Orally administered heat-killed Lactobacillus paracasei MCC1849 enhances antigen-specific IgA secretion and induces follicular helper T cells in mice

Abstract

Antigen-specific immunoglobulin (Ig) A plays a major role in host defense against infections in gut mucosal tissue. Follicular helper T (Tfh) cells are located in germinal centers and promote IgA production via interactions with germinal center B cells. Several studies have demonstrated that some lactic acid bacteria (LAB) strains activate the host's acquired immune system, inducing IgA secretion in the intestine. However, the precise molecular mechanisms underlying the effects of LAB on IgA production and Tfh cells are not fully resolved. Lactobacillus paracasei MCC1849 is a probiotic strain isolated from the intestine of a healthy adult. In this study, we investigated the effects of orally administered heat-killed MCC1849 on IgA production in the intestine and on Tfh cell induction in vivo. We found that orally administered MCC1849 induced antigen-specific IgA production in the small intestine, serum and lungs. We also observed that MCC1849 increased the proportion of IgA+ B cells and Tfh cells in Peyer's patches (PPs). In addition, MCC1849 increased the gene expression of IL-12p40, IL-10, IL-21, STAT4 and Bcl-6 associated with Tfh cell differentiation. These results suggest that orally administered MCC1849 enhances antigen-specific IgA production and likely affects Tfh cell differentiation in PPs.

Fig 1. Effects of Lactobacillus paracasei MCC1849 on IL-12 production.

Murine splenocytes were cultured with various heat-killed Lactobacillus strains (10 μg/ml) for 2 days. Data shown are the mean ± SD of the levels of IL-12p70 which are the representative of three independent experiments.

Fig 2. Effects of Lactobacillus paracasei MCC1849 on total IgA production in mice.

(A) Mice were treated with or without MCC1849 for 5 weeks. Total IgA concentrations in homogenized small intestine and serum samples were determined via ELISA. (B) The proportions of IgA⁺ B220⁺ cells and IgA⁺ plasmablasts (IgA⁺ B220^- cells) in PPs were analyzed via FCM. (C) Gene expression related to the differentiation of IgA⁺ cells was measured via real-time RT-PCR analysis. The level of gene expression was normalized to that of GAPDH mRNA expression in the control group. Data are shown as the mean ± SD. N = 16. *p<0.05, **p<0.01 compared to the control.

Fig 3. Effects of Lactobacillus paracasei MCC1849 on total IgA and OVA-specific IgA production in OVA-immunized mice.

(A, B) Mice were treated with or without MCC1849 for 5 weeks. All mice were orally immunized on days 14, 21, and 28 with OVA and cholera toxin. On day 35, mice were euthanized and dissected. Data show the total IgA and OVA-specific IgA concentrations in homogenized small intestine, small-intestine lavage fluid, homogenized colon, colon contents, serum and lung samples. AU: arbitrary unit. Data are shown as the mean ± SD. N = 14. Data are representative three independent experiments. *p<0.05, **p<0.01 compared to the control.

Fig 4. Effects of Lactobacillus paracasei MCC1849 on the population of IgA-related cells in the intestinal tissues of OVA-immunized mice.

(A, B and C) The proportion of IgA⁺ B220⁺ cells and IgA⁺ plasmablasts (IgA⁺ B220^- cells) in PP, MLN and LP cell samples were analyzed via FCM. Data are shown as the mean ± SD. N = 14. Data are representative two independent experiments. *p<0.05, **p<0.01 compared to the control.

Fig 5. Effects of Lactobacillus paracasei MCC1849 on the population of T cells and gene expression in PP cells in OVA-immunized mice.

(A) The proportion of Tfh cells (CD4⁺ CXCR5⁺ PD-1^high cells) and Th1 cells (CD4⁺ T-bet⁺ cells) in PPs were analyzed via FCM. (B) Gene expression related to T cell differentiation was measured via real-time RT-PCR analysis. The level of gene expression was normalized to that of GAPDH mRNA expression in the control group. Data are shown as the mean ± SD. n = 14. Data are representative two independent experiments. *p<0.05, **p<0.01 compared to the control.