Oral versus intravenous iron replacement therapy distinctly alters the gut microbiota and metabolome in patients with IBD

Abstract

Objective: Iron deficiency is a common complication in patients with IBD and oral iron therapy is suggested to exacerbate IBD symptoms. We performed an open-labelled clinical trial to compare the effects of per oral (PO) versus intravenous (IV) iron replacement therapy (IRT).

Design: The study population included patients with Crohn's disease (CD; N=31), UC (N=22) and control subjects with iron deficiency (non-inflamed, NI=19). After randomisation, participants received iron sulfate (PO) or iron sucrose (IV) over 3 months. Clinical parameters, faecal bacterial communities and metabolomes were assessed before and after intervention.

Results: Both PO and IV treatments ameliorated iron deficiency, but higher ferritin levels were observed with IV. Changes in disease activity were independent of iron treatment types. Faecal samples in IBD were characterised by marked interindividual differences, lower phylotype richness and proportions of Clostridiales. Metabolite analysis also showed separation of both UC and CD from control anaemic participants. Major shifts in bacterial diversity occurred in approximately half of all participants after IRT, but patients with CD were most susceptible. Despite individual-specific changes in phylotypes due to IRT, PO treatment was associated with decreased abundances of operational taxonomic units assigned to the species Faecalibacterium prausnitzii, Ruminococcus bromii, Dorea sp. and Collinsella aerofaciens. Clear IV-specific and PO-specific fingerprints were evident at the level of metabolomes, with changes affecting cholesterol-derived host substrates.

Conclusions: Shifts in gut bacterial diversity and composition associated with iron treatment are pronounced in IBD participants. Despite similar clinical outcome, oral administration differentially affects bacterial phylotypes and faecal metabolites compared with IV therapy.

Trial registration number: clinicaltrial.gov (NCT01067547).

Keywords: ANEMIA; IBD CLINICAL; INFLAMMATORY BOWEL DISEASE; INTESTINAL BACTERIA; IRON DEFICIENCY.

Figure 1

Clinical  design and outcomes. (A) Anaemic subjects diagnosed with IBD and control anaemic participants without GI disorders were enrolled in the study and randomised into the oral or intravenous iron replacement therapy (IRT) group. Faecal samples were collected at baseline and after the end of intervention (3 months later). Samples were analysed by high-throughput 16S rRNA gene sequencing and Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR-mass spectrometry (MS)) for analysis of bacterial communities and metabolites, respectively. (B) IRT restores iron storage, independently of the route of administration. (C) IBD activity was assessed with Modified Harvey Bradshaw Index (HBI) for CD and Partial Mayo (pMayo) score for UC. Values are % changes. (D) % changes in quality of life per participants category. Quality of life was scored according to Shorten IBD Questionnaire (SIBDQ) and Euro Quality 5 Dimensions Visual Analogue Scale (EQ5D). CD, Crohn's disease; CRP, C reactive protein; EQ5D, Euro Quality 5 Dimensions; IV, intravenous; NI, non-IBD control anaemic subjects; PO, per oral; SIBDQ, Shorten IBD Questionnaire.

Figure 2

IBD-specific features of faecal microbiota and metabolomes. (A) NMDS plot based on generalised UniFrac distances, including all patients before and after iron replacement therapy, indicating that bacterial communities in patients with CD are most heterogeneous and distant from UC and control subjects. (B) Shannon effective diversity boxplots display decreased numbers of dominant molecular species in patients with IBD. (C) Relative abundances of dominant bacterial taxa with significant difference between subject categories. The numbers of samples, in which the given bacteria were detected, are shown in brackets below categories. (D) Score plot of orthogonal partial least squares discriminant analysis (OPLS-DA) classification of non-inflamed (NI) control anaemic participants (blue circles) and patients with CD (red circles) at baseline. Learning the model resulted in one predictive and one orthogonal component (x-axis and y-axis, respectively). (E) Top features mostly responsible for classification in the corresponding OPLS-DA model in (D). The features were chosen according to the values of variable importance in projection scores. P values obtained by Mann–Whitney test are shown. (F) Score plot of OPLS-DA classification of NI control anaemic participants (blue bars) and patients with UC (green bars) at baseline. Learning the model resulted in only one predictive component (x-axis). The y-axis indicates individual samples in each group. (G) Top features mostly responsible for classification in the corresponding OPLS-DA model in (F). ANOVA, analysis of variance; B, baseline; CD, Crohn's disease; NI, non-IBD control anaemic subjects; NMDS, non-metric multidimensional scaling. *< 0.05; ***< 0.01.

Figure 3

The gut environment in patients with IBD versus control anaemic subjects is more sensitive to overtime changes associated with iron replacement therapy (IRT). (A) NMDS plot of phylogenetic distances showing overall changes in bacterial diversity before and after treatment (connecting lines). (B) Boxplots depict the distribution of generalised UniFrac distances per patient category, indicating that samples before and after IRT are more similar in non-inflamed (NI) control subjects than in patients with IBD. (C) Wards’ clustering dendrogram calculated from phylogenetic distances between all samples. Paired samples (located next to each other before and after IRT for one given individual) are marked with black bars (inner ring). Disease categories were colour-coded in the outer ring as follows: red, Crohn's disease (CD); green, UC and blue, NI. Percentages of paired samples per patient category are shown in the barplot. The p value was obtained using a χ² test. (D) Influence of iron treatment on classification between control and CD participants. The position of points corresponding to the samples from after IRT (blue and red triangles) was predicted using the orthogonal partial least squares discriminant analysis (OPLS-DA) model built for classification of NI and CD samples at baseline (blue and red circles). ANOVA, analysis of variance; NMDS, non-metric multidimensional scaling.

Figure 4

PO iron treatment is associated with specific features in faecal samples. (A) Iron and total sulfur content in faeces. (B) Relative abundance of phylotypes that reacted differently to intravenous (IV) or oral iron therapy. IRT groups were compared using Mann–Whitney test. Prior to testing, individual counts <0.5% were zeroed, and only prevalent and dominant molecular species (ie, detected in at least 30% of the participants (n=124) and with a median sequence abundance >1% in at least one group) were considered. (C) Score plot of orthogonal partial least squares discriminant analysis (OPLS-DA) classification of IV-IRT (beige) versus PO-IRT (brown). Learning the model resulted in one predictive and one orthogonal component (x-axis and y-axis, respectively). (D) Top features responsible for classification in the OPLS-DA model are shown as boxplots. B, baseline; IRT, iron replacement therapy; OTU, operational taxonomic unit; PO, per oral.