Phagocytosis of lactobacilli with different susceptibility to human β defensins.

SZYMANOWSKI, F; HUGO, A. A; TYMCZYSZYN, E. E, GÓMEZ-ZAVAGLIA, A; PÉREZ, P. F.

Congreso; X Congreso Argentino de Microbiologia General (SAMIGE); 2014

Phagocytic cells play a key role in host´s defenses and constitute a link between innate and adaptive immune responses. The activity of phagocytic cells is crucial to reduce bacterial load at the site of infection and also for antigen processing and presentation. Antimicrobial peptides such as defensins are involved in the inactivation of internalized microorganisms and the differential interaction of microorganisms with those antibacterial molecules could modifies immune response. Human β defensins (HBDs) are cationic antimicrobial peptides secreted by epithelial cells of different mucosa. Positive charged defensins interact with negatively charged bacterial surface structures thus increasing membrane permeability and leading to cell death. Considering that lactic acid bacteria (LAB) have demonstrated an interesting probiotic potential, differences in the interaction of LAB with defensins could modify the fate of internalized microorganisms and cell response. Since we have demonstrated that related lactic acid bacteria (i. e. Lactobacillus delbrueckii subsp. lactis CIDCA 133 and L. delbrueckii subsp. bulgaricus CIDCA 331) have different susceptibility to human defensins, in the present work we sought to gain insight on the effect of HBD-1 and HBD-2 on the interaction between lactobacilli and cultured phagocytic cells. Bacteria were incubated with HBD-1 and HBD-2 at different concentrations. Membrane permeabilization was evaluated by flow cytometry (FACS) after propidium iodine (PI) staining. Zeta potential was determined with a Z meter system. Phagocytosis was evaluated by FACS analysis incubating FITC-labeled bacteria at different multiplicities of infection (m.o.i) with THP-1 cells differentiated with phorbol myristate acetate (PMA). Trypan blue was used as quenching solution. Bacterial adhesion was assesed by viable counts.Strain CIDCA 331 was more susceptible to HBD-2 than strain CIDCA 133. Indeed percentages of PI (+) cells were 2. 6 to 3.5-fold higher for strain CIDCA 331 at 4 and 8 ug/ml respectively. Both lactobacilli were unaffected by HBD-1. The surface potential of both strains were negative, being -5,07(mV) for strain CIDCA 133 and -28,9 mV for strain 331. Bacterial adhesion and internalization by THP-1 cells were higher for strain 133 at all the moi tested. Results indicate that strain CIDCA 331 is more susceptible to HBD-2 than strain CIDCA133. These findings could be ascribed to differences in bacterial surface charges that correlate with the lower glycolipid/phospolipid ratio of the membrane of strain CIDCA 331. This characteristic determines higher negative surface charge. Strain CIDCA 133, which is more resistant to HBD-2, showed higher interaction with THP-1 cells than strain CIDCA 331 and it was endocytosed in high amounts by THP-1 cells.