Early work in rodents highlighted the gut microbiota's importance in metabolic disease, including Type II Diabetes Mellitus (T2DM) and obesity. Glucagon-like peptide-1 (GLP-1), an incretin secreted by L-cells lining the gastrointestinal epithelium, has important functions: promoting insulin secretion, insulin sensitivity, and β-cell mass, while inhibiting gastric emptying and appetite. We set out to identify microbial strains with GLP-1 stimulatory activity as potential metabolic disease therapeutics. Over 1500 human-derived strains were isolated from healthy individuals and screened for GLP-1 modulation by incubating bacterial cell-free supernatants with NCI H716 L-cells. Approximately 45 strains capable of increasing GLP-1 were discovered. All GLP-1 positive strains were identified as Staphylococcus epidermidis by 16S rRNA sequencing. Mass spectrometry analysis identified a 3 kDa peptide, Hld (delta-toxin), present in GLP-1 positive supernatants but absent in GLP-1 neutral supernatants. Studies in NCI-H716 cells and human jejunal enteroids engineered to make more enteroendocrine cells demonstrated that Hld alone is sufficient to enhance GLP-1 secretion. When administered in high-fatfed mice, Hld-producing S. epidermidis significantly reduced markers associated with obesity and T2DM. Further characterization of Hld suggests GLP-1 stimulatory action of Hld occurs 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 metabolic disease. Metabolic disorders, including Type 2 Diabetes Mellitus (T2DM) and obesity, pose a serious public health concern both nationally and globally. According to the Centers for Disease Control and Prevention (CDC), 9.4% of the population of the United States has Diabetes, including 25.2% of those aged 65 years or older 1. Obesity is also a major health concern, with more than one-third of American adults considered obese 2. Current treatment approaches, including intensive lifestyle modifications, diet intervention, and pharmacologics, have proven unsuccessful in controlling the global increase of metabolic disorders. Therefore, a novel approach to combat metabolic disorders is needed. A number of research groups have recently demonstrated the role of the human gut microbiota in metabolism and metabolic disease, leading to the attempt to develop microbial therapeutics. Bäckhed et al. initially spearheaded this research; using germ-free rodents, it was demonstrated that when the microbiota of conventionally raised animals was transplanted into germ-free rodents, the latter developed an increase in adiposity and insulin resistance 3. Studies by a number of different groups also demonstrated the importance of the microbiota in metabolism, described in a recent review 4. Specifically, the gut microbiota has been shown to play an important role in gut hormone modulation, including GLP-1. Administration of prebiotics, non-digestible food ingredients that stimulate the grow...
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. IMPORTANCE Humans 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.
19Early work in germ-free rodents highlighted the gut microbiota's importance in 20 metabolic disease, including Type II Diabetes Mellitus (T2DM) and obesity. Glucagon-21 like peptide-1 (GLP-1) is an incretin secreted by enteroendocrine L-cells lining the 22 gastrointestinal epithelium. GLP-1 has important functions including promoting insulin 23 secretion, insulin sensitivity, and β-cell mass, while inhibiting gastric emptying and 24 appetite. We set out to elucidate how the microbiota can modulate GLP-1 secretion, 25with the goal to identify microbial strains with GLP-1 stimulatory activity as a metabolic 26 disease therapeutic. Over 1500 human-derived strains were isolated from fecal, breast 27 milk, and colon and intestinal biopsy samples from healthy individuals. . CC-BY-NC-ND 4.0 International license It is made available under a was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.
12Metabolic diseases, including Type 2 Diabetes and obesity, have become 13 increasingly prevalent global health concerns. Studies over the past decade have 14 established connections between the gastrointestinal microbiota and host metabolism, 15 but the mechanisms behind these connections are only beginning to be understood. We 16were interested in identifying microbes that have the ability to modulate the levels of the 17 incretin hormone glucagon like peptide 1 (GLP-1). Using a human derived cell line that 18 is capable of secreting GLP-1 in response to stimulatory ligands (NCI-H716), we 19 identified supernatants from several bacterial isolates that were capable of decreasing 20 GLP-1 levels, including several strains of Enterococcus faecalis. We further identified 21 the secreted protease GelE, an established virulence factor from E. faecalis, as being 22 responsible for GLP-1 inhibition via direct cleavage of GLP-1 by GelE. Finally, we 23 demonstrated that E. faecalis supernatants can disrupt a colonic epithelial monolayer 24 and cleave GLP-1 in a gelE dependent manner. This work suggests that a secreted 25 factor from an intestinal microbe can traverse the epithelial barrier and impact levels of 26 an important intestinal hormone. 27 28 Importance 29Humans have a complex and interconnected relationship with their 30 gastrointestinal microbiomes, yet our interest in the microbiome tends to focus on overt 31 pathogenic or probiotic activities, leaving the roles that commensal species may have 32 on host physiology and metabolic processes largely unexplored. Commensal organisms 33 in the microbiome produce and secrete many factors that have an opportunity to interact 34 3 with the gastrointestinal tract and host biology. Here we show that a secreted protease 35 from E. faecalis, GelE, is able to degrade the gastrointestinal hormone GLP-1, which is 36 responsible for regulating glucose homeostasis and appetite in the body. The disruption 37 of natural GLP-1 signaling by GelE may have significant consequences for maintaining 38 healthy blood glucose levels and in the development of metabolic disease. Furthermore, 39 this work deepens our understanding of specific host-microbiome interactions. 40 with a focus on the interaction of the microbiome with GI hormone peptides, in particular 57 the nutrient-stimulated incretin Glucagon-like Peptide-1 (GLP-1). Already a therapeutic 58 target for Type 2 Diabetes (T2D), GLP-1 is an integral signaling hormone responsible 59 for promoting insulin secretion and satiety, while decreasing glucagon secretion and 60 gastric emptying. Early work showed that addition of the prebiotic oligofructose to the 61 diets of rats on a high-fat diet increased GLP-1 levels measured from the portal vein, in 62 addition to protecting from weight gain (4). Additional studies demonstrated that 63 5 administration of Akkermansia muciniphila could reverse high-fat diet induced metabolic 64 disorders, and that this activity was mediated at least partially by an outer membrane 65 protein purified f...
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