Some lactic acid bacteria play an important role in the immune system with potential benefits to the host. However, detailed mechanisms of immune modulation exerted by probiotics remain to be clarified. Since immune response changes in a time-related manner in some cases, we monitored changes in mRNA levels in the spleen of mice during 14 d feeding with Lactobacillus brevis KB290 (KB290). Female BALB/c mice, aged 9 weeks, commenced a diet containing KB290 (3 × 109colony-forming units/g) or starch for a period of 1, 4, 7 or 14 d. Cytotoxic activity of the resulting splenocytes against YAC-1 cells was measured using flow cytometry. The activity was found to be significantly higher in the treated group on days 1 and 7. The highest activity appeared on day 4, but was not statistically significantly different. Gene expression profiles were analysed using DNA microarray. Gene Ontology (GO) terms related to the immune process were significantly enriched in the up-regulated gene set on days 1, 4 and 7, and GO terms related to the cellular process were enriched in the down-regulated gene set on days 4 and 7. Although the up-regulated genes involved in antigen processing and presentation for stimulation of CD8+ cytotoxic T cells were not observed on day 14, some genes involved in T-cell and natural killer cell activation remained up-regulated until day 14. For the majority of the genes tested, RT-PCR analysis was used to verify the results obtained from the DNA microarray analysis. The sequential gene expression profiling reflected changes in cytotoxic activity during KB290 feeding.

Lactic acid bacteria confer a variety of health benefits. Here, we investigate the mechanisms by which Lactobacillus brevis KB290 (KB290) enhances cell-mediated cytotoxic activity. Female BALB/c mice aged 9 weeks were fed a diet containing KB290 (3 × 109 colony-forming units/g) or starch for 1 d. The resulting cytotoxic activity of splenocytes against YAC-1 cells was measured using flow cytometry and analysed for gene expression using DNA microarray technology. KB290 enhanced the cell-mediated cytotoxic activity of splenocytes. DNA microarray analysis identified 327 up-regulated and 347 down-regulated genes that characterised the KB290 diet group. The up-regulated genes were significantly enriched in Gene Ontology terms related to immunity, and, especially, a positive regulation of T-cell-mediated cytotoxicity existed among these terms. Almost all the genes included in the term encoded major histocompatibility complex (MHC) class I molecules involved in the presentation of antigen to CD8+ cytotoxic T cells. Marco and Signr1 specific to marginal zone macrophages (MZM), antigen-presenting cells, were also up-regulated. Flow cytometric analysis confirmed that the proportion of MZM was significantly increased by KB290 ingestion. Additionally, the over-represented Kyoto Encyclopedia of Genes and Genomes pathways among the up-regulated genes were those for natural killer (NK) cell-mediated cytotoxicity and antigen processing and presentation. The results for the selected genes associated with NK cells and CD8+ cytotoxic T cells were confirmed by quantitative RT-PCR. These results suggest that enhanced cytotoxic activity could be caused by the activation of NK cells and/or of CD8+ cytotoxic T cells stimulated via MHC class I presentation.