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Clinical and Vaccine Immunology, September 2007, p. 1138-1148, Vol. 14, No. 9
1071-412X/07/$08.00+0     doi:10.1128/CVI.00079-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Aggregating Phenotype in Lactobacillus crispatus Determines Intestinal Colonization and TLR2 and TLR4 Modulation in Murine Colonic Mucosa

Department of Histology, Microbiology, and Medical Biotechnologies,¹ Department of Gastroenterological Sciences,² Department of Human Anatomy and Physiology, University of Padua, Padua, Italy,³ AAT-Advanced Analytical Technologies S.r.l., Piacenza, Italy⁴

Received 5 February 2007/ Returned for modification 18 April 2007/ Accepted 5 July 2007

The colonic microbiota is a major modulator of the mucosal immune system; therefore, its manipulation through supplementation with probiotics may significantly affect the host's immune responses. Since different probiotics seem to exert various effects in vivo, we tested the relevance of the autoaggregation phenotype on the intestinal persistence of lactobacilli and their ability to modulate the host's innate immune responses. After 14 days of diet supplementation, the aggregating strain Lactobacillus crispatus M247 but not aggregation-deficient isogenic mutant MU5 was recovered from the feces and colonic mucosa of mice. This observation was confirmed by strain-specific PCR amplification and by Lactobacillus-specific denaturing gradient gel electrophoresis analysis. Indeed, L. crispatus M247 increased Toll-like receptor 2 (TLR2) mRNA levels, while it reduced TLR4 mRNA and protein levels in the colonic mucosa, whereas MU5 was ineffective. In colonic epithelial cells (CMT-93 cells) L. crispatus M247 but not MU5 induced time-dependent extracellular signal-regulated kinase-1 (ERK1) tyrosine phosphorylation and TLR modulation, which were abolished in the presence of PD98059 (an ERK1 inhibitor). To assess the functional relevance of probiotic-induced TLR modulation, we determined the consequences of L. crispatus preexposure on TLR4 (lipopolysaccharide [LPS]) and TLR2 [Pam₃Cys-Ser-(Lys)₄] ligand-mediated effects in intestinal epithelial cells. Preexposure to L. crispatus M247 blunted LPS-induced interleukin-6 (IL-6) release and inhibition of CMT-93 migration over a wound edge, whereas it enhanced TLR2-mediated IL-10 up-regulation. In summary, the aggregation phenotype is required for L. crispatus persistence in the colon and for modulation of TLR2/TLR4 expression through an ERK-dependent pathway. We speculate that the aggregation phenotype in L. crispatus M247 is required to temper epithelial cell responsiveness to bacterial endotoxins, which thus affects the evolution of intestinal inflammatory processes.

Published ahead of print on 18 July 2007.