MicroRNA Expression and Intestinal Permeability in Children Living in a Slum Area of Bangladesh.

Introduction: MicroRNAs (miRNAs) are small, non-coding RNAs that post-transcriptionally regulate gene expression. Changes in miRNA expression have been reported in a number of intestinal diseases, in both tissue samples and readily accessible specimens like stools. Pathogenic infections, diet, toxins, and other environmental factors are believed to influence miRNA expression. However, modulation of miRNAs in humans is yet to be thoroughly investigated. In this study, we examined the expression levels of two human miRNAs (miRNA-122 and miRNA-21) in stool samples of a group of Bangladeshi children who had an altered/increased intestinal permeability (IIP). Methods: Stool samples were collected from children with IIP (L:M > 0.09) and normal intestinal permeability (NIP) (L:M ≤ 0.09). Quantitative PCR was performed to quantify the levels of miRNA-122 and miR-21 in stools. Commercial ELISA kits were used to measure gut inflammatory markers Calprotectin and REG1B. Serum samples were tested using Human Bio-Plex Pro Assays to quantify IL-1ß, IL-2, IL-5, IL-10, IL-13, IFN-γ, and TNF-α. Total nucleic acid extracted from stool specimens were used to determine gut pathogens using TaqMan Array Card (TAC) system real-time polymerase chain reaction. Results: The expression levels of miRNA-122 (fold change 11.6; p < 0.001, 95% CI: 6.14-11.01) and miR-21 (fold change 10; p < 0.001, 95% CI: 5.05-10.78) in stool were upregulated in children with IIP than in children with normal intestinal permeability (NIP). Significant correlations were observed between stool levels of miR-122 and miR-21 and the inflammatory cytokines IL-1ß, IL-2, IFN-γ, and TNF-α (p < 0.05). Children with IIP were frequently infected with rotavirus, Campylobacter jejuni, Bacteroides fragilis, adenovirus, norovirus, astrovirus, and various Escherichia coli strains (ETEC_STh, ETEC_STp, EAEC_aaiC, EAEC_aatA) (p < 0.001). miR-122 significantly correlated with the fecal inflammatory biomarkers REG1B (p = 0.015) and Calprotectin (p = 0.030), however miR-21 did not show any correlation with these fecal biomarkers.

Fecal MicroRNAs as Potential Biomarkers for Screening and Diagnosis of Intestinal Diseases.

MicroRNAs (miRNAs) are a class of conserved endogenous, small non-coding RNA molecules with a length of 18-25 nucleotides that regulate gene expression by RNA interference processes, including mRNA chopping, mRNA deadenylation, and translation inhibition. miRNAs maintain the physiological functions of the intestine and are instrumental in gut pathogenesis. miRNAs play an important role in intercellular communication and are present in all body fluids, including stools with different composition and concentrations. However, under diseased conditions, miRNAs are aberrantly expressed and act as negative regulators of gene expression. The stable and differentially expressed miRNAs in stool enables miRNAs to be used as potential biomarkers for screening of various intestinal diseases. In this review, we summarize the expressed miRNA profile in stool and highlight miRNAs as biomarkers with potential clinical and diagnostic applications, and we aim to address the prospects for recent advanced techniques for screening miRNA in diagnosis and prognosis of intestinal disorders.

Screening for coeliac disease in children and adults living in a slum of Dhaka, Bangladesh.

BACKGROUND AND OBJECTIVE: Serological screening with a confirmation through biopsy has improved the understanding of coeliac disease (CD) epidemiology worldwide. Prevalence of CD in Bangladesh is not yet explored and therefore, we aimed to assess the seroprevalence of CD in slum-dwelling malnourished children and adults in Dhaka. METHODS: Serum samples were collected from three different cohorts: stunted (length-for-age Z-scores (LAZ) <-2) and at risk of stunting children (LAZ <-1 to -2) and malnourished adults (body mass index <18.5 kg/m2). Samples from all the participants were assessed for anti-tissue transglutaminase antibody (tTG-IgA) and total serum IgA by ELISA. Positive tTG-IgA and randomly selected low IgA values were reconfirmed using anti-tTG-IgG and gliadin IgG ELISA. CD was diagnosed when second screening tests were found positive and the participants were further investigated by small bowel biopsy. RESULTS: A total of 818 participants (240 stunted, 272 at risk of stunting children and 306 malnourished adults) were enrolled in the study. Overall, anti-tTG-IgA was positive in 5/818 (0.6%; 95% CI 0.25% to 1.46%). Of the five positive cases, anti-tTG-IgG and gliadin IgG were found positive in only one participant. Duodenal biopsy of positive participant revealed characteristic lesions of CD. Randomly selected low IgA values were found negative in tTG-IgG and gliadin IgG for all the participants. No participant was found total IgA deficient. CONCLUSION: The incidence of coeliac autoimmunity is low in malnourished slum dwellers regardless of age in Bangladesh. It is important to investigate the nationwide prevalence to reveal the definite picture.

Role of Eosinophils and Tumor Necrosis Factor Alpha in Interleukin-25-Mediated Protection from Amebic Colitis.

The parasite Entamoeba histolytica is a cause of diarrhea in infants in low-income countries. Previously, it was shown that tumor necrosis factor alpha (TNF-α) production was associated with increased risk of E. histolytica diarrhea in children. Interleukin-25 (IL-25) is a cytokine that is produced by intestinal epithelial cells that has a role in maintenance of gut barrier function and inhibition of TNF-α production. IL-25 expression was decreased in humans and in the mouse model of amebic colitis. Repletion of IL-25 blocked E. histolytica infection and barrier disruption in mice, increased gut eosinophils, and suppressed colonic TNF-α. Depletion of eosinophils with anti-Siglec-F antibody prevented IL-25-mediated protection. In contrast, depletion of TNF-α resulted in resistance to amebic infection. We concluded that IL-25 provides protection from amebiasis, which is dependent upon intestinal eosinophils and suppression of TNF-α.IMPORTANCE The intestinal epithelial barrier is important for protection from intestinal amebiasis. We discovered that the intestinal epithelial cytokine IL-25 was suppressed during amebic colitis in humans and that protection could be restored in the mouse model by IL-25 administration. IL-25 acted via eosinophils and suppressed TNF-α. This work illustrates a previously unrecognized pathway of innate mucosal immune response.

Role of Serum Amyloid A, Granulocyte-Macrophage Colony-Stimulating Factor, and Bone Marrow Granulocyte-Monocyte Precursor Expansion in Segmented Filamentous Bacterium-Mediated Protection from Entamoeba histolytica.

Intestinal segmented filamentous bacteria (SFB) protect from ameba infection, and protection is transferable with bone marrow dendritic cells (BMDCs). SFB cause an increase in serum amyloid A (SAA), suggesting that SAA might mediate SFB's effects on BMDCs. Here we further explored the role of bone marrow in SFB-mediated protection. Transient gut colonization with SFB or SAA administration alone transiently increased the H3K27 histone demethylase Jmjd3, persistently increased bone marrow Csf2ra expression and granulocyte monocyte precursors (GMPs), and protected from ameba infection. Pharmacologic inhibition of Jmjd3 H3K27 demethylase activity during SAA treatment or blockade of granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling in SFB-colonized mice prevented GMP expansion, decreased gut neutrophils, and blocked protection from ameba infection. These results indicate that alteration of the microbiota and systemic exposure to SAA can influence myelopoiesis and susceptibility to amebiasis via epigenetic mechanisms. Gut microbiota-marrow communication is a previously unrecognized mechanism of innate protection from infection.

Interleukin-25 Mediated Induction of Angiogenin-4 Is Interleukin-13 Dependent.

The intestinal surface is directly exposed to both commensal microorganisms as well as pathogens with a single layer of epithelium separating luminal microorganisms from internal tissues. Antimicrobial peptides play a crucial role in allowing epithelial cells to contain in the lumen beneficial and pathogenic microorganisms. The commensal dependent, epithelial produced, Th2 cytokine IL-25 can induce IL-13 and potentially the antimicrobial peptide angiogenin-4. Here we show that IL-13 downstream of IL-25 is required to induce angiogenin-4. IL-25 mediated induction of angiogenin-4 is furthermore not dependent on IL-22 or IL-17.

Malnutrition and helminth infection affect performance of an interferon gamma-release assay.

OBJECTIVE: We sought to compare the tuberculin skin test (TST) to the QuantiFERON-TB Gold In-Tube assay (QFT-IT) and assess the effects of malnourishment and intestinal helminth infection on QFT-IT results. METHODS: In this population-based cross-sectional study from Dhaka, Bangladesh, we screened children for latent tuberculosis infection with the QFT-IT and TST. We assess the agreement between the TST and QFT-IT, risk factors associated with indeterminate QFT-IT results, and magnitude of interferon γ (IFN-γ) production. RESULTS: Three hundred and two children (aged 11-15.3 years) were enrolled, including 93 (30.8%) who were malnourished. Of 251 participants who provided stool samples, 117 (46.6%) were infected with Ascaris lumbricoides and/or Trichuris trichiura. TST results were positive (≥10 mm) for 101 (33.4%) children and negative for 201 (66.6%) children. QFT-IT results were positive for 107 (35.4%) children, negative for 121 (40.1%) children, and indeterminate for 74 (24.5%) children. Agreement between the tests was moderate (κ = 0.55 [95% confidence interval: 0.44-0.65]; P < .0001) when excluding indeterminate results. Children with indeterminate QFT-IT results were separately compared with children with positive and negative QFT-IT results; malnutrition (P = .0006 and .0003), and helminth infection (P = .05 and .02), and the statistical interaction between these 2 terms (P = .03 and .004) were associated with indeterminate results. Higher levels of IFN-γ in response to tuberculosis antigens were associated with positive TST results (P < .0001); lower levels were associated with malnutrition (P = .02). CONCLUSIONS: Malnutrition and helminth infections were associated with indeterminate QFT-IT results. Therefore, the presence of such conditions may limit the interpretability of QFT-IT results in children.