The results are as follows: Figure 5 As shown, the results indicate that CD14 interacts significantly with a variety of immune-related proteins, supporting its important role in neuromyelitis optica (NMO).

[0030] This network analysis provides potential targets for subsequent biomarker validation and clinical applications.

[0031] Example 4: ELISA Validation and ROC Curve Analysis ELISA verification: The difference in CD14 protein expression in the cerebrospinal fluid of NMO patients and healthy controls was further verified by ELISA. The results showed that the CD14 protein level in NMO patients was significantly higher than that in the healthy control group, and the difference was statistically significant.

[0032] ROC curve analysis: The AUC value of CD14 protein was calculated, and the ROC curve analysis results are as follows: Figure 7 As shown, the AUC value of the CD74 ROC curve was 0.7214. ROC curve analysis showed that CD14 had good discriminative ability in the diagnosis of NMO, with an AUC value of 0.7214, demonstrating its potential as a biomarker for neuromyelitis optica.

[0033] Comparative experiments and data analysis Comparative experiment: The method of this invention was compared with traditional NMO diagnostic methods (such as imaging examinations and routine CSF analysis), and the optimal cutoff value was found to be 64.89 ng / mL, at which point the sensitivity was 64.29% and the specificity was 90%. The results show that the proteomics-based diagnostic method of this invention has higher sensitivity and specificity in early diagnosis.

[0034] Data Analysis: This invention successfully screened CD14 as a potential biomarker for NMO through differential analysis, pathway enrichment analysis, and PPI network construction, and the data supported its feasibility in clinical diagnosis through ELISA validation.

Claims

1. A biomarker for the diagnosis of neuromyelitis optica, characterized in that, The biomarker is CD14 protein.

2. A biomarker for the diagnosis of neuromyelitis optica according to claim 1, characterized in that, The CD14 protein was derived from a cerebrospinal fluid sample.

3. A biomarker for the diagnosis of neuromyelitis optica according to claim 1, characterized in that, The amino acid sequence of the CD14 protein is shown in SEQ ID NO:

1.

4. The use of the biomarker CD14 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 CD14 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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