Establishing Classification Tree Models in Rheumatoid Arthritis Using Combination of Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry and Magnetic Beads

Abstract

Background: There is no simple method for early diagnosis and evaluation of rheumatoid arthritis (RA). This study aimed to determine potential biomarkers and establish diagnostic patterns for RA using proteomic fingerprint technology combined with magnetic beads.Methods: The serum protein profiles of 97 RA patients and 76 healthy controls (HCs) were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) with weak cationic exchange (WCX) magnetic beads. Samples were randomly divided into training (83 RA patients and 56 HCs) and test sets (14 RA patients and 20 HCs). Patients were classified according to their Disease Activity Score: in remission, n = 28; with low disease activity, n = 17; with moderate disease activity, n = 21; with high disease activity, n = 31. There are 44 RA patients alone, 22 RA patients with interstitial lung disease (RA-ILD), 18 RA patients with secondary Sjögren's syndrome (RA-sSS), 6 RA patients with osteonecrosis of the femoral head (RA-ONFH), and 7 RA patients with other complications. Eleven patients were treated with etanercept only for half a year, after which their serum protein profiles were detected. The proteomic pattern was identified by Biomarker Patterns Software, and the potential biomarkers for RA diagnosis were further identified and quantified by enzyme-linked immunosorbent assay.Results: The diagnostic pattern with four potential protein biomarkers, mass-to-charge (m/z) 3,448.85, 4,716.71, 8,214.29, and 10,645.10, could accurately recognize RA patients from HCs (specificity, 91.57%; sensitivity, 92.86%). The test set were correctly classified by this model (sensitivity, 95%; specificity, 100%). The components containing the four biomarkers were preliminarily retrieved through the ExPasy database, including the C-C motif chemokine 24 (CCL24), putative metallothionein (MT1DP), sarcolipin (SLN), and C-X-C motif chemokine 11 (CCXL11). Only the CCL24 level was detected to have a significant decrease in the serum of RA patients as compared with HCs (p < 0.05). No significant difference was found in others, but a decreasing trend consistent with the down-regulation of the four biomarkers detected by MALDI-TOF-MS was observed. The diagnostic models could effectively discriminate between RA alone and RA with complications (RA-ILD: m/z 10,645.10 and 12,595.86; RA-sSS: m/z 6,635.62 and 33,897.72; RA-ONFH: m/z 2,071.689). The classification model, including m/z 1,130.776, 1,501.065, 2,091.198, and 11,381.87, could distinguish between RA patients with disease activity and those in remission. RA with low disease activity could be efficiently discriminated from other disease activity patients by specific protein biomarkers (m/z 2,032.31, 2,506.214, and Z9286.495). Two biomarkers (m/z 2,032.31 and 4,716.71) were applied to build the classification model for RA patients with moderate and high disease activities. Biological markers for etanercept (m/z 2,671.604064, 5,801.840579, 8,130.195641, and 9,286.49499) were observed between the responder (n = 7) and non-responder groups (n = 4) (p < 0.05).Conclusion: We successfully established a series of diagnostic models involving RA and RA with complications as well as assessed disease activity. Furthermore, we found that CCL24 may be a valuable auxiliary diagnostic indicator for RA. These results provide reference values for clinical practice in the future.

Proteomics: Potential techniques for discovering the pathogenesis of connective tissue diseases-interstitial lung disease

“…However, studies on 14-3-3ϵ in pulmonary fibrosis, especially in CTD-ILD, are scarce. Similarly, the level of transthyretin in the BALF of patients with SSc-ILD was elevated, as detected by proteomic methods (15,18,20). One study found that transthyretin stimulates the production of collagen I and immunoglobulin-binding proteins in fibroblasts, which participate in endoplasmic reticulum stress activation and profibrosis through mitochondrial oxidative stress in cardiac amyloidosis (44).…”

“…In addition, Landi’s team found that 14-3-3ϵ was increased in the BALF proteome of SSc-ILD patients compared with nonsmokers ( 18 , 20 ). In inflammatory and autoimmune diseases, 14-3-3-ϵ, as a component of the TNFR2 complex, restricts the activation of NF-κB through PI3K/Akt/mTOR signaling and stimulates the activation of C/EBP-β, thus guiding the plasticity of macrophages ( 43 ).…”

Development and validation of a decision tree classification model for the essential hypertension based on serum protein biomarkers

“…MALDI-TOF MS technology combined with magnetic beads has been widely used in the study of a variety of diseases and has led to the discovery of a variety of protein biomarkers, including for cancer ( 12 ), immune diseases ( 13 ), and endocrine diseases ( 14 ). MALDI-TOF MS technology combined with magnetic beads can directly detect all kinds of complex biological samples (e.g., urine, blood, cerebrospinal fluid, tissue biopsies, and even tear samples).…”