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1.
Microbiology (Reading) ; 170(9)2024 Sep.
Article in English | MEDLINE | ID: mdl-39302176

ABSTRACT

Streptococcus thermophilus holds promise as a chassis for producing and secreting heterologous proteins. Used for thousands of years to ferment milk, this species has generally recognized as safe (GRAS) status in the USA and qualified presumption of safety (QPS) status in Europe. In addition, it can be easily genetically modified thanks to its natural competence, and it secretes very few endogenous proteins, which means less downstream processing is needed to purify target proteins, reducing costs. Extracellular degradation of heterologous proteins can be eliminated by introducing mutations that inactivate the genes encoding the bacterium's three major surface proteases. Here, we constructed an inducible expression system that utilizes a peptide pheromone (SHP1358) and a transcriptional regulator (Rgg1358) involved in quorum-sensing regulation. We explored the functionality of a complete version of the system, in which the inducer is produced by the bacterium itself, by synthesizing a luciferase reporter protein. This complete version was assessed with bacteria grown in a chemically defined medium but also in vivo, in the faeces of germ-free mice. We also tested an incomplete version, in which the inducer had to be added to the culture medium, by synthesizing luciferase and a secreted form of elafin, a human protein with therapeutic properties. Our results show that, in our system, protein production can be modulated by employing different concentrations of the SHP1358 inducer or other SHPs with closed amino acid sequences. We also constructed a genetic background in which all system leakiness was eliminated. In conclusion, with this new inducible expression system, we have added to the set of tools currently used to produce secreted proteins in S. thermophilus, whose myriad applications include the delivery of therapeutic peptides or proteins.


Subject(s)
Bacterial Proteins , Quorum Sensing , Recombinant Proteins , Streptococcus thermophilus , Quorum Sensing/genetics , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Streptococcus thermophilus/genetics , Streptococcus thermophilus/metabolism , Animals , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Mice , Gene Expression Regulation, Bacterial , Trans-Activators/genetics , Trans-Activators/metabolism , Pheromones/metabolism , Pheromones/genetics
2.
Ann Rheum Dis ; 83(3): 312-323, 2024 Feb 15.
Article in English | MEDLINE | ID: mdl-38049981

ABSTRACT

OBJECTIVES: Alterations in tryptophan (Trp) metabolism have been reported in inflammatory diseases, including rheumatoid arthritis (RA). However, understanding whether these alterations participate in RA development and can be considered putative therapeutic targets remains undetermined.In this study, we combined quantitative Trp metabolomics in the serum from patients with RA and corrective administration of a recombinant enzyme in experimental arthritis to address this question. METHODS: Targeted quantitative Trp metabolomics was performed on the serum from 574 previously untreated patients with RA from the ESPOIR (Etude et Suivi des POlyarthrites Indifférenciées Récentes) cohort and 98 healthy subjects. A validation cohort involved 69 established patients with RA. Dosages were also done on the serum of collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA) mice and controls. A proof-of-concept study evaluating the therapeutic potency of targeting the kynurenine pathway was performed in the CAIA model. RESULTS: Differential analysis revealed dramatic changes in Trp metabolite levels in patients with RA compared with healthy controls. Decreased levels of kynurenic (KYNA) and xanthurenic (XANA) acids and indole derivatives, as well as an increased level of quinolinic acid (QUIN), were found in the serum of patients with RA. They correlated positively with disease severity (assessed by both circulating biomarkers and disease activity scores) and negatively with quality-of-life scores. Similar profiles of kynurenine pathway metabolites were observed in the CAIA and CIA models. From a mechanistic perspective, we demonstrated that QUIN favours human fibroblast-like synoviocyte proliferation and affected their cellular metabolism, through inducing both mitochondrial respiration and glycolysis. Finally, systemic administration of the recombinant enzyme aminoadipate aminotransferase, responsible for the generation of XANA and KYNA, was protective in the CAIA model. CONCLUSIONS: Altogether, our preclinical and clinical data indicate that alterations in the Trp metabolism play an active role in the pathogenesis of RA and could be considered as a new therapeutic avenue.


Subject(s)
Arthritis, Experimental , Arthritis, Rheumatoid , Humans , Animals , Mice , Tryptophan/therapeutic use , Kynurenine/therapeutic use , Biomarkers , Arthritis, Experimental/pathology
3.
Microb Cell Fact ; 23(1): 172, 2024 Jun 12.
Article in English | MEDLINE | ID: mdl-38867272

ABSTRACT

There is increasing evidence that probiotic and commensal bacteria play a role in substrate metabolism, energy harvesting and intestinal homeostasis, and may exert immunomodulatory activities on human health. In addition, recent research suggests that these microorganisms interact with vitamins and minerals, promoting intestinal and metabolic well-being while producing vital microbial metabolites such as short-chain fatty acids (SCFAs). In this regard, there is a flourishing field exploring the intricate dynamics between vitamins, minerals, SCFAs, and commensal/probiotic interactions. In this review, we summarize some of the major hypotheses beyond the mechanisms by which commensals/probiotics impact gut health and their additional effects on the absorption and metabolism of vitamins, minerals, and SCFAs. Our analysis includes comprehensive review of existing evidence from preclinical and clinical studies, with particular focus on the potential interaction between commensals/probiotics and micronutrients. Finally, we highlight knowledge gaps and outline directions for future research in this evolving field.


Subject(s)
Bacteria , Fatty Acids, Volatile , Gastrointestinal Microbiome , Minerals , Probiotics , Vitamins , Probiotics/metabolism , Humans , Vitamins/metabolism , Minerals/metabolism , Fatty Acids, Volatile/metabolism , Bacteria/metabolism , Symbiosis , Animals
4.
Gut ; 72(7): 1296-1307, 2023 07.
Article in English | MEDLINE | ID: mdl-36270778

ABSTRACT

OBJECTIVE: The extent to which tryptophan (Trp) metabolism alterations explain or influence the outcome of inflammatory bowel diseases (IBDs) is still unclear. However, several Trp metabolism end-products are essential to intestinal homeostasis. Here, we investigated the role of metabolites from the kynurenine pathway. DESIGN: Targeted quantitative metabolomics was performed in two large human IBD cohorts (1069 patients with IBD). Dextran sodium sulphate-induced colitis experiments in mice were used to evaluate effects of identified metabolites. In vitro, ex vivo and in vivo experiments were used to decipher mechanisms involved. Effects on energy metabolism were evaluated by different methods including Single Cell mEtabolism by profiling Translation inHibition. RESULTS: In mice and humans, intestinal inflammation severity negatively correlates with the amount of xanthurenic (XANA) and kynurenic (KYNA) acids. Supplementation with XANA or KYNA decreases colitis severity through effects on intestinal epithelial cells and T cells, involving Aryl hydrocarbon Receptor (AhR) activation and the rewiring of cellular energy metabolism. Furthermore, direct modulation of the endogenous tryptophan metabolism, using the recombinant enzyme aminoadipate aminotransferase (AADAT), responsible for the generation of XANA and KYNA, was protective in rodent colitis models. CONCLUSION: Our study identified a new mechanism linking Trp metabolism to intestinal inflammation and IBD. Bringing back XANA and KYNA has protective effects involving AhR and the rewiring of the energy metabolism in intestinal epithelial cells and CD4+ T cells. This study paves the way for new therapeutic strategies aiming at pharmacologically correcting its alterations in IBD by manipulating the endogenous metabolic pathway with AADAT.


Subject(s)
Colitis , Inflammatory Bowel Diseases , Humans , Animals , Mice , Tryptophan/metabolism , Inflammatory Bowel Diseases/drug therapy , Colitis/chemically induced , Colitis/drug therapy , Colitis/metabolism , Intestines , Inflammation
5.
Cell Mol Life Sci ; 78(4): 1191-1206, 2021 Feb.
Article in English | MEDLINE | ID: mdl-32979054

ABSTRACT

Most cervical cancer (CxCa) are related to persistent infection with high-risk human papillomavirus (HR-HPV) in the cervical mucosa, suggesting that an induction of mucosal cell-mediated immunity against HR-HPV oncoproteins can be a promising strategy to fight HPV-associated CxCa. From this perspective, many pre-clinical and clinical trials have proved the potential of lactic acid bacteria (LAB) genetically modified to deliver recombinant antigens to induce mucosal, humoral and cellular immunity in the host. Altogether, the outcomes of these studies suggest that there are several key factors to consider that may offer guidance on improvement protein yield and improving immune response. Overall, these findings showed that oral LAB-based mucosal HPV vaccines expressing inducible surface-anchored antigens display a higher potential to induce particularly specific systemic and mucosal cytotoxic cellular immune responses. In this review, we describe all LAB-based HPV vaccine investigations by reviewing databases from international studies between 2000 and 2020. Our aim is to promote the therapeutic HPV vaccines knowledge and to complete the gaps in this field to empower scientists worldwide to make proper decisions regarding the best strategies for the development of therapeutic HPV vaccines.


Subject(s)
Gastrointestinal Microbiome/genetics , Lactobacillales/genetics , Microorganisms, Genetically-Modified/genetics , Papillomavirus Infections/genetics , Female , Gastrointestinal Microbiome/immunology , Humans , Immunity, Mucosal/genetics , Immunity, Mucosal/immunology , Lactobacillales/immunology , Microorganisms, Genetically-Modified/immunology , Papillomaviridae/drug effects , Papillomaviridae/immunology , Papillomaviridae/pathogenicity , Papillomavirus Infections/immunology , Papillomavirus Infections/prevention & control , Papillomavirus Infections/virology , Papillomavirus Vaccines/immunology , Papillomavirus Vaccines/therapeutic use , Uterine Cervical Neoplasms/immunology , Uterine Cervical Neoplasms/metabolism , Uterine Cervical Neoplasms/prevention & control , Uterine Cervical Neoplasms/virology , Vagina/immunology , Vagina/microbiology , Vagina/virology
6.
FASEB J ; 33(4): 4741-4754, 2019 04.
Article in English | MEDLINE | ID: mdl-30608881

ABSTRACT

Lipidomic techniques can improve our understanding of complex lipid interactions that regulate metabolic diseases. Here, a serum phospholipidomics analysis identified associations between phosphatidylglycerols (PGs) and gut microbiota dysbiosis. Compared with the other phospholipids, serum PGs were the most elevated in patients with low microbiota gene richness, which were normalized after a dietary intervention that restored gut microbial diversity. Serum PG levels were positively correlated with metagenomic functional capacities for bacterial LPS synthesis and host markers of low-grade inflammation; transcriptome databases identified PG synthase, the first committed enzyme in PG synthesis, as a potential mediator. Experiments in mice and cultured human-derived macrophages demonstrated that LPS induces PG release. Acute PG treatment in mice altered adipose tissue gene expression toward remodeling and inhibited ex vivo lipolysis in adipose tissue, suggesting that PGs favor lipid storage. Indeed, several PG species were associated with the severity of obesity in mice and humans. Finally, despite enrichment in PGs in bacterial membranes, experiments employing gnotobiotic mice colonized with recombinant PG overproducing Lactococcus lactis showed limited direct contribution of microbial PGs to the host. In summary, PGs are inflammation-responsive lipids indirectly regulated by the gut microbiota via endotoxins and regulate adipose tissue homeostasis in obesity.-Kayser, B. D., Lhomme, M., Prifti, E., Da Cunha, C., Marquet, F., Chain, F., Naas, I., Pelloux, V., Dao, M.-C., Kontush, A., Rizkalla, S. W., Aron-Wisnewsky, J., Bermúdez-Humarán, L. G., Oakley, F., Langella, P., Clément, K., Dugail, I. Phosphatidylglycerols are induced by gut dysbiosis and inflammation, and favorably modulate adipose tissue remodeling in obesity.


Subject(s)
Adipose Tissue/metabolism , Dysbiosis/metabolism , Inflammation/metabolism , Obesity/metabolism , Phosphatidylglycerols/metabolism , Animals , Female , Humans , Lipidomics/methods , Lipolysis/physiology , Male , Metagenomics/methods , Mice
7.
Environ Microbiol ; 21(11): 4020-4031, 2019 11.
Article in English | MEDLINE | ID: mdl-31325218

ABSTRACT

Antimicrobial peptides secreted by intestinal immune and epithelial cells are important effectors of innate immunity. They play an essential role in the maintenance of intestinal homeostasis by limiting microbial epithelium interactions and preventing unnecessary microbe-driven inflammation. Pancreatitis-associated protein (PAP) belongs to Regenerating islet-derived III proteins family and is a C-type (Ca+2 dependent) lectin. PAP protein plays a protective effect presenting anti-inflammatory properties able to reduce the severity of colitis, preserving gut barrier and epithelial inflammation. Here, we sought to determine whether PAP delivered at intestinal lumen by recombinant Lactococcus lactis strain (LL-PAP) before and after chemically induced colitis is able to reduce the severity in two models of colitis. After construction and characterization of our recombinant strains, we tested their effects in dinitro-benzenesulfonic-acid (DNBS) and Dextran sulfate sodium (DSS) colitis model. After the DNBS challenge, mice treated with LL-PAP presented less severe colitis compared with PBS and LL-empty-treated mice groups. After the DSS challenge, no protective effects of LL-PAP could be detected. We determined that after 5 days administration, LL-PAP increase butyrate producer's bacteria, especially Eubacterium plexicaudatum. Based on our findings, we hypothesize that a treatment with LL-PAP shifts the microbiota preventing the severity of colon inflammation in DNBS colitis model. These protective roles of LL-PAP in DNBS colitis model might be through intestinal microbiota modulation.


Subject(s)
Antimicrobial Cationic Peptides/therapeutic use , Colitis/drug therapy , Gastrointestinal Microbiome/drug effects , Lactococcus lactis/metabolism , Pancreatitis-Associated Proteins/therapeutic use , Animals , Antimicrobial Cationic Peptides/metabolism , Benzenesulfonates/toxicity , Colitis/chemically induced , Colitis/pathology , Colon/metabolism , Dextran Sulfate/toxicity , Disease Models, Animal , Inflammation , Mice , Mice, Inbred C57BL , Pancreatitis-Associated Proteins/metabolism , Peptides/metabolism
8.
Appl Microbiol Biotechnol ; 102(24): 10703-10711, 2018 Dec.
Article in English | MEDLINE | ID: mdl-30310964

ABSTRACT

Efficient delivery of antigens to the gut-associated lymphoid tissue (GALT) is the most critical step for the induction of mucosal immunity by oral vaccines. As M cells are the main portal for luminal antigens into the GALT, the M cell-targeting of antigens affords a promising strategy toward the development of effective oral vaccines. Lactococcus lactis is a fascinating recombinant host for oral vaccines, as they survive and produce antigens in the gut and have a particularly safe profile for human use. In this study, we developed and evaluated an M cell-targeting oral immunization system using recombinant L. lactis strains. For the purpose, we generated an L. lactis strain that secretes a model antigen fused with the OmpH ß1α1 domain of Yersinia enterocolitica, which has been shown to bind to a complement C5a receptor on the M cell surface. As the model antigen, Staphylococcus aureus nuclease was used for fusion, resulting in L. lactis-expressing Nuc-OmpH (LL/Nuc-OmpH). Ex vivo intestinal loop assays showed that the amount of Nuc-OmpH taken up into Peyer's patches was more than that of the unfused nuclease (Nuc). In addition, oral administration of the recombinant L. lactis strains to mice demonstrated that LL/Nuc-OmpH-induced nuclease-specific fecal IgA and serum IgG titers were significantly higher than those induced by LL/Nuc. These results indicate that OmpH works as an M cell-targeting molecule when fused with antigens secreted from L. lactis and that the M cell-targeting strategy affords a promising platform for L. lactis-based mucosal immunization.


Subject(s)
Deoxyribonucleases/administration & dosage , Immunity, Mucosal , Lactococcus lactis/metabolism , Peyer's Patches/immunology , Administration, Oral , Animals , Antigens/metabolism , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/immunology , Deoxyribonucleases/genetics , Deoxyribonucleases/metabolism , Female , Intestinal Mucosa/cytology , Intestinal Mucosa/immunology , Lactococcus lactis/genetics , Mice, Inbred C57BL , Microorganisms, Genetically-Modified , Recombinant Proteins/administration & dosage , Recombinant Proteins/genetics , Recombinant Proteins/metabolism
9.
Microb Cell Fact ; 16(1): 79, 2017 May 08.
Article in English | MEDLINE | ID: mdl-28482838

ABSTRACT

The aim of this review is to summarize the effect in host energy metabolism of the production of B group vitamins and short chain fatty acids (SCFA) by commensal, food-grade and probiotic bacteria, which are also actors of the mammalian nutrition. The mechanisms of how these microbial end products, produced by these bacterial strains, act on energy metabolism will be discussed. We will show that these vitamins and SCFA producing bacteria could be used as tools to recover energy intakes by either optimizing ATP production from foods or by the fermentation of certain fibers in the gastrointestinal tract (GIT). Original data are also presented in this work where SCFA (acetate, butyrate and propionate) and B group vitamins (riboflavin, folate and thiamine) production was determined for selected probiotic bacteria.


Subject(s)
Energy Metabolism , Fatty Acids, Volatile/metabolism , Probiotics/metabolism , Symbiosis , Vitamins/metabolism , Adenosine Triphosphate/biosynthesis , Adenosine Triphosphate/metabolism , Animals , Butyrates/metabolism , Dietary Fiber/metabolism , Fatty Acids, Volatile/biosynthesis , Fermentation , Food , Gastrointestinal Microbiome/physiology , Gastrointestinal Tract/microbiology , Humans , Mice , Vitamins/biosynthesis
10.
Microb Cell Fact ; 16(1): 27, 2017 Feb 13.
Article in English | MEDLINE | ID: mdl-28193209

ABSTRACT

BACKGROUND: Mucositis is one of the most relevant gastrointestinal inflammatory conditions in humans, generated by the use of chemotherapy drugs, such as 5-fluoracil (5-FU). 5-FU-induced mucositis affects 80% of patients undergoing oncological treatment causing mucosal gut dysfunctions and great discomfort. As current therapy drugs presents limitations in alleviating mucositis symptoms, alternative strategies are being pursued. Recent studies have shown that the antimicrobial pancreatitis-associated protein (PAP) has a protective role in intestinal inflammatory processes. Indeed, it was demonstrated that a recombinant strain of Lactococcus lactis expressing human PAP (LL-PAP) could prevent and improve murine DNBS-induced colitis, an inflammatory bowel disease (IBD) that causes severe inflammation of the colon. Hence, in this study we sought to evaluate the protective effects of LL-PAP on 5-FU-induced experimental mucositis in BALB/c mice as a novel approach to treat the disease. RESULTS: Our results show that non-recombinant L. lactis NZ9000 have antagonistic activity, in vitro, against the enteroinvasive gastrointestinal pathogen L. monocytogenes and confirmed PAP inhibitory effect against Opportunistic E. faecalis. Moreover, L. lactis was able to prevent histological damage, reduce neutrophil and eosinophil infiltration and secretory Immunoglobulin-A in mice injected with 5-FU. Recombinant lactococci carrying antimicrobial PAP did not improve those markers of inflammation, although its expression was associated with villous architecture preservation and increased secretory granules density inside Paneth cells in response to 5-FU inflammation. CONCLUSIONS: We have demonstrated for the first time that L. lactis NZ9000 by itself, is able to prevent 5-FU-induced intestinal inflammation in BALB/c mice. Moreover, PAP delivered by recombinant L. lactis strain showed additional protective effects in mice epithelium, revealing to be a promising strategy to treat intestinal mucositis.


Subject(s)
Antigens, Neoplasm/genetics , Antigens, Neoplasm/metabolism , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Ileitis/prevention & control , Lactococcus lactis/genetics , Lactococcus lactis/physiology , Lectins, C-Type/genetics , Lectins, C-Type/metabolism , Mucositis/prevention & control , Animals , Antibiosis , Antigens, Neoplasm/pharmacology , Biomarkers, Tumor/pharmacology , Disease Models, Animal , Enterococcus faecalis/physiology , Fluorouracil , Humans , Ileitis/chemically induced , Ileitis/drug therapy , Inflammatory Bowel Diseases/drug therapy , Inflammatory Bowel Diseases/prevention & control , Intestinal Mucosa/metabolism , Intestine, Small/immunology , Intestine, Small/microbiology , Intestine, Small/pathology , Lactococcus lactis/metabolism , Listeria monocytogenes/physiology , Mice , Mice, Inbred BALB C , Mucositis/chemically induced , Mucositis/drug therapy , Mucositis/microbiology , Pancreatitis-Associated Proteins
11.
Appl Microbiol Biotechnol ; 100(1): 385-396, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26476654

ABSTRACT

Probiotics are live microorganisms which when administered in adequate amounts, confer health benefits on the host. Their use is more and more widespread for both prevention and treatment of diseases, including traveler's diarrhea and inflammatory bowel diseases (IBDs). In this work, we isolated and characterized novel candidate probiotic strains from pulque (xaxtle), a traditional Mexican alcoholic fermented beverage. A total of 14 strains were obtained from xaxtle samples isolated from three different Mexican regions. Species identification was performed by biochemical methods and 16S rRNA gene targeted PCR. The isolates belonged to the Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus brevis, and Lactobacillus composti phylogenetic groups, with L. brevis being the most dominant group. Bacteria were tested for lysozyme, low pH, and bile acid resistance. Moreover, the strains were tested for adherence to human intestinal epithelial cells and screened for their immunomodulatory properties using a cellular model. Selected bacterial strains with anti-inflammatory properties were then tested in vivo in a dinitro-benzene sulfonic acid (DNBS)-induced chronic colitis mouse model, and weight loss, gut permeability, and cytokine profiles were measured as readouts of inflammation. One of the selected strains, Lactobacillus sanfranciscensis LBH1068, improved mice health as observed by a reduction of weight loss, significant decreases in gut permeability, and cytokine modulation. Altogether, our results highlighted the potential of lactobacilli isolated from pulque and in particular the strain L. sanfranciscensis LBH1068 as a novel probiotic to treat IBD.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/therapeutic use , Beverages/microbiology , Lactobacillus/classification , Lactobacillus/isolation & purification , Probiotics/pharmacology , Probiotics/therapeutic use , Animals , Anti-Inflammatory Agents/isolation & purification , Bacterial Adhesion , Cell Line , Cluster Analysis , Colitis/chemically induced , Colitis/drug therapy , Colitis/pathology , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , DNA, Ribosomal/chemistry , DNA, Ribosomal/genetics , Dinitrofluorobenzene/analogs & derivatives , Dinitrofluorobenzene/toxicity , Disease Models, Animal , Epithelial Cells/microbiology , Humans , Lactobacillus/physiology , Mexico , Mice , Molecular Sequence Data , Phylogeny , Probiotics/isolation & purification , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , Treatment Outcome
12.
Microbiology (Reading) ; 161(Pt 4): 708-18, 2015 Apr.
Article in English | MEDLINE | ID: mdl-25635270

ABSTRACT

The vaginal microbiota of healthy, fertile women is dominated by lactobacilli. As a defence mechanism, these bacteria produce H2O2 to discourage colonization of the vagina by undesirable micro-organisms. In particular, Lactobacillus jensenii CECT 4306 is a strong producer of H2O2 and has been found to protect itself from the bactericidal effects of this compound through the activity of extracellular peroxidases. However, this peroxidase activity is dependent on the presence of Fe(3+), which is found in elevated concentrations in the vaginal mucosa as a consequence of the menstrual discharge. The aim of the present work was to evaluate whether Fe(3+) is able to modulate other potential probiotic properties of strain 4306. We found that Fe(3+) enhances the adhesion of L. jensenii CECT 4306 to mucin and to HT-29 and HT-29 MTX cells, and, in addition, improves the anti-inflammatory profile, as judged by an increase in the ratio of IL-10/IL-12p70 that were secreted by macrophages. A comparison of total, secreted and surface proteins produced in the presence and absence of Fe(3+) revealed significant differences in the concentration of the moonlighting protein glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In conclusion, Fe(3+) seems to improve the probiotic characteristics of L. jensenii CECT 4306, and future research of the interactions of this strain with its vaginal environment may reveal further information about different aspects of its probiotic potential.


Subject(s)
Iron/metabolism , Lactobacillus/metabolism , Probiotics , Vagina/microbiology , Bacterial Adhesion , Bacterial Proteins , Cell Line , Female , Ferric Compounds/metabolism , Humans , Immunomodulation , Microbiota , Mucous Membrane/immunology , Mucous Membrane/microbiology , Proteome , Proteomics , Vagina/immunology
13.
BMC Microbiol ; 15: 67, 2015 Mar 21.
Article in English | MEDLINE | ID: mdl-25888448

ABSTRACT

BACKGROUND: The human gut houses one of the most complex and abundant ecosystems composed of up to 10(13)-10(14) microorganisms. The importance of this intestinal microbiota is highlighted when a disruption of the intestinal ecosystem equilibrium appears (a phenomenon called dysbiosis) leading to an illness status, such as inflammatory bowel diseases (IBD). Indeed, the reduction of the commensal bacterium Faecalibacterium prausnitzii (one of the most prevalent intestinal bacterial species in healthy adults) has been correlated with several diseases, including IBD, and most importantly, it has been shown that this bacterium has anti-inflammatory and protective effects in pre-clinical models of colitis. Some dysbiosis disorders are characterized by functional and physiological alterations. Here, we report the beneficial effects of F. prausnitzii in the physiological changes induced by a chronic low-grade inflammation in a murine model. Chronic low-grade inflammation and gut dysfunction were induced in mice by two episodes of dinitro-benzene sulfonic acid (DNBS) instillations. Markers of inflammation, gut permeability, colonic serotonin and cytokine levels were studied. The effects of F. prausnitzii strain A2-165 and its culture supernatant (SN) were then investigated. RESULTS: No significant differences were observed in classical inflammation markers confirming that inflammation was subclinical. However, gut permeability, colonic serotonin levels and the colonic levels of the cytokines IL-6, INF-γ, IL-4 and IL-22 were higher in DNBS-treated than in untreated mice. Importantly, mice treated with either F. prausnitzii or its SN exhibited significant decreases in intestinal permeability, tissue cytokines and serotonin levels. CONCLUSIONS: Our results show that F. prausnitzii and its SN had beneficial effects on intestinal epithelial barrier impairment in a chronic low-grade inflammation model. These observations confirm the potential of this bacterium as a novel probiotic treatment in the management of gut dysfunction and low-grade inflammation.


Subject(s)
Clostridiales/immunology , Enteritis/pathology , Enteritis/prevention & control , Animals , Benzenesulfonates/toxicity , Colon/pathology , Cytokines/analysis , Disease Models, Animal , Enteritis/chemically induced , Mice , Permeability , Serotonin/analysis
14.
Microb Cell Fact ; 14: 204, 2015 Dec 21.
Article in English | MEDLINE | ID: mdl-26691177

ABSTRACT

BACKGROUND: The L-arabinose isomerase is an intracellular enzyme which converts L-arabinose into L-ribulose in living systems and D-galactose into D-tagatose in industrial processes and at industrial scales. D-tagatose is a natural ketohexose with potential uses in pharmaceutical and food industries. The D-galactose isomerization reaction is thermodynamically equilibrated, and leads to secondary subproducts at high pH. Therefore, an attractive L-arabinose isomerase should be thermoactive and acidotolerant with high catalytic efficiency. While many reports focused on the set out of a low cost process for the industrial production of D-tagatose, these procedures remain costly. When compared to intracellular enzymes, the production of extracellular ones constitutes an interesting strategy to increase the suitability of the biocatalysts. RESULTS: The L-arabinose isomerase (L-AI) from Lactobacillus sakei was expressed in Lactococcus lactis in fusion with the signal peptide of usp45 (SP(Usp45)). The L-AI protein and activity were detected only in the supernatant of the induced cultures of the recombinant L. lactis demonstrating the secretion in the medium of the intracellular L. sakei L-AI in an active form. Moreover, we showed an improvement in the enzyme secretion using either (1) L. lactis strains deficient for their two major proteases, ClpP and HtrA, or (2) an enhancer of protein secretion in L. lactis fused to the recombinant L-AI with the SP(Usp45). Th L-AI enzyme secreted by the recombinant L. lactis strains or produced intracellularly in E. coli, showed the same functional properties than the native enzyme. Furthermore, when mice are fed with the L. lactis strain secreting the L-AI and galactose, tagatose was produced in vivo and reduced the glycemia index. CONCLUSIONS: We report for the first time the secretion of the intracellular L-arabinose isomerase in the supernatant of food grade L. lactis cultures with hardly display other secreted proteins. The secreted L-AI originated from the food grade L. sakei 23 K was active and showed the same catalytic and structural properties as the intracellular enzyme. The L. lactis strains secreting the L-arabinose isomerase has the ability to produce D-tagatose in vivo and conferred an anti-hyperglycemic effect to mice.


Subject(s)
Aldose-Ketose Isomerases/metabolism , Hexoses/metabolism , Hypoglycemic Agents/therapeutic use , Animals , Hypoglycemic Agents/administration & dosage , Mice
15.
Microb Cell Fact ; 14: 26, 2015 Feb 26.
Article in English | MEDLINE | ID: mdl-25889561

ABSTRACT

BACKGROUND: Different studies have described the successful use of recombinant lactic acid bacteria (recLAB) to deliver anti-inflammatory molecules at the mucosal level to treat Inflammatory Bowel Disease (IBD). METHODS: In order to identify the best strategy to treat IBD using recLAB, we compared the efficacy of different recombinant strains of Lactococcus lactis (the model LAB) secreting two types of anti-inflammatory molecules: cytokines (IL-10 and TGF-ß1) and serine protease inhibitors (Elafin and Secretory Leukocyte Protease Inhibitor: SLPI), using a dextran sulfate sodium (DSS)-induced mouse model of colitis. RESULTS: Our results show that oral administration of recombinant L. lactis strains expressing either IL-10 or TGF-ß1 display moderate anti-inflammatory effects in inflamed mice and only for some clinical parameters. In contrast, delivery of either serine protease inhibitors Elafin or SLPI by recLAB led to a significant reduction of intestinal inflammation for all clinical parameters tested. Since the best results were obtained with Elafin-producing L. lactis strain, we then tried to enhance Elafin expression and hence its delivery rate by producing it in a L. lactis mutant strain inactivated in its major housekeeping protease, HtrA. Strikingly, a higher reduction of intestinal inflammation in DSS-treated mice was observed with the Elafin-overproducing htrA strain suggesting a dose-dependent Elafin effect. CONCLUSIONS: Altogether, these results strongly suggest that serine protease inhibitors are the most efficient anti-inflammatory molecules to be delivered by recLAB at the mucosal level for IBD treatment.


Subject(s)
Interleukin-10/metabolism , Lactococcus lactis/metabolism , Serine Proteinase Inhibitors/metabolism , Transforming Growth Factor beta/metabolism , Administration, Oral , Animals , Colitis/microbiology , Colitis/pathology , Colitis/therapy , Disease Models, Animal , Elafin/genetics , Elafin/metabolism , Gene Expression/drug effects , Interleukin-10/genetics , Mice , Mice, Inbred C57BL , Nisin/pharmacology , Secretory Leukocyte Peptidase Inhibitor/genetics , Secretory Leukocyte Peptidase Inhibitor/metabolism , Serine Proteinase Inhibitors/genetics , Transforming Growth Factor beta/genetics
16.
Biotechnol Lett ; 37(6): 1203-11, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25868531

ABSTRACT

OBJECTIVES: To evaluate the effects of a recombinant strain of Lactococcus lactis secreting bioactive heme oxygenase-1 (LL-HO-1) in a model of hyperoxia-induced lung injury in rat pups RESULTS: Intranasal administration with LL-HO-1 significantly reduced hyperoxia-induced lung injury as demonstrated by a decreased wet/dry ratio, myeloperoxidase activity in lung tissue, tumor necrosis factor alpha levels in bronchoalveolar lavage (BAL) fluid, and attenuated lung injury scores. Although expression of HO-1 and NADPH oxidase-1 and production of superoxide in lung tissue were not affected by LL-HO-1 treatment, HO-1 levels in nasal mucosa and interleukin-10 concentrations in BAL fluid significantly increased compared with the hyperoxia control group CONCLUSIONS: Intranasal administration of LL-HO-1 protects against hyperoxia-induced lung damage, apparently through attenuation of inflammation.


Subject(s)
Acute Lung Injury/prevention & control , Heme Oxygenase-1/metabolism , Hyperoxia/complications , Lactococcus lactis/enzymology , Pneumonia/prevention & control , Acute Lung Injury/chemically induced , Acute Lung Injury/pathology , Administration, Intranasal , Animals , Animals, Newborn , Bronchoalveolar Lavage Fluid/chemistry , Heme Oxygenase-1/genetics , Interleukin-10/analysis , Lactococcus lactis/genetics , Lung/pathology , Peroxidase/analysis , Pneumonia/chemically induced , Pneumonia/pathology , Rats , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Severity of Illness Index
17.
Mol Microbiol ; 89(3): 518-31, 2013 Aug.
Article in English | MEDLINE | ID: mdl-23772975

ABSTRACT

Most bacteria of the genus Streptococcus are opportunistic pathogens, and some of them produce extracellular DNases, which may be important for virulence. Genome analyses of Streptococcus agalactiae (GBS) neonate isolate NEM316 revealed the presence of seven genes putatively encoding secreted DNases, although their functions, if any, are unknown. In this study, we observed that respiration growth of GBS led to the extracellular accumulation of a putative nuclease, identified as being encoded by the gbs0661 gene. When overproduced in Lactococcus lactis, the protein was found to be a divalent cation-requiring, pH-stable and heat-stable nuclease that we named Nuclease A (NucA). Substitution of the histidine(148) by alanine reduced nuclease activity of the GBS wild-type strain, indicating that NucA is the major nuclease ex vivo. We determined that GBS is able to degrade the DNA matrix comprising the neutrophil extracellular trap (NET). The nucA(H148A) mutant was impaired for this function, implicating NucA in the virulence of GBS. In vivo infection studies confirmed that NucA is required for full infection, as the mutant strain allowed increased bacterial clearance from lung tissue and decreased mortality in infected mice. These results show that NucA is involved in NET escape and is needed for full virulence.


Subject(s)
Bacterial Proteins/metabolism , Deoxyribonucleases/metabolism , Neutrophils/immunology , Streptococcal Infections/immunology , Streptococcus agalactiae/pathogenicity , Amino Acid Sequence , Amino Acid Substitution , Animals , Bacterial Proteins/genetics , Deoxyribonucleases/genetics , Humans , Immune Evasion , Lung/microbiology , Mice , Molecular Sequence Data , Neutrophils/microbiology , Streptococcal Infections/microbiology , Streptococcus agalactiae/enzymology , Streptococcus agalactiae/genetics , Toll-Like Receptor 9/immunology , Virulence
18.
Appl Environ Microbiol ; 80(3): 869-77, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24242245

ABSTRACT

The aims of this study were to develop strains of lactic acid bacteria (LAB) having both immunomodulatory and antioxidant properties and to evaluate their anti-inflammatory effects both in vitro, in different cellular models, and in vivo, in a mouse model of colitis. Different Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains were cocultured with primary cultures of mononuclear cells. Analysis of the pro- and anti-inflammatory cytokines secreted by these cells after coincubation with candidate bacteria revealed that L. delbrueckii subsp. bulgaricus CRL 864 and S. thermophilus CRL 807 display the highest anti-inflammatory profiles in vitro. Moreover, these results were confirmed in vivo by the determination of the cytokine profiles in large intestine samples of mice fed with these strains. S. thermophilus CRL 807 was then transformed with two different plasmids harboring the genes encoding catalase (CAT) or superoxide dismutase (SOD) antioxidant enzymes, and the anti-inflammatory effects of recombinant streptococci were evaluated in a mouse model of colitis induced by trinitrobenzenesulfonic acid (TNBS). Our results showed a decrease in weight loss, lower liver microbial translocation, lower macroscopic and microscopic damage scores, and modulation of the cytokine production in the large intestines of mice treated with either CAT- or SOD-producing streptococci compared to those in mice treated with the wild-type strain or control mice without any treatment. Furthermore, the greatest anti-inflammatory activity was observed in mice receiving a mixture of both CAT- and SOD-producing streptococci. The addition of L. delbrueckii subsp. bulgaricus CRL 864 to this mixture did not improve their beneficial effects. These findings show that genetically engineering a candidate bacterium (e.g., S. thermophilus CRL 807) with intrinsic immunomodulatory properties by introducing a gene expressing an antioxidant enzyme enhances its anti-inflammatory activities.


Subject(s)
Antioxidants/metabolism , Immunologic Factors/metabolism , Inflammation/immunology , Metabolic Engineering , Streptococcus thermophilus/immunology , Animals , Catalase/genetics , Catalase/metabolism , Cells, Cultured , Colitis/chemically induced , Colitis/immunology , Colitis/microbiology , Colitis/pathology , Cytokines/metabolism , Disease Models, Animal , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/microbiology , Mice , Streptococcus thermophilus/genetics , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism , Trinitrobenzenesulfonic Acid/administration & dosage , Trinitrobenzenesulfonic Acid/toxicity
19.
J Clin Gastroenterol ; 48 Suppl 1: S12-7, 2014.
Article in English | MEDLINE | ID: mdl-25291117

ABSTRACT

BACKGROUND: Oral treatment with Lactococcus lactis strains secreting the anti-inflammatory cytokine interleukin (IL)-10 has previously shown success as a therapy for inflammatory bowel diseases (IBD). GOALS: Our aim was to compare the protective effects of IL-10, delivered by recombinant lactoccoci using 2 novel expression systems, in a murine colitis model mimicking the relapsing nature of IBD. The first system is based on a Stress-Inducible Controlled Expression system for the production and delivery of heterologous proteins at mucosal surfaces and the second allows the delivery to the host cells of an il-10 cDNA cassette, harbored in a eukaryotic DNA expression vector (pValac). STUDY: Colitis was induced in female BALB/c mice by intrarectal injection of 2,4,6-trinitrobenzenesulphonic acid (TNBS). Mice that recovered received one of the bacteria treatments or saline solution orally during 14 days. Colitis was reactivated 25 days after the first TNBS injection with a second TNBS challenge. Three days after colitis reactivation, cytokine profiles and inflammation in colon samples were evaluated. RESULTS: Animals (N=9) receiving L. lactis strains secreting IL-10 using Stress-Inducible Controlled Expression system or delivering pValac:il-10 plasmid showed lower weight loss (P<0.005), lower damage scores (P<0.005), and immune activation in their large intestines compared with inflamed nontreated mice. CONCLUSIONS: Our results confirm the protective effect of IL-10 delivered either as a protein or as a cDNA in a colitis model mimicking the relapsing nature of IBD and provides a step further in the "proof-of-concept" of genetically engineered bacteria as a valid system to deliver therapeutic molecules at mucosal level.


Subject(s)
Colitis/prevention & control , Colon/microbiology , Genetic Vectors , Interleukin-10/biosynthesis , Intestinal Mucosa/microbiology , Lactobacillus/metabolism , Probiotics , Trinitrobenzenesulfonic Acid , Animals , Colitis/chemically induced , Colitis/genetics , Colitis/immunology , Colitis/metabolism , Colitis/microbiology , Colitis/pathology , Colon/immunology , Colon/metabolism , Colon/pathology , Disease Models, Animal , Female , Inflammation Mediators/metabolism , Interleukin-10/genetics , Interleukin-10/immunology , Intestinal Mucosa/immunology , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Lactobacillus/genetics , Lactobacillus/immunology , Mice, Inbred BALB C , Severity of Illness Index , Time Factors , Weight Loss
20.
Comput Struct Biotechnol J ; 23: 1-9, 2024 Dec.
Article in English | MEDLINE | ID: mdl-38094217

ABSTRACT

Inflammatory bowel disease (IBD) is a group of disorders characterized by an inflammation of the gastrointestinal tract (GIT) and represents a major social and economic burden. Despite ongoing research into the etiology and pathophysiology of this multifactorial disease, treatment options remain limited. From this perspective, the gut microbiota has emerged as a potential player in the pathogenesis of IBD, and animal and human studies support this hypothesis. Indeed, the human gut is one of the most complex ecological communities (composed of 1013-1014 microorganisms) that plays a critical role in human health by influencing normal physiology and disease susceptibility through its collective metabolic activities and host interactions. In addition, live probiotic bacteria present in some food products (which transit through the GIT) have been shown to interact with the host immune system and confer several health benefits. The aim of this review is to provide an overview of the link between Faecalibacterium duncaniae and Escherichia coli and IBD, highlighting the main areas of research in this field. An ecological perspective on the gut microbiota may offer new insights for the development of clinical therapies targeting this bacterial community to improve human health.

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