Original Article
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World J Gastroenterol. Jan 7, 2014; 20(1): 163-174
Published online Jan 7, 2014. doi: 10.3748/wjg.v20.i1.163
Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases
Tomasz Dawiskiba, Stanisław Deja, Agata Mulak, Adam Ząbek, Ewa Jawień, Dorota Pawełka, Mirosław Banasik, Agnieszka Mastalerz-Migas, Waldemar Balcerzak, Krzysztof Kaliszewski, Jan Skóra, Piotr Barć, Krzysztof Korta, Kornel Pormańczuk, Przemyslaw Szyber, Adam Litarski, Piotr Młynarz
Tomasz Dawiskiba, Jan Skóra, Piotr Barć, Krzysztof Korta, Kornel Pormańczuk, Przemyslaw Szyber, Department of Vascular, General and Transplantation Surgery, Wroclaw Medical University, 50-556 Wroclaw, Poland
Stanisław Deja, Faculty of Chemistry, Opole University, 45-040 Opole, Poland
Agata Mulak, Department of Gastroenterology and Hepatology, Wroclaw Medical University, 50-556 Wroclaw, Poland
Adam Ząbek, Ewa Jawień, Piotr Młynarz, Bioorganic Chemistry Group, Department of Chemistry, Wroclaw University of Technology, 50-370 Wroclaw, Poland
Dorota Pawełka, Waldemar Balcerzak, Krzysztof Kaliszewski, First Department and Clinic of General, Gastroenterological and Endocrinological Surgery, Wroclaw Medical University, 50-369 Wrocław, Poland
Mirosław Banasik, Department and Clinic of Nephrology and Transplantation Medicine, Wroclaw Medical University, 50-556 Wroclaw, Poland
Agnieszka Mastalerz-Migas, Department of Family Medicine, Wroclaw Medical University, 51-141 Wroclaw, Poland
Adam Litarski, Department of Urology, 4th Military Clinical Hospital, 50-981 Wroclaw, Poland
Author contributions: Dawiskiba T and Młynarz P designed the research; Dawiskiba T supervised the research; Mulak A coordinated the collection of human material; Mulak A, Pawełka D, Balcerzak W, Kaliszewski K, Mastalerz-Migas A, Skóra J, Barć P, Korta K, Pormańczuk K and Szyber P were involved in the collection of human material from inflammatory bowel diseases patients and contributed to database construction; Litarski A collected human material from healthy control subjects; Deja S, Ząbek A, Jawień E and Młynarz P performed the major part of the experimental work and performed the chemometrics and the statistical analysis; Dawiskiba T, Deja S, Ząbek A and Młynarz P analyzed the data; Dawiskiba T and Banasik M performed the extensive literature search; Dawiskiba T, Deja S, Mulak A, Ząbek A and Młynarz P prepared the manuscript; Mulak A and Młynarz P revised the manuscript critically.
Supported by Wroclaw Medical University Research Program, Grant Number 15/Pbmn
Correspondence to: Tomasz Dawiskiba, MD, PhD, Department of Vascular, General and Transplantation Surgery, Wroclaw Medical University, 213 Borowska Street, 50-556 Wroclaw, Poland. tomasz.dawiskiba@umed.wroc.pl
Telephone: +48-71-7332002 Fax: +48-71-7332009
Received: October 24, 2013
Revised: November 21, 2013
Accepted: December 5, 2013
Published online: January 7, 2014
Processing time: 88 Days and 2.7 Hours
Abstract

AIM: To evaluate the utility of serum and urine metabolomic analysis in diagnosing and monitoring of inflammatory bowel diseases (IBD).

METHODS: Serum and urine samples were collected from 24 patients with ulcerative colitis (UC), 19 patients with the Crohn’s disease (CD) and 17 healthy controls. The activity of UC was assessed with the Simple Clinical Colitis Activity Index, while the activity of CD was determined using the Harvey-Bradshaw Index. The analysis of serum and urine samples was performed using proton nuclear magnetic resonance (NMR) spectroscopy. All spectra were exported to Matlab for preprocessing which resulted in two data matrixes for serum and urine. Prior to the chemometric analysis, both data sets were unit variance scaled. The differences in metabolite fingerprints were assessed using partial least-squares-discriminant analysis (PLS-DA). Receiver operating characteristic curves and area under curves were used to evaluate the quality and prediction performance of the obtained PLS-DA models. Metabolites responsible for separation in models were tested using STATISTICA 10 with the Mann-Whitney-Wilcoxon test and the Student’s t test (α = 0.05).

RESULTS: The comparison between the group of patients with active IBD and the group with IBD in remission provided good PLS-DA models (P value 0.002 for serum and 0.003 for urine). The metabolites that allowed to distinguish these groups were: N-acetylated compounds and phenylalanine (up-regulated in serum), low-density lipoproteins and very low-density lipoproteins (decreased in serum) as well as glycine (increased in urine) and acetoacetate (decreased in urine). The significant differences in metabolomic profiles were also found between the group of patients with active IBD and healthy control subjects providing the PLS-DA models with a very good separation (P value < 0.001 for serum and 0.003 for urine). The metabolites that were found to be the strongest biomarkers included in this case: leucine, isoleucine, 3-hydroxybutyric acid, N-acetylated compounds, acetoacetate, glycine, phenylalanine and lactate (increased in serum), creatine, dimethyl sulfone, histidine, choline and its derivatives (decreased in serum), as well as citrate, hippurate, trigonelline, taurine, succinate and 2-hydroxyisobutyrate (decreased in urine). No clear separation in PLS-DA models was found between CD and UC patients based on the analysis of serum and urine samples, although one metabolite (formate) in univariate statistical analysis was significantly lower in serum of patients with active CD, and two metabolites (alanine and N-acetylated compounds) were significantly higher in serum of patients with CD when comparing jointly patients in the remission and active phase of the diseases. Contrary to the results obtained from the serum samples, the analysis of urine samples allowed to distinguish patients with IBD in remission from healthy control subjects. The metabolites of importance included in this case up-regulated acetoacetate and down-regulated citrate, hippurate, taurine, succinate, glycine, alanine and formate.

CONCLUSION: NMR-based metabolomic fingerprinting of serum and urine has the potential to be a useful tool in distinguishing patients with active IBD from those in remission.

Keywords: Metabolomics; Inflammatory bowel disease; Ulcerative colitis; Crohn's disease; Proton nuclear magnetic resonance spectroscopy; Urine; Serum; Partial least-squares-discriminant analysis

Core tip: This study demonstrates that proton nuclear magnetic resonance -based metabolic fingerprinting of human serum and urine combined with multivariate data analysis could be a useful tool for differentiation between the active and remission phase of the disease that is of great importance in inflammatory bowel diseases (IBD) monitoring. At the same time, the results indicate that this diagnostic method has rather a weak potential in the differential diagnosis of IBD.