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Applied and Environmental Microbiology, August 2008, p. 4746-4755, Vol. 74, No. 15
0099-2240/08/$08.00+0     doi:10.1128/AEM.00412-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Suppressive Effect on Activation of Macrophages by Lactobacillus casei Strain Shirota Genes Determining the Synthesis of Cell Wall-Associated Polysaccharides

Emi Yasuda, Masaki Serata, and Tomoyuki Sako^*

Yakult Central Institute for Microbiological Research, Yaho, Kunitachi, Tokyo 186-8650, Japan

Received 19 February 2008/ Accepted 9 June 2008

Although many Lactobacillus strains used as probiotics are believed to modulate host immune responses, the molecular natures of the components of such probiotic microorganisms directly involved in immune modulation process are largely unknown. We aimed to assess the function of polysaccharide moiety of the cell wall of Lactobacillus casei strain Shirota as a possible immune modulator which regulates cytokine production by macrophages. A gene survey of the genome sequence of L. casei Shirota hunted down a unique cluster of 10 genes, most of whose predicted amino acid sequences had similarities to various extents to known proteins involved in biosynthesis of extracellular or capsular polysaccharides from other lactic acid bacteria. Gene knockout mutants of eight genes from this cluster resulted in the loss of reactivity to L. casei Shirota-specific monoclonal antibody and extreme reduction of high-molecular-mass polysaccharides in the cell wall fraction, indicating that at least these genes are involved in biosynthesis of high-molecular-mass cell wall polysaccharides. By adding heat-killed mutant cells to mouse macrophage cell lines or to mouse spleen cells, the production of tumor necrosis factor alpha, interleukin-12 (IL-12), IL-10, and IL-6 was more stimulated than by wild-type cells. In addition, these mutants additively enhanced lipopolysaccharide-induced IL-6 production by RAW 264.7 mouse macrophage-like cells, while wild-type cells significantly suppressed the IL-6 production of RAW 264.7. Collectively, these results indicate that this cluster of genes of L. casei Shirota, which have been named cps1A, cps1B, cps1C, cps1D, cps1E, cps1F, cps1G, and cps1J, determine the synthesis of the high-molecular-mass polysaccharide moiety of the L. casei Shirota cell wall and that this polysaccharide moiety is the relevant immune modulator which may function to reduce excessive immune reactions during the activation of macrophages by L. casei Shirota.

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* Corresponding author. Mailing address: Yakult Central Institute for Microbiological Research, 1796 Yaho, Kunitachi, Tokyo 186-8650, Japan. Phone: 81-42-577 89 60. Fax: 81-42-577 30 20. E-mail: tomoyuki-sako{at}yakult.co.jp

Published ahead of print on 13 June 2008.

E.Y., M.S., and T.S. contributed equally to this work.

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Applied and Environmental Microbiology, August 2008, p. 4746-4755, Vol. 74, No. 15
0099-2240/08/$08.00+0     doi:10.1128/AEM.00412-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Lebeer, S., Vanderleyden, J., De Keersmaecker, S. C. J. (2008). Genes and Molecules of Lactobacilli Supporting Probiotic Action. Microbiol. Mol. Biol. Rev. 72: 728-764 [Abstract] [Full Text]