Metabolic Disturbances in Adult-Onset Still's Disease Evaluated Using Liquid Chromatography/Mass Spectrometry-Based Metabolomic Analysis

Abstract

Objective: Liquid chromatography/mass spectrometry (LC/MS)-based comprehensive analysis of metabolic profiles with metabolomics approach has potential diagnostic and predictive implications. However, no metabolomics data have been reported in adult-onset Still's disease (AOSD). This study investigated the metabolomic profiles in AOSD patients and examined their association with clinical characteristics and disease outcome.

Methods: Serum metabolite profiles were determined on 32 AOSD patients and 30 healthy controls (HC) using ultra-performance liquid chromatography (UPLC)/MS analysis, and the differentially expressed metabolites were quantified using multiple reactions monitoring (MRM)/MS analysis in 44 patients and 42 HC. Pure standards were utilized to confirm the presence of the differentially expressed metabolites.

Results: Eighteen differentially expressed metabolites were identified in AOSD patents using LC/MS-based analysis, of which 13 metabolites were validated by MRM/MS analysis. Among them, serum levels of lysoPC(18:2), urocanic acid and indole were significantly lower, and L-phenylalanine levels were significantly higher in AOSD patients compared with HC. Moreover, serum levels of lysoPC(18:2), PhePhe, uridine, taurine, L-threonine, and (R)-3-Hydroxy-hexadecanoic acid were significantly correlated with disease activity scores (all p<0.05) in AOSD patients. A different clustering of metabolites was associated with a different disease outcome, with significantly lower levels of isovalerylsarcosine observed in patients with chronic articular pattern (median, 77.0AU/ml) compared with monocyclic (341.5AU/ml, p<0.01) or polycyclic systemic pattern (168.0AU/ml, p<0.05).

Conclusion: Thirteen differentially expressed metabolites identified and validated in AOSD patients were shown to be involved in five metabolic pathways. Significant associations of metabolic profiles with disease activity and outcome of AOSD suggest their involvement in AOSD pathogenesis.

Fig 1

A different clustering of metabolic ions was shown in the supervised orthogonal partial least squares-discriminant analysis (OPLS-DA) score plot (A1 and A2) from 32 patients with adult-onset Still’s disease (AOSD), 30 healthy subjects (control group), and quality control (QC). The comparisons in serum levels of the differentially expressed metabolites between AOSD patients and control group (B-N). For the box plots, the bottom and top of the boxes represent the 25^th and 75^th percentile, respectively. The top and bottom bars represent the entire stretch of the data points for the subjects, except the extreme points which are indicated with exact values of data. The hyphen within box indicates the median value. *p<0.05, **p<0.01, ***p<0.001, versus the control group.

Fig 2

A different clustering of metabolic ions was shown in the OPLS-DA score plot from 14 active AOSD patients, 18 inactive AOSD patients, and 30 healthy controls (A1). A different clustering of metabolic ions was shown in AOSD patients with different activity scores (A2). The correlation between serum levels of the altered metabolites and AOSD activity scores (B-N).

Fig 3

A different clustering of metabolic ions in the OPLS-DA score plots from AOSD patients with different patterns of disease outcome (A). The comparisons in serum levels of the differentially expressed metabolites in AOSD patients with different patterns of disease outcome (B-N). *p<0.05, versus monocyclic systemic pattern or polycyclic systemic pattern. The explanations for box plots s are as those described in Fig 1 legend.

Fig 4. Schematic representation depicts the metabolic imbalance in patients with adult-onset Still’s disease (AOSD).

(↑) represents up-regulated changes observed in AOSD; (↓) represents down-regulated changes observed in AOSD.