ImmunoTox Letter

December 2018

Report on the 25th Annual Meeting of the Japanese Society of Immunotoxicology(JSIT2018)

Keiko NOHARA
President of the 25th JSIT Annual Meeting
National Institute for Environmental Studies
The 25th Annual Meeting of the Japanese Society of Immunotoxicology (JSIT) was held on
September 18th and 19th, 2018 at International Congress Center in Tsukuba.

As the main themeof this meeting “Discussing and deepening our insight into the interaction between the immune
system and the environment” calls for, enthusiastic discussions were carried among participants
on a variety of immune reactions themselves and in relation to environments inside and outsidebodies.
The three topics featured in this meeting were “gut microbiota”, “cancer immunotherapy” and“epigenetics”. The first two topics were desired by many members and epigenetics was one I
proposed.

I appreciate all the outstanding speakers who kindly accepted our request taking timeout of their busy schedule. All the sessions and talks were very impressive and instructive.
On the first day in Educational Lecture, Dr Akihiko Yoshimura (Keio University School ofMedicine) gave a talk titled “Immune modulation by epigenetic modification of regulatory Tregs”.
In the Symposium session “Gut microbiota and immune diseases - A new perspective on
immunotoxicology”, four talks were given: “Association between gut microbiota andenvironmental chemicals” by Dr Rie Yanagisawa (National Institute for Environmental Studies),“Relation between gut microbiome and allergic diseases in childhood: clinical perspective” by Dr
Naoki Shimojo (Graduate School of Medicine, Chiba University), “Involvement of epithelial
indigenous flora in allergy and inflammation” by Dr Akira Shibuya (TARA, Tsukuba University)and “Microbiota in immune disorders” by Dr Kiyoshi Takeda (Immunology Frontier ResearchCenter, Osaka University).
On the second day, Special lecture was given by Dr B Page Lawrence (University of Rochester
School of Medicine and Dentistry, USA). She was a representative of the researcher exchange
program between the ITSS, SOT and JSIT. Her talk “Developmental exposure alters cellular
processes critical for T cell functions, and affects some T cell properties across generations” wasvery new for many audience and exciting.
We were also provided and enjoyed video messages from ITSS executives, President Dr Jamie
Dewitt (East Carolina University), Postdoctoral representative Dr Alessandro Venosa (UniversityNo.2（17）42

ImmunoTox Letterof Pennsylvania), and Student representative Dr Alexa Murray (The State University of NewJersey).
In the workshop “Development of cancer immunotherapy and safety assessment of immune
checkpoint inhibitors”, three speakers gave talks: Dr Hiroshi Shiku (Department of Immuno-Gene
Therapy/Personalized Cancer Immunotherapy, Mie University) on “Guidances for Developmentof Cancer Immunotherapy”, Dr Kazuhiko Taguchi (Bristol-Myers Squibb K.K.) on “Nonclinicalassessment of CTLA-4 and PD-1 inhibitors for predicting adverse events in clinical studies”, andDr Minoru Satoh (Department of Clinical Nursing, University of Occupational andEnvironmental Health) on “Autoantibodies as biomarkers for predicting risk to develop
autoimmune diseases following treatment with immune checkpoint inhibitors”. After their talks,
the issues were further discussed as a panel discussion.
As a new program, we had International session, in which Dr Khaled Hossain (University ofRajshahi, Bangladesh ） and Dr Myint Myint Nyein (University of Medicine 1, Myanmar)presented human data obtained in their countries.
On the second day, we had Award ceremony to honor Dr Reiko Teshima (Okayama Universityof Science) for the JSIT Research Award and Dr Kiyoshi Kushima (Astellas Pharma Inc, Astellas
Research Institute of America) for the prize of Encouragement 2018. Dr Teshima gave award
lecture titled “Immunotoxicological evaluation of food allergens”.

Dr Kushima talked on“Promotion of Developmental ImmunoToxicology (DIT) assessment and Adverse Outcome
Pathway (AOP)”.
On the night after the first day of meeting, we
had a social gathering.
Young executives of JSIT performed a dance
to a famous J-pop number “Choo ChooTRAIN”, which was a fun surprise.
The meeting gathered more than 130
participants.

I would like to sincerely

appreciate all who supported our meeting.
Thank you very much.

No.2（18）43

ImmunoTox Letter

The Best Presentation Award

IL-17-induced mRNA stabilization dictates the expression level of IκB-ζ inkeratinocytes
Ryuta Muromoto
Department of Immunology,
Faculty of Pharmaceutical Sciences, Hokkaido University
Interleukin-17A (IL-17) is an immune cell-derived cytokine that acts on various types of cells,
including epidermal keratinocytes, and induces production of antimicrobial peptides andchemokines to induce antibacterial and antifungal defense responses. An excess of this IL-17
action leads to inflammatory skin disease such as psoriasis. Recently, we demonstrated that IκBζ, a nuclear IκB protein that modulates NF-κB-dependent transcription, is IL-17-inducible andmediates IL-17-induced responses. However, the mechanism controlling IκB-ζ expression in IL17-stimulated cells remained unclear. It has been reported that the 3'-untranslated region (3'UTR)
downstream of the protein coding sequence of IκB-ζ mRNA has an influence on the stability ofIκB-ζ mRNA. In this study, we investigated whether and how the posttranscriptional regulationof mRNA stability is involved in the IL-17-mediated induction of IκB-ζ. A reporter plasmid(LUC-IκB-ζ 3'UTR), in which a 3'UTR sequence derived from murine IκB-ζ mRNA was ligateddownstream of a luciferase, was used to examine the effects of IL-17 stimulation. The IL-17
stimulation showed a significant enhancement of the activity of 3’UTR-regulated reporter. On theother hand, IL-17 could not activate the activity of a luciferase reporter plasmid that is expressed
under the control of the promoter region upstream of the transcription start site of the IκB-ζ gene.
These data suggested that IL-17 enhances the stability of the IκB-ζ mRNA post-transcriptionally.
In addition, our flow cytometric assay using fluorescent protein (Venus)-based reporter fused tothe truncated IκB-ζ 3’UTR (1 - 150 nt from stop codon) showed that this portion of IκB-ζ 3’UTR
can respond sufficiently to IL-17 stimulation. This portion has two characteristic stem-loop
structures that are conserved between species and required for the recognition and degradation bya ribonuclease, Regnase-1/MCPIP1 (encoded by ZC3H12A). We mutated the two stem-loops inVenus-IκB-ζ 3’UTR reporter to disrupt these secondary structures and found that the loop-mutant
reporter shows enhanced expression than the control intact reporter under unstimulated condition
while it does not respond to IL-17. Knockdown of endogenous Regnase-1 in HaCaT cells by
siRNA resulted in an accumulation of basal levels of IκB-ζ mRNA. These results suggest thatRegnase-1-mediated degradation of IκB-ζ mRNA occurs constitutively. An IL-17 signaling
adaptor protein ACT1 has been shown to mediate IL-17-induced upregulation of IκB-ζ inNo.2（19）44

ImmunoTox Letter

keratinocytes, although it has not yet been examined whether the IκB-ζ 3’UTR-mediated mRNAstabilization is affected by ACT1 in IL-17-treated cells. We observed that IL-17-induced
expression of Venus-IκB-ζ-3’UTR reporter was abrogated by ACT1 knockdown. In addition,overexpression of ACT1 counteracted the suppressing effect of Regnase-1 on the expression ofVenus-IκB-ζ 3’UTR reporter. These results suggest that IL-17/ACT1 signaling counteracts theconstitutively occurring Regnase-1-mediated degradation of IκB-ζ mRNA. These findings would
improve our understanding of the molecular mechanisms of IL-17 action in immunity and chronic
inflammation.

No.2（20）45

ImmunoTox Letter

The Student and Young Scientists Award

nSP50-inducible hepatic damages would worsen via acquired immune system
Shun-ichi Eto1,21
Laboratory of Toxicology and Safety Science,Grad. Sch. Pharm. Sci., Osaka Univ.
2Interdisciprinary Program for Biomedical Sciences, Osaka Univ.
It’s my honor to win the best presentation award among the young scientists at the 25th Annual
Meeting of the Japanese Society of Immunotoxicology. I would like to appreciate the selection
committee and the researchers, who gave me lots of advices, from the bottom of my heart. Now,
I’ll introduce the awarded research topic.
In recent years, the use of nanoparticles has been expanding in various industries such as
medicine, foods, and cosmetics because of their innovative functions. On the other hand, theirunexpected biological effects resulted from the refinement of the particle size have been
concerned. From this viewpoint, safety analysis of nanoparticles has been studied intensively.
And we revealed that silver nanoparticles could trigger metal allergy-like symptoms bysensitization of themselves, which is not induced by silver ion sensitization (T. Hirai et al., Nat.
Nanotechnol., 2016). Given that the hazard induced by multiple exposures may be different
from one by single exposure, hazard analysis focused on the multiple exposures-induced
acquired immune systems is required. Thus, we attempted to assess the effect of amorphoussilica nanoparticles (nSPs) on acquired immune responses. As a result, we showed that
pretreatments of nSPs could worsen nSPs-inducible hepatic damage though there were nodifferences in immunodeficient mice. Moreover, this exacerbation disappeared by depletion ofT cells. These results suggest that the pretreatment of nSPs would worsen nSPs-inducible
hepatic damages through the acquired immune system, especially the cell-mediated immunity.
Considering that we are exposed to nanoparticles many times on a daily basis, this exacerbationof acute toxicity has crucial meaning, that is, pre-exposure to nanoparticles has potential totrigger the transition of NOAEL (No Observable Adverse Effect Level) to lower doses. Now,
we try to elucidate the mechanism of this exacerbation for developing optimal design ofnanoparticles. We consider that our researches would contribute not only to the sustainable useof nanoparticles but also to the advancement of immunotoxicology.

No.2（21）46

ImmunoTox Letter

The Student and Young Scientists Award

Facilitated antigen sensitization on the skin by triacylglycerol in an FITC-induced
contact hypersensitivity mouse model.
〇Masato Tsutsumi1, Kota Sekiguchi1, Erina Ogawa1,Kohta Kurohane1, Yasuyuki Imai1
1Lab. Microbiology and Immunology,
School of Pharmaceutical Sciences, University of Shizuoka
Aim: Triacyl glycerol (TAG) is glycerol ester with fatty acids. TAG is used in cosmetics and we havechances to contact with it on the skin. However, effects of TAG on cutaneous immunity are not wellestablished. We investigated the effects of TAG on skin sensitization in an FITC-induced contacthypersensitivity mouse model. This model has been employed to reveal adjuvant effect of chemicals,such as dibutyl phthalate (DBP).
Materials and Methods: BALB/c mice were sensitized with FITC in acetone in the presence orabsence of a test compound on days 0 and 7. On day 14, mice were challenged with FITC on an ear
auricle to assess ear-swelling responses. Migration of FITC positive dendritic cells (FITC+DC) todraining lymph nodes was analyzed by flow cytometer after staining with phycoerythrin (PE)conjugated anti-CD11c.
Results and Discussion: TAG with short (carbon number C4 and C6) and medium (C8 and C10)
chain fatty acids enhanced skin sensitization to FITC. These TAGs promoted FITC+DC migration
to draining lymph node. Short chain TAGs appeared to exhibit stronger effects than the medium
chain ones. In contrast, triolein did not enhance skin sensitization. Triolein suppressed the enhancing
effect of tributyrin on FITC sensitization. These results suggest that TAG with different types offatty acids causes different effects on the skin immunity.

No.2（22）47

ImmunoTox LetterImmunotoxicological Research

CpG-ODN contributes to Flucloxacillin-induced acute liver injury
through FasL/Fas mediated pathway
Yuying Gao
Laboratory of Biopharmaceutics,
Graduate School of Pharmaceutical Sciences, Chiba University

Idiosyncratic drug induced liver injury (IDILI) is often the major cause for drug withdraw which
has been reported mostly immune-mediated. Despite of the rarity, IDILI sometimes would
induce serious outcomes, even liver injury requiring transplantation or death. However, thepathogenesis of IDILI has not been clearly explained so far, which partially because of the lackof appropriate animal models. Thus, developing valid animal models is essential for better
understanding the onset mechanisms.
Flucloxacillin (FLUX) is a Beta-lactam antibiotic of penicillin class which could induce rarely
liver injury. Patients may develop liver injury on days 1 to 45 within FLUX treatment. So we
tried to co-administrate FLUX and CpG-ODN (toll-like receptor 9 agonist) to C57BL/6J mice
to reproduce early phase of FLUX-induced liver injury in animals, and investigate its
pathogenesis in liver. Then we detected that co-administration of CpG-ODN with FLUX
significantly elevated plasma ALT level in mice, while treatment of FLUX or CpG-ODN alone
was insufficient to induce an ALT increase. Moreover, the co-administration also increased
expression of Fas ligand on NKT cells and Fas on hepatocytes. When applying FasL-mutated
gld/gld mice, no obvious hepatotoxicity was observed despite the co-administration of CpGODN with FLUX. Above, we developed an animal model of FLUX-induced acute liver injury
by co-treatment of CpG-ODN. These results suggest that CpG-ODN accelerates FLUX-induced
liver injury through FasL/Fas-mediated apoptosis pathway between NKT cells and hepatocytes,
which plays an important role in this animal model.

No.2（23）48

ImmunoTox Letter

Real Voices of International Immunotoxicologists

What the other researchers think about? How can I do more exciting research in the future?
Most of the young Japanese immunotoxicologists may hope to listen to some kind of commentsfrom the experts in this field, in particular from the outside of Japan. This time, we interviewed
Dr. B. Paige Lawrence from University of Rochester, USA. Let’s go to listen to her voice. What
will you feel and learn?

B. Paige Lawrence, Ph.D.
Chair, Department of Environmental Medicine
Director, Environmental Health Science Center
Professor of Environmental Medicine and of
Microbiology & Immunology School of Medicine &Dentistry University of Rochester
Rochester, NY 14642 U.S.A.

Q1. What was the most impressive event for you in your trip to Japan this time?
The opportunity to travel, experience new places, and meet new people is one of the many
wonderful aspects of being a scientist. My entire visit to Japan was very pleasant and memorable.
One thing that particularly stands out is the kindness of strangers. Shortly after I arrived, Dr. KeikoNohara’s daughter, Asuna and her husband Yasunobu, whom I had never met, took me on awonderful walking tour to see little bit of Tokyo, including the Asakusa Kannon (Sensoji) Temple.
They also took me out for a wonderful dinner and I was able to try okonomiyaki (which isdelicious!). Also, before I departed for the airport, Dr. Tomoki Fukuyama took me to an ancient
Shinto shrine on Mount Tsukuba, and for wonderful lunch at a restaurant nearby. This was arelaxing and very special way to end my trip to Japan, and I am grateful for this excursion.
Strangers no more, through their generosity they helped make the world a smaller place.

No.2（24）49

ImmunoTox Letter

Another impressive aspect of my visit to Japan was the conference reception and banquet. This
was a special meeting for the JSIT, as they were celebrating their 25th year, which is a major
milestone. There was a special banquet and award ceremony at the Hotel Grand Shinonome inTsukuba. Along with celebrating the numerous achievements of JSIT members and welcoming
new members, several young investigators received awards. The evening was also filled withlively and warm conversation, but what really impressed me was that throughout the evening
several members of the JSIT displayed remarkable talents including dancing, theatrical readings,and musical performances. These were among the highlights, not only because art transcends
language, but because the level of encouragement, sharing, and fun that these performances added
to the evening was entirely special. The overall ambiance at the reception radiated cheerfulness
and camaraderie. It was a wonderful reminder that, no matter how stressful we find our jobs, there
is much also happiness, and it is important to take the time to share, to celebrate, and to laughtogether.

No.2（25）50

ImmunoTox Letter

Q2. What is the most exciting thing in your career to date.
Scientific discovery and figuring something out are very rewarding. However, the most gratifying
and exciting thing in my career is the success of the students, postdocs, and the many other typesof learners who have spent time in my lab. Mentoring is a privilege and a joy. Mentoring helps
me remember to pause and reflect, be a more creative scientist, and a better communicator. It also
keeps me invigorated as a scientist, as many ideas and perspectives are always better than one. I
believe that it is safe to say I have gained more from the people whom I have had the opportunity
to mentor than they have learned from me.
Q3. What are the things you are doing energetically, right now?
I am fortunate to have a fantastic team in my lab, and to work with wonderful collaborators. Oneof our main undertakings is to understand how signaling through the aryl hydrocarbon receptor(AHR) influences the development and function of the immune system. Much of the field isfocusing on teasing out how the AHR regulates the function of the fully mature immune system,
and we are interested in this as well. However, we have also extended research in some newdirections, such as illuminating how AHR activation during early life changes the function of theoffspring’s immune system, and influences subsequent generations. By integrating genetic,
pharmacological, immunological, and bioinformatic approaches, we are defining specific changes
to different immune cell types, and uncovering the causal mechanisms by which they are affected.
We also bidirectionally integrate research using model organisms and human populations,discovering associations between early life and longitudinal exposures to persistent organic
pollutants and changes in immune function in infants, young children, and adolescents.
Q4. What is required for breakthrough in immunotoxicology research in the future, do you
think?
I think that the future of immunotoxicology research is very exciting. Whether one’s passion istherapeutic agents, pollutants, or nutritional factors, the field is poised to start tackling how coexposures influence the immune system’s function and contribute to disease. This will lead tohuge breakthroughs because, to date, pharmacology and toxicology research and applications of
this research have generally focused on single exposures. This is a very important approach, andshould continue. Yet by expanding the complexity of experimental model systems, we will attaina new level of understanding of how exogenous factors influence—for better and for worse—theimmune system. The implications for improving health are tremendous, as we will be better able
to predict and reduce risks, and also to improve or target new therapies.

No.2（26）51