Iskanje po repozitoriju

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Povzetek: Introduction: With growing evidence of clinical efficacy of probiotics in various diseases, safety concerns have arisen regarding the therapeutic use of live probiotic bacteria, especially in critically ill, immunocompromised, and pediatric populations. Serious probiotic-related adverse effects have been reported in these patients, including bloodstream infection and sepsis. This has led to an increased interest in developing postbiotics (non-viable bacterial products) that may exert beneficial effects on the host without the risks associated with administration of live microorganisms. The aim of this study was to explore postbiotic potential of recombinant Lactococcus lactis bacteria that have been engineered to display interleukin 6 (IL6)-targeting affibody (ZIL6) on their surface and are intended for treatment of inflammatory intestinal diseases. Methods: Five different killing treatments were applied to kill bacteria (heat, ethanol, sonication, UV, and gamma irradiation) and their effect on bacterial viability, morphology and functionality was examined in vitro using a combination of different techniques, including microscopy, flow cytometry, immunoassays and cell-based reporter assay. Results: The results showed that ZIL6 affibody displayed on L. lactis via non-covalent anchoring withstood the treatments applied to kill bacteria and remained functional after the loss of microbial viability. The degree of functionality was dependent on the type of treatment. Heat-killed cells retained 50% of the activity of live strain, while most of the activity was preserved after exposure of bacteria to ethanol, sonication, UV and gamma irradiation. The applied treatments varied in killing efficacy, whereby ethanol and heat rendered bacteria non-viable, UV and gamma irradiation yielded non-replicative cells, whereas sonication was ineffective in killing L. lactis. Among non-viable cells, ethanol-killed bacteria exhibited the greatest activity and showed high maximum binding capacity of 200 ng IL6 per mg dry cell weight, possessed strong nanomolar affinity for IL6, and inhibited up to 78% of IL6-induced STAT3 signaling. Conclusion: The study demonstrates that functional non-viable bacterial cells can be derived from the recombinant L. lactis with therapeutic proteins displayed on their surface and provides a good foundations for further studies of their postbiotic potential in adjunctive therapy of inflammatory intestinal diseases.
Ključne besede: affibody, probiotics, postbiotics, Lactococcus lactis
Objavljeno v DiRROS: 03.11.2025; Ogledov: 187; Prenosov: 97
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Povzetek: Bacterial biofilms attach to various surfaces and represent an important clinical and public health problem, as they are highly recalcitrant and are often associated with chronic, nonhealing diseases and healthcare-associated infections. Antibacterial agents are often not sufficient for their elimination and have to be combined with mechanical removal. Mechanical forces can be generated by actuating nonspherical (anisotropic) magnetically responsive nanoparticles in a rotating magnetic field. We have thus prepared anisotropic superparamagnetic nanochains in the size range of 0.5−1 μm by magnetically assembling several iron oxide nanoparticle clusters and coating them with a layer of silica with different shell morphologies: smooth, moderately rough, and highly rough. The silica surface was additionally functionalized with carboxylic groups to increase colloidal stability. The efficacy of the nanochains in biofilm removal was studied systematically with three different model nonpathogenic bacterial species Escherichia coli, Lactococcus lactis, and Pseudomonas fragi; two different magnetic field strengths; two stirring speeds; and two treatment durations. All bacterial species were engineered to express fluorescent proteins to enable quantification of biofilm removal by colony-forming unit count and fluorescence measurements. Nanochains removed >90% of Gram-negative E. coli and P. fragi with a stronger magnetic field, and <90% of Gram-positive L. lactis with a weaker magnetic field. Surface roughness of nanochains, duration, and stirring speed also affected removal, but the effect could not be generalized. In contrast to their effects on biofilms, the functionalized nanochains showed no toxicity to Caco-2 intestinal epithelial cells, regardless of whether magnetomechanical force was employed or not. In summary, we demonstrated that remotely controlled spatial movement of nanoparticles can generate sufficient mechanical forces to disperse attached biofilms while retaining safety in an epithelial cell model.
Ključne besede: bacterial biofilm, magnetomechanical detachment, magnetic nanoparticles, nanochains
Objavljeno v DiRROS: 09.09.2025; Ogledov: 271; Prenosov: 118
Celotno besedilo (6,84 MB)
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