Abstract
The present study investigated the in vitro antibacterial, antibiofilm and anti-Quorum Sensing (anti-QS) activities of canine bone marrow mesenchymal stem cell-conditioned media (cBM MSC CM) containing all secreted factors <30 K, using a disc diffusion test (DDT), spectrophotometric Crystal Violet Assay (SCVA) and Bioluminescence Assay (BA) with QS-reporter Escherichia coli JM109 pSB1142. The results show a sample-specific bacterial growth inhibition (zones varied between 7–30 mm), statistically significant modulation of biofilm-associated Staphylococcus aureus and Escherichia coli bioluminescence (0.391 ± 0.062 in the positive control to the lowest 0.150 ± 0.096 in the experimental group, cf. 11,714 ± 1362 to 7753 ± 700, given as average values of absorbance A550 ± SD versus average values of relative light units to growth RLU/A550 ± SD). The proteomic analysis performed in our previous experiment revealed the presence of several substances with documented antibacterial, antibiofilm and immunomodulatory properties (namely, apolipoprotein B and D; amyloid-β peptide; cathepsin B; protein S100-A4, galectin 3, CLEC3A, granulin, transferrin). This study highlights that cBM MSC CM may represent an important new approach to managing biofilm-associated and QS signal molecule-dependent bacterial infections. To the best of our knowledge, there is no previous documentation of canine BM MSC CM associated with in vitro antibiofilm and anti-QS activity.
Highlights
As antibiotic resistance has increased rapidly, thereby greatly limiting the medications available to treat chronic bacterial infections in clinical practice, the development of a new antimicrobial agent, especially one effective against multidrug resistant pathogens [1] and/or bacteria living in biofilm with adaptation resistance, is an urgent issue [2].As multipotent adult stem cells with the capacity to differentiate into multiple cell types [3,4] and with a paracrine function, mesenchymal stem cells (MSCs) may be one of the options offering new prospects for the prevention and treatment of infections, for limiting the selection and spread of antibiotic resistance, or for potentially acting as drug delivery vehicles and enhancing the effectiveness of conventional antimicrobials
Escherichia coli (E. coli) JM109 pSB1142: luminescence-based Quorum Sensing (QS)-reporter strain responding to long-chain acyl homoserine lactone (AHL) (C10–C14) carrying the lasR and lasI promoter of Pseudomonas aeruginosa (P. aeruginosa) fused to the luxCDABE cassette from Photorhabdus luminescens was cultured in Brain Heart Infusion (BHI) agar (Oxoid, Basingstoke, Hampshire, UK) at 37 ◦C and maintained in tetracycline 20 μg/mL
All six experimental canine bone marrow mesenchymal stem cell (cBM MSC) Conditioned Media (CM) groups showed a statistically significant bioluminescence inhibition with a varying force of action (11,714 ± 1362 in P. aeruginosa CFCS (PC) group to the lowest 7753 ± 700 in experimental cBM MSC CM Alaskan Malamute (AM), and/or to the highest 10,294 ± 387 in experimental cBM MSC CM L; given as average values of RLU/A550 ± standard deviation (SD); p value < 0.001; p value < 0.05). These results indicate a bioluminescence reduction in QS-related E. coli JM109 pSB1142 ranging from 11% to 34% (Table 2). These results suggest the capacity of cBM MSC CM to modify and degrade AHL autoinducers, thereby attenuating QS-dependent virulence in Gram negative P. aeruginosa
Summary
As antibiotic resistance has increased rapidly, thereby greatly limiting the medications available to treat chronic bacterial infections in clinical practice, the development of a new antimicrobial agent, especially one effective against multidrug resistant pathogens [1] and/or bacteria living in biofilm with adaptation resistance, is an urgent issue [2].As multipotent adult stem cells with the capacity to differentiate into multiple cell types [3,4] and with a paracrine function, mesenchymal stem cells (MSCs) may be one of the options offering new prospects for the prevention and treatment of infections, for limiting the selection and spread of antibiotic resistance, or for potentially acting as drug delivery vehicles and enhancing the effectiveness of conventional antimicrobials. Interest in mesenchymal stem cells (MSCs) for regenerative and reparative therapies, both in human and animals, is emerging, as the current treatment options for several conditions often do not result in either the desired clinical outcome or the patients’ return to normal function [10]. Canine MSCs have been evaluated in some experimental and preclinical studies on the efficacy and safety testing of novel treatments for humans, since dogs are considered as potential animal models for human disease research (based on several reasons, such as a relatively long life span, large body mass and natural disease onset) [11] and have a prevalent role in the development of new therapeutic cell-based approaches. Research on canine-derived MSCs may provide insight into stem cell therapy not just for canines, but for humans as well [13]
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