基于靶向氨基酸代谢组学探讨乙型病毒性肝炎相关慢加急性肝衰竭瘀黄证亚型的生物学基础

Targeted amino acid metabolomics unveils the biological underpinnings of subtypes of jaundice in hepatitis B-related acute-on-chronic liver failure

目的:从氨基酸代谢角度探讨乙型肝炎病毒相关慢加急性肝衰竭(HBV-ACLF)湿热瘀黄证和气虚瘀黄证的生物学基础。方法:采用靶向氨基酸代谢组学分析HBV-ACLF湿热瘀黄证组(20例)与气虚瘀黄证组(20例)患者的血清氨基酸代谢谱,寻找两种证型的氨基酸代谢差异特征,分析不同证型中氨基酸代谢与炎症细胞因子表达的相关性。结果:湿热瘀黄证患者体内异亮氨酸表达水平显著高于气虚瘀黄证[(10.36±4.47)μg/ml vs(7.76±4.08)μg/ml,P=0.016],但色氨酸则表达相反[(10.26±4.95)μg/ml vs(14.51±7.32)μg/ml,P=0.014];湿热瘀黄证患者白细胞介素(IL)-6显著高于气虚瘀黄证[10.15(3.88,24.42)pg/ml vs 7.73(3.93,18.83)pg/ml,P=0.017],而IL-2、IL-10显著低于气虚瘀黄证[4.40(2.88,8.55)pg/ml vs 6.60(2.80,15.50)pg/ml,P=0.033;7.19(4.52,15.04)pg/ml vs 8.21(2.19,14.64)pg/ml,P=0.015];多种氨基酸代谢与两种证型的炎症细胞因子表达密切相关,尤其是色氨酸代谢与支链氨基酸代谢;湿热瘀黄证中氨基酸代谢主要与促炎细胞因子IL-2、IL-6相关,按照相关系数大小依次为天冬氨酸、谷氨酸、异亮氨酸、亮氨酸、蛋氨酸、酪氨酸、丝氨酸、赖氨酸、谷氨酰胺、色氨酸、苯丙氨酸;气虚瘀黄证中氨基酸代谢与肿瘤坏死因子α(TNF-α)、IL-2、IL-6、IL-10相关,按照相关系数大小依次为天冬氨酸、甘氨酸、丙氨酸、苏氨酸、色氨酸、丝氨酸。异亮氨酸及色氨酸代谢不仅在两种证型间差异有统计学意义,且差异性关联炎症细胞因子的表达。结论:HBV-ACLF湿热瘀黄证和气虚瘀黄证患者在氨基酸代谢及炎症反应中均具有异质性,且两者之间相互关联。色氨酸以及支链氨基酸代谢可能对HBV-ACLF的免疫炎症反应具有重要调节作用,与湿热瘀黄证和气虚瘀黄证的生物学基础密切相关。

Objective:To explore the biological basis of damp-heat stasis-jaundice syndrome and Qi-deficiency stasis-jaundice syndrome in patients with hepatitis B virus-associated acute-on-chronic liver failure(HBV-ACLF)from the perspective of amino acid metabolism.Methods:Targeted amino acid metabolomics was used to analyze the serum samples of HBV-ACLF patients with damp-heat stasis-jaundice syndrome(n=20)and Qi-deficiency stasis-jaundice syndrome(n=20).The different characteristics of amino acid metabolism and the correlation with the expression of inflammatory cytokines between syndromes were explored.Results:There were differences in tryptophan and isoleucine metabolism between damp-heat stasis-jaundice syndrome and Qi-deficiency stasis-jaundice syndrome in patients with HBV-ACLF.The expression level of isoleucine in HBV-ACLF damp-heat stasis-jaundice syndrome was higher than that in Qi-deficiency stasis-jaundice syndrome[(10.36±4.47)μg/ml vs(7.76±4.08)μg/ml,P=0.016],but the expression of tryptophan was opposite[(10.26±4.95)μg/ml vs(14.51±7.32)μg/ml,P=0.014].The level of IL-6 in damp-heat stasis-jaundice syndrome was significantly higher than that in Qi-deficiency stasis-jaundice syndrome[10.15(3.88,24.42)pg/ml vs 7.73(3.93,18.83)pg/ml,P=0.017],while the levels of interleukin-2(IL-2)and interleukin-10(IL-10)were significantly lower than those in Qi-deficiency stasis-jaundice syndrome[4.40(2.88,8.55)pg/ml vs 6.60(2.80,15.50)pg/ml,P=0.033;IL-10:7.19(4.52,15.04)pg/ml vs 8.21(2.19,14.64)pg/ml,P=0.015].A variety of amino acid metabolism is closely related to the expression of inflammatory cytokines of the two syndromes,especially tryptophan metabolism and branched-chain amino acid metabolism.Amino acid metabolism in damp-heat stasis-jaundice syndrome is mainly related to pro-inflammatory cytokines IL-2 and IL-6,including aspartic acid,glutamic acid,isoleucine,leucine,methionine,tyrosine,serine,lysine,glutamine,tryptophan,and phenylalanine.Amino acid metabolism in Qi-deficiency stasis jaundice syndrome is related to tumor necrosis factorα(TNF-α),IL-2,IL-6,and IL-10,including aspartic acid,glycine,alanine,threonine,tryptophan,and serine.It is suggested that the expression of isoleucine and tryptophan metabolism is not only significantly different between the two syndromes,but also closely related to the expression of inflammatory cytokines between syndrome types.Conclusion:Patients with HBV-ACLF with damp-heat jaundice and Qi deficiency jaundice both exhibit heterogeneity in amino acid metabolism and inflammatory responses,and there is a mutual connection between them.Tryptophan and branched-chain amino acid metabolism may play a significant regulatory role in the immune-inflammatory response of HBV-ACLF and are closely related to the biological basis of damp-heat jaundice and Qi deficiency jaundice.