Microencapsulation of Lactic Acid Bacteria Improves the Gastrointestinal Delivery and in situ Expression of Recombinant Fluorescent Protein
| Autor: | Nina D. Coelho-Rocha, Camila P. de Castro, Luis C. L. de Jesus, Sophie Y. Leclercq, Savio H. de Cicco Sandes, Alvaro C. Nunes, Vasco Azevedo, Mariana M. Drumond, Pamela Mancha-Agresti |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Microbiology (medical) pExu vector lcsh:QR1-502 Microbiology lcsh:Microbiology law.invention DNA vaccination 03 medical and health sciences chemistry.chemical_compound In vivo law Fluorescence microscope sodium alginate encapsulation biology Chemistry Transfection biology.organism_classification Lactic acid 030104 developmental biology Biochemistry Recombinant DNA mCherry reporter protein mCherry recombinant Lactococcus lactis Bacteria DNA delivery |
| Zdroj: | Frontiers in Microbiology, Vol 9 (2018) |
| ISSN: | 1664-302X |
| DOI: | 10.3389/fmicb.2018.02398 |
| Popis: | The microencapsulation process of bacteria has been used for many years, mainly in the food industry and, among the different matrixes used, sodium alginate stands out. This matrix forms a protective wall around the encapsulated bacterial culture, increasing its viability and protecting against environmental adversities, such as low pH, for example. The aim of the present study was to evaluate both in vitro and in vivo, the capacity of the encapsulation process to maintain viable lactic acid bacteria (LAB) strains for a longer period of time and to verify if they are able to reach further regions of mouse intestine. For this purpose, a recombinant strain of LAB (L. lactis ssp. cremoris MG1363) carrying the pExu vector encoding the fluorescence protein mCherry [L. lactis MG1363 (pExu:mCherry)] was constructed. The pExu was designed by our group and acts as a vector for DNA vaccines, enabling the host cell to produce the protein of interest. The functionality of the pExu:mCherry vector, was demonstrated in vitro by fluorescence microscopy and flow cytometry after transfection of eukaryotic cells. After this confirmation, the recombinant strain was submitted to encapsulation protocol with sodium alginate (1%). Non-encapsulated, as well as encapsulated strains were orally administered to C57BL/6 mice and the expression of mCherry protein was evaluated at different times (0-168 h) in different bowel portions. Confocal microscopy showed that the expression of mCherry was higher in animals who received the encapsulated strain in all portions of intestine analyzed. These results were confirmed by qRT-PCR assay. Therefore, this is the first study comparing encapsulated and non-encapsulated L. lactis bacteria for mucosal DNA delivery applications. Our results showed that the microencapsulation process is an effective method to improve DNA delivery, ensuring a greater number of viable bacteria are able to reach different sections of the bowel. |
| Databáze: | OpenAIRE |
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