In Vitro Characterization of Lactococcus Lactis Subsp. Lactis as a protein delivery vehicle using TEM β-Lactamase as a model protein and C33-A and Caco-2 as cell lines
Gagan Kaushal, Doctoral Candidate, College of Pharmacy and Health Sciences
Jun Shao, Department of Pharmacy and Administrative Sciences, College of Pharmacy and Health Sciences
Oral delivery of protein and peptide drugs is always challenging due to poor stability and erratic absorption. In order to overcome these problems a delivery system that can protect a protein molecule and also prolong its stay at the site of absorption can be helpful.
In our study, we have used probiotic Lactococcus lactis as a novel oral protein delivery system. L. lactis is a probiotic bacteria with Generally Regarded as Safe (GRAS) status. Lactococcus lactis subsp. lactis was previously transformed by a special plasmid so that they can secrete TEM - Lactamase. The in vitro cultured monolayers of two cell lines, namely Caco-2 (Human Colorectal adeno carcinoma) and C33-A (Human Cervix Carcinoma) were used to mimic the epithelial membranes of the intestine and vagina. The absorption of TEM -Lactamase through these monolayers was studied. The potential toxicity of the bacterial delivery system on the cell monolayers was also investigated by TEER, radio-labeled-mannitol and trypan blue exclusion method.
Protein delivered by the bacteria was more efficient than by solution (increased by 27% + 0.14% in Caco-2 case; 50% + 0.06% in C33-A case). It was found that the integrity of monolayers was maintained within 6 hours after the addition of the bacterial delivery systems. Transport of mannitol through Caco-2 cell monolayer was significantly increased and the transport of propranolol through Caco-2 cell monolayer was significantly decreased in the presence of L. lactis. The presence of L. lactis did not change the transport of propranolol and mannitol through the monolayers for six hours, and then slightly decreased the transport of propranolol but slightly increased the transport of mannitol through C-33A monolayer.
C-33A monolayer was more permeable to proteins than Caco-2 monolayer, suggesting vaginal route may provide advantages over oral route for protein delivery.