7 results on '"Ana L. Carvalho"'
Search Results
2. Extracellular vesicles produced by the human gut commensal bacterium Bacteroides thetaiotaomicron elicit anti-inflammatory responses from innate immune cells
- Author
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Sonia Fonseca, Ana L. Carvalho, Ariadna Miquel-Clopés, Emily J. Jones, Rokas Juodeikis, Régis Stentz, and Simon R. Carding
- Subjects
extracellular vesicles ,Bacteroides ,anti-inflammatory response ,innate immune tolerance ,BMDM ,THP-1 cells ,Microbiology ,QR1-502 - Abstract
Bacterial extracellular vesicles (BEVs) produced by gut commensal bacteria have been proposed to play an important role in maintaining host homeostasis via interactions with the immune system. Details of the mediators and pathways of BEV-immune cell interactions are however incomplete. In this study, we provide evidence for the anti-inflammatory and immunomodulatory properties of extracellular vesicles produced by the prominent human gut commensal bacterium Bacteroides thetaiotaomicron (Bt BEVs) and identify the molecular mechanisms underlying their interaction with innate immune cells. In mice treated with colitis-inducing dextran sodium sulfate (DSS) there was some indication that Bt BEVs improved survival, weight loss, disease activity and increased IL-10 production. Pre-treatment (conditioning) of murine bone marrow derived monocytes (BMDM) with Bt BEVs resulted in higher ratio of IL-10/TNFα production after an LPS challenge when compared to LPS pre-conditioned or non-conditioned BMDM. Using the THP-1 monocytic cell line the interactions between Bt BEVs and monocytes/macrophages were shown to be mediated primarily by TLR2. Histone (H3K4me1) methylation analysis showed that Bt BEVs induced epigenetic reprogramming which persisted after infectious challenge, as revealed by increased levels of H3K4me1 in Bt BEV-conditioned LPS-challenged BMDM. Collectively, our findings highlight the important role of Bt BEVs in maintaining host immune homeostasis and raise the promising possibility of considering their use in immune therapies.
- Published
- 2022
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3. Regulation of blood–brain barrier integrity by microbiome-associated methylamines and cognition by trimethylamine N-oxide
- Author
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Egle Solito, Michael Müller, Matthew G. Pontifex, Simon McArthur, Tom Snelling, Ana L. Carvalho, M. Areeb Anis-Alavi, Ildefonso Rodríguez-Ramiro, Sonia Fonseca, Lesley Hoyles, David Vauzour, Simon R. Carding, Khadija S. Jelane, Robert C. Glen, Hoyles, L., Pontifex, M. G., Rodriguez-Ramiro, I., Anis-Alavi, M. A., Jelane, K. S., Snelling, T., Solito, E., Fonseca, S., Carvalho, A. L., Carding, S. R., Muller, M., Glen, R. C., Vauzour, D., Mcarthur, S., McArthur, Simon [0000-0001-8521-1808], Apollo - University of Cambridge Repository, Medical Research Council (MRC), and Alzheimer's Research UK
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DISRUPTION ,INVOLVEMENT ,Microbiology (medical) ,Trimethylamine ,Trimethylamine N-oxide ,Gut flora ,Blood–brain barrier ,Microbiology ,Microbial ecology ,chemistry.chemical_compound ,Methylamines ,Mice ,Cognition ,1108 Medical Microbiology ,medicine ,Choline ,Animals ,The microbiota–gut–brain axis ,ACCUMULATION ,TRANSGENIC MICE ,Mammals ,Science & Technology ,IDENTIFICATION ,0602 Ecology ,Tight junction ,biology ,The microbiota���gut���brain axis ,Microbiota ,Research ,QR100-130 ,PERFORMANCE ,biology.organism_classification ,Cell biology ,METABOLITE ,medicine.anatomical_structure ,Blood���brain barrier ,chemistry ,Blood-Brain Barrier ,ANNEXIN A1 ,FATTY-ACIDS ,DIETARY CHOLINE ,Life Sciences & Biomedicine ,0605 Microbiology ,Astrocyte - Abstract
Background Communication between the gut microbiota and the brain is primarily mediated via soluble microbe-derived metabolites, but the details of this pathway remain poorly defined. Methylamines produced by microbial metabolism of dietary choline and l-carnitine have received attention due to their proposed association with vascular disease, but their effects upon the cerebrovascular circulation have hitherto not been studied. Results Here, we use an integrated in vitro/in vivo approach to show that physiologically relevant concentrations of the dietary methylamine trimethylamine N-oxide (TMAO) enhanced blood-brain barrier (BBB) integrity and protected it from inflammatory insult, acting through the tight junction regulator annexin A1. In contrast, the TMAO precursor trimethylamine (TMA) impaired BBB function and disrupted tight junction integrity. Moreover, we show that long-term exposure to TMAO protects murine cognitive function from inflammatory challenge, acting to limit astrocyte and microglial reactivity in a brain region-specific manner. Conclusion Our findings demonstrate the mechanisms through which microbiome-associated methylamines directly interact with the mammalian BBB, with consequences for cerebrovascular and cognitive function.
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- 2021
4. Use of bioengineered human commensal gut bacteria-derived microvesicles for mucosal plague vaccine delivery and immunization
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Régis Stentz, Phillip Brown, Andrea Telatin, M. Stock, S. Holmes, Simon G. P. Funnell, Udo Wegmann, N. J. Walker, Ethel Diane Williamson, Simon R. Carding, Emily J. Jones, Ana L. Carvalho, Mike Dennis, Ariadna Miquel-Clopés, and W. A. Butcher
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0301 basic medicine ,non‐human primates ,Yersinia pestis ,0302 clinical medicine ,humoral immunity ,Immunology and Allergy ,antibodies ,LcrV ,Cells, Cultured ,Immunity, Cellular ,biology ,Cell Death ,Antibodies, Bacterial ,3. Good health ,Plague vaccine ,Original Article ,Antibody ,outer membrane vesicles ,Pneumonic plague ,gut bacteria ,Pore Forming Cytotoxic Proteins ,Immunology ,Bioengineering ,Bubonic plague ,Microbiology ,03 medical and health sciences ,Immune system ,Antigen ,medicine ,Animals ,Humans ,Transport Vesicles ,Administration, Intranasal ,Antigens, Bacterial ,Plague ,Plague Vaccine ,biology.organism_classification ,medicine.disease ,Gastrointestinal Microbiome ,Immunity, Humoral ,Immunoglobulin A ,Editor's Choice ,mucosal vaccine ,Bacteroides thetaiotaomicron ,030104 developmental biology ,Bacterial Outer Membrane ,Immunoglobulin G ,biology.protein ,Macaca ,030215 immunology - Abstract
Plague caused by the Gram-negative bacterium, Yersinia pestis, is still endemic in parts of the world today. Protection against pneumonic plague is essential to prevent the development and spread of epidemics. Despite this, there are currently no licensed plague vaccines in the western world. Here we describe the means of delivering biologically active plague vaccine antigens directly to mucosal sites of plague infection using highly stable microvesicles (outer membrane vesicles; OMVs) that are naturally produced by the abundant and harmless human commensal gut bacterium Bacteroides thetaiotaomicron (Bt). Bt was engineered to express major plague protective antigens in its OMVs, specifically Fraction 1 (F1) in the outer membrane and LcrV (V antigen) in the lumen, for targeted delivery to the gastrointestinal (GI) and respiratory tracts in a non-human primate (NHP) host. Our key findings were that Bt OMVs stably expresses F1 and V plague antigens, particularly the V antigen, in the correct, immunogenic form. When delivered intranasally V-OMVs elicited substantive and specific immune and antibody responses, both in the serum [immunoglobulin (Ig)G] and in the upper and lower respiratory tract (IgA); this included the generation of serum antibodies able to kill plague bacteria. Our results also showed that Bt OMV-based vaccines had many desirable characteristics, including: biosafety and an absence of any adverse effects, pathology or gross alteration of resident microbial communities (microbiotas); high stability and thermo-tolerance; needle-free delivery; intrinsic adjuvanticity; the ability to stimulate both humoral and cell-mediated immune responses; and targeting of primary sites of plague infection.
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- 2019
5. Bioengineering commensal bacteria-derived outer membrane vesicles for delivery of biologics to the gastrointestinal and respiratory tract
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Régis Stentz, Eleanor G. Bentley, Janine L. Coombes, Udo Wegmann, Katherine Gil-Cordoso, Simon R. Carding, Khoon S. Kok, James P. Stewart, Ariadna Miquel-Clopés, Sonia Fonseca, Ana L. Carvalho, Anja Kipar, Kathryn Cross, Sanaria Al Katy, University of Zurich, and Carding, Simon R
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0301 basic medicine ,Commensal bacteria ,Histology ,10184 Institute of Veterinary Pathology ,mucosal drug delivery ,2722 Histology ,Virus ,Microbiology ,1307 Cell Biology ,03 medical and health sciences ,0302 clinical medicine ,Immune system ,Antigen ,medicine ,Secretion ,mucosal vaccines ,lcsh:QH573-671 ,therapeutic proteins ,biology ,lcsh:Cytology ,Human gastrointestinal tract ,Cell Biology ,biology.organism_classification ,bacterial microvesicles ,3. Good health ,030104 developmental biology ,medicine.anatomical_structure ,Salmonella enterica ,030220 oncology & carcinogenesis ,570 Life sciences ,Bacterial outer membrane ,Bacteria ,outer membrane vesicles ,Research Article - Abstract
Gram-negative bacteria naturally produce and secrete nanosized outer membrane vesicles (OMVs). In the human gastrointestinal tract, OMVs produced by commensal Gram-negative bacteria can mediate interactions amongst host cells (including between epithelial cells and immune cells) and maintain microbial homeostasis. This OMV-mediated pathway for host-microbe interactions could be exploited to deliver biologically active proteins to the body. To test this we engineered the Gram-negative bacterium Bacteroides thetaiotaomicron (Bt), a prominent member of the intestinal microbiota of all animals, to incorporate bacteria-, virus- and human-derived proteins into its OMVs. We then used the engineered Bt OMVs to deliver these proteins to the respiratory and gastrointestinal (GI)-tract to protect against infection, tissue inflammation and injury. Our findings demonstrate the ability to express and package both Salmonella enterica ser. Typhimurium-derived vaccine antigens and influenza A virus (IAV)-derived vaccine antigens within or on the outer membrane of Bt OMVs. These antigens were in a form capable of eliciting antigen-specific immune and antibody responses in both mucosal tissues and systemically. Furthermore, immunisation with OMVs containing the core stalk region of the IAV H5 hemagglutinin from an H5N1 strain induced heterotypic protection in mice to a 10-fold lethal dose of an unrelated subtype (H1N1) of IAV. We also showed that OMVs could express the human therapeutic protein, keratinocyte growth factor-2 (KGF-2), in a stable form that, when delivered orally, reduced disease severity and promoted intestinal epithelial repair and recovery in animals administered colitis-inducing dextran sodium sulfate. Collectively, our data demonstrates the utility and effectiveness of using Bt OMVs as a mucosal biologics and drug delivery platform technology.
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- 2019
6. Use of genetically modified bacteria for drug delivery in humans: Revisiting the safety aspect
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Udo Wegmann, Martin Stocks, Ana L. Carvalho, and Simon R. Carding
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0301 basic medicine ,Gene Transfer, Horizontal ,Science ,Transgene ,030106 microbiology ,Genetically modified bacteria ,Article ,Microbiology ,03 medical and health sciences ,Drug Delivery Systems ,Animals ,Bacteroides ,Humans ,Transgenes ,2. Zero hunger ,Gastrointestinal tract ,Multidisciplinary ,biology ,Thymineless death ,biology.organism_classification ,Gastrointestinal Tract ,Drug delivery ,Horizontal gene transfer ,Medicine ,Microbial Interactions ,Genetic Engineering ,Bacteria - Abstract
The use of live, genetically modified bacteria as delivery vehicles for biologics is of considerable interest scientifically and has attracted significant commercial investment. We have pioneered the use of the commensal gut bacterium Bacteroides ovatus for the oral delivery of therapeutics to the gastrointestinal tract. Here we report on our investigations of the biological safety of engineered B. ovatus bacteria that includes the use of thymineless death as a containment strategy and the potential for the spread of transgenes in vivo in the mammalian gastrointestinal tract. We demonstrate the ability of GM-strains of Bacteroides to survive thymine starvation and overcome it through the exchange of genetic material. We also provide evidence for horizontal gene transfer in the mammalian gastrointestinal tract resulting in transgene-carrying wild type bacteria. These findings sound a strong note of caution on the employment of live genetically modified bacteria for the delivery of biologics.
- Published
- 2017
7. A Novel Tightly Regulated Gene Expression System for the Human Intestinal Symbiont Bacteroides thetaiotaomicron
- Author
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Régis Stentz, Karin Overweg, Nikki Horn, Udo Wegmann, Simon R. Carding, and Ana L. Carvalho
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0301 basic medicine ,Genetics ,Microbiology (medical) ,Reporter gene ,inducible promoter ,030106 microbiology ,lcsh:QR1-502 ,mannan ,Biology ,Microbiology ,Reverse genetics ,lcsh:Microbiology ,Ribosomal binding site ,expression system ,03 medical and health sciences ,Bacteroides thetaiotaomicron ,Terminator (genetics) ,Gene expression ,Multiple cloning site ,Methods ,PUL ,Gene - Abstract
There is considerable interest in studying the function of Bacteroides species resident in the human gastrointestinal (GI)-tract and the contribution they make to host health. Reverse genetics and protein expression techniques, such as those developed for well-characterised Escherichia coli cannot be applied to Bacteroides species as they and other members of the Bacteriodetes phylum have unique promoter structures. The availability of useful Bacteroides-specific genetic tools is therefore limited. Here we describe the development of an effective mannan-controlled gene expression system for Bacteroides thetaiotaomicron containing the mannan-inducible promoter–region of an α-1,2-mannosidase gene (BT_3784), a ribosomal binding site designed to modulate expression, a multiple cloning site to facilitate the cloning of genes of interest, and a transcriptional terminator. Using the Lactobacillus pepI as a reporter gene, mannan induction resulted in an increase of reporter activity in a time- and concentration-dependent manner with a wide range of activity. The endogenous BtcepA cephalosporinase gene was used to demonstrate the suitability of this novel expression system, enabling the isolation of a His-tagged version of BtCepA. We have also shown with experiments performed in mice that the system can be induced in vivo in the presence of an exogenous source of mannan. By enabling the controlled expression of endogenous and exogenous genes in B. thetaiotaomicron this novel inducer-dependent expression system will aid in defining the physiological role of individual genes and the functional analyses of their products.
- Published
- 2016
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