Butyrate-producing gut bacteria are reduced in patients with active inflammatory bowel disease (IBD), supporting the hypothesis that butyrate supplementation may be beneficial in this setting. Nonetheless, earlier studies suggest that the oxidation of butyrate in IBD patients is altered. We propose that inflammation may decrease epithelial butyrate consumption. Non-IBD controls and IBD patients were recruited for the study. Stool samples were used for short-chain fatty acid and bacterial butyryl CoA:acetate CoA-transferase quantification. Colonic biopsies and ex vivo differentiated epithelial organoids (d-EpOCs) treated with butyrate and/or tumor necrosis factor alpha (TNFα) were used for analyzing the expression of transporters MCT1 and ABCG2, metabolic enzyme ACADS, and butyrate receptor GPR43, and for butyrate metabolism and consumption assays. We observed that lower stool content of butyrate-producing bacteria in active IBD patients did not correlate with decreased butyrate concentrations. Indeed, the intestinal epithelial expression of MCT1, ABCG2, ACADS, and GPR43 was altered in active IBD patients. Nonetheless, d-EpOCs derived from IBD patients showed SLC16A1 (gene encoding for MCT1 protein), ABCG2, ACADS, and GPR43 expression levels comparable to controls. Moreover, IBD- and non-IBD-derived d-EpOCs responded similarly to butyrate, as assessed by transcriptional regulation. TNFα significantly altered SLC16A1, ABCG2, and GPR43 transcription in d-EpOCs, mimicking the expression profile observed in biopsies from active IBD patients and resulting in reduced butyrate consumption. We provide evidence that the response to butyrate is not intrinsically altered in IBD patients. However, TNFα renders the epithelium less responsive to this metabolite, defeating the purpose of butyrate supplementation during active inflammation.

The implementation of successful strategies to achieve an HIV cure has become a priority in HIV research. However, the current location and size of HIV reservoirs is still unknown since there are limited tools to evaluate HIV latency in viral sanctuaries such as gut-associated lymphoid tissue (GALT). As reported in the so called “Boston Patients”, despite undetectable levels of proviral HIV-1 DNA in blood and GALT, viral rebound happens in just few months after ART interruption. This fact might imply that current methods are not sensitive enough to detect residual reservoirs. Showing that, it is imperative to improve the detection and quantification of HIV-1 reservoir in tissue samples. Herein, we propose a novel non-enzymatic protocol for purification of Lamina Propria Leukocytes (LPL) from gut biopsies combined to viral HIV DNA (vDNA) quantification by droplet digital PCR (ddPCR) to improve the sensitivity and accuracy of viral reservoir measurements (LPL-vDNA assay). Endoscopic ileum biopsies were sampled from 12 HIV-1-infected cART-suppressed subjects. We performed a DTT/EDTA-based treatment for epithelial layer removal followed by non-enzymatic disruption of the tissue to obtain lamina propria cell suspension (LP). CD45⁺ cells were subsequently purified by flow sorting and vDNA was determined by ddPCR. vDNA quantification levels were significantly higher in purified LPLs (CD45⁺) than in bulk LPs (p<0.01). The levels of vDNA were higher in ileum samples than in concurrent PBMC from the same individuals (p = 0.002). As a result of the increased sensitivity of this purification method, the Poisson 95% confidence intervals of the vDNA quantification data from LPLs were narrower than that from bulk LPs. Of note, vDNA was unambiguously quantified above the detection limit in 100% of LPL samples, while only in 58% of bulk LPs. We propose an innovative combined protocol for a more sensitive detection of the HIV reservoir in gut-associated viral sanctuaries, which might be used to evaluate any proposed eradication strategy.

Background Some limited studies of coeliac disease have shown higher frequency of coeliac disease in infancy and adolescence than in adulthood. This finding has remained unnoticed and not adequately demonstrated. Aim To assess whether there are age and gender differences in coeliac disease prevalence. Methods A total of 4230 subjects were included consecutively (1 to ≥80 years old) reproducing the reference population by age and gender. Sample size was calculated assuming a population‐based coeliac disease prevalence of 1:250. After an interim analysis, the paediatric sample was expanded (2010 children) due to high prevalence in this group. Anti‐transglutaminase and antiendomysial antibodies were determined and duodenal biopsy was performed if positive. Log‐linear models were fitted to coeliac disease prevalence by age allowing calculation of percentage change of prevalence. Differences between groups were compared using Chi‐squared test. Results Twenty‐one subjects had coeliac disease (male/female 1:2.5). Coeliac disease prevalence in the total population was 1:204. Coeliac disease prevalence was higher in children (1:71) than in adults (1:357) (P = 0.00005). A significant decrease of prevalence in older generations was observed [change of prevalence by age of −5% (95% CI: −7.58 to −2.42%)]. In the paediatric expanded group (1–14 years), a decrease of coeliac disease prevalence was also observed [prevalence change: −17% (95% CI: −25.02 to −6.10)]. Conclusions The prevalence of coeliac disease in childhood was five times higher than in adults. Whether this difference is due to environmental factors influencing infancy, or latency of coeliac disease in adulthood, remains to be demonstrated in prospective longitudinal studies.

Innate lymphoid cells (ILCs) are highly plastic and predominantly mucosal tissue-resident cells that contribute to both homeostasis and inflammation depending on the microenvironment. The discovery of naïve-like ILCs suggests an ILC differentiation process that is akin to naïve T cell differentiation. Delineating the mechanisms that underlie ILC differentiation in tissues is crucial for understanding ILC biology in health and disease. Here, we showed that tonsillar ILCs expressing CD45RA lacked proliferative activity, indicative of cellular quiescence. CD62L distinguished two subsets of CD45RA ⁺ ILCs. CD45RA ⁺ CD62L ⁺ ILCs (CD62L ⁺ ILCs) resembled circulating naïve ILCs because they lacked the transcriptional, metabolic, epigenetic, and cytokine production signatures of differentiated ILCs. CD45RA ⁺ CD62L ⁻ ILCs (CD62L ⁻ ILCs) were epigenetically similar to CD62L ⁺ ILCs but showed a transcriptional, metabolic, and cytokine production signature that was more akin to differentiated ILCs. CD62L ⁺ and CD62L ⁻ ILCs contained uni- and multipotent precursors of ILC1s/NK cells and ILC3s. Differentiation of CD62L ⁺ and CD62L ⁻ ILCs led to metabolic reprogramming including up-regulation of genes associated with glycolysis, which was needed for their effector functions after differentiation. CD62L ⁻ ILCs with preferential differentiation capacity toward IL-22–producing ILC3s accumulated in the inflamed mucosa of patients with inflammatory bowel disease. These data suggested distinct differentiation potential of CD62L ⁺ and CD62L ⁻ ILCs between tissue microenvironments and identified that manipulation of these cells is a possible approach to restore tissue-immune homeostasis.

Lymphocytic (LC) and collagenous (CC) colitis are the two major forms of microscopic colitis (MC). The aim of this study was to identify similarities and differences in their mucosal immune characteristics. Colonic biopsies from 15 CC, 8 LC, and 10 healthy controls were collected. Mucosal lymphocytes were assessed by flow cytometry. Tissue gene expression and protein levels were determined by real-time PCR and ELISA, respectively. LC patients had lower numbers of CD4 ⁺ and double-positive CD4 ⁺ CD8 ⁺ mucosal T lymphocytes, and higher numbers of CD8 ⁺ and CD4 ⁺ TCRγδ ⁺ mucosal T cells, compared with controls and CC patients. Regulatory Treg (CD4 ⁺ CD25 ⁺ FOXP3 ⁺ ) and double-negative (CD3 ⁺ CD4 ^- CD8 ^- ) T cell percentages were higher in both CC and LC compared with controls, coupled with higher levels of the anti-inflammatory IL-10, both at mRNA and protein levels. By contrast, Th1 and Th17 cells were lower in both CC and LC, although gene expression of Th1/Th17 cytokines was higher in both. CC and LC share some regulatory and effector mechanisms, but not others. Higher IL-10 levels and higher Treg and double-negative T cell percentages, found in both CC and LC, could be responsible for the lack of progression of structural damage and the blockade of proinflammatory cytokine production. However, CC and LC are revealed as separate, independent entities, as they show clearly different mucosal lymphocyte profiles, which could be caused by different luminal triggers of the two diseases. Hence, CC and LC are two closely related but independent intestinal disorders.

An increase in CD3+TCRγδ+ and a decrease in CD3− intraepithelial lymphocytes (IEL) is a characteristic flow cytometric pattern of celiac disease (CD) with atrophy. The aim was to evaluate the usefulness of both CD IEL cytometric pattern and anti-TG2 IgA subepithelial deposit analysis (CD IF pattern) for diagnosing lymphocytic enteritis due to CD. Two-hundred and five patients (144 females) who underwent duodenal biopsy for clinical suspicion of CD and positive celiac genetics were prospectively included. Fifty had villous atrophy, 70 lymphocytic enteritis, and 85 normal histology. Eight patients with non-celiac atrophy and 15 with lymphocytic enteritis secondary to Helicobacter pylori acted as control group. Duodenal biopsies were obtained to assess both CD IEL flow cytometric (complete or incomplete) and IF patterns. Sensitivity of IF, and complete and incomplete cytometric patterns for CD diagnosis in patients with positive serology (Marsh 1+3) was 92%, 85 and 97% respectively, but only the complete cytometric pattern had 100% specificity. Twelve seropositive and 8 seronegative Marsh 1 patients had a CD diagnosis at inclusion or after gluten free-diet, respectively. CD cytometric pattern showed a better diagnostic performance than both IF pattern and serology for CD diagnosis in lymphocytic enteritis at baseline (95% vs 60% vs 60%, p = 0.039). Analysis of the IEL flow cytometric pattern is a fast, accurate method for identifying CD in the initial diagnostic biopsy of patients presenting with lymphocytic enteritis, even in seronegative patients, and seems to be better than anti-TG2 intestinal deposits.