Clinical significance of serum diamine oxidase activity in inflammatory bowel disease

To the Editor: The intestinal mucosa serves as a major anatomic and functional barrier to potentially harmful intraluminal components such as bacteria and several antigens. The intestinal barrier is formed by epithelial cells and the junctional complex, including the tight junction (TJ) complex. Alterations of the composition of TJs was reported for both Crohn's disease (CD) and ulcerative colitis (UC) and consisted of upregulated expression of the pore-forming TJ protein claudin-2.1,2 In addition, the impaired permeability may represent the early onset of inflammatory bowel disease (IBD) because increased intestinal epithelial permeability precedes clinical relapse in patients with asymptomatic CD and in some healthy close relatives of the patients.3 These data suggest that small intestinal permeability plays a critical role in the disease onset of IBD and an examination of small intestinal permeability would be useful for diagnosing IBD and predicting disease relapse. We note with great interest the “Importance of disrupted intestinal barrier in inflammatory bowel disease” by Salim et al.4 In that review the authors highlighted the importance of intestinal barrier function in the pathogenesis of IBD and techniques for evaluating intestinal permeability in vivo and in vitro. However, it is not easy to evaluate small intestinal permeability in vivo, despite several modalities, such as the cellobiose/mannitol permeability test or 51Cr-EDTA test. Diamine oxidase (DAO) is an enzyme that catalyzes the oxidation of diamines such as histamine, putrescine, and cadaverine.5 In humans and rodents, DAO is specifically located at the apical end of mature villous cells with high activity and its activity reflects the integrity and maturity of the small intestinal mucosa. Several studies of humans and animals revealed that DAO activity in serum inversely correlates with intestinal permeability of small intestine. Ayuso et al6 reported that disease activity in patients with UC was related to the nonsynonymous single nucleotide polymorphism (SNP) in the DAO. On the other hand, a recent study demonstrated that a nonsynonymous SNP of the histamine-degrading enzyme DAO is not useful for assessing susceptibility to CD.7 Therefore, the significance of measuring DAO for IBD patients remains unclear. We measured serum DAO activity levels in patients with IBD and evaluated the clinical significance of DAO in IBD. Human serum samples were obtained from 13 patients with active CD (eight men and five women; mean age 35 ± 9 years), 42 patients (32 men and 10 women; mean age 36 ± 10 years) with inactive CD, 12 patients with active UC (five men and seven women; mean age 42 ± 20 years), 31 patients with inactive UC (19 men and 12 women; mean age 47 ± 20 years), and 17 healthy volunteers (14 men and 3 women; mean age 36 ± 10 years). All samples were centrifuged within 1 hour of collection and sera were stored at −80°C until assayed. The disease activities of patients with CD or UC were determined based on the Crohn's Disease Activity Index (CDAI) and Clinical Activity Index (CAI), respectively. A CDAI score greater than 150 and CAI score greater than 4 were defined active CD and active UC, respectively. Serum DAO activity was measured by enzyme-linked immunosorbent assay (ELISA) system according to the methods reported previously by Takagi et al.8 Informed consent was obtained from all patients and volunteers and the experimental design was approved by the Kyoto University Hospital Ethics Committee. Continuous variables were expressed as the mean (SE) and compared by paired t-test. P < 0.05 was considered statistically significant. Pearson correlation coefficient analysis was used to examine the relationship between serum DAO activity and C-reactive protein (CRP) or white blood cell count (WBC). Serum DAO activity in patients with CD (8.5 ± 2.4 units/L) was significantly lower than that in control subjects (10.3 ± 3.1 units/L) (Fig. 1A). In addition, serum DAO activity in patients with UC (8.9 ± 2.4 units/L) was significantly lower than that in control subjects (Fig. 1). There were no significant differences between patients with active CD (8.9 ± 2.2 units/L) and inactive CD (8.4 ± 2.5 units/L) or in patients with active UC (9.0 ± 2.7 units/L) and inactive UC (8.8 ± 2.3 units/L) (Fig. 1B). Furthermore, the disease phenotype of CD and UC was not associated with serum DAO activity (Fig. 1C). Serum DAO activity was not significantly correlated with CRP or WBC (Fig. 2). (A) Serum DAO activity in patients with IBD. Serum samples were obtained from patients with active CD (n = 13), inactive CD (n = 42), active UC (n = 12), inactive UC (n = 31), and healthy volunteers as normal controls (n = 17). Statistically significant compared to normal controls (P < 0.05). NS, not significant. (B) Comparison of serum DAO activity between patients with active and inactive CD or in patients with active and inactive UC. (C) Comparison of serum DAO activity among disease phenotype of CD and UC. Correlation between serum DAO activity and clinical parameters. This study first demonstrates that serum DAO activity is significantly lower in patients with CD and UC regardless of the level of disease activity. Of note, in patients with UC in whom small intestinal inflammation was not involved, serum DAO activity was significantly lower than that in healthy controls. These findings strongly indicate that small intestinal permeability is strongly involved in the pathophysiology of not only CD but also UC. A recent animal study showed that reducing small intestinal permeability using AT-1001 (a zonulin peptide inhibitor) attenuates colonic inflammation of interleukin-10 gene-deficient mice.9 Another study demonstrated increased small intestinal permeability in both senescence accelerated mice-prone and mdr1a-deficient mice prior to the development of colitis.10 In patients with IBD, a clinical trial with a CCR9 antagonist, which selectively blocks the migration of T cells into the small intestine, reduced disease activity in CD patients with colonic involvement.11 These data suggested that immune dysregulation in the small intestine would result in its increased epithelial permeability, which is clinically important as an etiologic event of human IBD. Another important finding of this study is that serum DAO activity was decreased in both UC and CD patients who were in remission. In this regard, we must focus on regulating mucosal permeability of the small intestine as a future therapeutic strategy for IBD patients Additionally, known genetic susceptibility variants of IBD together with the serum level of DAO activity may allow risk assessment of disease onset. In conclusion, measurement of serum DAO activity can be an easy and convenient modality for evaluating small intestinal permeability. Reduced serum DAO activity in patients with IBD suggests the importance of mucosal permeability of the small intestine as a pathogenic factor and its measurement together with serology, clinical factors, and genetics might be useful for predicting the disease onset of IBD. Further prospective studies with this modality are required.