Therapeutic microvesicles of probiotic bacteria
A probiotic, therapeutic technology, applied in the field of therapeutic microbubbles, can solve the problem that the time of emergence is not immediate
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Embodiment 1
[0158] Example 1 - Altered enzymatic activity associated with MV production
[0159]Lactobacillus reuteri DSM 17938 was cultured and subjected to different inducing treatments at specific time points. Responses to these inducing treatments were determined using enzymatic assays and compared to the responses obtained with control treatments. The inductive treatment involves inducing stress to the bacteria during their growth phase and then measuring the effect on enzymatic activity in the bacterial conditioned medium.
[0160] Materials and Methods
[0161] Culture / Sample Collection
[0162] Lactobacillus reuteri DSM 17938 was inoculated from a frozen stock into 25 mL of de Man-Rogosa-Sharpe (MRS) medium under normal culture conditions, ie anaerobically grown overnight at 37°C. Bacteria (20 mL) were then re-inoculated into 200 mL MRS, and different inducing treatments were applied, as described in more detail below. Bacterial samples at 5000 x g Centrifuge for 10 min, tr...
Embodiment 2
[0179] Example 2 - Specific culture conditions are associated with increased MV production
[0180] L. reuteri DSM 17938 was cultured and subjected to different inducing treatments. The amount of MV produced as a result of these inductive treatments was determined using Nanoparticle Tracking Analysis (NTA), and the results were compared to the responses obtained with control treatments. Inductive treatments involved oxygen treatment and sucrose treatment, and the effect on MV production was measured in bacterial conditioned media.
[0181] Materials and Methods
[0182] oxygen treatment
[0183] Lactobacillus reuteri DSM 17938 was cultured under normal culture conditions, i.e. anaerobically in bottles / flasks at 37°C in de Man-Rogosa-Sharpe (MRS) medium. Oxygen treatment was induced by vigorous shaking of the bacterial culture for 24 hours. As a control, Lactobacillus reuteri DSM 17938 was cultured under normal culture conditions, i.e. anaerobic culture in MRS medium at 3...
Embodiment 3
[0195] Example 3 - Isolated Bacterial Microvesicles Recapitulate Bacterial Effects on Intestinal Motility
[0196] Materials and Methods
[0197] animal
[0198] Adult male Swiss Webster mice (6-8 weeks) were obtained from Charles River Laboratories (Wilmington, MA, USA). Animals were housed 4-5 / cage under a 12 hour light / dark cycle and provided food and water ad libitum. Subsequent procedures following cervical dislocation were performed in vitro according to the McMaster Animal Ethics Research Board (AREB) (permit 16-08-30).
[0199] Tissue Flotation Bath Records
[0200]Tissue flotation bath recordings were performed as described in Wu et al. (2013). Jejunum and colon segments with a minimum length of 4 cm were extracted and fixed in a 20 mL tissue flotation bath filled with oxygenated Krebs at 34°C. The buccal end of the segment was cannulated and the contents flushed from the lumen by gravity perfusion of Krebs inflated with carbon gas using a Mariotte bottle. O...
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