Biology subjects

Tomaro-Duchesneau, C.

Publications and source records attributed to Tomaro-Duchesneau, C..

3 recordsLinked to original sources

bioRxiv2019

Degradation of the incretin hormone Glucagon-Like Peptide-1 (GLP-1) by Enterococcus faecalis metalloprotease GelE

Metabolic diseases, including Type 2 Diabetes and obesity, have become increasingly prevalent global health concerns. Studies over the past decade have established connections between the gastrointestinal microbiota and host metabolism, but the mechanisms behind these connections are only beginning to be understood. We were interested in identifying microbes that have the ability to modulate the levels of the incretin hormone glucagon like peptide 1 (GLP-1). Using a human derived cell line that is capable of secreting GLP-1 in response to stimulatory ligands (NCI-H716), we identified supernatants from several bacterial isolates that were capable of decreasing GLP-1 levels, including several strains of Enterococcus faecalis. We further identified the secreted protease GelE, an established virulence factor from E. faecalis, as being responsible for GLP-1 inhibition via direct cleavage of GLP-1 by GelE. Finally, we demonstrated that E. faecalis supernatants can disrupt a colonic epithelial monolayer and cleave GLP-1 in a gelE dependent manner. This work suggests that a secreted factor from an intestinal microbe can traverse the epithelial barrier and impact levels of an important intestinal hormone.\n\nImportanceHumans have a complex and interconnected relationship with their gastrointestinal microbiomes, yet our interest in the microbiome tends to focus on overt pathogenic or probiotic activities, leaving the roles that commensal species may have on host physiology and metabolic processes largely unexplored. Commensal organisms in the microbiome produce and secrete many factors that have an opportunity to interact with the gastrointestinal tract and host biology. Here we show that a secreted protease from E. faecalis, GelE, is able to degrade the gastrointestinal hormone GLP-1, which is responsible for regulating glucose homeostasis and appetite in the body. The disruption of natural GLP-1 signaling by GelE may have significant consequences for maintaining healthy blood glucose levels and in the development of metabolic disease. Furthermore, this work deepens our understanding of specific host-microbiome interactions.

microbiology↗

bioRxiv2019

Human intestinal enteroids with inducible neurogenin-3 expression as a novel model of gut hormone secretion

BackgroundEnteroendocrine cells (EECs) are specialized epithelial cells that produce molecules vital for intestinal homeostasis, but due to their limited numbers, in-depth functional studies have remained challenging. Human intestinal enteroids (HIEs) that are derived from intestinal crypt stem cells are a biologically relevant in vitro model of the intestinal epithelium. HIEs contain all intestinal epithelial cell types; however, like the intestine, HIEs spontaneously produce few EECs, which limits their study.\n\nMethodsTo increase the number of EECs in HIEs, we used lentivirus transduction to stably engineer jejunal HIEs with doxycycline-inducible expression of neurogenin-3 (NGN3), a transcription factor that drives EEC differentiation (tetNGN3-HIEs). We examined the impact of NGN3 induction on EECs by quantifying the increase in the enterochromaffin cells and other EEC subtypes. We functionally assessed secretion of serotonin and EEC hormones in response to norepinephrine and rotavirus infection.\n\nResultsTreating tetNGN3-HIEs with doxycycline induced a dose-dependent increase of chromogranin A (ChgA)-positive and serotonin-positive cells, demonstrating increased enterochromaffin cell differentiation. Despite increased ChgA-positive cells, other differentiated cell types of the epithelium remained largely unchanged by gene expression and immunostaining. RNA sequencing of doxycycline-induced tetNGN3- HIEs identified increased expression of key hormones and enzymes associated with several other EEC subtypes. Doxycycline-induced tetNGN3-HIEs secreted serotonin, monocyte chemoattractant protein-1, glucose-dependent insulinotropic peptide, peptide YY, and ghrelin in response to norepinephrine and rotavirus infection, further supporting the presence of multiple EEC types.\n\nConclusionsWe have combined HIEs and inducible-NGN3 expression to establish a flexible in vitro model system for functional studies of EECs in enteroids and advance the molecular and physiological investigation of EECs.\n\nSynopsisEnteroendocrine cells have low abundance but exert widespread effects on gastrointestinal physiology. We engineered human intestinal enteroids with inducible expression of neurogenin-3, resulting in increased enteroendocrine cells and facilitating investigations of host responses to the dynamic intestinal environment.

microbiology↗

bioRxiv2018

Discovery of a bacterial peptide as a modulator of GLP-1 and metabolic disease

Early work in germ-free rodents highlighted the gut microbiotas importance in metabolic disease, including Type II Diabetes Mellitus (T2DM) and obesity. Glucagon-like peptide-1 (GLP-1) is an incretin secreted by enteroendocrine L-cells lining the gastrointestinal epithelium. GLP-1 has important functions including promoting insulin secretion, insulin sensitivity, and {beta}-cell mass, while inhibiting gastric emptying and appetite. We set out to elucidate how the microbiota can modulate GLP-1 secretion, with the goal to identify microbial strains with GLP-1 stimulatory activity as a metabolic disease therapeutic. Over 1500 human-derived strains were isolated from fecal, breast milk, and colon and intestinal biopsy samples from healthy individuals. In vitro screening for GLP-1 modulation was performed by incubating bacterial cell-free supernatants with NCI H716 human L-cells. Approximately 45 strains capable of increasing GLP-1 levels, measured by ELISA, were discovered. Interestingly, all positive strains were identified as Staphylococcus epidermidis by 16S rRNA sequencing. Non-GLP-1 stimulatory S. epidermidis strains were also identified. Mass spectrometry analysis identified a 3 kDa peptide, termed GLP-1 stimulating peptide (GspA), present in GLP-1 positive but absent in GLP-1 neutral S. epidermidis. Studies in human L-cells and intestinal enteroids demonstrated that GspA alone is sufficient to enhance GLP-1 secretion. When administered in high-fat-fed mice, GspA-producing S. epidermidis significantly reduced markers associated with obesity and T2DM, including adiposity and hyperinsulinemia. Further characterization of GspA suggests a GLP-1 stimulatory action via calcium signaling. The presented results identify a novel host-microbe interaction which may ultimately lead to the development of a microbial peptide-based therapeutic for obesity and T2DM.\n\nImportanceThe human gastrointestinal microbiota has been shown to modulate metabolic disease, including Type II Diabetes Mellitus and obesity, through mechanisms involving gut hormone secretion. We initiated this study to identify bacterial strains that can stimulate one of these hormones, glucagon-like peptide-1. We first identified that some strains of Staphylococcus epidermidis have such stimulatory activity. We then found that these strains could be used in a mouse model of high-fat feeding to reduce markers associated with metabolic disease, including adiposity and elevated insulin levels. We also identified the peptide from S. epidermidis that stimulates glucagon-like peptide-1 and propose a mode of action through calcium signaling. This newly identified microbial-derived peptide and host-microbe interaction provide a promising therapeutic approach against Type II Diabetes Mellitus and obesity.

microbiology↗