Tokyo University of Science
Elucidation that fatty acid metabolites produced by intestinal lactic acid bacteria exhibit anti-inflammatory effects and alleviate inflammatory bowel disease – Analysis of the efficacy of components derived from edible oil at the genetic, cellular, and individual levels –
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[Research summary and points]
Dendritic cells (*1), a type of immune cell, are activated in response to pathogen-derived components, but excessive activation is known to lead to inflammatory and autoimmune diseases.
We have revealed that γKetoC (*2), a fatty acid produced by intestinal lactic acid bacteria Lactobacillus plantarum when it metabolizes edible oil, suppresses the inflammatory response of dendritic cells. We found that γKetoC exhibits anti-inflammatory effects by activating G protein-coupled receptors (GPCRs, *3), which are fatty acid receptors, and NRF2 (*4), a master transcription factor for oxidative stress responses.
Using a mouse model, we demonstrated that oral intake of γKetoC alleviates the pathology of inflammatory bowel disease.
Further development of this research is expected to lead to the development of therapeutic drugs and preventive methods for
immune-related diseases.
[Image: [Research overview]
Laboratory members including Professor Chiharu Nishiyama of the Department of Biosystems Engineering, Faculty of Advanced Engineering, Tokyo University of Science, Professor Jun Ogawa and Associate Professor Shigenobu Kishino of the Graduate School of Agriculture, Kyoto University, Professor Masayuki Yamamoto, Director of the Tohoku Medical Megabank Organization, Tohoku University, and members of the Tokyo University of Science In joint research with Professor Manabu Ichihara of the Department of Pharmaceutical Sciences, Faculty of Pharmaceutical Sciences, University, we investigated the effects of metabolites of intestinal bacteria on immune responses. We found that γKetoC exhibits the best anti-inflammatory activity compared to metabolites. Mechanistic analysis suggests that activation of GPCR and NRF2 suppresses the production of inflammatory cytokines (*5), and oral intake of γKetoC alleviates the pathology of inflammatory bowel disease model mice. In fact, it was revealed that the effect of γKetoC was not observed in NRF2-deficient mice.
It is known that polyunsaturated fatty acids undergo metabolic conversion such as hydroxylation and saturation by enzymes possessed by intestinal lactic acid bacteria, such as Lactobacillus plantarum, and are converted into other fatty acids. In recent years, it has been discovered that substances metabolized by these intestinal bacteria exhibit various beneficial physiological functions for the host (anti-inflammatory and antioxidant effects, functions to protect the intestinal epithelial barrier, etc.) . However, how these metabolites affect the immune response remains unknown. Therefore, this research group prepared immune cells from the spleen and bone marrow of mice and analyzed the correlation between multiple fatty acid metabolites and immune cell activity from various angles, including the genetic, cellular, and individual levels.
Research results revealed that enone fatty acids inhibit T cell proliferation and dendritic cell activation, and that γKetoC exhibits the most excellent anti-inflammatory effect. Since γ-linolenic acid, the starting material for γKetoC, did not exhibit these properties, it can be said that it was converted into a substance with beneficial functions by the action of intestinal bacteria. Additionally, experiments using GPCR agonists and G protein inhibitors suggested that Gq-type GPCRs are involved in part of the effects of γKetoC on dendritic cells. Furthermore, γKetoC has the effect of activating the NRF2 pathway in dendritic cells, and the anti-inflammatory effect of γKetoC was attenuated in NRF2-deficient dendritic cells. In an experiment in which γKetoC was orally administered to colitis model mice, it was found that γKetoC had the effect of improving
pathological conditions, and that γKetoC had no effect in
NRF2-deficient mice.
These results demonstrate that γKetoC exhibits an anti-inflammatory effect against excessive immune responses, and that the stimulation of GPCR and NRF2 is involved in this mechanism. The results of this research are useful findings for understanding the immune response mechanism, and include proposals for realistic and specific treatment strategies such as more effective intake methods of active
ingredients, and are useful for the treatment and prevention of immune-related diseases. You are expected to contribute.
In addition, γKetoC, which showed high anti-inflammatory activity in this study, has structural characteristics that are different from ω3 systems such as docosahexaenoic acid (DHA) and icosapentaenoic acid (IPA), and its effects on physiological activity and health are still unknown. It is also worth noting that it was an oil component that had not been sufficiently investigated. In the future, by conducting research targeting various types of oil components, we will discover and develop components other than γKetoC that are effective in preventing and alleviating immune-related diseases, as well as intestinal bacteria involved in their metabolism. It is expected that this will lead to
The results of this research were published online in the
international academic journal “Frontiers in Immunology” on April 30, 2024.
[Research background]
In the intestine, various secondary metabolites are produced by the action of bacteria using substances derived from food components as substrates. Recent research has shown that fatty acids in food are converted into derivatives such as hydroxy fatty acids, oxo fatty acids, enone fatty acids, and saturated fatty acids through the catalytic action of enzymes from intestinal lactic acid bacteria L. plantarum, and these metabolic intermediates have metabolic improving effects. It has been found that they have positive effects on the health of the host. However, the effects of these fatty acid metabolites on immune responses remained unknown.
This research group has been conducting research on the effects of food components and intestinal bacterial metabolites on immune response regulation. In January 2024, it was revealed that short-chain fatty acids produced from dietary fiber suppress allergic diseases via mast cells (*1). Now, by making full use of analysis at the
individual, cellular, and genetic levels, we are conducting research with the aim of elucidating how the oil components metabolized by the action of intestinal bacteria affect the immune response. I did. *1: Tokyo University of Science Press Release February 1, 2024 “Elucidating the mechanism of action of short-chain fatty acids to suppress allergies – Demonstrating the effectiveness of dietary fiber against allergies at the molecular level”
URL: https://www.tus.ac.jp/today/archive/20240201_2581.html
[Details of research results]
Hydroxy fatty acids, oxo fatty acids, and enone fatty acids were prepared from polyunsaturated fatty acids (*6) using a convertase derived from the intestinal lactic acid bacterium L. plantarum (synthesized by Associate Professor Kishino and Professor Ogawa), and each fatty acid was We investigated whether it affects the immune response.
When examined using immune cells prepared from mouse spleen, hydroxy fatty acids did not have any effect on immune activity, but treatment with enone fatty acids (KetoC, αKetoC, γKetoC) inhibited antigen stimulation-induced cytokine secretion. was found to decrease significantly. In addition, cytokine secretion was not suppressed by linoleic acid, α-linolenic acid, and γ-linolenic acid, which are the starting materials for these metabolisms, suggesting that fatty acids acquired new functions through metabolic conversion. . Furthermore, as a result of further analysis using various isolated immune cells, enone fatty acids are associated with inflammatory reactions caused by various bacterial body components such as LPS (lipopolysaccharoid), and especially inflammatory cytokines from dendritic cells. It was found to strongly suppress secretion.
Next, we used γKetoC, which showed the strongest activity, to investigate the mechanism by which the anti-inflammatory effect is exerted. First, in order to verify the involvement of GPCRs known as long-chain fatty acid receptors, we treated dendritic cells with GW9508, an agonist for Gq-type GPCRs, and found that the production of inflammatory cytokines was suppressed in a dose-dependent manner. I understand. Since GPR120, one of the GW9508-reactive GPCRs, is expressed in dendritic cells, γKetoC was expected to suppress LPS-induced dendritic cell activation via GPR120. Therefore, when we added an inhibitor of Gq protein a subunit when treating dendritic cells with γKetoC, we found that the anti-inflammatory effect of γKetoC was reduced by some cytokines (TNF-a). Although partially involved, the results also suggest the existence of other points of action.
Next, we conducted an experiment to confirm whether γKetoC induces antioxidant effects through activation of NRF2 in dendritic cells, and found that in γKetoC-treated dendritic cells, NRF2 protein levels and NRF2 target gene An increase in mRNA levels was confirmed. Therefore, in order to analyze the possibility that NRF2 is involved in the effect of γKetoC, we conducted an analysis using NRF2-deficient mice (established by Professor Yamamoto). As a result, we found that the suppressive effect of γKetoC on IL-6 and IL-12p40 production among LPS-induced inflammatory cytokine secretions was weakened in dendritic cells from NRF2-deficient mice. These results suggest that γKetoC negatively regulates LPS-induced IL-6 and IL-12p40 production by stimulating the NRF2 pathway.
Using dextran sodium sulfate (DSS)-induced colitis model mice, we verified the protective effect of γKetoC on in vivo inflammatory responses. Oral administration of γKetoC to wild-type mice suppressed the increase in disease activity index (DAI) scores and significantly reduced colon atrophy due to fibrosis. Furthermore, extending the administration period of γKetoC also significantly alleviated body weight loss, reduced colon tissue damage and inflammatory cell infiltration, and increased serum levels of TNF-α, IL-6, and IL-12p40. A tendency to be suppressed was also confirmed. On the other hand, in NRF2-deficient mice, administration of γKetoC did not improve the pathology. These results suggest that oral administration of γKetoC alleviates enteritis via the NRF2 pathway.
This series of experiments suggested that γKetoC, which is produced by intestinal lactic acid bacteria’s metabolism of γ-linolenic acid, is converted into a new compound in the intestinal tract, which exerts anti-inflammatory effects. Gamma-linolenic acid has been reported to be effective against autoimmune diseases and allergies, but the present results suggest that rather than gamma-linolenic acid itself, its metabolite, gammaKetoC, may hold the key. It also presents. It is expected that future research progress will reveal more effective ways to ingest active ingredients.
Commenting on the results of this research, Professor Nishiyama said, “I hope it will be widely recognized that intestinal bacteria produce substances that are useful for our health.I also hope that it will be widely recognized that intestinal bacteria produce substances that are useful for our health. We will conduct detailed analyzes at the individual, cellular, and genetic levels to prove that this has a significant impact on immune-related diseases. I would like to contribute to the development of this technology.”
*This research was supported by the Japan Society for the Promotion of Science Grant-in-Aid for Scientific Research (B) (23H02167, 20H02939), Special Researcher (DC2), Special Researcher Grant (21J12113), Japanese Society of Immunology “Kibo” Project, Tokyo Science This work was supported by the University President’s Research Promotion Grant, the Tojuro Iijima Memorial Foundation for the Promotion of Food Science, the Kaiun Mishima Memorial Foundation Academic Research Grant, and the Takeda Science Foundation Life Science Research Grant. 【term】
*1 Dendritic cells
They are white blood cells derived from bone marrow cells and have dendrites. After recognizing foreign substances from the outside and taking them in, they transmit information to lymphocytes such as T cells. When transmitting information between cells, they produce proteins called cytokines.
*2 γKetoC
10-oxo-cis-6,trans-11-octadecadienoic acid. A type of enone fatty acid obtained from γ-linolenic acid through bacterial metabolism. *3 G protein-coupled receptor (GPCR)
A protein located on the cell membrane that senses external stimuli (neurotransmitters, hormones, etc.) and initiates specific signal transduction pathways within the cell. It is also a very important target for pharmaceuticals, and many drugs function through their actions on GPCRs.
*4 NRF2
A master transcription factor that induces the expression of defense genes in vivo. Activated by oxidative stress. Its activity is regulated by the stress sensor Keap1 protein.
*5 Cytokine
A protein secreted during information transmission between cells. There are different types such as interleukins and chemokines. Inflammatory cytokines control inflammatory responses by acting on cells that trigger defense responses.
*6 Polyunsaturated fatty acids
An unsaturated fatty acid with two or more double bonds. It contains many substances known to have beneficial effects on the body, such as alpha-linolenic acid, DHA (docosahexaenoic acid), and IPA
(icosapentaenoic acid).
[Paper information]
Magazine name: Frontiers in Immunology
Paper title: The gut lactic acid bacteria metabolite, 10-oxo-cis-6, trans-11-octadecadienoic acid, suppresses inflammatory bowel disease in mice by modulating the NRF2 pathway and GPCR-signaling
Author: Miki Ando, Kazuki Nagata, Ryuki Takeshita, Naoto Ito, Sakura Noguchi, Natsuki Minamikawa, Naoki Kodama, Asuka Yamamoto, Takuya Yashiro, Masakazu Hachisu, Gaku Ichihara, Shigenobu Kishino, Masayuki Yamamoto, Jun Ogawa, and Chiharu Nishiyama
DOI: 10.3389/fimmu.2024.1374425
URL: https://doi.org/10.3389/fimmu.2024.1374425
【Presenter】
Miki Ando* Tokyo University of Science, Faculty of Advanced
Engineering, Nishiyama Laboratory, Master’s degree completed (Master’s degree completed in 2022)
Kazuki Nagata* Assistant Professor, Nishiyama Laboratory, Faculty of Advanced Engineering, Tokyo University of Science
Ryusei Takeshita* Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Undergraduate graduate (Graduated in 2023)
Naoto Ito Tokyo University of Science, Faculty of Advanced
Engineering, Nishiyama Laboratory, 3rd year doctoral student Ohka Noguchi, Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, 2nd year master’s student Natsumi Minamikawa Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Master’s degree completed (Master’s degree completed in 2023)
Naoki Kodama Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Master’s degree completed (Master’s degree completed in 2021)
Aihi Yamamoto Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Undergraduate Graduate (Graduated in 2017)
Takuya Yashiro, Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Lecturer (at the time)
Masakazu Yasu, Tokyo University of Science, Faculty of Advanced Engineering, Nishiyama Laboratory, Lecturer (at the time)
Manabu Ichihara, Professor, Faculty of Pharmaceutical Sciences, Tokyo University of Science
Shigenobu Kishino Associate Professor, Graduate School of Agriculture, Kyoto University
Masayuki Yamamoto Professor, Tohoku University, Director, Tohoku Medical Megabank Organization
Jun Ogawa Professor, Graduate School of Agriculture, Kyoto University Chiharu Nishiyama Professor, Faculty of Advanced Engineering, Tokyo University of Science
*First author (Ando, Nagata, and Takeshita are the same)