Serum Metabolomics Study Of Patients With AS And Different TCM Constitutions Using Gas Chromatography And Mass Spectrometry

Objective:Serum samples from healthy controls,patients with ankylosing spondylitis(AS)during the remission and acute stages,and main constitutions were analyzed by gas chromatography-mass spectrometry(GC-MS)to explore the serum metabolic features associated with AS and provide new ideas for clinical diagnosis and treatment of AS.Methods:GC-MS was employed for serum metabolic profiing analysis,and multivariate statistical analysis of serum metabolic data was performed,including principal component analysis(PCA)and partial least squares-discriminant analysis(PLS-DA).Serum differential metabolites between healthy controls and AS patients were picked out based on the variable importance(VIP value)of OPLS-DA model and t-test.Moreover,the metabolites associated with the disease development were screened out.The constitution classification of AS patients was according to the China Association of Chinese Medicine standard of 2006,and the self-test table made by Wang Qi.Results:(1)Different serum metabolic profiles of healthy controls and AS patients were observed and 24 metabolites were picked out which were potentially associated with the disease.As compared to healthy controls,18 of 24 metabolites were down-regulated in AS patients,including 2-Aminobutyrate,ethanolamine,glucose,creatinine,1,5-Anhydrosorbitol and various amino acids(valine,alanine,isoleucine,serine,glycine,threonine,tyrosine,tryptophan,leucine,proline,methionine,pyroglutamic acid and phenylalanine),and other 6 metabolites were up-regulated in patients,including 1,3-propanediol,porphine,propylene glycol,glycerol-3-phosphate,ribofuranose,hexadecanoic acid and so on.(2)The levels of CRP and ESR were much higher in patients during the acute stage than those during the remission stage,and there were 14 significantly different metabolites between patients during different stages.Among these metabolites,ribofuranose,2-hydroxybutyric acid,dodecanoic acid,hexadecanoic acid,linoleic acid,stearic acid,ribonic acid and pyroglutamic acid were negatively correlated with CRP;ribofuranose,linoleic acid,ribonic acid,serine and pyroglutamic acid were negatively correlated with ESR.These metabolites were mainly involved in energy metabolism and amino acid metabolism,and they may be highly related to the progression of AS.(3)According to the constitution measurement score,four types of biased constitution of AS patients were found,including yang-insufficiency constitution,qi-insufficiency constitution(can develop into yang insufficiency),phlegm-dampness constitution and blood-stasis constitution.The AS patients with different constitutions have distinct serum metabolic characteristics.A variety of metabolites related to oxidant stress were decreased in phlegm-dampness patients.The level of tyrosine was much lower in yang-deficiency patients.There are small metabolic differences between patients with blood-stasis and mild constitutions.Conclusions:As compared with the healthy controls,metabolites associated with the energy metabolism and body immunity(amino acids,glucose,fatty acids,etc.)were significantly changed in AS patients.The fatty acids were up-regulated,while glucose and amino acids were down-regulated in patients during the remission stage.During the acute stage,the increased energy consumption may lead to continued decrease of glucose;more fatty acids were transferred into the mitochondria for supporting energy,thus the level of serum fatty acids were further reduced.During the acute stage of AS,the CRP and ESR index were increased,and some fatty acids and amino acids were found to be highly correlated with them.And these metabolites may be associated with the disease activity and can provide more evidence for the evaluation of the disease progression and treatment.AS patients with different types of constitutions had distinct serum metabolic characteristics.Compared with the patients with mile constitution,the oxidative stress-mediated injury in the phlegm-dampness patients became more serious;yang-deficiency patients were with lower level of tyrosine,which may affect the synthesis of thyroxine;small metabolic differences between blood-stasis and mild patients were found.This study provides biological evidence for the syndrome differentiation dependent treatment system at the molecular level..