Department of Chemistry, University of Louisville, Louisville, Kentucky.
Alcohol Research Center, University of Louisville School of Medicine, Louisville, Kentucky.
Am J Physiol Gastrointest Liver Physiol. 2023 Feb 1;324(2):G142-G154. doi: 10.1152/ajpgi.00228.2022. Epub 2022 Dec 13.
Excess alcohol intake causes millions of deaths annually worldwide. Asymptomatic early-stage, alcohol-associated liver disease (ALD) is easily overlooked, and ALD is usually only diagnosed in more advanced stages. We explored the possibility of using polar urine metabolites as biomarkers of ALD for early-stage diagnosis and functional assessment of disease severity by quantifying the abundance of polar metabolites in the urine samples of healthy controls ( = 18), patients with mild or moderate liver injury ( = 21), and patients with severe alcohol-associated hepatitis ( = 25). The polar metabolites in human urine were first analyzed by untargeted metabolomics, showing that 209 urine metabolites are significantly changed in patients, and 17 of these are highly correlated with patients' model for end-stage liver disease (MELD) score. Pathway enrichment analysis reveals that the caffeine metabolic pathway is the most affected in ALD. We then developed a targeted metabolomics method and measured the concentration of caffeine and its metabolites in urine using internal and external standard calibration, respectively. The described method can quantify caffeine and its 14 metabolites in 35 min. The results of targeted metabolomics analysis agree with the results of untargeted metabolomics, showing that 13 caffeine metabolites are significantly decreased in patients. In particular, the concentrations of 1-methylxanthine, paraxanthine, and 5-acetylamino-6-amino-3-methyluracil are markedly decreased with increased disease severity. We suggest that these three metabolites could serve as functional biomarkers for differentiating early-stage ALD from more advanced liver injury. Our study using both untargeted and targeted metabolomics reveals the caffeine metabolic pathway is dysregulated in ALD. Three caffeine metabolites, 1-methylxanthine, paraxanthine, and 5-acetylamino-6-amino-3-methyluracil, can differentiate the severity of early-stage ALD.
过量饮酒每年导致全球数百万人死亡。无症状的早期酒精相关性肝疾病（ALD）很容易被忽视，而且通常只有在更晚期才被诊断出。我们通过定量分析健康对照组（n = 18）、轻度或中度肝损伤患者（n = 21）和重度酒精相关性肝炎患者（n = 25）尿液样本中的极性代谢物，探讨了将极性尿液代谢物用作 ALD 早期诊断和疾病严重程度功能评估的生物标志物的可能性。首先通过非靶向代谢组学分析人类尿液中的极性代谢物，结果表明患者尿液中有 209 种代谢物发生了显著变化，其中 17 种与患者终末期肝病模型（MELD）评分高度相关。通路富集分析显示，咖啡因代谢通路在 ALD 中受影响最大。然后我们开发了一种靶向代谢组学方法，分别使用内部和外部标准校准来测量尿液中咖啡因及其代谢物的浓度。所描述的方法可以在 35 分钟内定量分析咖啡因及其 14 种代谢物。靶向代谢组学分析的结果与非靶向代谢组学的结果一致，表明患者尿液中 13 种咖啡因代谢物的浓度显著降低。特别是随着疾病严重程度的增加，1-甲基黄嘌呤、副黄嘌呤和 5-乙酰氨基-6-氨基-3-甲基尿嘧啶的浓度明显降低。我们建议这三种代谢物可以作为区分早期 ALD 和更晚期肝损伤的功能生物标志物。我们使用非靶向和靶向代谢组学的研究表明，咖啡因代谢通路在 ALD 中失调。三种咖啡因代谢物，1-甲基黄嘌呤、副黄嘌呤和 5-乙酰氨基-6-氨基-3-甲基尿嘧啶，可以区分早期 ALD 的严重程度。
