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Lactococcus lactis Delivery of Surface Layer Protein A Protects Mice from Colitis by Re-Setting Host Immune Repertoire.

ABSTRACT: Inflammatory bowel disease (IBD) is characterized by gastrointestinal inflammation comprised of Crohn's disease and ulcerative colitis. Centers for Disease Control and Prevention report that 1.3% of the population of the United States (approximately 3 million people) were affected by the disease in 2015, and the number keeps increasing over time. IBD has a multifactorial etiology, from genetic to environmental factors. Most of the IBD treatments revolve around disease management, by reducing the inflammatory signals. We previously identified the surface layer protein A (SlpA) of Lactobacillus acidophilus that possesses anti-inflammatory properties to mitigate murine colitis. Herein, we expressed SlpA in a clinically relevant, food-grade Lactococcus lactis to further investigate and characterize the protective mechanisms of the actions of SlpA. Oral administration of SlpA-expressing L. lactis (R110) mitigated the symptoms of murine colitis. Oral delivery of R110 resulted in a higher expression of IL-27 by myeloid cells, with a synchronous increase in IL-10 and cMAF in T cells. Consistent with murine studies, human dendritic cells exposed to R110 showed exquisite differential gene regulation, including IL-27 transcription, suggesting a shared mechanism between the two species, hence positioning R110 as potentially effective at treating colitis in humans.

SUBMITTER: Arukha AP

PROVIDER: S-EPMC8470720 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Host lysozyme-mediated lysis of Lactococcus lactis facilitates delivery of colitis-attenuating superoxide dismutase to inflamed colons.

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| S-EPMC4485081 | biostudies-literature

Mucosal delivery of Lactococcus lactis carrying an anti-TNF scFv expression vector ameliorates experimental colitis in mice.

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| S-EPMC6593574 | biostudies-literature

Oral delivery of Lactococcus lactis that secretes bioactive heme oxygenase-1 alleviates development of acute colitis in mice.

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Food-grade delivery system for controlled gene expression in Lactococcus lactis.

Project description:A food-grade system for the delivery of desired genes to Lactococcus lactis, their inducible expression, and their transfer to related strains was established. Based on the thermosensitive pG(+)host replicon, two types of plasmid vectors were constructed which contained sections of either the chromosomal leu operon of L. lactis or the tel operon from the lactococcal sex factor. Genes cloned into the leu or tel sequences of these vectors were delivered to the homologous regions of the chromosome or the sex factor through two single crossovers, leading to integration of the recombinant plasmids and subsequent excision of the vector portions. Inducible transcription of integrated genes was achieved by using the nisin-controlled expression (NICE) system. To establish the signal transduction genes nisRK in L. lactis, the vectors pLNG1363 (targeted to the chromosome) and pUK500 (targeted to the sex factor) were constructed. Fusions of six different peptidase genes (pep) from Lactobacillus delbrueckii with the nisin-inducible promoter P(nisA) were delivered to the sex factor with derivatives of the vector pUK300. Food-grade recombinants of L. lactis were constructed which had the nisRK genes and individual P(nisA)::pep fusions integrated either separately into the chromosome and the sex factor or simultaneously into the sex factor. With both types of recombinants, expression of P(nisA)::pep fusions after induction with nisin was demonstrated. Depending on the loci used for integration of nisRK, variable induction rates were observed. Furthermore, an engineered sex factor carrying a P(nisA)::pepI fusion was transfered by conjugation between two strains of L. lactis at a frequency of 4 x 10(-4).

| S-EPMC129891 | biostudies-literature

PpiA, a surface PPIase of the cyclophilin family in Lactococcus lactis.

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Screening for New Surface Anchoring Domains for <i>Lactococcus lactis</i>.

Project description:The display of recombinant proteins on bacterial surfaces is a developing research area with a wide range of potential biotechnological applications. The lactic acid bacterium Lactococcus lactis is an attractive host for such surface display, and a promising vector for in vivo delivery of bioactive proteins. Surface-displayed recombinant proteins are usually anchored to the bacterial cell wall through anchoring domains. Here, we investigated alternatives to the commonly applied lactococcal lysine motif (LysM)-containing surface anchoring domain, the C-terminus of AcmA (cAcmA). We screened 15 anchoring domains of lactococcal or phage origins that belong to the Pfam categories LPXTG, LysM, CW_1, Cpl-7, WxL, SH3, and ChW, which can provide non-covalent or covalent binding to the cell wall. LPXTG, LysM, the duplicated CW_1 and SH3 domains promoted significant surface display of two model proteins, B domain and DARPin I07, although the display achieved was lower than that for the reference anchoring domain, cAcmA. On the other hand, the ChW-containing anchoring domain of the lactococcal phage AM12 endolysin (cAM12) demonstrated surface display comparable to that of cAcmA. The anchoring ability of cAM12 was confirmed by enabling non-covalent heterologous anchoring of the B domain on wild-type bacteria, as well as anchoring of CXCL8-binding evasin-3, which provided potential therapeutic applicability; both were displayed to an extent comparable to that of cAcmA. We have thereby demonstrated the effective use of different protein anchoring domains in L. lactis, with ChW-containing cAM12 the most promising alternative to the established approaches for surface display on L. lactis.

| S-EPMC6700490 | biostudies-literature

Targeting IL-6 by engineered Lactococcus lactis via surface-displayed affibody.

Project description:BackgroundDysregulated production of interleukin (IL)-6 is implicated in the pathology of inflammatory bowel disease (IBD). Neutralization of IL-6 in the gut by safe probiotic bacteria may help alleviate intestinal inflammation. Here, we developed Lactococcus lactis with potent and selective IL-6 binding activity by displaying IL-6-specific affibody on its surface.ResultsAnti-IL-6 affibody (designated as ZIL) was expressed in fusion with lactococcal secretion peptide Usp45 and anchoring protein AcmA. A high amount of ZIL fusion protein was detected on bacterial surface, and its functionality was validated by confocal microscopy and flow cytometry. Removal of IL-6 from the surrounding medium by the engineered L. lactis was evaluated using enzyme-linked immunosorbent assay. ZIL-displaying L. lactis sequestered recombinant human IL-6 from the solution in a concentration-dependent manner by up to 99% and showed no binding to other pro-inflammatory cytokines, thus proving to be highly specific for IL-6. The removal was equally efficient across different IL-6 concentrations (150-1200 pg/mL) that were found to be clinically relevant in IBD patients. The ability of engineered bacteria to capture IL-6 from cell culture supernatant was assessed using immunostimulated human monocytic cell lines (THP-1 and U-937) differentiated into macrophage-like cells. ZIL-displaying L. lactis reduced the content of IL-6 in the supernatants of both cell lines in a concentration-dependent manner by up to 94%. Dose response analysis showed that bacterial cell concentrations of 107 and 109 CFU/mL (colony forming units per mL) were required for half-maximal removal of recombinant and macrophage-derived IL-6, respectively.ConclusionThe ability of ZIL-displaying L. lactis to bind pathological concentrations of IL-6 at common bacterial doses suggests physiological significance.

| S-EPMC9287920 | biostudies-literature

Lactococcus lactis stringent response

Project description:The stringent response was defined in Lactococcus lactis through transcript profiling after the addition of a chemical inductor, the norvaline. Gene expression was measured in the exponential growth phase (reference sample) and at 1.6 h after norvaline addition. Four hundred and sixty one differentially expressed genes were identified and constituted the stringent response regulon. Keywords: stress response, time course

2009-04-28 | GSE10254 | GEO

LmrR Lactococcus lactis MG1363

Project description:Comparison of L. lactis NZ9000 ΔlmrR versus L. lactis NZ9000 wild type Keywords: Transcription profiling

2007-10-04 | GSE9168 | GEO

Recombinant Lactococcus lactis expressing porcine insulin-like growth factor I ameliorates DSS-induced colitis in mice.

Project description:BackgroundInsulin-like growth factor I (IGF-I) is one important family of growth factors, which plays key role in intestinal growth, regeneration, and damage repair. However, the low natural abundance of IGF-I limits its research opportunities and practical application in the fields of medicine and animal husbandry. In this study, a tandem repeat strategy was used to express three copies of the same pIGF-I3 protein in L. lactis. The activity of recombinant pIGF-I3 (rpIGF-I3) was further examined by a mouse model of dextran sulfate sodium (DSS)-induced colitis. In addition, the potential of recombinant L. lactis expressing pIGF-I3 to reduce inflammatory disease was evaluated.ResultspIGF-I3 could be expressed in L. lactis by the detection of SDS-PAGE and Western blot. Experimental colitis was induced in BALB/c mice by administration of 5 % DSS in drinking water, and the clinical symptoms were observed in DSS-treated mice. Oral administration of recombinant L. lactis expressing pIGF-I3 improved the colonic architecture, and significantly reduced the increase of colonic damage score (P < 0.05). Furthermore, recombinant L. lactis expressing pIGF-I3 treatment significantly reduced serum DAO activity and colonic MPO level, and elevated colonic occludin level compared to the DSS group (P < 0.05).ConclusionsThe pIGF-I3 expressed in L. lactis has good biological activity, and oral administration of recombinant L. lactis expressing pIGF-I3 attenuated the symptoms and development of DSS-induced colitis in mice. These suggested that L. lactis could be a potential host bacterium for production and delivery of IGF-I against intestinal diseases.

| S-EPMC4774141 | biostudies-literature

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