Introduction Diagnosis of overt hepatic encephalopathy (HE) in patients with cirrhosis often relies on subjective clinical examination. Arterial ammonia correlates poorly with HE grades and a blood marker for diagnosis and monitoring is lacking. Metabolic profiling by nuclear magnetic resonance (NMR) spectroscopy of blood correlates with Model for End Stage Liver Disease Score (MELD)1 and may offer diagnostic biomarkers in overt HE.
Methods Thirty-seven patients with cirrhosis and differing grades of HE were identified. Arterial blood was drawn for arterial ammonia, blood gas analysis and NMR spectroscopy. HE was diagnosed and graded by West-Haven criteria and Trail Making (A and B) Tests. 1H NMR spectroscopy was performed in a Bruker 600 MHz Avance spectrometer using a Carr-Purcell-Meiboom-Gill sequence prior to multivariate analysis using orthogonal partial least squares discriminant analysis (OPLS-DA).
Results 24 male and 13 female patients, 21 with alcohol-related cirrhosis, 5 viral hepatitis, 11 mixed/other causes, with median (range) age 57 (35–75) years made up the study cohort. Fifteen patients had no HE while 15 had grades 1–2 and 7 grades 3–4. Median MELD score was 14 (4–32) and median arterial ammonia level 106 (19–268 μmol/L). Ammonia level correlated weakly with HE grade (Kendall's τ=0.238, p=0.040) but not with MELD score (τ=0.010, p=0.118). No multivariate models using HE grade as a categorical (OPLSDA) or continuous (OPLS) variables resulted in validity (eg, OPLS model: R2(Y)=0.489, Q2(Y)=−0.091). A 2-component OPLS model using ammonia as a Y variable identified multiple metabolites correlating with arterial ammonia (overall R2(Y)=0.566, Q2(Y)=0.394, cross-validated ANOVA p=0.0008). Metabolites associated with high ammonia levels included pyruvate, 3-hydroxybutyrate, glutamate, phenylalanine and an unassigned resonance at 2.43 ppm, while the resonances associated with lipoproteins correlated negatively with ammonia.
Conclusion Plasma 1H NMR spectroscopy did not discriminate effectively between grades of HE in this cohort of patients with cirrhosis and overt HE. Multiple metabolites correlate with arterial ammonia level with some overlap from the metabolic pathways implicated in high MELD patients. Alternative metabolic profiling techniques such as mass spectroscopy may be required to assist in diagnosis in these patients.
Competing interests I Coltart: None declared, M McPhail Grant/Research Support from: Wellcome Trust, UK, E Want: None declared, K Veselkov: None declared, E Holmes: None declared, J Wendon: None declared, S Taylor-Robinson: None declared, W Bernal: None declared, D Shawcross: None declared.
Reference 1. Amathieu R, et al. Metabolomic approach by 1H NMR spectroscopy of serum for the assessment of chronic liver failure in patients with cirrhosis. J Proteome Res 2011;10:3239–45.
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