Integrative transcriptomics–metabolomics reveals sphingolipid metabolic reprogramming in traditional Chinese medicine syndrome differentiation of rheumatoid arthritis

Abstract

To investigate sphingolipid metabolism for differentiating heat-dampness syndrome from kidney-liver deficiency syndrome in Rheumatoid Arthritis (RA), identify metabolic biomarkers, and integrate transcriptomic data to elucidate molecular mechanisms. 40 RA patients (kidney-liver deficiency syndrome, n=20; heat-dampness syndrome, n=20) were enrolled and serum untargeted metabolomics profiling was performed using ultra-performance liquid chromatography-tandem mass spectrometry. A least absolute shrinkage and selection operator logistic regression diagnostic model was constructed, and its performance was assessed by receiver operating characteristic curve and decision curve analyses. Three independent transcriptomic datasets were downloaded from the Gene Expression Omnibus database (GSE55235: 10 RA synovium, 10 osteoarthritis synovium, and 10 normal synovium; GSE93272: RA whole blood and healthy control whole blood; GSE17755: RA peripheral blood mononuclear cells and healthy control peripheral blood mononuclear cells), and differential expression and tissue-specific regulation of sphingolipid-metabolism-related genes were analyzed. Immune cell infiltration was estimated using marker gene set mean expression scoring, and Spearman correlation analysis was used to construct metabolite-clinical indicator networks. Sphingosine kinase 1 activity (sphingosine-1-phosphate/C17-sphinganine ratio) was significantly higher in the heat-dampness syndrome group than in the kidney-liver deficiency syndrome group (P = 3.45×10 -6 ). C17-sphinganine was significantly negatively correlated with sphingosine-1-phosphate (r = -0.555). C17-sphinganine alone distinguished kidney-liver deficiency syndrome from heat-dampness syndrome with an area under the curve of 0.945 and an accuracy of 95%. Sphingosine-1-phosphate lyase 1 was markedly upregulated in rheumatoid arthritis synovium compared with normal synovium (log2 fold change = 1.124, P = 2.17×10 -4 ) and also significantly upregulated in peripheral blood albeit with a smaller effect size (log 2 fold change = 0.134, P = 8.44×10 -4 ), demonstrating tissue-specific magnitude-difference regulation. Sphingosine kinase 1 showed a negative correlation trend with neutrophil infiltration. C8-ceramide-1-phosphate was negatively correlated with C-reactive protein (r = -0.47) and positively correlated with mean corpuscular hemoglobin (r = 0.51). C17-sphinganine is a core metabolic biomarker for distinguishing kidney-liver deficiency syndrome from heat-dampness syndrome, enabling precise syndrome classification with a single measurement. The tissue-specific magnitude-difference regulation of sphingosine-1-phosphate lyase 1 and the sphingosine kinase 1-neutrophil correlative trend provide novel insights into how sphingolipid metabolism participates in traditional Chinese medicine syndrome differentiation in RA.