The influence of cell-free Lactobacillus rhamnosus GG supernatant on the phagocytic activity of macrophages
James E. Vincenti
Teesside University, Middlesbrough, Tees Valley, TS1 3BA, UK
27 Sept 2009
10 Mar 2010
11 Apr 2010
macrophage, probiotics, Lactobacillus rhamnosus GG, Escherichia coli, gentamicin protection assay
Bacteria such as Lactobacillus spp. are classed as probiotics and are already in mainstream use both in hospitals and the consumer market. They have been shown to confer a beneficial effect upon gastrointestinal health when administered in sufficient dosage; however, little is known on how this is achieved. Past studies have shown probiotics improve a number of pathological conditions by interacting with the host immune system and have demonstrated an increase in lysozyme and production of reactive oxygen species in mononuclear phagocytes. With the aim to understand further the bacteria–host interactions, this study endeavoured to test for the up-regulation of phagocytic activity. A gentamicin protection assay was utilized to quantify Escherichia coli ingested by murine macrophages (J774) at 15, 30 and 60 min periods. A multiplicity of infection of 100:1 was adopted with a bacterial concentration of 1 × 107 CFU ml−1 and a macrophage cell number of 1 × 105 ml−1. The assay was performed in DMEM alone or DMEM supplemented with either 20 µg ml−1 lipopolysaccharide (LPS) or 10% cell-free Lactobacillus rhamnosus GG (LGG) supernatant. Viable bacterial cells recovered from lysed macrophages at 15 min were negligible for all treatments. Following 30 min incubation, bacterial recovery was observed with 6.4, 5.3 and 3.8 E. coli recovered per macrophage from the control, LPS and LGG supernatant treatments, respectively. After 60 min incubation E. coli recovery remained the same for the control group, but declined by approximately an order of magnitude to 0.6 and 0.5 E. coli per macrophage for the LPS and LGG supernatant treatments. The reduced recovery of E. coli cells from macrophages, when treated with LPS or LGG supernatants, suggests these compounds modulate macrophage activity by enhancing phagocytic digestion of bacterial cells.