An ex-vivo Human Intestinal Model to Study Entamoeba histolytica Pathogenesis

Amoebiasis (a human intestinal infection affecting 50 million people every year) is caused by the protozoan parasite Entamoeba histolytica. To study the molecular mechanisms underlying human colon invasion by E. histolytica, we have set up an ex vivo human colon model to study the early steps in amoebiasis. Using scanning electron microscopy and histological analyses, we have established that E. histolytica caused the removal of the protective mucus coat during the first two hours of incubation, detached the enterocytes, and then penetrated into the lamina propria by following the crypts of Lieberkühn. Significant cell lysis (determined by the release of lactodehydrogenase) and inflammation (marked by the secretion of pro-inflammatory molecules such as interleukin 1 beta, interferon gamma, interleukin 6, interleukin 8 and tumour necrosis factor) were detected after four hours of incubation. Entamoeba dispar (a closely related non-pathogenic amoeba that also colonizes the human colon) was unable to invade colonic mucosa, lyse cells or induce an inflammatory response. We also examined the behaviour of trophozoites in which genes coding for known virulent factors (such as amoebapores, the Gal/GalNAc lectin and the cysteine protease 5 (CP-A5), which have major roles in cell death, adhesion (to target cells or mucus) and mucus degradation, respectively) were silenced, together with the corresponding tissue responses. Our data revealed that the signalling via the heavy chain Hgl2 or via the light chain Lgl1 of the Gal/GalNAc lectin is not essential to penetrate the human colonic mucosa. In addition, our study demonstrates that E. histolytica silenced for CP-A5 does not penetrate the colonic lamina propria and does not induce the host's pro-inflammatory cytokine secretion

Anti-Hu antibodies activate enteric and sensory neurons.

IgG of type 1 anti-neuronal nuclear antibody (ANNA-1, anti-Hu) specificity is a serological marker of paraneoplastic neurological autoimmunity (including enteric/autonomic) usually related to small-cell lung carcinoma. We show here that IgG isolated from such sera and also affinity-purified anti-HuD label enteric neurons and cause an immediate spike discharge in enteric and visceral sensory neurons. Both labelling and activation of enteric neurons was prevented by preincubation with the HuD antigen. Activation of enteric neurons was inhibited by the nicotinic receptor antagonists hexamethonium and dihydro-β-erythroidine and reduced by the P2X antagonist pyridoxal phosphate-6-azo (benzene-2,4-disulfonic acid (PPADS) but not by the 5-HT3 antagonist tropisetron or the N-type Ca-channel blocker ω-Conotoxin GVIA. Ca(++) imaging experiments confirmed activation of enteric neurons but not enteric glia. These findings demonstrate a direct excitatory action of ANNA-1, in particular anti-HuD, on visceral sensory and enteric neurons, which involves nicotinic and P2X receptors. The results provide evidence for a novel link between nerve activation and symptom generation in patients with antibody-mediated gut dysfunction

Excitability and synaptic transmission in the enteric nervous system: Does diet play a role?

© Springer International Publishing Switzerland 2016. Changes in diet are a challenge to the gastrointestinal tract which needs to alter its processing mechanisms to continue to process nutrients and maintain health. In particular, the enteric nervous system (ENS) needs to adapt its motor and secretory programs to deal with changes in nutrient type and load in order to optimise nutrient absorption. The nerve circuits in the gut are complex, and the numbers and types of neurons make recordings of specific cell types difficult, time-consuming, and prone to sampling errors. Nonetheless, traditional research methods like intracellular electrophysiological approaches have provided the basis for our understanding of the ENS circuitry. In particular, animal models of intestinal inflammation have shown us that we can document changes to neuronal excitability and synaptic transmission. Recent studies examining diet-induced changes to ENS programming have opted to use fast imaging techniques to reveal changes in neuron function. Advances in imaging techniques using voltage- or calcium-sensitive dyes to record neuronal activity promise to overcome many limitations inherent to electrophysiological approaches. Imaging techniques allow access to a wide range of ENS phenotypes and to the changes they undergo during dietary challenges. These sorts of studies have shown that dietary variation or obesity can change how the ENS processes information-in effect reprogramming the ENS. In this review, the data gathered from intracellular recordings will be compared with measurements made using imaging techniques in an effort to determine if the lessons learnt from inflammatory changes are relevant to the understanding of diet-induced reprogramming

Colonic Biopsies to Assess the Neuropathology of Parkinson's Disease and Its Relationship with Symptoms

The presence of Lewy bodies and Lewy neurites (LN) has been demonstrated in the enteric nervous system (ENS) of Parkinson's disease (PD) patients. The aims of the present research were to use routine colonoscopy biopsies (1) to analyze, in depth, enteric pathology throughout the colonic submucosal plexus (SMP), and (2) to correlate the pathological burden with neurological and gastrointestinal (GI) symptoms.A total of 10 control and 29 PD patients divided into 3 groups according to disease duration were included. PD and GI symptoms were assessed using the Unified Parkinson's Disease Rating Scale part III and the Rome III questionnaire, respectively. Four biopsies were taken from the ascending and descending colon during the course of a total colonoscopy. Immunohistochemical analysis was performed using antibodies against phosphorylated alpha-synuclein, neurofilaments NF 220 kDa (NF) and tyrosine hydroxylase (TH). The density of LN, labeled by anti-phosphorylated alpha-synuclein antibodies, was evaluated using a quantitative rating score. Lewy pathology was apparent in the colonic biopsies from 21 patients and in none of the controls. A decreased number of NF-immunoreactive neurons per ganglion was observed in the SMP of PD patients compared to controls. The amount of LN in the ENS was inversely correlated with neuronal count and positively correlated with levodopa-unresponsive features and constipation.Analysis of the ENS by routine colonoscopy biopsies is a useful tool for pre-mortem neuropathological diagnosis of PD, and also provides insight into the progression of motor and non-motor symptoms

Aging of the mammalian gastrointestinal tract: a complex organ system

Gastrointestinal disorders are a major cause of morbidity in the elderly population. The gastrointestinal tract is the most complex organ system; its diverse cells perform a range of functions essential to life, not only secretion, digestion, absorption and excretion, but also, very importantly, defence. The gastrointestinal tract acts not only as a barrier to harmful materials and pathogens but also contains the vast number of beneficial bacterial populations that make up the microbiota. Communication between the cells of the gastrointestinal tract and the central nervous and endocrine systems modifies behaviour; the organisms of the microbiota also contribute to this brain–gut–enteric microbiota axis. Age-related physiological changes in the gut are not only common, but also variable, and likely to be influenced by external factors as well as intrinsic aging of the cells involved. The cellular and molecular changes exhibited by the aging gut cells also vary. Aging intestinal smooth muscle cells exhibit a number of changes in the signalling pathways that regulate contraction. There is some evidence for age-associated degeneration of neurons and glia of the enteric nervous system, although enteric neuronal losses are likely not to be nearly as extensive as previously believed. Aging enteric neurons have been shown to exhibit a senescence-associated phenotype. Epithelial stem cells exhibit increased mitochondrial mutation in aging that affects their progeny in the mucosal epithelium. Changes to the microbiota and intestinal immune system during aging are likely to contribute to wider aging of the organism and are increasingly important areas of analysis. How changes of the different cell types of the gut during aging affect the numerous cellular interactions that are essential for normal gut functions will be important areas for future aging research

Neurogenic mechanisms in bladder and bowel ageing

The prevalence of both urinary and faecal incontinence, and also chronic constipation, increases with ageing and these conditions have a major impact on the quality of life of the elderly. Management of bladder and bowel dysfunction in the elderly is currently far from ideal and also carries a significant financial burden. Understanding how these changes occur is thus a major priority in biogerontology. The functions of the bladder and terminal bowel are regulated by complex neuronal networks. In particular neurons of the spinal cord and peripheral ganglia play a key role in regulating micturition and defaecation reflexes as well as promoting continence. In this review we discuss the evidence for ageing-induced neuronal dysfunction that might predispose to neurogenic forms of incontinence in the elderly

First Investigation of the Microbiology of the Deepest Layer of Ocean Crust

We would like to thank Frederick (Rick) Colwell for input on molecular analyses in low biomass environments, Donna Blackman, Benoît Ildefonse, Adélie Delacour, and Gretchen Früh-Green for discussions regarding geological and geochemical aspects of this manuscript, and the Integrated Ocean Drilling Program Expeditions 304/305 Science Party. We would also like to thank Captain Alex Simpson and the entire crew of the JOIDES Resolution.Conceived and designed the experiments: OUM MRF SJG. Performed the experiments: OUM TN MR JDVN AM. Analyzed the data: OUM TN MR JDVN AM. Contributed reagents/materials/analysis tools: TN MR JZ MRF SJG. Wrote the paper: OUM.The gabbroic layer comprises the majority of ocean crust. Opportunities to sample this expansive crustal environment are rare because of the technological demands of deep ocean drilling; thus, gabbroic microbial communities have not yet been studied. During the Integrated Ocean Drilling Program Expeditions 304 and 305, igneous rock samples were collected from 0.45-1391.01 meters below seafloor at Hole 1309D, located on the Atlantis Massif (30 °N, 42 °W). Microbial diversity in the rocks was analyzed by denaturing gradient gel electrophoresis and sequencing (Expedition 304), and terminal restriction fragment length polymorphism, cloning and sequencing, and functional gene microarray analysis (Expedition 305). The gabbroic microbial community was relatively depauperate, consisting of a low diversity of proteobacterial lineages closely related to Bacteria from hydrocarbon-dominated environments and to known hydrocarbon degraders, and there was little evidence of Archaea. Functional gene diversity in the gabbroic samples was analyzed with a microarray for metabolic genes (“GeoChip”), producing further evidence of genomic potential for hydrocarbon degradation - genes for aerobic methane and toluene oxidation. Genes coding for anaerobic respirations, such as nitrate reduction, sulfate reduction, and metal reduction, as well as genes for carbon fixation, nitrogen fixation, and ammonium-oxidation, were also present. Our results suggest that the gabbroic layer hosts a microbial community that can degrade hydrocarbons and fix carbon and nitrogen, and has the potential to employ a diversity of non-oxygen electron acceptors. This rare glimpse of the gabbroic ecosystem provides further support for the recent finding of hydrocarbons in deep ocean gabbro from Hole 1309D. It has been hypothesized that these hydrocarbons might originate abiotically from serpentinization reactions that are occurring deep in the Earth's crust, raising the possibility that the lithic microbial community reported here might utilize carbon sources produced independently of the surface biosphere.Yeshttp://www.plosone.org/static/editorial#pee