基于LC-MS的除湿胃苓汤加减治疗脾虚湿蕴型慢性湿疹的血清代谢组学研究

A study on serum metabolomics of treatment for chronic eczema with spleen deficiency and dampness accumulation syndrome by modified Chushi Weiling Decoction based on LC-MS

目的 基于液相色谱-质谱法(LC-MS)研究除湿胃苓汤加减治疗脾虚湿蕴型慢性湿疹的代谢机制。方法 选取10例脾虚湿蕴型慢性湿疹患者给予除湿胃苓汤加减治疗并收集患者治疗前后血清,分别纳入患者治疗前血清组、患者治疗后血清组;另选取10例健康无湿疹症状及病史的健康人血清作为健康对照血清组进行对照。比较治疗前后慢性湿疹患者的湿疹面积及严重程度指数(EASI)评分。将三组应用LC-MS进行血清样品的分离和数据采集,以主成分分析(PCA)、偏最小二乘法判别分析(PLS-DA)和正交偏最小二乘判别分析(OPLS-DA)法筛选出相关差异代谢物,通过京都基因与基因组百科全书(KEGG)和人体代谢组数据库(HMDB)等开放数据库对差异的标志物进行代谢通路分析。结果 患者治疗后血清组的EASI评分(1.81±0.45)分明显低于患者治疗前血清组的(8.16±1.41)分,差异有统计学意义(t=13.567, P=0.000<0.01)。三组血清样本的代谢模式存在显著差异,基于变量重要性投影(VIP)>1.7和P<0.05的原则筛选出12个代谢标志物,苹果酸、油酰基肉碱显著上升, L-蛋氨酸、丙酰基肉碱、脱氧胆酸、焦谷氨酸、甲基半胱氨酸、谷氨酸肉碱、L-苏氨酸、L-异苏氨酸、L-亮氨酸、L-缬氨酸显著下降;经除湿胃苓汤加减治疗后,血清中这12种代谢物水平均出现显著回调、接近健康对照血清组正常值。进一步的代谢富集分析显示D-谷氨酰胺和D-谷氨酸代谢途径和缬氨酸、亮氨酸和异亮氨酸生物合成途径是主要的影响通路。结论 除湿胃苓汤加减可能通过调控氨基酸及其衍生代谢等途径调节机体能量代谢和免疫功能来治疗脾虚湿蕴型慢性湿疹。

Objective To study the serum metabolomics of treatment for chronic eczema with spleen deficiency and dampness accumulation syndrome by modified Chushi Weiling Decoction based on liquid chromatography-mass spectrometry(LC-MS).Methods A total of 10 patients with chronic eczema with spleen deficiency and dampness accumulation syndrome were selected and treated with modified Chushi Weiling Decoction.Serum samples were collected before and after treatment and included into the pretreatment serum group and the posttreatment serum group.Serum from 10 healthy people without eczema symptoms and history were selected as healthy control group.The eczema area and severity index(EASI)score of chronic eczema patients before and after treatment were compared.Serum samples of the three groups were separated and data collected by LC-MS,and relevant differential metabolites were screened by principal component analysis(PCA),partial least square discriminant analysis(PLS-DA)and orthogonal partial least square discriminant analysis(OPLS-DA).Metabolic pathway analysis of markers of differences was performed using open databases such as Kyoto Encyclopedia of Genes and Genomes(KEGG)and Human Metabolome Database(HMDB).Results The EASI score of the posttreatment serum group was(1.81±0.45)points,which was significantly lower than that of(8.16±1.41)points of the pretreatment serum group,and the difference was statistically significant(t=13.567,P=0.000<0.01).There were significant differences in the metabolic patterns of serum samples from the three groups.12 metabolic markers were selected based on the principle of variable importance in the projection(VIP)>1.7 and P<0.05,and malic acid and oleylcarnitine were significantly increased.L-methionine,propionylcarnitine,deoxycholic acid,pyroglutamic acid,methylcysteine,glutamatecarnitine,L-threonine,L-allothreonine,L-leucine and L-valine were significantly decreased.After treatment with modified Chushi Weiling Decoction,the serum levels of these 12 metabolites showed significant regression,close to the normal value of healthy control group.Further metabolic enrichment analysis showed that the metabolic pathways of D-glutamine and D-glutamate and the biosynthesis pathways of valine,leucine and allothreonine were the main influencing pathways.Conclusion Modified Chushi Weiling Decoction may treat chronic eczema with spleen deficiency and dampness accumulation syndrome by regulating energy metabolism and immune function through amino acids and its derived metabolism.