Metabolic control of T cell immune response through glycans in inflammatory bowel disease.

Mucosal T lymphocytes from patients with ulcerative colitis (UC) were previously shown to display a deficiency in branched N-glycosylation associated with disease severity. However, whether this glycosylation pathway shapes the course of the T cell response constituting a targeted-specific mechanism in UC remains largely unknown. In this study, we demonstrated that metabolic supplementation of ex vivo mucosal T cells from patients with active UC with N-acetylglucosamine (GlcNAc) resulted in enhancement of branched N-glycosylation in the T cell receptor (TCR), leading to suppression of T cell growth, inhibition of the T helper 1 (Th1)/Th17 immune response, and controlled T cell activity. We further demonstrated that mouse models displaying a deficiency in the branched N-glycosylation pathway (MGAT5-/-, MGAT5+/-) exhibited increased susceptibility to severe forms of colitis and early-onset disease. Importantly, the treatment of these mice with GlcNAc reduced disease severity and suppressed disease progression due to a controlled T cell-mediated immune response at the intestinal mucosa. In conclusion, our human ex vivo and preclinical results demonstrate the targeted-specific immunomodulatory properties of this simple glycan, proposing a therapeutic approach for patients with UC.

The prognostic impact of TERT promoter mutations in glioblastomas is modified by the rs2853669 single nucleotide polymorphism.

Human hotspot TERT promoter (TERTp) mutations have been reported in a wide range of tumours. Several studies have shown that TERTp mutations are associated with clinicopathological features; in some instances, TERTp mutations were considered as biomarkers of poor prognosis. The rs2853669 SNP, located in the TERT promoter region, was reported to modulate the increased TERT expression levels induced by the recurrent somatic mutations. In this study we aimed to determine the frequency and prognostic value of TERTp mutations and TERT rs2853669 SNP in 504 gliomas from Portuguese and Brazilian patients. TERTp mutations were detected in 47.8% of gliomas (216/452). Glioblastomas (GBM) exhibited the highest frequency of TERTp mutations (66.9%); in this glioma subtype, we found a significant association between TERTp mutations and poor prognosis, regardless of the population. Moreover, in a multivariate analysis, TERTp mutations were the only independent prognostic factor. Our data also showed that the poor prognosis conferred by TERTp mutations was restricted to GBM patients carrying the rs2853669 A allele and not in those carrying the G allele. In conclusion, the presence of TERTp mutations was associated with worse prognosis in GBM patients, although such association depended on the status of the rs2853669 SNP. The status of the rs2853669 SNP should be taken in consideration when assessing the prognostic value of TERTp mutations in GBM patients. TERTp mutations and the rs2853669 SNP can be used in the future as biomarkers of glioma prognosis.

Genetic Variants of the MGAT5 Gene Are Functionally Implicated in the Modulation of T Cells Glycosylation and Plasma IgG Glycome Composition in Ulcerative Colitis.

OBJECTIVES: The impact of genetic variants (single nucleotide polymorphisms [SNPs]) in the clinical heterogeneity of ulcerative colitis (UC) remains unclear. We showed that patients with UC exhibit a deficiency in MGAT5 glycogene transcription in intestinal T cells associated with a hyperimmune response. Herein, we evaluated whether MGAT5 SNPs might functionally impact on T cells glycosylation and plasma IgG glycome in patients with UC, as well as in UC clinical outcomes. METHODS: Three selected MGAT5 SNPs (rs3814022, rs4953911, and rs1257220), previously associated with severity of autoimmune disease or with plasma glycome composition in healthy individuals, were functionally evaluated in patients with UC through analysis of MGAT5 mRNA levels in colonic (n = 14) and circulating (n = 24) T cells and through profiling the plasma IgG Fc glycosylation (n = 152). MGAT5 SNPs were genotyped in 931 patients with UC from 2 European cohorts and further associated with patients' prognosis. Targeted next-generation sequencing for MGAT5 coding and regulatory regions was also performed. RESULTS: MGAT5 SNPs were shown to be functionally associated with low transcription levels of MGAT5 in colonic and circulating T cells from patients with UC and with agalactosylation of IgGs, often associated with a proinflammatory phenotype. The SNPs rs3814022 and rs4953911 were further associated with the need of biologics. Next-generation sequencing data further revealed a combination of MGAT5 SNPs that stratify patients with UC according to their severity. DISCUSSION: Our results revealed that MGAT5 SNPs have a phenotypic impact on T cells glycosylation and in plasma IgG glycome composition associated with UC pathogenesis. MGAT5 SNPs display a tendency in the association with a worse disease course in patients with UC.

The Effect of Inflammatory Status on Butyrate and Folate Uptake by Tumoral (Caco-2) and Non-Tumoral (IEC-6) Intestinal Epithelial Cells.

OBJECTIVE: Colorectal cancer (CRC) is the second leading cause of cancer death in occidental countries. Chronic inflammatory bowel disease (crohn's disease and ulcerative colitis) is associated with an increased risk for CRC development. The aim of this work was to investigate the relationship between inflammatory status and absorption of nutrients with a role in CRC pathogenesis. MATERIALS AND METHODS: In this experimental study, we evaluated the in vitro effect of tumour necrosis factor-alpha (TNF-α), interferon-γ (IF-γ), and acetylsalicylic acid on 14C-butyrate (14C- BT), 3H-folic acid (3H-FA) uptake, and on proliferation, viability and differentiation of Caco-2 and IEC-6 cells in culture. RESULTS: The proinflammatory cytokines TNF-α and INF-γ were found to decrease uptake of a low concentration of 14C-BT (10 µM) by Caco-2 (tumoral) and IEC-6 (normal) intestinal epithelial cell lines. However, the effect of TNF-α and INF-γ in IEC-6 cells is most probably related to a cytotoxic and antiproliferative impact. In contrast, INF-γ increases uptake of a high concentration (10 mM) of 14C-BT in Caco-2 cells. The anticarcinogenic effect of BT (10 mM) in these cells is not affected by the presence of this cytokine. On the other hand, acetylsalicylic acid stimulates 14C-BT uptake by Caco-2 cells and potentiates its antiproliferative effect. Finally, both TNF-α and INF-γ cause a significant decrease in 3H-FA uptake by Caco-2 cells. CONCLUSION: The inflammatory status has an impact upon cellular uptake of BT and FA, two nutrients with a role in CRC pathogenesis. Moreover, the anti-inflammatory acetylsalicylic acid potentiates the anticarcinogenic effect of BT in Caco-2 cells by increasing its cellular uptake.

The effect of oxidative stress upon the intestinal epithelial uptake of butyrate.

Our aim was to investigate the effect of oxidative stress upon butyrate uptake at the intestinal epithelial level. For this, IEC-6 cells were treated with tert-butylhydroperoxide 3000µM (tBOOH), which increased levels of oxidative stress biomarkers, while maintaining cellular viability. The effect of tBOOH upon uptake of [(14)C]butyrate ([(14)C]BT) (10µM) can be summarized as follows: (a) it caused a reduction in the intracellular accumulation of [(14)C]BT over time, (b) it strongly reduced total [(14)C]BT uptake but did not affect Na(+)-independent uptake of [(14)C]BT, and (c) it did not affect the kinetics of [(14)C]BT uptake at 37°C, but increased uptake at 4°C. Moreover, tBOOH increased the efflux of [(14)C]BT not mediated by breast cancer resistance protein. We thus conclude that tBOOH strongly inhibits Na(+)-coupled monocarboxylate cotransporter 1 (SMCT1)-mediated, but not H(+)-coupled monocarboxylate transporter (MCT1)-mediated butyrate uptake; moreover, it increases uptake and efflux of butyrate by passive diffusion. tBOOH did not affect the mRNA expression levels of MCT1 and SMCT1 nor their cell membrane insertion. Rather, its effect was dependent on extracellular signal regulated kinase 1/2 and protein tyrosine kinase activation and on the generation of reactive oxygen species by NADPH and xanthine oxidases and was partially prevented by the polyphenols quercetin and resveratrol. In conclusion, tBOOH is an effective inhibitor of SMCT1-mediated butyrate transport in non-tumoral intestinal epithelial cells. Given the important role played by butyrate in the intestine, this mechanism may contribute to the procarcinogenic and proinflammatory effect of oxidative stress at this level.