IDO1 and FOXP3: Possible Immune-Regulating Genes Alleviating Cedar Pollinosis Via L. Plantarum YIT 0132.

Japanese cedar pollinosis (JCPsis) is a common allergic disease which affects 30%–40% of the Japanese population.1 Although lactic acid bacteria (LAB) have been reported to alleviate JCPsis,2 the underlying mechanism remains unknown. We previously discovered that Lactiplantibacillus (Lactobacillus) plantarum YIT 0132 (LP0132) alleviated JCPsis symptoms and suppressed the reduction in regulatory T cells (Tregs) during pollen dispersion.3 Moreover, stratified analysis implicated that increased Treg levels alleviated symptoms via LP0132.3 Here, we further identified the immunoregulatory genes that correlated with the symptoms by LP0132 intake. The participants consumed LP0132 fermented citrus juice or placebo (unfermented citrus juice) for 8 weeks (125 mL/day). Peripheral blood collected from 36 background-matched participants in trial3 (Table S1) was used for microarray analysis; genes significantly fluctuating from enrolment were analysed using upstream and pathway analyses in the LP0132 group. Genes that correlated with changes in symptom scores were identified (Figure S1), and symptom scores were stratified by fold-change gene expression. Data from the placebo group were used to confirm the specificity of LP0132 consumption. Of the 33,327 probes (17,222 genes), 216 and 264 in the LP0132 and placebo groups, respectively, significantly increased by more than 1.25-fold or decreased by less than 0.80-fold (Figure S2). Thirty-three commonly altered genes in both the groups included eosinophil-associated genes—IL5RA and CLC (Table S2)—as described in the European cohorts.4 Pathway analysis in the LP0132 group identified the ‘bile secretion’ pathway (Table S3) and demonstrated a significant reduction of FXR (NR1H4), which reportedly induces Tregs (Figure S3)5; however, its expression did not correlate with any symptom scores. Upstream analysis predicted genes that significantly altered the downstream genes (Table 1A, Table S4). Of the genes identified, IDO1, which catalyses tryptophan to kynurenine, positively correlated with mean-Δ total ocular symptom score (TOSS) from weeks 1 to 4 only in the LP0132 group (Table S5), and participants with enhanced IDO1 expression had higher symptom scores, particularly in the LP0132 group (Figure 1A). In contrast, HSD3B1, involved in adrenal androgen production, negatively correlated with mean-ΔTOSS, and the subgroup with high HSD3B1 was associated with lower symptom scores (Figure 1B). However, these associations were also observed in the placebo group. An extensive search without cut-off value demonstrated the number of significantly altered genes was found to be 1648 and 3252 in the LP0132 and placebo groups, respectively. Among the identified genes (Table 1B), FOXP3, master regulator of Treg function, negatively correlated with mean-ΔTOSS only in the LP0132 group (Table S5). Moreover, the subgroup with high FOXP3 expression had lower symptom scores (Figure 1C) only in the LP0132 group. IDO1 in dendritic cells is known to promote Treg cell differentiation, while high serum IDO1 activity has been reported in patients with perennial allergic rhinitis.6 Our results are consistent with the latter, suggesting a differential role of IDO1 in lymphoid tissue and peripheral blood and that decreased IDO1 expression may be associated with responsiveness to LP0132. However, no data could explain why this association was observed in an LP0132-specific manner. In contrast, enhanced FOXP3 expression correlated with mild symptoms only in the LP0132 group. This supports our hypothesis that LAB, such as LP0132, alleviate symptoms of seasonal allergic rhinitis via Tregs. This study lacked a comparison of results in people without JCPsis and was unable to clarify the gene expression profile in the respective immune cells, including Tregs. Local changes in the intestinal and nasal mucosa remain unclear. In conclusion, the downregulation of IDO1 and upregulation of FOXP3 may be associated with alleviated JCPsis symptoms by LP0132 intake, which provides new insights into the immunological effects of LAB. NK, SS, NHM and KN designed and developed the protocol for this trial. YK, NK, SS, SK and KN conducted the experiment, analyzed the data, interpreted the findings, and drafted the manuscript. NHM supported the interpretation of the findings and reviewed the manuscript. This study was funded by the Yakult Honsha Co., Ltd. We thank all the participants of this trial. We are also grateful to the staff of Shimoshizu National Hospital for clinical support; T. Uetake and K. Fujiura of CX Wellness Co., Ltd. for their support with subject management and data collection; M. Miyagawa of Takumi Information Technology Corporation; K. Rokutan of Tokushima University; J. Nakayama of LSI Medience Corporation; S. Nakamura of Riken; C. Nonaka, K. Nakamori, and T. Igarashi of Yakult Honsha Co., Ltd.; and K. Miyazaki of Yakult Central Institute for their helpful advice. NK, SK and NHM are employees of Yakult Honsha Co. Ltd. The authors declare no conflicts of interest. Research data are not shared. Appendix S1 Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.