Spontaneous and Microbiota-Driven Degradation of Anthocyanins in an In Vitro Human Colon Model.

SCOPE: The consumption of dietary anthocyanins is associated with various health benefits. However, anthocyanins are poorly bioavailable, and most ingested anthocyanins will enter the colon where they are degraded to small phenolic metabolites that are the main absorbed forms. Little is known about the processes of anthocyanin degradation in the gut and the role of the human gut microbiota. This study aims to determine the contribution of spontaneous and microbiota-dependent degradation of anthocyanins in the human colon. METHODS AND RESULTS: Purified anthocyanin extracts from black rice and bilberry were incubated in an in vitro human fecal-inoculated pH-controlled colon model over 24 h and anthocyanins were analyzed using HPLC-DAD. The study shows that the loss of anthocyanins occurs both spontaneously and as a consequence of metabolism by the gut microbiota. The study observes that there is high variability in spontaneous degradation but only modest variation in total degradation, which included the microbiota-dependent component. The degradation rate of anthocyanins is also shown to be dependent on the B-ring substitution pattern and the type of sugar moiety, both for spontaneous and microbiota-dependent degradation. CONCLUSION: Anthocyanins are completely degraded in a model of the human colon by a combination of spontaneous and microbiota-dependent processes.

Transcriptional and Post-Translational Regulation of Junctional Adhesion Molecule-B (JAM-B) in Leukocytes under Inflammatory Stimuli.

Junctional adhesion molecules (JAMs; comprising JAM-A, -B and -C) act as receptors for viruses, mediate cell permeability, facilitate leukocyte migration during sterile and non-sterile inflammation and are important for the maintenance of epithelial barrier integrity. As such, they are implicated in the development of both communicable and non-communicable chronic diseases. Here, we investigated the expression and regulation of JAM-B in leukocytes under pathogen- and host-derived inflammatory stimuli using immunoassays, qPCR and pharmacological inhibitors of inflammatory signalling pathways. We show that JAM-B is expressed at both the mRNA and protein level in leukocytes. JAM-B protein is localised to the cytoplasm, Golgi apparatus and in the nucleus around ring-shaped structures. We also provide evidence that JAM-B nuclear localisation occurs via the classical importin-α/ß pathway, which is likely mediated through JAM-B protein nuclear localisation signals (NLS) and export signals (NES). In addition, we provide evidence that under both pathogen- and host-derived inflammatory stimuli, JAM-B transcription is regulated via the NF-κB-dependent pathways, whereas at the post-translational level JAM-B is regulated by ubiquitin-proteosome pathways. Anaphase-promoting ubiquitin ligase complex (APC/C) and herpes simplex virus-associated ubiquitin-specific protease (HAUSP/USP) were identified as candidates for JAM-B ubiquitination and de-ubiquitination, respectively. The expression and regulation of JAM-B in leukocytes reported here is a novel observation and contrasts with previous reports. The data reported here suggest that JAM-B expression in leukocytes is under the control of common inflammatory pathways.

Microbiomes in physiology: insights into 21st-century global medical challenges.

NEW FINDINGS: What is the topic of this review? The role of the gut microbiome in physiology and how it can be targeted as an effective strategy against two of the most important global medical challenges of our time, namely, metabolic diseases and antibacterial resistance. What advances does it highlight? The critical roles of the microbiome in regulating host physiology and how microbiome analysis is useful for disease stratification to enable informed clinical decisions and develop interventions such as faecal microbiota transplantation, prebiotics and probiotics. Also, the limitations of microbiome modulation, including the potential for probiotics to enhance antimicrobial resistance gene reservoirs, and that currently a 'healthy microbiome' that can be used as a biobank for transplantation is yet to be defined. ABSTRACT: The human gut microbiome is a key factor in the development of metabolic diseases and antimicrobial resistance, which are among the greatest global medical challenges of the 21st century. A recent symposium aimed to highlight state-of-the-art evidence for the role of the gut microbiome in physiology, from childhood to adulthood, and the impact this has on global disease outcomes, ageing and antimicrobial resistance. Although the gut microbiome is established early in life, over time the microbiome and its components including metabolites can become perturbed due to changes such as dietary habits, use of antibiotics and age. As gut microbial metabolites, including short-chain fatty acids, secondary bile acids and trimethylamine-N-oxide, can interact with host receptors including G protein-coupled receptors and can alter host metabolic fluxes, they can significantly affect physiological homoeostasis leading to metabolic diseases. These metabolites can be used to stratify disease phenotypes such as irritable bowel syndrome and adverse events after heart failure and allow informed decisions on clinical management and treatment. While strategies such as use of probiotics, prebiotics and faecal microbiota transplantation have been proposed as interventions to treat and prevent metabolic diseases and antimicrobial resistance, caution must be exercised, first due to the potential of probiotics to enhance antimicrobial resistance gene reservoirs, and second, a 'healthy gut microbiome' that can be used as a biobank for transplantation is yet to be defined. We highlight that sampling other parts of the gastrointestinal tract may produce more representative data than the faecal microbiome alone.

Development and Validation of a LC-MS/MS Technique for the Analysis of Short Chain Fatty Acids in Tissues and Biological Fluids without Derivatisation Using Isotope Labelled Internal Standards.

The gut microbiota is critical to the maintenance of physiological homeostasis and as such is implicated in a range of diseases such as colon cancer, ulcerative colitis, diabetes, cardiovascular diseases, and neurodegenerative diseases. Short chain fatty acids (SCFAs) are key metabolites produced by the gut microbiota from the fermentation of dietary fibre. Here we present a novel, sensitive, and direct LC-MS/MS technique using isotopically labelled internal standards without derivatisation for the analysis of SCFAs in different biological matrices. The technique has significant advantages over the current widely used techniques based on sample derivatization and GC-MS analysis, including fast and simple sample preparation and short LC runtime (10 min). The technique is specific and sensitive for the quantification of acetate, butyrate, isobutyrate, isovalerate, lactate, propionate and valerate. The limits of detection were all 0.001 mM except for acetate which was 0.003 mM. The calibration curves for all the analytes were linear with correlation coefficients r2 > 0.998. The intra- and inter-day precisions in three levels of known concentrations were <12% and <20%, respectively. The quantification accuracy ranged from 92% to 120%. The technique reported here offers a valuable analytical tool for use in studies of SCFA production in the gut and their distribution to host tissues.

The use of an in-vitro batch fermentation (human colon) model for investigating mechanisms of TMA production from choline, L-carnitine and related precursors by the human gut microbiota.

PURPOSE: Plasma trimethylamine-N-oxide (TMAO) levels have been shown to correlate with increased risk of metabolic diseases including cardiovascular diseases. TMAO exposure predominantly occurs as a consequence of gut microbiota-dependent trimethylamine (TMA) production from dietary substrates including choline, carnitine and betaine, which is then converted to TMAO in the liver. Reducing microbial TMA production is likely to be the most effective and sustainable approach to overcoming TMAO burden in humans. Current models for studying microbial TMA production have numerous weaknesses including the cost and length of human studies, differences in TMA(O) metabolism in animal models and the risk of failing to replicate multi-enzyme/multi-strain pathways when using isolated bacterial strains. The purpose of this research was to investigate TMA production from dietary precursors in an in-vitro model of the human colon. METHODS: TMA production from choline, L-carnitine, betaine and γ-butyrobetaine was studied over 24-48 h using an in-vitro human colon model with metabolite quantification performed using LC-MS. RESULTS: Choline was metabolised via the direct choline TMA-lyase route but not the indirect choline-betaine-TMA route, conversion of L-carnitine to TMA was slower than that of choline and involves the formation of the intermediate γ-BB, whereas the Rieske-type monooxygenase/reductase pathway for L-carnitine metabolism to TMA was negligible. The rate of TMA production from precursors was choline > carnitine > betaine > γ-BB. 3,3-Dimethyl-1-butanol (DMB) had no effect on the conversion of choline to TMA. CONCLUSION: The metabolic routes for microbial TMA production in the colon model are consistent with observations from human studies. Thus, this model is suitable for studying gut microbiota metabolism of TMA and for screening potential therapeutic targets that aim to attenuate TMA production by the gut microbiota. TRIAL REGISTRATION NUMBER: NCT02653001 ( http://www.clinicaltrials.gov ), registered 12 Jan 2016.

Relationship between pemphigus and American tegumentary leishmaniasis: insights from serological and genetic profiles.

Background: Antibodies against Leishmania peptides (Lbr-peps) and desmogleins (Dsgs) have been reported in pemphigus foliaceus (PF) and leishmaniasis patients, respectively. We aimed to compare serological and genetic features in a Brazilian region endemic for American tegumentary leishmaniasis (ATL) and pemphigus. Methods: Commercial anti-Dsg ELISA and in-house ELISA with Lbr-peps were used to determine the serological profile, in addition to immunoblotting (IB) and indirect immunofluorescence (IIF) assays. HLA-DRB1 and -DQA1/DQB1 alleles were characterized by PCR combined with sequence-specific oligonucleotide probes (PCR-SSOP). The serological and genetic profiles were compared using 78 PF, 62 pemphigus vulgaris (PV) and 58 ATL patients against 163 and 1592 healthy controls, respectively. Results: Some ATL patients showed positive results for anti-Dsg1 and/or anti-Dsg3 antibodies. They also revealed 130, 160 and/or 230 kDa epidermal peptides in IB. Moreover, some ATL samples exhibited pemphigus or a bullous pemphigoid pattern in IIF. ELISA and IB assays showed Lbr-peps in pemphigus patients. HLA-DQA1*01 and -DQA1*01:02 were protective and susceptibility alleles for ATL, respectively, but the opposite for pemphigus. Conclusions: Anti-Dsgs in ATL may represent epiphenomena. Anti-Lbr-pep antibodies in pemphigus suggest a previous infection. A differential association of the HLA profile may contribute to the lack of co-association between pemphigus and ATL.

Evaluation by imaging methods of cochlear implant candidates: radiological and surgical correlation.

UNLABELLED: Evaluation by imaging methods is critical in the preoperative care of cochlear implant (CI) surgery, providing safety to surgeons when indicating and performing this procedure. The ideal imaging study consists of an association between Computed Tomography (CT) and Magnetic Resonance Imaging (MRI). AIM: To investigate the accuracy of imaging studies as predictors of possible complications of surgery. STUDY DESIGN: A cross-sectional investigation. MATERIAL AND METHOD: The medical records of 104 patients undergoing CI surgery between May 2003 and October 2006 were studied. The preoperative muldisciplinary selection process included CT associated or not with MRI. RESULTS: The final sample was composed of 100 patients after 4 patients with no records of radiological exams were excluded. Patients were divided into two groups. The accuracy of group A (CT only) was 69.69%, the sensitivity was 36.36%, the specificity was 86.36%, the Positive Predictive Value (PPV) was 57.14%, and the Negative Predictive Value (NPV) was 73.07%; the accuracy of group B (CT and MRI) was 80.59%, the sensitivity was 38.46%, the specificity was 90.74%, the PPV was 50.0%, and the NPV was 85.96%. CONCLUSION: The preoperative radiological evaluation by CI was effective in identifying anatomic abnormalities, allowing surgeons to avoid, or at least be aware of, possible complications. This study demonstrated that CT and MRI were superior to CT alone.

Avaliação por imagem nos candidatos ao implante coclear: correlação radiológico-cirúrgica / Evaluation by imaging methods of cochlear implant candidates: radiological and surgical correlation

No pré-operatório da cirurgia de Implante Coclear (IC) o estudo por imagem adquire importância fundamental, proporcionando ao cirurgião segurança na indicação e realização da cirurgia. A avaliação ideal compreende a associação entre Tomografia Computadorizada (TC) e Ressonância Nuclear Magnética (RNM). OBJETIVO: Avaliar a acurácia dos exames de imagem como preditores de possíveis complicações durante o ato cirúrgico. Desenho do Estudo: Corte transversal. MATERIAL E MÉTODOS: Foram avaliados por prontuários, 104 pacientes submetidos à cirurgia de IC de maio de 2003 a outubro de 2006. Estes passaram por uma seleção multidisciplinar pré-operatória, e radiológica por TC associada ou não à RNM. RESULTADOS: Excluiu-se 4 pacientes sem registro de exames radiológicos, resultando 100 pacientes. Estes foram divididos em dois grupos: grupo A (TC) apresentou acurácia 69,69 por cento, sensibilidade 36,36 por cento; especificidade 86,36 por cento; Valor Preditivo Positivo (VPP) 57,14 por cento; Valor Preditivo Negativo (VPN) 73,07 por cento e o B (TC e RNM) acurácia 80,59 por cento, sensibilidade 38,46 por cento; especificidade 90,74 por cento; VPP 50,0 por cento; VPN 85,96 por cento. CONCLUSÕES: A avaliação radiológica pré-operatória do IC mostrou-se importante, sendo capaz de identificar alterações anatômicas, evitando, ou ao menos, preparando o cirurgião para possíveis complicações. Esse estudo demonstrou que a avaliação associada com TC e RNM pôde ser considerada superior à TC isoladamente.

Evaluation by imaging methods is critical in the preoperative care of cochlear implant (CI) surgery, providing safety to surgeons when indicating and performing this procedure. The ideal imaging study consists of an association between Computed Tomography (CT) and Magnetic Resonance Imaging (MRI). AIM: To investigate the accuracy of imaging studies as predictors of possible complications of surgery. Study Design: A cross-sectional investigation. MATERIAL AND METHOD: The medical records of 104 patients undergoing CI surgery between May 2003 and October 2006 were studied. The preoperative muldisciplinary selection process included CT associated or not with MRI. RESULTS: The final sample was composed of 100 patients after 4 patients with no records of radiological exams were excluded. Patients were divided into two groups. The accuracy of group A (CT only) was 69.69 percent, the sensitivity was 36.36 percent, the specificity was 86.36 percent, the Positive Predictive Value (PPV) was 57.14 percent, and the Negative Predictive Value (NPV) was 73.07 percent; the accuracy of group B (CT and MRI) was 80.59 percent, the sensitivity was 38.46 percent, the specificity was 90.74 percent, the PPV was 50.0 percent, and the NPV was 85.96 percent. CONCLUSION: The preoperative radiological evaluation by CI was effective in identifying anatomic abnormalities, allowing surgeons to avoid, or at least be aware of, possible complications. This study demonstrated that CT and MRI were superior to CT alone.