WPI Immunology Frontier Research Center 2023- 2024

Osaka University

Immunology
Frontier
ResearchCenterAnnual Reportof IFReC
2023 - 2024

Edit : Research Planning & Management Office, IFReC
Published in June, 2024

Osaka University

Copyright : Immunology Frontier Research Center (IFReC ), Osaka University

CO NTENTS

Message from the Director
As the Director of the Immunology Frontier Researchorganized the second “International School on Advanced

Center (WPI-IFReC) at Osaka University, I am very pleased

Immunology” in FY2023 leading to discussions about

to present the IFReC annual report for fiscal year 2023.

future collaborations. Additionally, IFReC co-organizedSince joining the WPI Academy in 2017, we at IFReC havetwoLaboratories ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・9pioneered a unique academic-industry partnership that

Symposium" in Finland and the "IFReC-Doherty Instituteunleashes new possibilities in collaborative research.

and Partners Immunology Symposium" in Osaka. Moreover,・ Immunology Groups ・
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IFReC goes beyond basic research and social engagement,

we successfully hosted "The International Symposium on

and aims to significantly contribute to our university's

Microbiology and Immunology," our 13th international

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educational system by developing graduate programs

symposium. These events brought together international

specifically for international students specializing inresearchers and facilitated high-level discussions on the・ Bioinformatics Groups ・
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immunology, with details to be announced in 2024.

frontiers in immunology.

Message from the Director ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・1
Organization ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・3

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Events & Outreach Activities ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・77
Information ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・89
Access Map ・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・・104

symposia:the"Finnish-Japanese

Immunology

After the COVID-19 pandemic began to subside in late

IFReC plays a central role in immunology and infectious

2022, Japan officially downgraded COVID-19 in May 2023

disease research, fostering collaborations among various

to the same category that includes seasonal influenza.

research departments and institutes such as the ResearchThis led to the relaxation or complete removal of various

Institute for Microbial Diseases (RIMD), the Center for

restrictive measures, allowing many regions to resume

Infectious Diseases Education and Research (CiDER) and thetheir pre-COVID-19 activities. Universities, for example,

Center for Advanced Modalities and DDS (CAMaD) at Osaka

have successfully transitioned back to in-person classes

University. We are committed in our efforts to continue

and events.

basic research in immunology and to seek ways to makemeaningful contributions to society, and through research

Building on the success of the inaugural event, IFReC
2

and ImmunoSensation (University of Bonn) again co-and education, we will drive the advancement of scienceand shape the future of immunology research worldwide.

Kiyoshi TAKEDA, MD/ PhD
Director
WPI Immunology Frontier Research Center
Osaka University

1Organization

Organization Chart
MEXT

Osaka University

WPI Initiative Program Committee

International Advanced
Research Institute (IARI)

JSPS

As of March 2024

Joint Research Chairs with the Pharmaceutical Companies
International
Scientific
Advisory
Board

Research
GroupsImmunology Groups• Innovative Drug Discovery in Immunology (Chugai Pharma)• Immune-Therapeutic Drug Discovery (Otsuka Pharma)• Innate Immunity• Innovative Drug Discovery in Host Defense

Imaging Groups
Informatics Groups
Core Instrumentation Facility

Unit for Combined Research Fields

Common Facilities

Animal Resource Center for Infectious Diseases
Network Administration OfficeDirector

Management
Committee

Board of Representatives

Administrative Office

DeputyDirectors

Administrative
Director

General Affairs Section
Administrative
Department• General Affairs Section• Accounting Section• Employment and acceptance procedures for researchers and staff• Social insurance (for part-time staff ), employment insurance• Various research support (MTA, patents, animal experiments, safety etc.)• Support for international researchers and students• IFReC Kishimoto Foundation Fellowship Program

Accounting Section
Research Planning and Management Office

Cooperative Institutions• Institute for Frontier Life and Medical Sciences, Kyoto University, Japan• RIKEN Center for Integrative Medical Sciences, Japan• University College London, UK• ImmunoSensation2 , Cluster of Excellence, the Rheinische Friedrich-Wilhelms-University of Bonn, Germany• Budget drafting / implementation / management• Acceptance and implementation of third-party funding• Purchasing and payment of payroll, travel expense and honorarium• Health insurance procedures• Management of facilities

Research Planning and Management Office• Research promotion and support (consultation for grants and patents, etc.)• Establishing research environments (facility and safety management, research agreement, etc.)• Fostering young scientists (Winter School, Advanced Postdoc Program, orientation, etc.)• Organizing scientific events (symposia, colloquia, seminars, etc.)• Public relations (publishing, website, outreach to citizens, etc.)• The Peter Doherty Institute for Infection and Immunity, the University of Melbourne, Australia

45Committees & Advisory Board for IFReC

The World Premier International Research Center Initiative (WPI)■■ Program Director
Akira UKAWA

As of November, 2023
WPI Program Director and Academy Director■■ Program Committee Members

6

Michinari HAMAGUCHI(Chairperson)

Director General, Strategic Center Biomedical Advanced Vaccine Research and Development for
Preparedness and Response(SCARDA), Japan Agency for Medical Research and Development (AMED)Councelor to the President, Japan Science and Technology Agency (JST)Mariko HASEGAWA

President, Japan Arts Council

Kazuhiko ISHIMURA

President, National Institute of Advanced Industrial Science and Technology

Takaaki KAJITA

Professor, Institute for Cosmic Ray Research, The University of Tokyo
Nobel laureate in Physics (2015)

Maki KAWAI

President, National Institutes of Natural Sciences

Motoko KOTANIExecutive Vice President, Tohoku University

Ryozo NAGAI

President, Jichi Medical University

Rita COLWELL

Distinguished University Professor, University of Maryland, USA

Richard DASHER

Director, US-Asia Technology Management Center, Stanford University, USA

Victor Joseph DZAU

President, National Academy of Medicine, USA

Pavel KABAT

Secretary-General, International Human Frontier Sciences Program Organization (HFSPO), France

Matthias KLEINER

University Professor, Technical University Dortmund, Germany

LIM Chuan Poh

Chairman, Singapore Food Agency (SFA)

Mona NEMER

Chief Science Advisor, Government of Canada

Jean ZINN-JUSTIN

Scientific adviser, Institute of Research into the Fundamental Laws of the Universe (IRFU/CEA), France

WPI Academy
In 2017, MEXT established the WPI Academy to be the vanguard in internationalizing and further renovating Japan’s research
environment. The WPI Academy is a much-anticipated upgrade of WPI institutes,and is expected to position Japan as a hub at the
pinnacle of international researcher circulation. In the decade ahead, the research institutes of WPI and WPI Academy will work
together to hold public relations and outreach activities.

■■ Program Officer for IFReC
Kouji MATSUSHIMA

Professor, Research Institute for Biomedical Sciences, Tokyo University of Science

International Scientific Advisory Board for IFReC
Lewis LANIER

University of California, San Francisco, USA

Anne O’GARRAThe Francis Crick Institute, UK

Jeffrey RAVETCH

Rockefeller University, USA

Art WEISS

University of California, San Francisco/Howard Hughes Medical Institute, USA

Joachim SCHULTZ

DZNE/LIMES Institute, University of Bonn, Germany

Kayo INABA

Japan Agency for Medical Research and Development, Japan

Kazuhiko YAMAMOTO

RIKEN Center for Integrative Medical Sciences, Japan

Osamu OHARA

Kazusa DNA Research Institute, Japan

Hiroshi KIYONO

Future Medicine Education and Research Organization at Chiba University, Japan

As of March, 2024

7

Laboratories
As of March 2024

Immunology

Host Defense
models.

Role of Nuclease-null Regnase-1

Shizuo Akira, MD/PhD

Our research is dedicated to unraveling the intricate
mechanisms of host defense against a spectrum of pathogens,encompassing both innate and acquired immunity. Additionally,
we are committed to developing efficacious therapeuticinterventions for immune-related disorders. A focal point of ourinvestigation lies in understanding the function of Regnase-1, an
RNA-binding protein (RBP) implicated in autoimmune diseasedevelopment. Specifically, we are exploring how Regnase-1
degrades a subset of inflammatory mRNAs, thus contributing to
the maintenance of a finely tuned immune response. The insights
gained from our recent research not only shed light on Regnase1's role in natural killer (NK) cells but also unveil its self-regulatory
mechanism and enzymatic activity in RNA degradation. Thisknowledge has the potential to pave the way for novel strategies
aimed at preventing autoimmune diseases.

Role of Regnase-1 in NK cells
Recent studies have emphasized the critical role of Regnase-1/
ZC3H12A in the function and metabolism of CD8+ T cells, as wellas in maintaining B cell balance to prevent immune-relateddisorders. However, how Regnase-1 regulates NK cell signaling
and their anti-tumor functions remains unclear. To investigate,
we used mice with NK cell-specific Regnase-1 knockout (Reg1ΔNK).
We found that knocking out Regnase-1 enhances the ability of NK
cells to fight tumors by increasing IFN-γ and cytolytic protein
production. Analysis of NK cells from the spleen and tumorinfiltrating NK cells revealed an activated subpopulation in
Reg1ΔNK-NK cells, with high expression of cytotoxic genes.
Additionally, we also found increased expression of CXCR6 in
Reg1ΔNK-NK cells, which typically remains low in healthy NK cells,
promoting their infiltration into tumors. The enhanced anti-

10■ Professor

Shizuo Akira■ Associate Professor

Kazuhiko Maeda■ Assistant Professor

Kiyoharu Fukushima■ Postdoctoral Fellow

3■ Research Assistant4■ Visiting Scientist

3■ Support Staff

4

Regnase-1 is an RNase that plays a crucial role in dampening
immune responses by breaking down inflammatory mRNAs. We
created mice with a single point mutation in the catalytic centerof Regnase-1's RNase domain (D141N), rendering it unable tofunction as an endonuclease. The D141N mutant mice developed
systemic inflammation, immune cell infiltration in multiple
organs, and progressive lung granuloma formation. In D141N
mutant mice, CD4+ T cells, which are mainly affected by thismutation, exhibited heightened activity in the mTORC1 pathway,
leading to autoimmune characteristics. RNA-seq analysis revealed
high expression of the serine-threonine kinase Pim2, which is
involved in the inflammation of the lungs in these mice. Inhibitionof Pim2 kinase activity improved granulomatous inflammation,
reduced immune cell infiltration and proliferation in the lungs,
and decreased adhesion molecule expression on CD4+ T cells. In

this study, we verified that Pim2 is indeed a target gene ofRegnase-1, suggesting that Pim2 is involved in facilitating theadhesion and migration of immune cells to inflamed tissues. Ourfindings shed light on the importance of Regnase-1's RNase
activity in immune regulation and highlight the potential oftargeting Pim2 as a therapeutic strategy for modulating abnormal
immune responses.

Exploration of Cellular Dynamism and Crosstalk inLung Fibrosis
Fibrosis is a life-threatening disease with unknown causes. The
activation of monocytes and macrophages is associated withfibrosis development, but the underlying pathogenesis remains
poorly understood. Currently, we are trying to understand thecomplex spatial and intercellular dynamics of fibrotic lesions andclarify how fibroblast activation are maintained in fibrotic lungs
in association with profibrotic immune cell populations.

tumor activity and tumor infiltration of Reg1ΔNK-NK cells werelinked to the IFN-γ signaling pathway. Blocking IFN-γ prevented
the accumulation of CXCR6+ Regnase-1-deficient NK cells intumors and CXCL16 expression in the tumor microenvironment.
Furthermore, Reg1ΔNK-NK cells showed increased expression ofoctamer-binding protein 2 (OCT2) and IκBζ, which form acomplex with NF-κB and bind to the Ifng promoter, enhancing its
transcription. These findings suggest that inhibiting Regnase-1 in
NK cells could enhance their anti-tumor activity and infiltration
into tumors. This could lead to innovative strategies, such as
Regnase-1 deleted chimeric antigen receptor (CAR)-NK therapy,
to improve NK cell-based immunotherapies against solid tumors.

Regulation of Regnase-1 3’UTR
Regnase-1 autoregulates its own expression by binding tostem-loop (SL) RNA structures in its 3’UTR and cleaving its mRNA.
To study the importance of Regnase-1-mediated autoregulationin immune responses, we created mice with a 2-bp deletion in
the target SL of the Regnase-1 3’UTR. This deletion prevented SLformation, reducing Regnase-1-mediated mRNA degradation.
The mutant mice showed normal hematopoietic celldifferentiation. Biochemically, the mutation increased Regnase-1
mRNA stability and levels of both Regnase-1 mRNA and protein inmouse embryonic fibroblasts (MEFs). Il6, a Regnase-1 target gene,
was consistently suppressed in mutant MEFs. Additionally,
Regnase-1 protein levels were higher in mutant MEFs comparedto wild-type MEFs both at baseline and after proinflammatory
cytokine stimulation. These findings suggest a negative feedback
loop for Regnase-1 expression and offer a unique model forstudying Regnase-1 overexpression in vivo. Further validation iscurrently underway in skin inflammation and lung inflammation

Figure.
The accumulation of NK cell and their heightened production of cytotoxic proteins within the tumor
microenvironment bolster the anti-tumor activity of Reg1ΔNK.

Recent Publications
1. Sun X, Nagahama Y, et al. Deletion of the mRNA endonuclease
Regnase-1 promotes NK cell anti-tumor activity via OCT2-dependenttranscription of Ifng. Immunity. (2024).
2. Lu Y, et al. CGRP sensory neurons promote tissue healing via
neutrophils and macrophages. Nature. 628:604-611 (2024).

4. Akira S and Maeda K. Control of RNA stability in immunity. Ann Rev
Immunol. 39:481-509 (2021).
5. Fukushima K, et al. Dysregulated expression of the nuclear exosome
targeting complex component Rbm7 in non-hematopoietic cells
licenses the development of fibrosis. Immunity. 52:542-556 (2020).

3. Kawai T, et al. Decoding Toll-like receptors: Recent insights andperspectives in innate immunity. Immunity. 57:649-673 (2024).

11

Immunology

Immunoglycobiology

Taroh Kinoshita, PhD
Yoshiko Murakami, MD/PhD (Co-PI)■ Professor

Taroh Kinoshita
Yoshiko Murakami

Our major interest has been biosynthesis and deficiencies ofglycosylphosphatidylinositols (GPIs), which are a class of
glycolipids that serve as membrane anchors for more than 150
different cell surface proteins in mammals, in addition to being
present as free glycolipids. In FY2023, we made the following
progress.

Role of ARV1 in GPI biosynthesis
ARV1 is an ER-resident membrane protein involved in the
homeostasis of various lipids including GPI. However, themechanism by which ARV1 regulates GPI biosynthesis has not yet
been fully elucidated. We reported last year that ARV1 associateswith PIGQ, a component of the initial enzyme complex in the GPI
biosynthesis pathway, and is required for the upregulation of GPI
biosynthesis (Liu Y-S et al, J. Cell Biol., 2023). We further extended
our study on ARV1’s role in the initial enzyme, GPI-Nacetylglucosamine transferase, and found that ARV1 is required
for the efficient usage of phosphatidylinositol (PI) by this enzyme
complex. In fact, the GPI-N-acetylglucosamine transferase
complex containing ARV1, but not the complex lacking ARV1,
had associated PI and used it to generate N-acetylglucosamine-PI
upon the addition of UDP-N-acetylglucosamine (Lu T et al,
manuscript in preparation).

Origin of alkyl-acyl glycerol in mammalian GPIs
PI moiety in GPIs is derived from cellular PI, however, the PI in
mammalian GPI’s is mainly in the alkyl-acyl form, whereas cellar PI
is usually in the diacyl form. The mechanism by which alkyl-acyl PI
moiety is generated has been unclear. We showed thatmammalian cells contain small amounts of alkyl-acyl PIs andpreferentially use them for GPI synthesis. Like other ether

12■ Research Assistant

1■ Visiting Scientist

2■ Support Staff

3phospholipids, the generation of alkyl-acyl PI is dependent on
the alkyl-phospholipid biosynthesis pathway present in theperoxisome, which starts from dihydroxyacetone phosphate (Li X
et al, manuscript in preparation).

Gene therapy of inherited GPI deficiency model mice
To establish effective therapies for inherited GPI deficiency, we
generated model mice by knocking out the Piga gene in thebrain. These model mice exhibited more severe symptoms thanthose observed in humans with inherited GPI deficiency. Genetherapy using Adeno-associated virus PHP.eB (AAV-PHP.eB),
aimed at systemically restoring normal Piga mRNA levels in themodel mice, was effective in prolonging lifespan and amelioratingsome symptoms (Murakami Y et al, Mol. Therapy — Methods Clin.
Dev., 2023). These results, together with our previously reported
gene therapy results with Pigo mutant model mice (Kuwayama R
et al, Nat Commun., 2022), suggest that AAV-mediated genereplacement is a promising approach for the effective treatment
of inherited GPI deficiency.

Figure.
Gene therapy in inherited GPI deficiency model mice. Mice with brain-specific KO of the Piga gene
were intravenously injected on days 1-2 with AAV-PHP.eB carrying PIGA cDNA. Due to the X-linkage
of Piga, heterozygous KO females become mosaic with GPI-expressing and -deficient cells in thebrain. Survival was extended in both males and females, and some neurological phenotypes wereameliorated. (cited from Murakami Y et al, Mol. Ther. Methods Clin. Dev., 14:32(1):101176 (2023))

Recent Publications
1. Murakami Y, Umeshita S, Imanishi K, Yoshioka Y, Ninomiya A, Sunabori
T, Likhite S, Koike M, Meyer KC and Kinoshita T. AAV-based genetherapy ameliorated central nervous system specific GPI defect inmouse models. Mol Ther Methods Clin Dev. 14:32(1):101176 (2023).
2. Liu IS, Wang Y, Zhou X, Zhang L, Gao XD, Murakami Y, Fujita M andKinoshita T. Accumulated precursors of specific GPI-anchored proteins
upregulate GPI biosynthesis with ARV1. J Cell Biol. 222: e202208159(2023).

4. Ishida M, Maki Y, Ninomiya A, et al. Ethanolamine-phosphate on thesecond mannose is a preferential bridge for some GPI-anchoredproteins. EMBO Rep. e54352 (2022).
5. Wang Y, Menon AK, Maki Y, et al. Genome-wide CRISPR screen
reveals CLPTM1L as a lipid scramblase required for efficient
glycosylphosphatidylinositol biosynthesis. Proc Natl Acad Sci USA.
119(14): e2115083119 (2022).

3. Kuwayama R, Suzuki K, Yoshioka Y, et al. Establishment of a mousemodel of inherited PIGO deficiency and therapeutic potential of AAVbased gene therapy. Nat Commun. 13:3107 (2022).

13

Immunology

Immunopathology

Atsushi Kumanogoh, MD/PhD

Our research team is involved in two approaches, that is, basic
and clinical immunology. As basic aspects of our projects, ourproposed study is the regulation of immune cell motility andmigratory behavior in vivo by soluble and membrane-bound‘immune guidance molecules’ such as semaphorins and their
receptors. Semaphorins were originally identified as axonguidance molecules that function during neuronal development.
However, cumulative evidence indicates that semaphorins also
participate in immune responses, both physiological andpathological, and they are now considered to be potential
diagnostic and/or therapeutic targets for a range of diseases.
Beyond such basic implications, we are trying to apply thefindings from this proposed study into the diagnosis/therapy forhuman immunological disorders, such as autoimmunity, allergy,
immune deficiency, cancer/metastasis, and neurodegenerative
diseases. We recently focus on how to apply the findings obtainedby bench work into the bedside. Under such research background,
we have performed single cell analysis on patients with ANCAassociated vasculitis.

■ Professor

Atsushi Kumanogoh■ Assistant Professor

Takahiro Kawasaki■ Research Assistant5■ Support Staff

5

The immunological basis of the clinical heterogeneity in
autoimmune vasculitis remains poorly understood. In this study,
we conduct single-cell transcriptome analyses on peripheral
blood mononuclear cells (PBMCs) from newly-onset patientswith microscopic polyangiitis (MPA). Increased proportions ofactivated CD14+ monocytes and CD14+ monocytes expressing
interferon signature genes (ISGs) are distinctive features of MPA.
Patient-specific analysis further classifies MPA into two groups.
The MPA-MONO group is characterized by a high proportion ofactivated CD14+ monocytes, which persist before and after
immunosuppressive therapy. These patients are clinically definedby increased monocyte ratio in the total PBMC count and have ahigh relapse rate. The MPA-IFN group is characterized by a highproportion of ISG+ CD14+ monocytes. These patients are clinicallydefined by high serum interferon-alpha concentrations and show
good response to immunosuppressive therapy. Our findings
identify the immunological phenotypes of MPA and provide
clinical insights for personalized treatment and accurate
prognostic prediction.

Recent Publications
1. Nishide M, Shimagammi H and A Kumanogoh A. Single-cell analysis inrheumatic and allergic diseases: Beyond the sea of data, challenges
for clinical application. Nat Rev Immunol. (in press).

4. Edahiro R, et al. Single-cell analyses and host genetics highlight therole of innate immune cells in COVID-19 severity. Nat Gen. 55: 753-767(2023).

2. Naito Y, et al. Tumor-derived semaphorin 4A improves PD-1-blocking
antibody efficacy by enhancing CD8(+) T cell cytotoxicity andproliferation. Sci Adv. 9: eade0718 (2023).

5. Tsujimoto K, et al. The lysosomal Ragulator complex activates NLRP3
inflammasome in vivo via HDAC6. EMBO J. 42: e111389 (2023).

3. Nishide M, et al. Single-cell multi-omics analysis identifies two distinct
phenotypes of newly-onset microscopic polyangiitis. Nat Commun.
14:5789 (2023).

14

15

Immunology

Immunochemistry

Hisashi Arase, MD/PhD■ Professor

Hisashi Arase■ Assistant Professor

Hui JinKazuki KishidaA) Misfolded proteins complexed with MHC class II
molecules are targets for autoimmune diseases.
MHC class II allelic polymorphisms are associated withsusceptibility to many autoimmune diseases. However, the roleof MHC class II molecules in autoimmune disease susceptibility
has remained unclear. We have found that misfolded cellular
autoantigens are rescued from protein degradation by MHC class
II molecules (Int. Immunol. 2013). Moreover, we discovered that
misfolded proteins complexed with MHC class II molecules serve
as targets for autoantibodies in autoimmune diseases such as
rheumatoid arthritis, antiphospholipid syndrome, and ANCAassociated vasculitis (PNAS 2014; Blood. 2015; Br. J. Dermatol.
2017; Arthritis Rheumatol. 2017; Arthritis Rheumatol. 2021). We
also found a strong correlation between autoantibody binding to
misfolded proteins transported to the cell surface by MHC class II
molecules and susceptibility to autoimmune diseases. Further
analyses revealed that self-antigens complexed with MHC II
molecules can abrogate self-tolerance, thereby inducing an
autoimmune response (Science Advances 2022). These findings
demonstrate that misfolded proteins, which would normally not
be exposed to the immune system, are involved in thepathogenicity of autoimmune diseases as ‘neo-self’ antigens(Figure 1).

■ Postdoctoral Fellow

2■ Research Assistant6■ Visiting Scientist

1■ Support Staff

4

Figure 1.
Misfolded proteins transported to the cell surface by MHC class II molecules are targets for autoantibodies.
Misfolded cellular proteins are generally degraded in the cells and are not transported to outside the cells. Therefore,
misfolded proteins transported to the cell surface by MHC class II molecules may be recognized as ‘neo-self’ antigens by
the immune system, which initiates an aberrant immune response to self-antigens (Int. Immunol. 2013; PNAS 2014; Blood
2015, Br. J. Dermatol. 2017; Arthritis Rheumatol. 2017, Arthritis Rheumatol. 2020; Arthritis Rheumatol. 2022; Science
Advances 2022).

B) Studies on host-pathogen interaction
The immune system has coevolved with infectious diseases,
which suggests that studies on host-pathogen interactions are
crucial for understanding the immune system. We have foundthat PILRα, a member of the inhibitory paired receptors, plays acrucial role in regulating the immune response (Nat. Immunol.
2012; Int. Immunol. 2015; Eur. J. Immunol. 2016), as well as in HSV1 infection (Cell 2008; J. Virol. 2009). Similarly, Siglec-4 andSiglec-7, which are also paired receptors, are implicated in VZV
infection (PNAS 2010; BBRC 2022). The LILR family represents
another group of paired receptors. We discovered that theactivation of LILRA2 is involved in detecting immunoglobulin
abnormalities in microbial infection (Nature Microbiology 2016).
Additionally, we found that RIFINs, which are products of themultigene family of Plasmodium falciparum, contribute to
immune evasion by binding to the inhibitory receptors LILRB1
and LILRB2 (Figure 1. Nature 2017; Nature 2020; BBRC 2021). From
our analysis of SARS-CoV-2 infection, we found that certain
antibodies against the spike protein cause conformationalchanges in the spike protein, thereby enhancing its infectivity(Cell 2021, Figure 2). This finding suggests that SARS-CoV-2 may
use antibodies to enhance its infectivity. These findings
underscore the crucial role of host-pathogen interactions inregulating infectious diseases.

Figure 2.
SARS-CoV-2 infectivity enhancing antibodies.
Some antibodies against NTD of spike protein induces conformational change in spike protein to induce the open formof RBD and enhance the infectivity of SARS-CoV-2 (Cell 2021).

Recent Publications
1. Jin H, Kishida K, Arase N, Matsuoka S, Nakai W, Kohyama M, Suenaga T,
Yamamoto K, Sasazuki T, Arase H. Abrogation of self-tolerance by
misfolded self-antigens complexed with MHC class II molecules. Sci
Adv. 8: eabj9867 (2022).
2. Liu Y, Soh WT, Tada A, Arakawa A, Matsuoka S, Nakayama EE, Li S, Ono
C, Torii S, Kishida K, Jin H, Nakai W, Arase N, Nakagawa A, Shindo Y,
Kohyama M, Nakagami H, Tomii K, Ohmura K, Ohshima S, Okada M,
MatsuuraY, Standley DM, Shioda T, Arase H. An infectivity-enhancing
site on the SARS-CoV-2 spike protein is targeted by COVID-19 patient
antibodies. Cell. 184:3452-3466 (2021).

16

3. Saito F, et al. Immune evasion of Plasmodium falciparum by RIFIN via
inhibitory receptors. Nature. 552:101-105 (2017).
4. Hiwa R, Ohmura K, Arase N, Jin H, Hirayasu K, Kohyama M, Suenaga T,
Saito F, Terao C, Atsumi T, Iwatani H, Mimori T, Arase H. Myeloperoxidase/
HLA Class II Complexes Recognized by Autoantibodies in Microscopic
Polyangiitis. Arthritis Rheumatol. 69:2069-2080 (2017).
5. Hirayasu K, Saito F, Suenaga T, Shida K, Arase N, Oikawa K, Yamaoka T,
Murota H, Chibana H, Nagai H, Nakamura Y, Katayama I, Colonna M,
Arase H. LILRA2 is an innate immune sensor for microbially cleaved
immunoglobulins. Nat Microbiol. 1:1-7 (2016).

17

Immunology

Immune Regulation

Tadamitsu Kishimoto, MD/PhD
Sujin Kang, PhD

18■ Professor

Tadamitsu Kishimoto■ Associate Professor

Sujin Kang

■ Assistant Professor

Hozaifa Metwally■ Research Assistant

1■ Visiting Scientist

1■ Support Staff

8

Gp130–HIF1α axis-induced vascular damage is prevented
by the short-term inhibition of IL-6 receptor signaling

Threonine Phosphorylation of STAT1 Restricts Interferon
Signaling and Promotes Innate Inflammatory Responses

Endothelial integrity is important for maintaining immune
homeostasis. Previously, we identified that interleukin (IL)-6trans-signaling induced endothelial activation in the context ofcytokine production and activation of the coagulation cascade.
Treatment with an anti-IL-6 Receptor (IL-6R) antibody, tocilizumab
inhibited endothelial activation and prevented the developmentof cytokine release syndrome (CRS). Protection against
endothelial damage is recognized as a frontline approach topreventing the progression of CRS. Accumulating evidence has
demonstrated that interleukin-6 (IL-6) promotes vascularendothelial damage during CRS, although the molecularmechanisms remain to be fully elucidated. Targeting IL-6 receptor
signaling delays CRS progression; however, current options arelimited by persistent inhibition of the immune system. Here, weshow that endothelial IL-6 trans-signaling promoted vasculardamage and inflammatory responses via hypoxia-inducible
factor-1α (HIF1α)–induced glycolysis. Using pharmacologicalinhibitors targeting HIF1α activity or mice with the genetic
ablation of gp130 in the endothelium, we found that inhibition of
IL-6R–HIF1α signaling in endothelial cells protected against
vascular injury caused by septic damage and provided survival
benefit in a mouse model of sepsis. In addition, we developed ashort half-life anti-IL-6R antibody (silent anti-IL-6R antibody) andfound that it was highly effective at augmenting survival forsepsis and severe burn by strengthening the endothelial
glycocalyx and reducing cytokine storm, and vascular leakage.
Together, our data advance the role of endothelial IL-6 transsignaling in the progression of CRS and indicate a potential
therapeutic approach for burns and sepsis (Figure 1).

The STAT1 working paradigm has focused on interferons (IFN)mediated JAK-Tyr701 phosphorylation and antiviral defense. We
have shown that lipopolysaccharide (LPS)-induced Thr749(equivalent to mouse Thr748) phosphorylation of STAT1, whichalters its target DNA motif, and stimulates the expression of IL12p40 and ARID5A, which increased the stability of IL6 mRNA
independently of the canonical IFN-JAK pathway in humanmacrophage. To investigate the physiological relevance of theThr748 phosphorylation of STAT1 in regulating the host’s defense
and inflammatory responses, we generated geneticallyengineered mice expressing phospho-deficient threonine748–
to-alanine (T748A) mutant Stat1. T748A mice were resistant to
LPS–induced lethality while exhibiting undisturbed IFN signaling
and total expression of Stat1. Notably, the T748A point-mutationof Stat1 recapitulates the safeguard effect of the genetic ablationof Stat1 following LPS-induced lethality, indicating that the Thr748
phosphorylation contributes inflammatory functionalities of
Stat1. LPS-induced Toll-like receptor 4 endocytosis activates acell-intrinsic IκB kinase (IKK)–mediated Thr748 phosphorylation of
Stat1, which promotes macrophages inflammatory response
while restricting the IFN and anti-inflammatory responses.
Moreover, we found that the Thr748 phosphorylation of STAT1 isimportant for maintaining intestinal epithelial barrier andsymbiotic gut microbiome composition. Following dextran
sodium sulfate (DSS)-induced colitis, T748A mice phenocopy the
detrimental pathology of the complete genetic ablation of Stat1
as evaluated by weight loss, colon lengths and histopathologicalanalysis. Together with the canonical tyrosine and serine, ouridentification of the Thr748 completes the phosphorylation
catalog of the transactivation domain of STAT1, and moregenerally other STAT proteins. Our findings show that Stat1
displays a phosphorylation-dependent modular functionality ininnate immune responses: IFN phospho-tyrosine dependent, andinflammatory phospho-threonine dependent, which provides
deeper insight on the combinatorial signaling logic of Stat1 inshaping cell-specific and context-dependent responses (Figure2).

Arid5a controls intestinal homeostasis by regulationof the cell cycle of plasmablasts
AT-rich interactive domain 5a (Arid5a) plays a crucial role in the
immune system through transcriptional and post-transcriptional
processes. Arid5a was abundantly expressed in B cells, but therole of Arid5a in B cells remains unclear. To investigate the role ofArid5a in B cells, we generated B cell-specific Arid5a knock-out

Figure 1.
Giving medication a short half-life anti-IL-6 to patients of Sepsis, ARDS, Burns, etc. is
expected to suppress vascular damage or secondary infection.

mice (Arid5aBKO). Immunoglobulin isotypes in the serum werecomparable between the control and Arid5aBKO mice, whereas
the concentration of IgA in feces was decreased in Arid5aBKO mice.
In addition, the percentage of IgA-secreting cells was significantly
reduced in the large intestine of Arid5aBKO mice, whereas no
significant differences in the small intestine and peyer`s patches.
By comparison of the frequency of antibody-secreting cells(ASCs) in the large intestine, we found that the frequency of ASCs
was decreased in Arid5aBKO mice. Deficiency of Arid5a in B cells
was confirmed to lead to a reduced frequency of plasmablast.
Mechanistically, Arid5a modulated the cell cycle of plasmablasts
by regulating the expression of Erdr1 upon stimulation withdifferentiation factors. Moreover, the deficiency of Arid5aaffected the IgA-microbiota axis and enhanced the susceptibility
to colitis. Collectively, the findings revealed that Arid5a played acrucial role in maintaining intestinal homeostasis by controllingthe cell cycle of plasmablasts.

Figure 2.
Threonine phosphorylation of STAT1 regulates the host’s inflammatory
responses.

Recent Publications
1. Kang S, Onishi S, Ling Z, Inoue H, Zhang Y, Chang H, Zhao H, Wang T,
Okuzaki D, Matsuura H, Takamatsu H, Oda J, Kishimoto T. Gp130-HIF1α
axis-induced vascular damage is prevented by the short-term
inhibition of IL-6 receptor signaling, PNAS. 121(2):e2315898120(2024).

3. Ishibashi T, Inagaki T, Okazawa M, Yamagishi A, Ohta-Ogo K, Asano R,
Masaki T, Kotani Y, Ding X, Chikaishi-Kirino T, Maedera N, Shirai M,
Hatakeyama K, Kubota Y, Kishimoto T, Nakaoka Y. IL-6/gp130 signalingin CD4+ T cells drives the pathogenesis of pulmonary hypertension.
PNAS. 121(16):e2315123121 (2024).

2. Metwally H, Elbrashy MM, Ozawa T, Okuyama K, White JT, Tulyeu J,
Søndergaard JN, Wing JB, Muratsu A, Matsumoto H, Ikawa M, Kishi H,
Taniuchi I and Kishimoto T. Threonine Phosphorylation of STAT1
Restricts Interferon Signaling and Promotes Innate Inflammatory
Responses. PNAS. 121(17):e2402226121 (2024).

4. Kishimoto T and Kang S. IL-6 Revisited: From Rheumatoid Arthritis to
CAR T Cell Therapy and COVID-19. Annu Rev Immunol. 40:323-348(2022).

19

Immunology

Mucosal Immunology

Kiyoshi Takeda, MD/PhD■ Professor

Kiyoshi Takeda■ Associate Professor

Hisako Kayama■ Assistant Professor

Ryu Okumura
Mari Murakami
Taiki Sakaguchi

Ulcerative colitis (UC) and Crohn’s disease (CD), clinicalphenotypes of inflammatory bowel disease (IBD), are chronic
disorders of the gastrointestinal tract involving inflammation andulceration. Genome-wide association studies have identified
more than 200 susceptible loci that are either shared by UC and
CD or related to one disease. Among them, the gene encoding
OTU deubiquitinase 3 (OTUD3) is located in the UC-associated1p36 locus (RNF186-OTUD3-PLA2G2E). In addition, the intronic
variant rs10799593 and the missense variant rs758098056 in
OTUD3 were identified as UC risk single nucleotide
polymorphisms. In patients with UC, dysbiosis is linked toalterations in the epigenome, transcriptional profile, andmetabolism of host cells as well as changes in the composition ofhost and microbial metabolites. However, it remains poorly
understood how UC risk genes interact with the dysbiotic
microbiota to influence the onset and/or progression of thedisease through modulating host cell physiology.

OTUD3 prevents the aggravation of colitis throughmodulation of the microbial cGAMP-STING-IFN-β
pathway in colonic fibroblasts.
In the murine and human colons, OTUD3 protein was
detectable in PDGFRα+ fibroblasts, whereas it was not detected inepithelial cells and CD45+ lamina propria cells. Accordingly,
Pdgfra-cre; Otud3f/f mice, but not LysM-cre; Otud3f/f, Villin-cre;Otud3f/f, and Cd4-cre; Otud3f/f mice, suffered from more severe
colitis accompanied by increase in the numbers of neutrophils,
eosinophils, CD64+ macrophages, and Ly6Cint CD11b+ dendritic
cells compared with Otud3f/f mice during administration of
dextran sodium sulfate (DSS). Ingenuity pathway analysis basedon the results from RNA-seq analysis using colonic fibroblasts

20■ Postdoctoral Fellow

2■ Research Assistant9■ Visiting Scientist

1■ Support Staff

1

exhibited significantly enriched upstream regulators of the genes
upregulated in the cells from Otud3-/- mice, which included
molecules related to the STING-IFN-β signaling pathway. Otud3deficient colonic fibroblasts stimulated with poly(dA:dT) showedenhanced phosphorylation of TBK1 and IRF3 compared withthose in wild-type cells. Additionally, IFN-β production in
response to poly(dA:dT) was increased in Otud3-deficientfibroblasts relative to wild-type cells. During DSS-induced colitis,the introduction of a Sting deficiency into Otud3-/- mice lessened
the clinical parameters and ameliorated large intestinal
pathology, indicating that OTUD3 negatively regulates the
STING-IFN-β signaling pathway in fibroblasts and thereby
prevents the aggravation of colitis.
Promoted transcription of interferon-stimulated genes (ISGs)
in fibroblasts from the colon of Otud3-/- mice was markedly
reduced by antibiotic treatment, implying that OTUD3
downregulates the microbiota-dependent activation of the
STING-IFN-β pathway in colonic fibroblasts. Bacteria trigger
STING activation without direct interaction with host cells
through secreting 3’3’-cGAMP. Elevated intracellular levels of
3’3’-cGAMP in colonic fibroblasts cultured in the presence of
3’3’-cGAMP became undetectable following treatment with thetransporter VRAC inhibitor DCPIB. 3’3’-cGAMP-induced
phosphorylation of TBK1 and IRF3 was accelerated in colonic
fibroblasts from Otud3-/- mice compared with that in cells fromwild-type mice. UC risk variant rs758098056 in the second exon of
OTUD3 (human OTUD3 (p. R90Q) corresponding to murine
OTUD3 (p. R89Q)), is located in the OTU domain of OTUD3. We
found that this missense variant links to reduced catalytic activityof OTUD3 against K27-linked polyubiquitination on STING.
Compared with wild-type mice, Otud3R89Q/R89Q mice showed more

severe disease manifestation with worsened large intestinal
pathology during DSS-induced colitis. In this context, peritoneal
injection of DCPIB reduced severe clinical parameters along withthe amelioration of large intestinal pathology in Otud3R89Q/R89Q
mice. Among patients with UC, carriers that were heterozygous
for the rs10799593 in the first intron of OTUD3 displayed low
expression of OTUD3 mRNA as well as OTUD3 protein in
fibroblasts from the non-inflamed site of the colon comparedwith non-carriers, whereas K27-linked STING polyubiquitination
was enhanced in colonic fibroblasts from rs10799593
heterozygous carriers. We found that 3’3’-cGAMP-induced
transcription of ISGs was upregulated in colonic fibroblasts fromrs10799593 heterozygous carriers. These findings indicate thatOTUD3 contributes to prevention of colitis progression by
inhibiting excessive activation of the cGAMP-STING-IFN-β
pathway in intestinal fibroblasts.

Promotion of 3’3’-cGAMP production in the UCassociated dysbiotic microbiota.
Because a similar allele frequency of rs10799593 was shown
between Japanese patients with UC and healthy controls (HCs),

we hypothesized that augmented 3’3’-cGAMP production in the
microbiota influences intestinal pathology in carriers of OTUD3
risk variants. We found that the enhanced colonization of cGAMPproducing bacteria species in UC patients. Accordingly, the fecal
concentration of 3’3’-cGAMP was high in patients with UC
compared with that in HCs. In addition, Otud3R89Q/R89Q mice
showed pathological features of UC in the colon after
transplantation of a fecal microbiome with the potential toproduce excessive cGAMP from UC patients. Collectively, theseresults demonstrate the critical role of OTUD3 in the maintenanceof intestinal homeostasis and highlight one mechanism ofinteraction between the host (OTUD3 in fibroblasts) andmicrobiota (STING agonist cGAMP) in UC development.
The evidence suggests that UC is an idiopathic, multifactorial,and progressive disorder linked to impaired quality-of-life and anincreased risk of fibrosis and colorectal cancer. Therefore, thedevelopment of a personalized and optimized approach forfacilitating mucosal healing and long-lasting remission isdesirable. Given our data in mice and humans, cGAMP transporter
VRAC and cGAMP-producing bacteria may be therapeutic targets
for patients with UC risk variants in OTUD3.

Figure.
OTUD3 prevents ulcerative colitis by modulating microbiota-mediated STING activation.
Left: OTUD3 contributes to maintenance of the large intestinal homeostasis by finely tunning
bacterial cGAMP-STING signaling in fibroblasts. Right: Ulcerative colitis (UC) risk variants in OTUD3
cooperate with dysbiotic microbiota-derived cGAMP to trigger disease progression.

Recent Publications
1. Nii T, Maeda Y, Motooka D, Naito M, Matsumoto Y, Ogawa T, OguroIgashira E, Kishikawa T, Yamashita M, Koizumi S, Kurakawa T, Okumura
R, Kayama H, Murakami M, Sakaguchi T, Das B, Nakamura S, Okada Y,
Kumanogoh A, Takeda K. Genomic repertoires linked with pathogenic
potency of arthritogenic Prevotella copri isolated from the gut ofrheumatoid arthritis patients. Annals Rheum. Dis. 82:621-629 (2023).

4. Tani H, Li B, Kusu T, Okumura R, Nishimura J, Okuzaki D, Motooka D,
Arakawa S, Yoshihara T, Ogino T, Tsai SH, Furuta Y, Muneta M, Nakamura
S, Fukusaki E, Yamamoto K, Yagita H, Kayama H and Takeda K: The ATPhydrolyzing ectoenzyme E-NTPD8 attenuates colitis throughmodulation of P2X4 receptor-dependent metabolism in myeloid cells.
Proc Natl Acad Sci USA. 118: e2100594118, (2021).

2. Yokoi T, Murakami M, Kihara T, Seno S, Arase M, Wing JB, Søndergaard
JN, Kuwahara R, Minagawa T, Oguro-Igashira E, Motooka D, Okuzaki D,
Mori R, Ikeda A, Sekido Y, Amano T, Iijima H, Ozono K, Mizushima T,
Hirota S, Ikeuchi H, Takeda K. Identification of a unique subset of tissue
resident memory CD4+ T cells in Crohn's disease. Proc Natl Acad Sci
USA. 120: e2204269120 (2023).

5. Morita N, Umemoto E, Fujita S, Hayashi A, Kikuta J, Kimura I, Haneda T,
Imai T, Inoue A, Mimuro H, Maeda Y, Kayama H, Okumura R, Aoki J,
Okada N, Kida T, Ishii M, Nabeshima R, Takeda K. GPR31-dependent
dendrite protrusion of intestinal CX3CR1+ cells by bacterial
metabolites. Nature 566:110-114 (2019).

3. Otake-Kasamoto Y, Kayama H, Kishikawa T, Shinzaki S, Tashiro T,
Amano T, Tani M, Yoshihara T, Li B, Tani H, Liu L, Hayashi A, Okuzaki D,
Motooka D, Nakamura S, Okada Y, Iijima H, Takeda K, Takehara T.
Lysophosphatidylserines derived from microbiota in Crohn's diseaseelicit pathological Th1 response. J Exp Med. 219: e20211291 (2022).

21

Immunology

Experimental Immunology
A

B

H165A (ZAC)

BALB/cBALB/c

H165A (ZAC)

H165A (ZAC)

BALB/c■ Professor

Shimon Sakaguchi■ Associate Professor

Kenji Ichiyama■ Assistant Professor

Arthritis Score

Norihisa Mikami
Yusuke Takeshima
Kelvin Chen Yigene
Tay Jian Wei Christopher■ Postdoctoral Fellow

3■ Research Assistant3■ Visiting Scientist

16■ Support Staff

4

6
5
4
3
2
1
0

BALB/c
H165A (ZAC)

05

W163C (SKG)
W163A

10 15 20
Age (week)% diarrheic mice

Shimon Sakaguchi, MD/PhD

60
40
20
0

25

W163C (SKG)
W163A

BALB/c
H165A (ZAC)

80

0

5

10 15 20
Age (week)

25

C
Tconv

TCR signal
amplification

Treg
TCR

TCR

ZAP-70attenuation

This laboratory studies: (i) the cellular and molecular basis of
immunologic self-tolerance, in particular the roles of regulatory T
cells (Tregs); (ii) the strategy for eliciting effective immune
responses to autologous tumor cells, or inducing immunologic
tolerance to organ transplants, by targeting Tregs; and (iii) the
cause and pathogenetic mechanism of autoimmune diseases byanalyzing the balance between Tregs and autoimmune T cells.
T cells mediate a variety of common autoimmune diseases
such as rheumatoid arthritis (RA) and type I diabetes. Both the
production of autoreactive conventional T (Tconv) cells throughT cell selection in the thymus and their pathogenic activation in
the periphery critically depend on T-cell receptor (TCR) signaling
upon recognition of self-peptide-bound major histocompatibility
complexes (self-pMHCs). In addition, depending on the strengthof the interaction between TCRs and self-pMHCs, somedeveloping T cells differentiate into Treg cells, which specifically
express the transcription factor Foxp3 and suppress the activationof pathogenic self-reactive T cells that have escaped thymic
negative selection. It remains to be determined, however, howqualitative or quantitative alteration of TCR signaling itself should
affect thymic generation of Treg and autoreactive Tconv cells andtheir peripheral functions to cause autoimmune diseases. Thereis accumulating evidence in humans and rodents that genetic
anomalies or variations in TCR-proximal signaling molecules,
such as ZAP-70 and LAT, and also in the molecules interactingwith them, such as PTPN22 and CBL family proteins, are causativeof and predisposing to a variety of autoimmune diseases. Amongthem, ZAP-70 mutations are unique in that they produce a wide
spectrum of immunological disorders encompassing immunodeficiency,
autoimmunity, immunopathology, and allergy in humans androdents, by strictly affecting the T cell compartment (Sakaguchi

22

et al., Nature 2003, Ito et al., Science 2014). Modulation of thestructure of ZAP-70 or the amount of its expression can therefore
be instrumental in deciphering how quantitative or qualitative
alteration of TCR signaling impacts on thymic production andperipheral activation of self-reactive Tconv as well as Treg cells,
consequently the balance between the two populations, to cause
a plethora of autoimmune/inflammatory diseases.
This year, we expressed in normal mice mutated ZAP-70
molecules with different affinities for the CD3 chains, or wild-type
ZAP-70 at graded expression levels under tetracycline-inducible
control (Tanaka A, Maeda S, et al., J. Exp. Med., 2023). Bothmanipulations reduced TCR signaling intensity to various extents
and thereby rendered those normally deleted self-reactive
thymocytes to become positively selected and form a highly
autoimmune TCR repertoire. The signal reduction more
profoundly affected Treg development and function because
their TCR signaling was further attenuated by Foxp3 thatphysiologically repressed the expression of TCR-proximal
signaling molecules, including ZAP-70, upon TCR stimulation.
Consequently, the TCR signaling intensity reduced to a criticalrange generated pathogenic autoimmune Tconv cells andconcurrently impaired Treg development/function, leading tospontaneous occurrence of autoimmune/inflammatory diseases,
such as autoimmune arthritis and inflammatory bowel disease.
The results provide a general model of how attenuation of TCR
signaling intensity via ZAP-70 and other TCR-proximal signaling
molecules, whether due to genetically-induced structuralanomalies or reduced expression of structurally intact forms, canbe causative of autoimmune and other immunological disorders
by affecting thymic development and peripheral function of
Tconv and Treg cells.

ZAP-70ZAP-70
LAT
MAPK

LAT

SLP76
NF-NB

Treg-intrinsic
repression of
Lck and ZAP-70

Lck

Lck

NFAT

Amount of signaling

MAPK

SLP76

NF-NB

NFAT

Amount of signaling

Figure.
Spontaneous development of autoimmune diseases in ZAP-70 mutated mice.
(A and B) Among ZAP-70 mutant strains (H165A (ZAC), W163C (SKG), W163A), only H165A (ZAC) micespontaneously develop autoimmune arthritis (A) and colitis with severe diarrhea (B).
(C) Key molecules of the TCR signaling pathway, such as Lck and ZAP-70, are specifically downregulated in Treg cells. TCR signal attenuation through ZAP-70 to a critical autoimmune range
differentially affects the development and function of Treg and Tconv cells, leading to spontaneousdevelopment of autoimmune diseases.

Recent Publications
1. Ichiyama K, Long J, Kobayashi Y, Horita Y, Kinoshita T, Nakamura Y,
Kominami C, Georgopoulos K, Sakaguchi S. Ikzf1 association with
Foxp3 for gene repression in Treg cells: disruption of the association
causes autoimmunity and promotes tumor immunity. Immunity.
(2024).
2. Tanaka A, Maeda S, Nomura T, Llamas-Covarrubias MA, Tanaka S, Jin L,
Lim EL, Morikawa H, Kitagawa Y, Akizuki S, Ito Y, Fujimori C, Hirota K,
Murase T, Hashimoto M, Higo J, Zamoyska R, Ueda R, Standley DM,
Sakaguchi N, Sakaguchi S. Construction of a T-cell receptor signaling
range for spontaneous development of autoimmune disease. J Exp
Med. 220(2):e20220386 (2023).

4. Tekguc M, James Badger Wing JB, Osaki M, Long J, Sakaguchi S. Tregexpressed CTLA-4 depletes CD80/CD86 by trogocytosis, releasing free
PD-L1 on antigen-presenting cells. Proc Natl Acad Sci U S A.
118(30):e2023739118 (2021).
5. Kawakami R, Kitagawa Y, Chen KY, Arai M, Ohara D, Nakamura Y, Yasuda
K, Osaki M, Mikami N, Lareau CA, Watanabe H, Kondo G, Hirota K,
Ohkura N, and Sakaguchi S. Coordinated activation of distinct Foxp3
enhancer elements for Treg development, maintenance, andimmunological self-tolerance. Immunity. 44(5):947-961 (2021).

3. Mimitou EP, Lareau CA, Chen KY, Zorzetto-Fernandes AL, Hao Y,
Takeshima Y, Luo W, Huang T, Yeung BZ, Papalexi E, Thakore PI,
Kibayashi T, Wing JB, Hata M, Satija R, Nazor KL, Sakaguchi S, Ludwig
LS, Sankaran VG, Regev A, Smibert P. Scalable, multimodal profiling ofchromatin accessibility, gene expression, and protein levels in single
cells. Nat Biotech. doi: 10.1038/s41587-021-00927-2 (2021).

23

Immunology

Lymphocyte Differentiation

Tomohiro Kurosaki, MD/PhD■ Professor

Tomohiro Kurosaki■ Postdoctoral Fellow

2■ Research Assistant

1■ Visiting Scientist

1■ Support Staff

4
Figure 1.
To track long-term survival of plasma cells, we sought to generate an experimental system that could
irreversibly label plasma cells in an inducible manner. To this end, we established a new mouse line inwhich plasma cells were labeled by a fluorescent reporter in response to a specific drug treatment.

Antibodies (Abs) are produced by terminally differentiatedplasma cells (PCs) and have distinct effector functions dependingon their isotype. While the majority of PCs generated in thesecondary lymphoid organs die within days after their formation(termed short-lived PCs; SLPCs), a small majority persists formonths, years, or even decades. These long-lived PCs (LLPCs) arefound in locations distinct from their generation site, often in thebone marrow. Hence, in order to distinguish between SLPCs and
LLPCs, previous studies determined the number of PCs residingin the bone marrow, considering them as LLPCs. However, since
it is known that this bone marrow population also includes
recently generated SLPCs, determining the exact number of
LLPCs is not possible using this approach. To overcome thelimitation of the current assay system, we developed an in vivo
time-stamping method for PCs, thereby allowing us to directly
measure the decay of the labeled bone marrow PCs (Figure).

Using this method, we first demonstrated that the homeostatic
PC pool in the bone marrow was continuously replenished byrecently arriving B220hiMHC-IIhi cells, a small fraction of whichthen matured into a B220loMHC-IIlo LLPC pool. Second, in the NPCGG immune response, contrary to expectation, we could not
find significant survival differences between GC-independentand -dependent NP+ PCs in the bone marrow. Third, in the bonemarrow, about 10-20% of the newly arrived SLPCs are maturated
to LLPCs and their longevity in the case of mouse is more than 1
year. Fourth, although it has been thought that IgM type LLPCs
do not exist, they exist, probably recognizing auto-antigens andmicrobiota. Finally, in contrast to NP+B220hiMHC-IIhi SLPCs in thebone marrow, the NP+B220loMHC-IIlo LLPCs with greater survival
potential were more likely to be immobilized in the niches,suggesting an association between adhesion strength andprovision of survival signals to PCs.

Figure 2.
Plasma cells right after arriving at bone marrow are B220hi MHC-IIhi, whereas those survived for long
period of time in bone marrow become B220lo MHC-IIlo. Bone marrow plasma cells exit cell cycle andacquire enhanced survival potential during progressive differentiation.

Recent Publications
1. Inoue T, Matsumoto Y, Kawai C, Ito M, Nada S, Okada M, Kurosaki T. Csk
restrains BCR-mediated ROS production and contributes to germinal
center selection and affinity maturation. J Exp Med. (2024).
2. Yada Y, Matsumoto M, Inoue T, Baba A, Higuchi R, Kawai C, Yanagisawa
M, Kitamura D, Ohga S, Kurosaki T, Baba Y. STIM-mediated calcium
influx regulates maintenance and selection of germinal center B cells.
J Exp Med. 221(1):e20222178 (2024).
3. Ise W, Kurosaki T. Tissues of origin matter to plasma cell longevity. Nat
Immunol. 25(2):194-195 (2024).

5. Koike T, Fujii K, Kometani K, Butler NS, Funakoshi K, Yari S, Kikuta J, Ishii
M, Kurosaki T, Ise W. Progressive differentiation toward the long-lived
plasma cell compartment in the bone marrow. J Exp Med.
220(2):e20221717 (2023).
6. Inoue T, Shinnakasu R, Kawai C, Yamamoto H, Sakakibara S, Ono C, Itoh
Y, Terooatea T, Yamashita K, Okamoto T, Hashii N, Ishii-Watabe A, Butler
NS, Matsuura Y, Matsumoto H, Otsuka S, Hiraoka K, Teshima T,
Murakami M, Kurosaki T. Antibody feedback contributes to facilitating
the development of Omicron-reactive memory B cells in SARS-CoV-2
mRNA vaccinees. J Exp Med. 220(2):e20221786 (2023).

4. Inoue T, Kurosaki T. Memory B cells. Nat Rev Immunol. 24(1):5-17(2024).

24

25

Malaria Immunology

Immunology

Cevayir Coban, MD■ Professor

Cevayir Coban■ Assistant Professor

Michelle S.J. Leeagainst the tumor, it is hypothesized that the iron balance isdisrupted by excessive iron consumption, possibly leading toincreased expression of LCN2 as an intracellular iron transporter.
We recently investigated the expressions of programmed cell
death ligand-1 (PD-L1) and LCN2 in breast cancers with various
molecular subtypes, along with their correlations with otherprognostic indicators, including Ki-67, lymph node metastasis,
histological grade, tumor-infiltrating lymphocyte (TILs)

accumulation, and necrosis. We found that there is an associationof LCN2 with known prognostic factors and molecular subtypes.
Moreover, significant elevations of LCN2 and PD-L1 expressions
were observed in triple-negative and HER2-positive breast
cancers. The findings from this research may contribute to theimmunotherapeutic application of LCN2 and its prognostic
significance in breast cancer management (Ekemen et al., Breast
Cancer: Targets and Therapy, 2024).

Jalal Alshaweesh

Our laboratory focuses on the elucidation of host-pathogen
interactions. We mainly work on malaria, but also cover several
other infectious diseases such as leishmaniasis and respiratory
infections. We study both innate and adaptive immune responses
against these diverse pathogens in order to develop successful
vaccines against them.

Adjuvant discovery and development platform
Adjuvants are considered essential vaccine components forenhancing vaccine responses. Recently, we have been involvedin the discovery of novel adjuvants as part of the AMED SCARDA
project led by Prof. Jun Kunisawa. We have systematicallyscreened innate and adaptive immune signaling molecules
involved in the mode of action (MOA) of adjuvants and vaccines.
One of the recent findings is to understand how the combinationof TLR9 and STING agonists synergistically induce innate andadaptive responses to generate robust anti-tumor responses(Temizoz et al., International Immunology, 2022). Our recent
projects have focused on investigating B cell development andthe pathways involved in germinal center (GC) formation for thegeneration of potent antibody responses against infections andduring vaccination. We found that TBK1, the well-known innate
immune signaling kinase that controls antiviral immune
responses and nucleic acid-mediated type I interferon responses,
is very important for the generation of GCs that confer sterile
immunity to reinfection (Lee et al., J Exp Medicine, 2022).

Elucidation of host-pathogen interactions
Our laboratory has been engaged in several aspects ofimmunopathology caused by Plasmodium parasites. Recently,
our team has been investigating the possible bone marrow

26■ Postdoctoral Fellow

1■ Research Assistant3

niches responsible for malaria-induced loss of memory. It is
becoming increasingly clear that perturbation of the bonemarrow microenvironment by infection and inflammation affects
hematopoiesis and may affect immune cell development. Our
recent findings demonstrate that the mesenchymal stromal
CXCL12-abundant reticular (CAR) cell population is reduced
during acute malaria infection (Lee et al., Int. Immunology, 2024).
The reduction of CXCL12 and IL-7 signals in the bone marrowaffects the lymphopoietic niche, which leads to the depletion ofcommon lymphoid progenitors, B cell progenitors, and mature B
cells, including plasma cells in the bone marrow. We found that
IFNγ is responsible for the upregulation of Sca1 on CAR cells, yet
the decline in CAR cell and B cell populations in the bone marrow
is IFNγ-independent. Interestingly, while B cell populations
declined, HSCs and multipotent progenitors increased with the
expansion of myelopoiesis and erythropoiesis. This suggests thatmalaria may affect host immunity by modulating the bonemarrow niche (Lee et al., Int. Immunology, 2024).

Infection and Cancer
Previously, we investigated the role of Lipocalin 2 (LCN2, also
known as siderocalin or neutrophil gelatinase-associatedlipocalin (NGAL)) in malaria infection that bolsters innate andadaptive immune responses to malaria infection throughmodulation of iron metabolism (Zhao et al., Cell Host Microbe,2012). LCN2 expression is also increased in cancer. In
carcinogenesis, in addition to the accumulation of somatic
mutations, stroma-associated immunity is an important regulatorof tumor growth. Tumor cells create a microenvironment byreleasing various mediators to maintain their presence andspread. Due to the infiltration of monocytes and leukocytes

Figure.
Acute malaria suppresses the B lymphocytic niche in the bone marrow through the alteration of
CXCL12-abundant reticular cells (Lee et al., International Immunology, 2024).

Recent Publications
1. Lee MSJ, Matsuo Dapaah J, Del Rosario Zorrilla C, Omatsu Y, Nagasawa
T, Uemura S, Iwama A, Ishii KJ, Coban C. Acute malaria suppresses the
B lymphocytic niche in the bone marrow through the alteration of
CXCL12-abundant reticular cells. Int Immunol. dxae012 (2024).
2. Ekemen S, Bilir E, Soultan HEA, Zafar S, Demir F, Tabandeh B, Toprak S,
Yapicier O, Coban C. The Programmed Cell Death Ligand 1 andLipocalin 2 Expressions in Primary Breast Cancer and Their Associations
with Molecular Subtypes and Prognostic Factors. Breast Cancer:
Targets and Therapy. 16: 1-13 (2024).

4. Lee MSJ, Inoue T, Ise W, et al. B cell intrinsic TBK1 is essential for
germinal center formation during infection and vaccination in mice. J
Exp Med. 219(2):e20211336 (2022).
5. Coban C. The host targeting effect of chloroquine in malaria. Curr
Opin Immunol. 66:98-107 (2020).

3. Becker HJ, Ishida R, Wilkinson AC, Kimura T, Lee MSJ, Coban C, Ota Y,
Tanaka Y, Roskamp M, Sano T, Tojo A, Kent DG, Yamazaki S. Controlling
genetic heterogeneity in gene-edited hematopoietic stem cells bysingle-cell expansion. Cell Stem Cell. 30(7):987-1000.e8 (2023).

27

Immunology

Vaccine Science

Ken J. Ishii, MD/PhD■ Professor

Ken J. Ishii■ Associate Professor

Kouji Kobiyama, Ph.D

■ Assistant Professor

Burcu Temizoz, Ph.D
Tomoya Hayashi, Ph.D

■ Postdoctoral Fellow

2■ Research Assistant4■ Visiting Scientist

4■ Support Staff

6

undesired type 2 immunity induced by STING agonists. We havepreviously shown that combining TLR9 and STING agonists
overcame limitations by synergistically inducing innate andadaptive IFNγ, making the combination an advantageous type 1
adjuvant, suppressing STING-mediated type 2 immunity andculminating in robust anti-tumor activities when used as atherapeutic agent for cancer immunotherapy in B16 F10 and
EG-7 mouse tumor models. Furthermore, we also found a potent
anti-tumor effect of the combination adjuvant in a Pan02
peritoneal dissemination model of pancreatic cancer only when
agonists for TLR9 and STING were administered locally. The
mechanisms of action involve both CD4 and CD8 T cells.
Mechanistically, IL-12 is crucial and synergistically coordinateswith type I IFNs to regulate the anti-tumor effect of the
combination. Synergism operates within individual antigenpresenting cells and is regulated at the level of promotor
activation, transcription and translation of IL-12. Such intracellulartranscriptional synergy may hold a key in successful cancer
immunotherapy and provide further insights into dual agonism
of innate immune sensors during host homeostasis and diseases.
In collaboration with Erasmus Medical Center, we have further
extended the applications of our combination adjuvant toneoantigen peptide cancer vaccines and found that K3 CpG+c-diAMP adjuvanted vaccine is able to induce potent Ag-specific Th1type and CD8+ T cell immune responses against synthetic long

Our laboratory aims to understand the immunologicalmechanisms involved in intra- and inter-cellular signaling
pathways that mediate the immunogenicity of successful
vaccines, as well as biological responses to adjuvants. This
understanding will aid in the development of new concepts,
modalities, and next-generation immuno-preventive and/or
therapeutic agents against infectious diseases, cancer, allergy,
and other non-communicable diseases.

DMXAA is a partial STING agonist competing for human
STING activation.
DMXAA is a mouse-specific stimulator of interferon gene
(STING) agonist that has shown STING-dependent anti-tumor
activity. Despite its inability to fully activate human STING,DMXAA has reached phase III in lung cancer clinical trials. The
mechanism by which DMXAA is effective against human lungcancer is currently unknown. In this study, we demonstrated thatDMXAA is a partial STING agonist that may interfere with agonistic
STING activation, which could explain its partial anti-tumor effectobserved in humans because STING was reported to be protumorigenic for lung cancer cells with low antigenicity.
Nonetheless, we developed a novel DMXAA derivative,
3-hydroxy-5-(4-hydroxybenzyl)-4-methyl-9H-xhanthen-9one(HHMX), which has the potential to strongly antagonize STING
pathway activation both in humans and mice. HHMX was foundto suppress abnormal responses caused by STING gain-offunction mutations that lead to STING-associated vasculopathy
with onset in infancy (SAVI) in vitro. Furthermore, HHMX
treatment was observed to reduce abnormal STING pathway
activity in peripheral blood mononuclear cells from SAVI patients.
Finally, in a SAVI mouse model, HHMX demonstrated a strong

28

peptides and neopeptide pools derived from melanoma andmesothelioma tumors. Indeed, K3 CpG+c-di-AMP adjuvant
formulation induced 10 times higher T cell responses against
neopeptides than the TLR3 agonist, which is currently the leading
adjuvant in clinical trials of neoantigen peptide vaccines.
Moreover, therapeutic vaccination with 20-mer peptide
combined with K3 CpG+c-di-AMP adjuvant synergizes in vivo
with α-PD-1 treatment to better control an ICB-resistant tumor.
Thus, our findings show that the TLR9+STING-adjuvanted vaccine
formulation generates robust T cell immunity against synthetic
long peptides and is a promising candidate to further improve
neoantigen vaccine platforms.

The 100 Days Mission
During the response to COVID-19, it became clear thatdeveloping safe, effective, and affordable vaccines, therapeutics,
and diagnostics for infectious diseases is crucial to saving lives.
Ken J. Ishii, as a scientific advisory member for CEPI, IPPS, andSTEG for G7, is committed to contributing to the 100 Days
Mission. The 100 Days Mission aims to prepare for a pandemic
threat by ensuring that approved, accurate, and rapid diagnostic
tests, initial therapeutic regimens and vaccines that have been
produced at large scale and ready for global deployment areavailable within the first 100 days of emergence of a newpathogen.

therapeutic effect by slowing disease progression (Figure 1).
These findings suggest that HHMX may have therapeuticpotential for STING-associated autoinflammatory diseases.

Challenges in developing personalized neoantigen
cancer vaccines
Promising results from the clinical data related to cancer
immunotherapy emphasized the importance of utilization of theimmune system for fighting against cancer. Vaccines have a
proven track record of promoting protective immune responses
against pathogens. As a result, vaccines targeting cancer
neoantigens are being advocated to direct and enhance immune
responses against tumors while minimizing the collateral damage
to the healthy tissue. Indeed, over one hundred clinical trials andextensive preclinical research to test various strategies fordiscovering neoantigens and formulating vaccines have been
performed up to now. However, it is important to acknowledge
that despite the efforts to develop neoantigen vaccines, mostclinical trials have not yet provided strong evidence of efficacy.
This review discusses the challenges that must be addressed tomaximize the potential of neoantigen vaccines in cancer therapy
by focusing primarily on the vaccine design and the tumor
microenvironment.

TLR9 and STING agonist combinations are robust cancer
immunotherapeutics and neopeptide cancer vaccine
adjuvants.
Agonists for TLR9 and stimulator of IFN genes (STING) havetherapeutic applications as anti-tumor agents and vaccine
adjuvants. However, their clinical use is limited due to the weak
IFN induction of the clinically available TLR9 agonist and the

Figure.
Mode of action of DMXAA and its derivative HHMX in human and mice.
DMXAA is a mouse-selective STING agonist, but only a partial agonist in humans, interfering with agonistic STING activation. Based on this data, a novel
DMXAA derivative, 3-hydroxy-5-(4-hydroxybenzyl)-4-methyl-9H-xhanthen-9one (HHMX), was developed. HHMX has the potential to become a therapeutic
agent for STING-associated autoinflammatory diseases, including SAVI, as it can antagonize the STING signaling pathway in both humans and mice.

Recent Publications
1. Temizoz B, Shibahara T, Hioki K, Hayashi T, Kobiyama K, Lee MSJ,
Surucu N, Sag E, Kumanogoh A, Yamamoto M, Gursel M, Ozen S,
Kuroda E, Coban C, Ishii KJ. 5,6-dimethylxanthenone-4-acetic acid
(DMXAA), a Partial STING Agonist, Competes for Human STING
Activation. Front Immunol. 15:1353336 (2024).
2. Katsikis PD, Ishii KJ, Schliehe C. Challenges in developing personalized
neoantigen cancer vaccines. Nat Rev Immunol. 24(3):213-227 (2024).
3. Kaku Y, Okumura K, Padilla-Blanco M, Kosugi Y, Uriu K, Hinay AA Jr,
Chen L, Plianchaisuk A, Kobiyama K, Ishii KJ. Genotype to Phenotype
Japan (G2P-Japan) Consortium; Zahradnik J, Ito J, Sato K. Virologicalcharacteristics of the SARS-CoV-2 JN.1 variant. Lancet Infect Dis.
24(2):e82 (2024).

4. Castro Eiro MD, Hioki K, Li L, Wilmsen MEP, Kiernan CH, BrouwersHaspels I, van Meurs M, Zhao M, de Wit H, Grashof DGB, van de Werken
HJG, Mueller YM, Schliehe C, Temizoz B, Kobiyama K, Ishii KJ, Katsikis
PD. TLR9 plus STING Agonist Adjuvant Combination Induces Potent
Neopeptide T Cell Immunity and Improves Immune Checkpoint
Blockade Efficacy in a Tumor Model. J Immunol. 212(3):455-465 (2024).
5. Dzau V, Swaminathan S, Baker C, Bright RA, Castillo J, Chuan TC,
Draghia-Akli R, Eardley-Patel R, Gao GF, Ishii K, Tebeje YK, Lambe T,
Machingaidze S, Røttingen JA, Shaligram U, Simão M, Swarup R,
Toussaint JF, Wairagkar NS. The 100 Days Mission: how a new medicalcountermeasures network can deliver equity and innovation. Lancet.
402(10412):1507-1510 (2023).

29

Immunology

Immunoparasitology
slowed tumor growth but less autoimmunity, targeting
T-bet+Foxp3+ cells appears to be safer than that targeting whole
Treg cells.
Collectively, the intersectional genetic labeling/depletion ofT-bet+Foxp3+ cells revealed their specific roles in suppressing
tumor immunity.

Masahiro Yamamoto, PhD

Regulatory T (Treg) cells expressing the transcription factor(TF) Foxp3 also express other TFs shared by Th subsets undercertain conditions. In 2023, to determine the roles of T-betexpressing Treg cells, we have reported generation of a mousestrain, called VeDTR mice, in which T-bet/Foxp3 double-positive
cells were engineered to be specifically labelled and depleted by
a combination of Cre- and Flp-recombinase-dependent gene
expression control. Characterization of T-bet+Foxp3+ cells using
the VeDTR mice revealed the high resistance under oxidative
stress, which was involved in T-bet+Foxp3+ cells to accumulate intumor tissues. Moreover, short-term depletion of T-bet+Foxp3+
cells only led to anti-tumor immunity but not autoimmunity,
whereas that of whole Treg cells did both.
To date, several methods to inducibly delete immune cell
populations have been developed using cytotoxic antibodies
and chemicals such as clodronate liposome, or by herpesvirus
thymidine kinases or DTR that are directly expressed under thecell-specific gene promoters and are able to render the cells
sensitive to ganciclovir or DT, respectively. DTR can also be
expressed indirectly using Cre recombinase-inducible mice
which allow their expression in various cell types upon lineagespecific Cre-mediated excision of a loxP-flanked stop cassette
under the Rosa26 promoter. However, these conventional
methods exhibit limited specificity toward the intended target
immune cells because most of immune cell types rarely definedby single gene but rather by intersectional expression of multiple
genes. To improve the specificity of Cre-mediated depletion ofmacrophage/monocyte subpopulations, transgenic mousesystems utlizing sequential promoter activities were developed.
Alternatively, intersectional genetic methods using the Cre/loxPand Flp/FRT double recombination systems or binary split Cre

30■ Professor

Masahiro Yamamoto■ Associate Professors

Miwa Sasai■ Postdoctoral Fellow

1■ Research Assistant2■ Support Staff

2

recombinases have been introduced to dissect various cell
subpopulations in brain. However, these intersectional genetic
strategies have not been developed for conditional and selective
depletion of immune cells. In the present study, we have createdthe VeDTR mouse line that can intersectionally label andconditionally deplete a cell-type-of-interest by YFP and DTR,
respectively.
T-bet+Foxp3+ cells showed higher suppressive activity thanT-bet-Foxp3+ cells. Moreover, TCR-stimulated T-bet+Foxp3+ cells
preferentially expressed perforin and anti-oxidant proteins.
Consistent with high concentrations of T-bet+Foxp3+ cells intumor tissues compared with spleens, it has been shown that
perforin expression in Treg cells is important for Treg cellmediated suppression of anti-tumor immunity. Preferential
expression of anti-oxidant proteins such as MT3 was also required
for Treg cell localization at tumor tissues that are shown tocontain increased concentrations of ROS, since MT3-deficient
Treg cells were sensitive to oxidative stress and failed toaccumulate in tumor tissues.
It was of note that whole Treg cell depletion in Foxp3-Cre/
VeDTR(ΔFRT) mice induced histopathology and morbidity,
whereas inducible T-bet+Foxp3+ cell depletion in Foxp3-Cre/
Tbx21-Flp/VeDTR(LF) mice did not lead to autoimmunity.
Although some intratumor CD45+ cell subsets such as NK cells
and CD11b+ cells were somewhat differentially affected bydepletion of T-bet+Foxp3+ cells or whole Treg cells, the resultant
slow tumor growth was comparable, suggesting that thedifference might seldomly influence anti-tumor immunity.
Indeed, antibody-mediated deletion of NK1.1+ cells alone did notrecover tumor growth in T-bet+Foxp3+ cell-depleted mice. From acancer immunotherapy perspective, considering the comparable

Figure.
The novel VeDTR mouse system specifically targets Th1-Treg cells.

Recent Publications
1. Okamoto M, Sasai M, Kuratani A, Okuzaki D, Arai M, Wing JB, Sakaguchi
S, Yamamoto M. A genetic method specifically delineates Th1-type
Treg cells and their roles in tumor immunity. Cell Rep. 42:112813(2023).

4. Sakaguchi N, Sasai M, Bando H, Lee Y, Pradipta A, Ma JS, Yamamoto M.
Role of Gate-16 and Gabarap in Prevention of Caspase-11-Dependent
Excess Inflammation and Lethal Endotoxic Shock. Front Immunol.
11:561948 (2020).

2. Tachibana Y, Hashizaki E, Sasai M, Yamamoto M. Host genetics
highlights IFN-γ-dependent Toxoplasma genes encoding secreted
and non-secreted virulence factors in in vivo CRISPR screens. Cell Rep.
42:e112592 (2023).

5. Bando H, Pradipta A, Iwanaga S, Okamoto T, Okuzaki D, Tanaka S,
Vega-Rodríguez J, Lee Y, Ma JS, Sakaguchi N, Soga A, Fukumoto S,
Sasai M, Matsuura Y, Yuda M, Jacobs-Lorena M, Yamamoto M. CXCR4
regulates Plasmodium development in mouse and humanhepatocytes. J Exp Med. 216:1733-1748 (2019).

3. Sasai M, Ma JS, Okamoto M, Nishino K, Nagaoka H, Takashima E,
Pradipta A, Lee Y, Kosako H, Suh PG, Yamamoto M. Uncovering a novelrole of PLCβ4 in selectively mediating TCR signaling in CD8+ but not
CD4+ T cells. J Exp Med. 218:e20201763 (2021).

31

Immunology

Biochemistry & Immunology
P2X7
ATP

K+

Xk

Shigekazu Nagata, PhD■ Professor■ Assistant Professor

Shigekazu Nagata?

Takaharu Sakuragi■ Postdoctoral Fellow

2■ Research Assistant3■ Visiting Scientist

1■ Support Staff

1

Ca2+

GDP

scrambling
phospholipids

Vps13a?

Figure 1.
Association of XK with VPS13A at the plasma membrane to scramble phospholipids.
VPS13A, a giant cytoplasmic protein (human VPS13A: 3174 aa), is associated with XK in the plasma membrane. P2X7 is a receptor for extracellular ATP andserves as a cation transporter. The binding of ATP to P2X7 activates the XK-VPS13A complex to scramble phospholipids by an unknown mechanism(s),leading to PtdSer-exposure and cell lysis.

The phospholipid composition of plasma membranes exhibits
an asymmetrical distribution across their inner and outer leaflets,with phosphatidylserine (PtdSer) predominantly localized in the
inner leaflet. This asymmetry is upheld by ATP-dependent
flippases, which facilitate the translocation of PtdSer from theouter to the inner leaflet. However, during apoptosis, cell fusion,and cell activation, the asymmetrical distribution of phospholipids
is disrupted by scramblases that non-specifically scramble
phospholipids between leaflets, leading to the exposure of
PtdSer on the cell surface.
Our previous studies (Segawa et al. Science 2014; Suzuki et al.
Nature 2010, Science 2013) have identified the key players in thisprocess: ATP11A, ATP11C, and CDC50A as flippases, TMEM16F asa Ca2+-dependent scramblase, XKR8 as a caspase-dependent
scramblase. We found a mutation of the TMEM16F gene inhuman Scott syndrome suffering hemophilia, which underscores
its significance in exposing PtdSer in the activated platelets,
crucial for activating blood clotting factors. Moreover, we've
uncovered XKR8, forming complexes with Basigin or Neuroplastin,
crucial for efficient clearance of dead cells and prevention of
autoimmunity (Suzuki et al. PNAS 2016; Kohno et al. PNAS 2019;
Yamashita et al. MCB 2019).
A high concentration of ATP (~ 4 mM) is present inside thecells, while its extracellular concentration is low (less than 30 nM).
However, its extracellular concentration increases by several
hundred μM in the inflamed tissues or tumor environment, wherea large amount of ATP is released from the cells undergoingnecrosis. ATP rapidly induces PtdSer-exposure in CD25+CD4+ T
cells and macrophages by binding to its receptor, P2X7, followedby necrosis and release of inflammatory cytokines. In the CRISPR/
Cas9 screening for the molecules involved in ATP-induced

32

PtdSer-exposure, we found that XK, a paralogue of XKR8, and the
VPS13A cytosolic lipid transporter, is required for the ATPinduced PtdSer-exposure downstream of P2X7. An unidentified
signal from the ATP-engaged P2X7 receptor seems to activate
the XK-VPS13A complex to scramble phospholipids in the plasma
membranes (Figure 1). Patients of neuroacanthocytosis, adisorder that affects erythrocytes and the central and peripheral
nervous system, carry a defect in Xk or Vps13a, indicating that the
XK- and VPS13A-mediated scrambling of phospholipids plays an
inevitable role in maintaining the homeostasis in hematopoietic
and nervous systems.
How phospholipids are translocated between lipid bilayers by
flippases and scramblases? We established the method to
produce a large amount of human XKR8-Basigin complex andsucceeded in purifying it to homogeneity. In 2021, we determined
its tertiary structure at a resolution of 3.8 Å (Sakuragi et al. Nat.
Struct. Mol. Biol. 2021). Its membrane-spanning region adopts a
cuboid-like structure. At least seven charged residues essential
for scramblase activity are placed from top to bottom as astaircase inside the molecule, providing a pathway for thetranslocation of phospholipids. The structure was recently refinedwith the complex embedded in lipid nanodisc (Sakuragi et al. J.
Biol. Chem. 2024)(Figure 2). This structure showed that the
C-terminal tail of XKR8 is engaged in intricate polar and van der
Waals interactions with a groove at its cytoplasmic surface of
XKR8. Point mutations to disrupt these interactions strongly
enhanced the scrambling activity of XKR8, indicating that thecytoplasmic tail region of XKR8 functions as a plug to prevent thescrambling of phospholipids.

(a)

(b)

Fab

BSG

XKR8

(c)

BSGȴ

(d)

8

9

Y261
F251

PC

6

F274
F381

11
R98

L185

nanodisc

11

XKR8

Y337

4

F382
P343

2
D71

R373

L377

11

10 Y338

W67

2

L189

5

N371

D129

4

3b

Figure 2.
The structure of the XKR8–basigin (BSG) complex. (a) a cryo-EM map of XKR8–BSG-anti BSG Fab complex in lipid nanodiscs. (b) A side view of the structureof the XKR8-Basigin complex. The helix 11 of XKR8 is shown in orange. The areas enclosed by the red (c) or green (d) box are viewed. Residues involved in
the interaction are shown as a stick model or a sphere model. Likely, hydrogen bonds are shown as dotted lines.

Recent Publications
1. Sakuragi T, Kanai R, Otani, M, Kikkawa M, Toyoshima C, and Nagata S.
The role of the C-terminal tail region as a plug to regulate XKR8 lipid
scramblase. J Biol Chem. 300:105755 (2024).

4. Ryoden Y, Segawa K, and Nagata S. Requirement of Xk and Vps13a for
the P2X7-mediated phospholipid scrambling and cell lysis in mouse T
cells. Proc Nat Acad Sci USA. 119:e2119286119 (2022).

2. Sakuragi T and Nagata S. Regulation of phospholipid distribution in
the lipid bilayer by flippases and scramblases. Nat Rev Mol Cell Biol.
24:576-596 (2023).

5. Sakuragi T, Kanai R, Tsutsumi A, Narita H, Onishi E, Nishino K, Miyazaki
T, Baba T, Kosako H, Nakagawa A, Kikkawa M, Toyoshima C, and Nagata
S. The tertiary structure of the human Xkr8-Basigin complex thatscrambles phospholipids at plasma membranes. Nat Struct Mol Biol.
28:825-834 (2021).

3. Ochiai Y, Suzuki C, Segawa K, Uchiyama Y, and Nagata S. Inefficientdevelopment of syncytiotrophoblasts in the Atp11a-deficient mouseplacenta. Proc Nat Acad Sci USA. 119: e2200582119 (2022).

33

Immunology

Molecular Neuroscience
Toshihide Yamashita, MD/PhD■ Professor

Toshihide Yamashita■ Associate Professor

Takahide Itokazu
Hiroshi Tsujioka■ Assistant Professor

Oluwaseun Fatoba
Takeshi Furumiya
Iyo Yorifuji
Yosuke Ohtake
Toru Nakanishi
Nao Shibuya
Abdellatif Abbaoui
Teruyuki Ishikura

Disorders of the central nervous system, such as cerebrovasculardiseases, cerebrospinal trauma, and encephalomyelitis, oftencause spatiotemporal changes in the nervous system and invarious biological systems, such as the immune system andvascular system. We have analyzed disorders of the neuralnetworks in the central nervous system and the subsequent
restoration process from the perspective of the functional
network of biological systems (Fig. 1). Further, we have analyzed
the mechanism by which the spatiotemporal dynamics in thosebiological systems control a series of processes (Fig. 2).
Particularly, the ultimate goal of this study is to elucidate themanner in which the control mechanism is affected by the
associations among the nervous system, immune system, andvascular system. Additionally, we aim to elucidate the processes
involved in the functioning of living organisms with neuralnetwork disorders within the central nervous system by observing
such disorders and their functional recovery process with respect
to the dynamics of the entire biological system and by conducting
a comprehensive analysis of the association between eachsystem.

■ Postdoctoral Fellow

1■ Research Assistant2■ Visiting Scientist

6■ Support Staff

2

Figure 1.
The mechanism of spontaneous functional recovery.

We observe the central nervous system as a single organ withina biological system. Further, studies from the perspective of how
the entire biological system is involved in disorders and recoveryof neural networks are scarce. By observing disorders in neuralnetworks and the biological reactions during the subsequent
recovery process as a “scrap-and-build” strategy, we aim toelucidate the mechanisms behind a series of reactions as well as
their significance that may potentially lead to a new and original
trend in Life Sciences.

Figure 2.
Biological systems that regulate rewiring of neural network after CNS injury.

Recent Publications
1. Shimizu M, Shiraishi N, Tada S, Sasaki T, Beck G, et al. RGMa collapses
the neuronal actin barrier against disease-implicated protein andexacerbates ALS. Sci. Adv. 9, eadg3193 (2023).

34

4. Fujita Y, Nakanishi T, Ueno M, Itohara S & Yamashita T. Netrin-G1
regulates microglial accumulation along axons and supports thesurvival of layer V neurons in the postnatal mouse brain. Cell Rep.
10:107580 (2020).

2. Iwamoto S, Itokazu T, Sasaki A, Kataoka H, Tanaka S, et al. RGMa signal
in macrophages induces neutrophil-related astrocytopathy in NMO.
An Neurol. 91:532-547 (2022).

5. Tanabe S & Yamashita T. B-1a lymphocytes promote oligodendrogenesis
during brain development. Nat. Neurosci. 21:506-516 (2018).

3. Ito M, Muramatsu R, Kato Y, Sharma B, Uyeda A, et al. Age-dependent
decline in myelination capacity is mediated by apelin-APJ signaling.
Nat Aging 1:284-294 (2021).

6. Kuroda M, Muramatsu R, Maedera N, Koyama Y, Hamaguchi M, et al.
Promotion of central nervous system remyelination by peripheral
FGF21. J Clin Invest. 127:3496-3509 (2017).

35

Immunology

Molecular Immunology
M. leprae

Sho Yamasaki, PhD
PGL-III■ Professor

Sho Yamasaki■ Associate Professor

Norikazu Yabuta■ Assistant Professor

Masamichi Nagae
Eri Ishikawa

Although leprosy (Hansen’s disease) is one of the oldest known
diseases, the pathogenicity of Mycobacterium leprae (M. leprae)remains enigmatic. Indeed, the cell wall components responsible
for the immune response against M. leprae are as yet largely
unidentified. We reveal here phenolic glycolipid-III (PGL-III) as an
M. leprae-specific ligand for the immune receptor Mincle. PGL-III
is a scarcely present trisaccharide intermediate in the biosynthetic
pathway to PGL-I, an abundant and characteristic M. leprae
glycolipid. Using activity-based purification, we identified PGL-III
as a Mincle ligand that is more potent than the well-known M.
tuberculosis trehalose dimycolate. The cocrystal structure of
Mincle and a synthetic PGL-III analogue revealed a uniquerecognition mode, implying that it can engage multiple Mincle
molecules. In Mincle-deficient mice infected with M. leprae,increased bacterial burden with gross pathologies were observed.
These results show that PGL-III is a noncanonical ligand
recognized by Mincle, triggering protective immunity.
https://pubs.acs.org/doi/full/10.1021/acscentsci.3c00040■ Postdoctoral Fellow

3■ Research Assistant6■ Visiting Scientist

8■ Support Staff

5

Immune protection
Rha

RO

R=

H

ImmuneevasionMincleGlc

Me

PGL-III

PGL-I

Stimulatory
ligandSilentRecent Publications
1. Ito E, Inuki S, Izumi Y, Takahashi M, Dambayashi Y, Ciacchi L, Awad W,
Takeyama A, Shibata K, Mori S, Mak JYW, Fairlie DP, Bamba T, Ishikawa
E, Nagae M, Rossjohn J, Yamasaki S. Sulfated bile acid is a host-derived
ligand for MAIT cells. Sci. Immunol. 9:eade6924 (2024).
2. Oka S, Watanabe M, Ito E, Takeyama A, Matsuoka T, Takahashi M, Izumi
Y, Arichi N, Ohno H, Yamasaki S, Inuki S. Archaeal Glycerolipids Are
Recognized by C-Type Lectin Receptor Mincle. J Am Chem Soc.
145:18538-18548 (2023).

4. Watanabe M, Motooka D, Yamasaki S. The kinetics of signaling throughthe common FcRγ chain determine cytokine profiles in dendritic cells.
Sci Signal. 16:eabn9909 (2023).
5. Schutt CR, Yamasaki S. Lectin recruites pathogenic bugs. J Exp Med.
220:e20221732 (2023).

3. Ishizuka S, van Dijk JHM, Kawakita T, Miyamoto Y, Maeda Y, Goto M, Le
Calvez G, Groot LM, Witte MD, Minnaard AJ, van der Marel GA, Ato M,
Nagae M, Codée JDC, Yamasaki S. PGL-III, a Rare Intermediate of
Mycobacterium leprae Phenolic Glycolipid Biosynthesis, Is a Potent
Mincle Ligand. ACS Cent Sci. 9:1388-1399 (2023).

36

37

Immunology

Stem Cell Biology and Developmental Immunology
cells were markedly reduced compared to those in contact withCXCL12-intact CAR cells. These results suggest that HSCs
detached from CXCL12-deficient CAR cells and attached to
CXCL12-intact CAR cells in the mutants, demonstrating thatCXCL12 attracts HSCs to CAR cells within BM.

Takashi Nagasawa, MD/PhD

Special microenvironments known as niches are essential forthe maintenance of hematopoietic stem cells (HSCs), which giverise to blood cells, and lympho-hematopoiesis within bonemarrow (BM). We isolated a chemokine, CXCL12 (SDF-1/PBSF) as
a molecule that stimulates the growth of B cell precursors(Nagasawa et al., PNAS 1994) and found that CXCL12 and its
receptor CXCR4 are essential for the colonization of BM by HSCs
during embryogenesis (Nagasawa et al., Nature 1996; Ara et al.,Immunity 2003), the maintenance of a pool of HSCs (Sugiyama etal., Immunity 2006), and the development of immune cells,
including B cells, plasmacytoid dendritic cells (pDCs), and NK cells
as well as vascular formation and cardiogenesis (Tachibana et al.,Nature 1998). Based on a key role of CXCL12 in HSC maintenance,
we identified a population of fibroblastic cells expressing CXCL12
at high levels, termed CXCL12-abundant reticular (CAR) cells
within BM (Sugiyama et al., Immunity 2006) and found that these
BM-CAR cells are the major producer of CXCL12 and SCF (Omatsu
et al., Immunity 2010), and the major cellular components ofniches for HSCs and immune cells, including B cells and plasma
cells (Tokoyoda et al., Immunity 2004; Sugiyama et al., Immunity
2006; Omatsu at al., Immunity 2010). Ding et al. showed that
leptin receptor-expressing (LepR+) cells overlap strongly with
CAR cells in BM (Ding et al., Nature 2012). Inconsistent with theclassical concept, we showed that numerous HSC niches remain
empty and that all BM-CAR cells create facultative niches for HSCs
(Shimoto et al., Blood 2017).
In addition, we showed that BM-CAR cells are mesenchymal
stem cells, which give rise to adipocytes and osteoblasts, and thattranscription factors, Foxc1 and Ebf3 are preferentially expressed
in BM-CAR cells and play a critical role in the formation andmaintenance of niches for HSCs and immune cells, inhibiting38■ Professor

Takashi Nagasawa■ Associate Professor

Yoshiki Omatsu■ Assistant Professor

Taichi Nakatani■ Research Assistant3■ Support Staff1

differentiation of BM-CAR cells into adipocytes and osteoblasts,
respectively (Omatsu et al., Nature 2014; Seike et al., Genes Dev.
2018). Furthermore, we showed that BM-CAR cells require Runx1
or Runx2 to prevent their fibrotic conversion and maintain HSCs
and hematopoiesis in adults (Omatsu et al., Nat Commun 2022).
In addition to the mouse, we revealed that the humancounterpart of CAR cells, which specifically expressed CXCL12,
Foxc1, and Ebf3, was the major component of nonhematopoietic
cells in human BM. This enabled the evaluation of their alterations
in various hematological disorders by flow cytometric andhistological analyses (Aoki et al., Br J Haematol 2021).
Ding et al. suggest that CAR/LepR+ cell-derived CXCL12 is notessential for the maintenance of HSCs since the number of HSCs
was unaltered in BM when CXCL12 was conditionally deleted
from LepR+ cells using LepR-Cre;CXCL12f/- mice (Ding et al., Nature2013). CXCL12 was deleted from only 70% of the CAR cells in the
mutants, probably due to lower LepR expression in early
postnatal bone marrow. Thus, we examined the role of CAR cellderived CXCL12 in the behavior and maintenance of HSCs.

CXCL12 attracts HSCs to BM-CAR cells within bonemarrow.
To visualize HSCs in the bone marrow sections, we generated
HSC-reporter mice, Evi1-GFP mice, and CXCL12-tdTomatof/f mice,
which enabled us to visualize CXCL12-intact and CXCL12-deficient
CAR cells. Control and Ebf3-CreERT2;CXCL12-tdTomatof/f;CXCL12f/mice were transplanted with BM cells from Evi1-GFP mice andinjected with tamoxifen three times. In these chimeric mice,
CXCL12-intact CAR cells were reduced by about 2-fold, but the
HSC numbers were unaltered. Histological analyses revealed thatthe frequencies of HSCs in contact with CXCL12-deficient CAR

BM-CAR cell-derived CXCL12 is essential for the
maintenance of HSCs and hematopoietic progenitors.
To examine the role of CXCL12 produced by BM-CAR cells,
Ebf3-CreERT2;CXCL12f/- mice were injected with tamoxifen eighttimes. qRT-PCR analysis revealed that CXCL12 was deleted from
more than 99.5% of the CAR cells (CXCL12ΔCAR mice). Flowcytometric analysis revealed that the numbers of phenotypic
HSCs, common lymphoid progenitors (CLPs), pro-B cells, pre-B
cells, mature B cells, pDCs, NK cells, granulocyte/macrophage
progenitors (GMPs), granulocytes, megakaryocyte/erythrocyte
progenitors (MEPs), and proerythroblasts were reduced in the BMof the mutants compared with control animals. The magnitude of
the reduction was greater in B cell progenitors compared with
HSCs, GMPs, and MEPs in CXCL12ΔCAR mice. Analyses based onlimiting-dilution, competitive repopulation assays revealed thatthe numbers of functional HSCs were markedly reduced in the
BM of CXCL12ΔCAR mice. These results indicate that CAR cell-

derived CXCL12 is essential for the maintenance of HSCs andhematopoietic progenitors.

The ability of HSCs to generate B cell progenitors wasmarkedly reduced in mice lacking CXCL12 in CAR cells.
Distinct subsets of HSCs, termed lymphoid-biased andmyeloid-biased HSCs, that are stably biased towards thegeneration of lymphoid or myeloid cells exist in the BM (MüllerSieburg et al., Blood 2002). A recent study showed that depletionof myeloid-biased HSCs in aged mice improved immune
responses to viral infection (Ross et al., Nature 2024). We
transplanted HSCs from CXCL12ΔCAR or control mice withcompetitor BM cells into wild-type mice and found that donor
contribution into B cell progenitors, i.e., % donor B cell progenitors
divided by % donor myeloid progenitors (donor pro-B/GMP and
pre-B/GMP reconstitution ratios), in recipients of HSCs fromCXCL12ΔCAR mice was markedly lower than that in recipients of
HSCs from control animals, indicating a stable myeloid bias of
HSCs from CXCL12ΔCAR mice. Thus, the ability of HSCs to generate
B cell progenitors was markedly reduced in CXCL12ΔCAR mice.
Together, our findings indicate that CAR cell-derived CXCL12
supports the localization of HSCs and the maintenance of HSCs,
especially lymphoid-biased HSCs, to produce the required
number of B cell progenitors.

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Lymphoid-biased
HSCs

2VWHREODVWVMyeloid progenitors

CAR cells
Foxc1hiEbf3/1hi
Runx2/1hi
CXCL12hiSCFhi

Ebf3/1CXCL12SCF(U\WKURLG0\HO
ORLG(U\WKURLG0\HORLG
SURJHQLWRUV&;&/

B cell progenitors

Runx2/1
Foxc1

+6&V

&$5 FHOO/\PSKRLG
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SURJHQLWRUVMyeloid progenitors

Myeloid-biased
HSCs)LEUREODVWV$GLSRF\WHV

Figure 1.
The functions of BM-CAR cells.
CAR cells are the major cellular component of non-hematopoietic cells in bone marrow(BM) characterized by several salient features in both mouse and human. The
transcription factors Foxc1, Ebf1/Ebf3, and Runx1/2 and cytokines, CXCL12 and SCF, arepreferentially expressed in CAR cells and critical for the formation and maintenance of
niches for HSCs and immune cells within the BM.

Figure 2.
Working model for how CXCL12 regulates HSCs to produce B cellprogenitors.
CAR cell-derived CXCL12 plays a critical role in the localization of HSCs
near their niches and the maintenance of HSCs, especially lymphoidbiased HSCs, in the BM.

Recent Publications
1. Nakatani T, Sugiyama T, Omatsu Y, Watanabe H, Kondoh G, Nagasawa T.
Ebf3+ niche-derived CXCL12 is required for the localization andmaintenance of hematopoietic stem cells. Nat Commun. 14(1):6402,(2023).
2. Omatsu Y, Aiba S, Maeta T, Higaki K, Aoki K, Watanabe H, Kondoh G,
Nishimura R, Takeda S, Chung UI, Nagasawa T. Runx1 and Runx2 inhibit
fibrotic conversion of cellular niches for hematopoietic stem cells. Nat
Commun. 13(1):2654 (2022).

4. Omatsu Y, Seike M, Sugiyama T, Kume T, Nagasawa, T. Foxc1 is a criticalregulator of haematopoietic stem/progenitor cell niche formation.
Nature. 508:536-540 (2014).
5. Sugiyama T, Kohara H, Noda M, Nagasawa T. Maintenance of thehematopoietic stem cell pool by CXCL12-CXCR4 chemokine signaling
in bone marrow stromal cell niches. Immunity 25:977-988 (2006).

3. Seike M, Omatsu Y, Watanabe H, Kondoh, G, Nagasawa, T. Stem cellniche-specific Ebf3 maintains the bone marrow cavity. Gen Dev. 32(56):359-372 (2018).

39

Immunology

Aging Biology
ileum, which in turn causes dysbiosis of the gut microbiota. This
discovery is expected to elucidate the causes of age-related
abnormalities in gut microbiota, which have long been unknown,
and to develop new methods to prevent abnormalities in gut
microbiota, thereby contributing to the extension of healthy life
expectancy.

Eiji Hara, PhD■ Professor

Eiji Hara■ Associate Professor

Shimpei Kawamoto■ Assistant Professor

Tomonori Matsumoto
Masahiro Wakita
Koyu Ito
Yumi Katayama

We all experience aging, a phenomenon in which our biological
functions deteriorate. We all want to avoid aging because it leads
to a decline in quality of life and the development of severe agerelated diseases such as cancer and dementia. Therefore, researchhas been conducted in many countries with aging populations,
including Japan, to extend a healthy life span by slowing theaging process. Recently, it has become clear that one of thecauses of aging is the increase of senescent cells, whichaccumulate in the body with age and secrete various inflammatory
substances, thereby accelerating the aging process. However, it
has yet to be understood why senescent cells accumulate in the
body with aging.
The gut microbiota in our intestinal tract, composed of anenormous number and variety of gut bacteria, plays an essential
role in maintaining our health. It has become clear thatunfavorable changes in the composition of the gut microbiota-dysbiosis- with aging are closely related to the development ofvarious age-related diseases. However, little is known about why
the dysbiosis of gut microbiota occurs with aging. We usedgenetically engineered mice in which senescent cells could be
visualized alive in vivo and kept them in a typical environment
with bacteria in the gut or a sterile environment with no bacteria
in the body for an extended period to observe changes in theaccumulation of senescent cells over time. As a result, we noticed
that mice bred in a normal environment showed an increase insenescent cells in the abdomen with aging, while mice bred in asterile environment showed no increase in senescent cells.
Furthermore, when the organs isolated from these mice were
examined, significant differences were observed mainly in theileal part of the gut. We then used single-cell RNA sequencing
analysis and other techniques to determine which cells in the40■ Postdoctoral Fellow

1■ Research Assistant3■ Support Staff

2

ileum are inducing cellular senescence. We found that a part ofgerminal center B cells, which are responsible for immunoglobulin
A (IgA) production in the gut, causes cellular senescence. IgA
regulates the composition of the gut microbiota by binding to
gut bacteria. Therefore, we hypothesized that cellular senescenceof germinal center B cells in the ileum with aging might lead todecreased IgA production and abnormal intestinal microflora.
We thus followed the same individual mice for two years andanalyzed in detail the age-related changes in IgA production andgut microbiota. The results showed that the production of IgA
decreases with age, and the binding of IgA to gut bacteriachanges, leading to a dysbiosis of gut microbiota. Furthermore,
the age-related decline in IgA production and diversity observedin wild-type mice was suppressed in mice where cellularsenescence was prevented. In addition, it was found that B cells
isolated from wild-type mice show a decrease in IgA production
capacity and a marked reduction in the ability to control the gut
microbiota as they age, while B cells isolated from mice with
reduced susceptibility to cellular senescence show no decrease in
IgA production capacity and maintain some ability to control thegut microbiota even as they age.
In conclusion, it is clear that in young mice, gut homeostasis is
maintained by controlling the composition of the gut microbiota
through IgA (Fig. 1 left), but when stimulated by gut bacteria overa long period during aging, the germinal center B cells in theileum cause cellular senescence, resulting in a decrease in IgAquantity and diversity, which in turn provokes dysbiosis in thegut microbiota (Fig. 1 right). This study revealed that the gut
microbiota, which is supposed to have a symbiotic relationshipwith the host, becomes stressful for the host in the long term,
inducing cellular senescence of germinal center B cells in the

Figure.
A model of IgA-mediated crosstalk between the gut microbiota and cellular senescence. In young mice, GC B cells induced by the gut microbiota
are appropriately selected in the PPs and differentiate into IgA plasma cells, secrete sufficient amounts of gut bacteria-specific IgA into the gut
lumen to regulate the gut microbiota and maintain symbiosis (left). However, during the ageing process, cellular senescence is induced in the GC
B cells of PPs and ILFs by continuous stimulation of the gut microbiota, leading to a decrease in the quantity and quality of IgA produced. This
provokes changes in the gut microbiota known as dysbiosis (right).

Recent Publications
1. Kawamoto S, Uemura K, Hori N, Takayasu L, et al. Bacterial induction of
B-cell senescence promotes age-related changes in the gut
microbiota. Nat Cell Biol. 25:865-876 (2023).

3. Okumura S, Konishi Y, Narukawa M, et al. Gut bacteria identified incolorectal cancer patients promote tumourigenesis via butyrate
secretion. Nat Commun. 12:5674 (2021).

2. Tsuji S, Minami S, Hashimoto R, et al. ARS-CoV-2 infection triggers
paracrine senescence and leads to a sustained senescence-associatedinflammatory response. Nat Aging. 2:115-124 (2022).

4. Wakita M, Takahashi A, Sano O, et al. A BET family protein degrader
provokes senolysis by targeting NHEJ and autophagy in senescentcells. Nat Commun. 11:1935 (2020).

41

Immunology

Signal Transduction

Nobuyuki Takakura, MD/PhD■ Professor

Nobuyuki Takakura■ Assistant Professor

Fumitaka Muramatsu
Weizhen Jia
Bal Zeynep
Keigo Akuta

Blood vessel formation is essential for organogenesis andorgan integrity. Process of blood vessel is divided into two
mechanisms. One is vasculogenesis, a do novo blood vessel
formation usually observed in embryos. In this process,mesodermal cells differentiate into endothelial cells (ECs) andECs form tube. Mural cells are recruited near ECs and adhere to
ECs for the structural stabilization. After birth, new blood vessels
are usually developed from preexisting blood vessels by the
sprouting angiogenesis. There are more than three types of ECs
for sprouting angiogenesis. Initially, tip ECs emerge for theguidance of migration direction of new branch. This cell typedoes not have ability to proliferate. Beneath tip ECs, stalk ECs
adhere and they have ability to generate high amount of ECs forelongation of new branch. Finally, phalanx ECs emerge to induce
maturation of new blood vessels by cell-cell adhesion throughVE-cadherin. Moreover, ECs from arteries, veins and capillaries are
genetically different. These suggest that ECs have heterogeneity;
however, how such heterogeneity of ECs is induced has not been
elucidated.
Tissue-resident stem cells play essential roles in tissue
homeostasis and regeneration. We have previously reported that
CD157-positive ECs in mice possess a vascular endothelial stem
cell potential, contributing to physiological turnover and■ Postdoctoral Fellow

2■ Research Assistant6■ Support Staff

2

regeneration of the vasculature in the liver. Recently, we also
found that ECs in the brain were divided into CD157-positive or-negative cells. We found that CD157-positive ECs showed higher
proliferation and were beneficial compared to CD157-negativeECs upon inoculation in a chronic cerebral hypoperfusion model.
We propose novel methods to improve the symptoms of chronic
cerebral hypoperfusion using CD157-positive ECs (Matuis et al.
Commun biol 2024). In this study, transplantation of multiple
organ-derived ECs into a chronic cerebral hypoperfusion modelin mice revealed the superior efficacy of brain-derived ECs. We
found that brain-derived vascular ECs express more Unc 5B thanother organs and angiogenesis was activated by netrin-1,
suggesting that the netrin-1 and Unc 5B systems may be involvedin brain-specific vascular system formation.
Moreover, we investigated transcription factors involved in thedevelopment or maintenance of vascular endothelial stem cells.
We found that six transcription factors (ATF3, Bhlhe40, Egr1, Egr2,
Elf3 and Klf4) increased CD157 in ECs and induced proliferation,
angiogenesis, and drug resistance in these cells. These findings
indicated that such transcription factors are useful clues forregenerative medicine relating to blood vessels and forunderstanding behavior of ECs (Konishi et al. Mol Cell Biol 2024).

Figure.
Fluorescence images of vasculature transplanted with GFP-positive ECs derived from brains of green
mice. We transplanted ECs into brains directly in a mouse model of chronic cerebral hypoperfusion.
Blood flow was overlapped by labeling with AngioSPARK 680. Such trafficking was poor for liverderived ECs and moderate for adipose-derived ECs. In addition, we injected tetramethylrhodaminelabeled dextran intravenously into each transplanted mouse and detected vascular permeability.
Leakage was not observed from vessels generated by the transplantation of brain ECs. However,leakage occurred from vessels generated by the transplantation of fat or liver-derived ECs. Taken
together, we concluded that brain-derived ECs generated adequate brain-specific blood vessels
upon transplantation and that ECs from organs other than the brain may not fully function as cerebral
vessels when transplanted ((Matuis et al. Commun biol 2024)).

Recent Publications
1. Matsui Y, Muramatsu F, Nakamura H, Noda Y, Matsumoto K, Kishima H,
Takakura N. Brain-derived endothelial cells are neuroprotective in achronic cerebral hypoperfusion mouse model. Commun Biol. 7:338(2024).

4. Naito H, Iba T, Wakabayashi T, Tai-Nagara I, Suehiro JI, Jia W, Eino D,
Sakimoto S, Muramatsu F, Kidoya H, Sakurai H, Satoh T, Akira S, Kubota
Y, Takakura N. TAK1 prevents endothelial apoptosis and maintains
vascular integrity. Dev Cell 48, 151-166 (2019).

2. Jia W, Kong L, Kidoya H, Naito H, Muramatsu F, Hayashi Y, Hsieh HY,
Yamakawa D, Hsu DK, Liu F-T, and Takakura N. Indispensable role of
Galectin-3 in promoting quiescence of hematopoietic stem cells. Nat
Commun. 12:2118 (2021).

5. Wakabayashi T, Naito H, Suehiro JI, Lin Y, Kawaji H, Iba T, Kouno T,
Ishikawa-Kato S, Furuno M, Takara K, Muramatsu F, Weizhen J, Kidoya
H, Ishihara K, Hayashizaki Y, Nishida K, Yoder MC, Takakura N. CD157
marks tissue-resident endothelial stem cells with homeostatic andregenerative properties. Cell Stem Cell. 22:384-397 (2018).

3. Kidoya H, Muramatsu F, Shimamura T, Jia W, Satoh T, Hayashi Y, Naito H,
Kunisaki Y, Arai F, Seki M, Suzuki Y, Osawa T, Akira S, Takakura N.
Regnase-1-mediated post-transcriptional regulation is essential forhematopoietic stem and progenitor cell homeostasis. Nat Commun.
10:1072 (2019).

42

43

Immunology

Cutaneous Immunology
autoantibodies against the α6β4 integrin's extracellular domain
in some patients, and these autoantibodies were capable ofinhibiting the binding of laminins 511 and 332 to α6β4 integrin.
Clinically, high antibody titres correlated with epithelial
exfoliation in both skin and small intestine, with reductions in

antibody levels corresponding to improvements in skin andgastrointestinal symptoms. This study highlights the significant
inhibitory effect of anti-α6β4 integrin autoantibodies on integrinlaminin binding and their broader systemic impact, affecting
multiple epithelial tissues.

Manabu Fujimoto, MD/PhD
Manabu Fujimoto■ Research Assistant4■ Support Staff

1

Increased anti-oxidative action compensates forcollagen tissue degeneration in vitiligo dermis

CD69 Is Indispensable for Development of Functional
Local Immune Memory in Murine Contact Hypersensitivity

Vitiligo, a common disorder causing loss of skin pigment,shows enhanced skin tightness in affected areas compared tonormal skin. Our study reveals that despite high oxidative stressin vitiligo, collagen production is maintained, with increased
expression of collagen-related genes and antioxidants in vitiligoderived fibroblasts. We observed more collagen fibers in vitiligo
lesions, decreased collagen degradation due to reduced matrix
metalloproteinases activity, and lower oxidative stress markers.
The NRF2 pathway, crucial for combatting oxidative stress, wasalso upregulated. These findings suggest that both anti-oxidative
actions and collagen integrity are preserved in vitiligo, offering
insights into its biochemical resilience.

This study explores the role of cell-surface molecules CD69 andCD103 in skin immune memory using a contact hypersensitivity(CHS) model in mice. By employing CD69-deficient (CD69KO) andCD103-deficient mice, the research observed the effects of thesemolecules on resident memory T (TRM) cells upon antigen reexposure. Findings indicate that while CD103 deficiency does not
significantly impair CHS, the absence of CD69 leads to a weakened
CHS response upon repeated challenges. Despite thedevelopment of TRM cell-like CD8 T cells at the antigen exposuresites in CD69KO mice, these cells fail to function as effective TRM
cells, as evidenced by diminished CHS responses mitigated by
CD8 neutralization or FTY720, implicating the involvement ofcirculating CD8 T cells. Additionally, macrophage infiltration andchemokine levels (Cxcl1, Cxcl2) were reduced in CD69KO
rechallenged sites, highlighting CD69's crucial role in facilitating
effective immune recall through TRM cell functionality andmacrophage recruitment.

IL-10–Producing Potency from Blood B Cells Correlateswith the Prognosis of Alopecia Areata
We investigated the role of B cells in the pathogenesis ofalopecia areata (AA), an autoimmune disorder that causes hairloss. While cytotoxic T cells have been identified as a major factor,
the involvement of B cells had not been studied. We found that B
cells from patients with AA produced more of the antiinflammatory cytokine IL-10 than B cells from healthy individuals.
When AA patients were divided into good prognosis and poor
prognosis groups based on hair regrowth, B cells from the good
prognosis group produced more IL-10 than those from the poor
prognosis group. CD8 T cells of patients with poor prognosis also
showed reduced responsiveness to B cell-induced upregulationof IFN-g–producing activity.

44■ Professor

Anti-α6β4 integrin autoantibodies inhibit the bindingof laminins to α6β4 integrin in patients with
pemphigoid and affect the gastrointestinal tract
This study investigates the role of anti-α6β4 integrin
extracellular domain autoantibodies in patients with pemphigoid,
exploring their impact on laminin-α6β4 integrin interactions andsystemic effects. It involved analyzing sera from 20 pemphigoid
patients to identify these autoantibodies and assess their ability
to inhibit laminin binding to α6β4 integrin, expressed in its active
conformation. The results confirmed the presence of IgG

Figure.
The infiltration of macrophages is impaired after the second CHS in CD69KO mice. (a) The relative expression levels of the indicated genes
evaluated by qPCR from day 29 ears of first CHS (denoted as L, white bars) and second CHS (denoted as R, gray bars) in WT (white dots) and69KO (red dots) mice, normalized to the housekeeping gene. n = 5–6 per group. (b) Top: representative immunofluorescence results
showing F4/80+ (left) and Ly6G/Ly6C+ (right) cells in day 29 right ears of WT and 69KO mice. Bar = 50 μm. Bottom: graphs showing thenumbers of F4/80+ (left) and Ly6G/Ly6C+ (right) cells per unit area in day 29 right ears of WT and 69KO mice. n = 3–5 per group. Each dot
represents the average from 4-unit areas of 3 specimens. CHS, contact hypersensitivity; NS, nonsignificant; WT, wild type.

Recent Publications
1. Connolly CM, et al. Idiopathic inflammatory myopathies: current
insights and future frontiers. The Lancet Rheumatology. Feb 6(2). doi:
10.1016/S.2665-9913(23)00322-3 (2024).

4. Nakai S, et al. CD69 Is Indispensable for Development of Functional
Local Immune Memory in Murine Contact Hypersensitivity. J Invest
Dermatol. Dec 21:S0022-202X(23)03190-1 (2023).

2. Arase N, Sasaoka Y, Narita J, Kiyohara E, Hashimoto K, Shinzaki S,
Nojima S, Takagi J, Fujimoto M. Anti-α6β4 integrin autoantibodies
inhibit the binding of laminins to α6β4 integrin in patients with
pemphigoid and affect the gastrointestinal tract. J Eur Acad Dermatol
Venereol. 38(2):404-412 (2024).

5. Matsumura Y, et al. IL-10Producing Potency from Blood B Cells
Correlates with the Prognosis of Alopecia Areata. J Invest Dermatol.
143(5):871-874.e5 (2023).

3. Wang WN, et al. Distinct Transcriptional Profiles in the Different
Phenotypes of Neurofibroma from the Same Subject with
Neurofibromatosis 1. J Invest Dermatol. 144(1):133-141 (2024).

45

Immunology

Innate Immune Systems
Rag2-/- mice as a novel mouse model for fibrosis research.

Kazuyo Moro, DDS/PhD■ Professor

Kazuyo Moro

■ Associate Professor

Yasutaka Motomura■ Assistant Professor

Takuya Yashiro
Satoshi Koga

Clinically, interstitial lung diseases (ILDs) are classified into two
main categories: idiopathic interstitial pneumonias (IIPs) andsecondary ILDs. IIPs can be further subdivided into seven diseases.
Pulmonary fibrosis is a specific type of ILD characterized byextracellular matrix deposition (such as collagen) in the alveolar
interstitium, causing dyspnea. Among the IIPs, idiopathic
pulmonary fibrosis (IPF), a progressive disease of unknown cause,
has poor prognosis, with no effective treatment. Although thepathomechanism of IPF is not yet fully elucidated, tissue repair in
response to repeated inflammation is thought to be involved inits development.
This year, we reported a novel mouse model for fibrosis
capable of elucidating the pathogenesis of pulmonary fibrosis
induced by internal factors. Fibrosis is a complex disease triggeredby both actors (such as smoking and environmental agents) andinternal factors (such as aging and autoimmune diseases) (Figure1). Previous studies predominantly utilized bleomycin-induced
mouse models to investigate fibrosis. Bleomycin induce DNA
damage in epithelial cells, leading to the activation of T cells,
neutrophils, and macrophages, ultimately resulting in TGFβdependent fibrosis (Figure 2 left). However, it has been recognizedthat this model is not ideal for comprehending fibrosis induced
by internal factors.
We discovered that Ifngr1-/-Rag2-/- mice, lacking the mechanisms
to suppress group 2 innate lymphoid cells (ILC2s), spontaneously
developed severe pulmonary fibrosis in an age-dependent
manner. The lungs of young Ifngr1-/-Rag2-/- mice showed normal
structures but displayed cellular infiltration with fibrin
accumulation at 15 weeks of age and obvious fibrosis was

46■ Postdoctoral Fellow

2■ Research Assistant1■ Support Staff

2

Figure 1.
External and internal risk factors for pulmonary fibrosis.

observed after 20 weeks of age. We expected that a detailed
comparison of the inflammatory and fibrotic phases of Ifngr1-/Rag2-/- mice, in which fibrosis develops without external stimuli,
would elucidate the mechanism of fibrosis caused by endogenous
factors.
Levels of surfactant protein D (SP-D), a common clinicalbiomarker reflecting disease activity, increased and the saturationof percutaneous oxygen (SpO2) levels significantly decreased inaged mice. In these mice, the IL-33hiIL-13hi ILC2 subpopulation
increased during the disease-onset phase before collagenproduction commenced. Although ILC2s are normally localizednear bronchioles and blood vessels, ILC2s were increased infibrotic areas along with IL-33 positive fibroblasts during fibrosis.
Defect in ILCs and IL-33 in Ifngr1-/-Rag2-/- mice prevented thedevelopment of fibrosis, indicating that IL-33-mediated activationof ILC2s is critical for fibrosis. Because Ifngr1-/-Il2rg-/-Rag2-/- micecongenitally lack ILC2s, we also confirmed that the acquired
elimination of ILC2s by neutralizing antibodies (anti-Thy-1
antibody) prevent fibrosis. ILC2s were found to directly induce
collagen production by fibroblasts in vitro, and pathogenic
fibroblasts began producing IL-33 in the chronic phase,
presumably establishing a positive feedback loop between
fibroblasts and ILC2s leading to irreversible fibrosis (Figure 2
right).
Finally, the increased expression levels of IL1RL1 (IL-33R) and
IL13, along with decreased expression of IFNGR1 were confirmed
in ILC2s from idiopathic pulmonary fibrosis patients, suggestingthat dysregulation of ILC2s may also cause endogenous fibrosis
in humans. These results highlighting the applicability of Ifngr1-/-

Figure 2.
The novel fibrosis model we present in this study consists of a mouse model in which fibrosis isinduced by the deletion of two genes, Ifngr1 and Rag2. In Ifngr1-/-Rag2-/- mice, ILC2s directly influence
fibroblasts, leading to the induction of collagen production. In addition, in the late stages of thedisease, fibroblasts show increased IL-33 expression, suggesting that a feedback loop between ILC2s
and IL-33 contributes to fibrosis exacerbations.

Recent Publications
1. Otaki N, Motomura Y, Terooatea T, Thomas Kelly S, Mochizuki M,
Takeno N, Koyasu S, Tamamitsu M, Sugihara F, Kikuta J, Kitamura H,
Shiraishi Y, Miyanohara J, Nagano Y, Saita Y, Ogura T, Asano K, Minoda
A, Moro K. Activation of ILC2s through constitutive IFNγ signaling
reduction leads to spontaneous pulmonary fibrosis. Nat Commun
14:8120 (2023).

3. Kobayashi T, Moro K. Tissue-Specific Diversity of Group 2 Innate
Lymphoid Cells in the Skin. Front Immunol 13:885642 (2022).

2. Irie M, Kabata H, Sasahara K, Kurihara M, Shirasaki Y, Kamatani T, Baba
R, Matsusaka M, Koga S, Masaki K, Miyata J, Araki Y, Kikawada T, Kabe Y,
Suematsu M, Yamagishi M, Uemura S, Moro K, Fukunaga K. Annexin A1
is a cell-intrinsic metalloregulator of zinc in human ILC2s. Cell rep
42:112610 (2023).

47

Immunology

Human Single Cell Immunology

James Wing, PhD■ Associate Professor

James Wing

■ Assistant Professor

Jonas N. Søndergaard■ Postdoctoral Fellow

3■ Research Assistant2■ Visiting Scientist

1■ Support Staff

1
Figure 1.
Schematic of “mass phenotyping” workflow, a novel system allowing hundreds of samples to be run
through a tri-panel design for detailed assessment of CD3+ (T-cells), CD3- (B, NK monocyte and DC)
and a unmanipulated linage panel. This system allows hundreds of samples to be captured in highresolution.

The human immune system is highly complex, comprising ofmany interacting cell types. While approaches such as single cell
sequencing (scRNAseq) have proven a critical advance in ourability to resolve this complexity, they are limited by the numberof cells they can utilise making it difficult to see rare cell types or
examine multiple samples over time. In contrast mass cytometry(CyTOF) can examine larger numbers of samples but is limited toonly 40 or so markers, which is insufficient to examine multiple
immune compartments in sufficient detail. To address this, wemade a novel workflow for mass cytometry “Mass phenotyping”
using this approach we can apply multiple specialised antibody
panels to individual samples to resolve over a hundred immune
markers in hundreds of human samples (Figure 1).
Using this approach, we examined large longitudinal cohorts
of COVID-19 or Bacterial sepsis patients and mRNA vaccine
recipients. This allowed us a wide overview of how the humanimmune system changes over time in these settings and a finecontrast of their similarities and differences using “Massphenotyping” to screen hundreds of samples and single cell RNA
sequencing to further investigate key differences. We primarily
focused on the specialised CD4 helper cells known as Tfh and Tph
and their interaction with antibody producing B-cells. During
infection and vaccination, B-cells play a crucial role in producing
antibodies and providing protection. However, our understandingof which B-cell types are most important remains incomplete.
Previously, "classical memory" B-cells interacting with Tfh helper
cells have been considered the primary responders to vaccines
and infections, but this notion has been limited by technicalchallenges that have hindered in-depth analysis and the ability to
examine large sample numbers over extended periods. By

48

applying our new methods to on large cohorts of COVID-19
patients and mRNA vaccine recipients, we discovered that several
forms of non-classical memory B-cells constituted the majority of
SARS-CoV2-specific B-cells at different time points. During theacute phase, we identified a novel subset termed "Activatedatypical" B-cells, which dominated the SARS-CoV2-specific B-cell
response in both COVID-19 infection and mRNA vaccination.
These cells were expanded at the same time as interferon
producing Tfh subgroup known as Tfh1. In contrast to the acutephase, we observed a distinct pattern during the resting phase of
the immune response. Our analysis revealed that a separate
group of CD23+IL4R+ non-classical B-cells contained the majorityof SARS-CoV2-specific memory (Figure 2). This finding highlights
the heterogeneity of B-cell subsets involved in maintaining longterm immunity against SARS-CoV-2 and suggests that nonclassical memory B-cells may play a more significant role thanpreviously appreciated. The identification of these CD23+IL4R+
non-classical B-cells as key contributors to SARS-CoV2-specificmemory provides valuable insights into the mechanisms
underlying long-term protection against the virus and may have
implications for the development of more effective vaccination
strategies. Since this is the first description of a role for both
Activated atypical memory and CD23+IL4R+ memory cells in
vaccination and infection this is a major finding that changes ourunderstanding of what types of cells respond to mRNA
vaccination and may prove critical to the development of more
effective vaccinations in the future.
This work is currently available as a preprint (Priest et al,
Research Square, 2024).

Figure 2.
Total and SARS-CoV2 specific B-cells following vaccination or SARS-CoV2 infection (COVID-19). While“classical” memory B-cells are the majority of B-cell memory we find that non-classical and atypicalB-cells dominate the SARS-CoV2 response.

Recent Publications
1. Okamoto M, Sasai M, Kuratani A, Okuzaki D, Arai M, Wing JB, Sakaguchi
S, Yamamoto M. A genetic method specifically delineates Th1-type
Treg cells and their roles in tumour immunity. Cell Rep. 42:7 (2023).

4. Shirai T, Nakai A, Ando E, et al Celastrol suppresses humoral immune
responses and autoimmunity by targeting the COMMD3/8 complex.
Sci Immunol. 8 (81) eadc9324. (2023).

2. Morita R, Kubota-Koketsu R, Lu X, Sasaki T, Nakayama EE, Liu Y-C,
Okuzaki D, Motooka D, Wing JB, et al. COVID-19 relapse associatedwith SARS-CoV-2 evasion from CD4+ T-cell recognition in an
agammaglobulinemia patient. iScience. 26:5 (2023).

5. Søndergaard JN, Tulyeu J, Edahiro R, …, Wing JB. A sex-biased
imbalance between Tfr, Tph and atypical B cells, determines antibody
responses in COVID-19 patients. Proc Natl Acad Sci USA. 120 (4)e2217902120 (2023).

3. Edahiro R, Shirai Y, Takeshima Y, et al. Single-cell analyses and hostgenetics highlight the role of innate immune cells in COVID-19
severity. Nat Gen. 55:753–767 (2023).

49

Immunology

Human Immunology (Single Cell Genomics)

Daisuke Okuzaki, PhD

We discovered a novel public antibody clonotype, namely PAN-CoV1804, that reacts with both the SARS-CoV-2 N protein andseveral self-antigens. We found that the PA-N-CoV1804 clonotype
underwent robust clonal expansion in a subset of COVID-19
patients. PA-N-CoV1804 was expressed exclusively inplasmablasts from COVID-19 patients. PA-N-CoV1804 harbors
numerous somatic mutations, similar to antiviral antibodies
derived from pre-existing memory cells for seasonal humancoronaviruses. However, this clonotype was strongly reactive to
the N protein of SARS-CoV-2 but not to that of seasonalcoronaviruses. Therefore, these clones appear to originate fromnaive B cells and develop through de novo clonal expansion after
SARS-CoV-2 infection. A manuscript describing this discovery iscurrently under peer review process.
In a joint research project conducted with Critical Care
Medicine at Osaka University, a platform was established to serve
as a guide for determining the necessity and duration of
mechanical ventilation for severe COVID-19 patients admitted to
the ICU at Osaka University Hospital. In a latent class analysis ofwhole blood RNA collected and sequenced from 40 patients, 45
genes were identified as effective markers for classifying thepatients into three classes. Interaction between microRNA andmRNAs was also studied through simultaneous measurement of
the mRNAs and microRNAs in the whole blood of severe COVID-19
patients. The integrated analysis revealed that, compared tointerferon signaling in healthy subjects, interferon signaling wasmore activated in the COVID-19 patients.
By the end of the pandemic, our research focus shifted to theimmune response following vaccination. In case of encephalitis

50■ Associate Professor

Daisuke Okuzaki■ Assistant Professor

Yuchen Liu■ Postdoctoral Fellow

2■ Visiting Scientist

2■ Support Staff

1

temporally associated with COVID-19 vaccination, single-cell
RNA sequencing (scRNA-seq) analysis was applied to elucidate
the distinct immune signature in the peripheral immune system.
Peripheral blood mononuclear cells (PBMCs) were analyzed using
scRNA-seq to clarify the cellular components of the patients inboth obtained data were compared to those acquired from ahealthy cohort. The scRNA-seq analysis identified a distinct
myeloid cell population in PBMCs during the acute phase of
encephalitis. This specific myeloid population was neither
detected in the remission phase of the disease nor in the healthy
cohort. Our findings illustrate the induction of a unique myeloid
subset in encephalitis temporally associated with COVID-19
vaccination. Further research into the dysregulated immune
signature of COVID-19 vaccination-associated autoimmunity,
including the cerebrospinal fluid (CSF) cells of central nervous
system (CNS), is required to clarify the pathogenic role of thismyeloid subset. The discovery was published in Frontiers in
Immunology Feb. 23, 2023.
During the pandemic, we conducted non-COVID-related
research that studied the transcriptome of Vibrio parahaemolyticus
using a new long-read sequencing method, known as direct RNA
sequencing. We developed a pipeline to process the data,
revealing a more complex transcriptome landscape thanpreviously understood, not only for Vibrio but for bacteria ingeneral. Additionally, our original studies on developing amethod for simultaneous detection of bacteria and eukaryotic
cells at the single-cell level, and for the detection of circRNA using
long-read sequencers, have the potential to advance research inmany fields.

Recent Publications
1. Kurosu T, Okuzaki D, Sakai Y, et al. Dengue virus infection induces
selective expansion of Vγ4 and Vγ6TCR γδ T cells in the small intestine
and a cytokine storm driving vascular leakage in mice. PLoS Negl Trop
Dis. 17(11): e0011743 (2023).

4. Ishikawa M, Shimada Y, Ozono T, et al. Single-cell RNA-seq analysis
identifi es distinct myeloid cells in a case with encephalitis temporally
associated with COVID-19 vaccination. Front Immunol. 14:998233(2023).

2. Edahiro R, Shirai Y, Takeshima Y, et al. Single-cell analyses and hostgenetics highlight the role of innate immune cells in COVID-19
severity. Nat Genet. 55(5):753-767 (2023).

5. Yamaguchi Y, Kato Y, Edahiro R, et al. Consecutive BNT162b2 mRNA
vaccination induces short-term epigenetic memory in innate immune
cells. JCI Insight. 7(22):e163347 (2022).

3. Namkoong H, Edahiro R, Takano T, et al. DOCK2 is involved in the hostgenetics and biology of severe COVID-19. Nature. 609(7928):754-760(2022).

51

Immunology

Immune Homeostasis

Yasutaka Okabe, PhD

Intra-abdominal infection is the second most common causeof life-threatening sepsis, and it requires prompt medical
treatment. Infection in the abdominal cavity (also reffered as theperitoneal cavity) can be caused by a pathological or traumatic
loss of intestinal wall integrity, cirrhosis, pancreatitis, abdominal
surgery, or peritoneal dialysis.
Macrophages in the peritoneal cavity are crucial component of
abdominal immunity. We found transcription factor GATA6 isselectively expressed in macrophages in the peritoneal cavity. We
reported that GATA6 acts as a master transcriptional regulator for
functional specialization of peritoneal macrophages, as thedeletion of GATA6 gene in peritoneal macrophages resulted in
the defect of peritoneal specific gene expression program. Thus,
GATA6-mediating program control macrophage positioning
within peritoneal cavity, local proliferation, and peritoneal
specific immune responses. In addition, we reported that retinoic
acid, which is a lipophilic molecule derived from vitamin A, playessential role in the induction of GATA6 gene in peritoneal
macrophages. We found retinoic acid nuclear receptors (RARs)
bind to the promoter of GATA6 gene in retinoic acid dependent
manner. In addition, the treatment of RAR inverse agonist(BMS493) abolished the expression of GATA6 gene in peritoneal
macrophages. Lastly, vitamin A deficiency of mice caused toreduce GATA6 gene expression.

■ Associate Professor

Yasutaka Okabe■ Postdoctoral Fellow

1■ Research Assistant2■ Support Staff

1

In addition to macrophages, the milky spots in omentum are
atypical lymphoid tissues that play a pivotal role in regulating
immune responses in the peritoneal cavity. The milky spots serve
as a central gathering places for collecting antigens and particles
from the peritoneal cavity, regulating the movement oflymphocytes, promoting the differentiation and self-renewal of
immune cells, and supporting the local germinal center reaction.
In addition, the milky spots exhibit unique developmental
characteristics that combine the features of secondary andtertiary lymphoid tissues. These structures are innately
programmed to form during fetal development; however, they
can also be formed postnatally in response to peritoneal irritationsuch as inflammation, infection, obesity, or tumor metastasis.
However, the mechanism underlying milky spot formation ispoorly understood. We identified a subset of omental fibroblastic
reticular cells (FRCs) that are characterized by the expression ofretinoic acid converting enzyme. We found these FRCs are
uniquely present in milky spots but not in lymph nodes.
Furthermore, these FRCs are essential for the recruitment ofcirculating lymphocytes to milky spots, which is mediated by the
induction of chemokine CXCL12 in a manner dependent on
retinoic acid. Thus, our study demonstrates the stromal-immune
cell interaction in the formation of nonclassical lymphoid tissues.

Figure.
Paraffin sections ofomentum fromWT mouse were stained with hematoxylin and eosin. Scale bar: 100 μm.

Recent Publications
1. Okabe Y. Development and organization of omental milky spots.
Immunological Reviews, in press.

3. Okabe Y. Immune Niche Within the Peritoneal Cavity. Curr Topic
Microbiol Immunol. 434:123-134 (2021).

2. Yoshihara T. & Okabe Y. Aldh1a2+ fibroblastic reticular cells regulate
lymphocyte recruitment in omental milky spots. J Exp Med. 220(5):e20221813 (2023).

52

53

Immunology

Cellular Immunotherapy
00*&$57FHOOUHFRJQL]HWKH
FRQIRUPDWLRQRIDSURWHLQ

Naoki Hosen, MD/PhD■ Professor

Naoki Hosen■ Associate Professor

Hisashi Kato

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UHFRJQL]HGJO\FRIRUPV RIDSURWHLQ

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00Kentaro Fukushima

1RUPDOOHXNRF\WHV

00

1RUPDOOHXNRF\WHV

Jiro Fujita■ Assistant ProfessorűƄƗŃŰƈƇ

Michiko Ichii

ŶƆƌŃŷƕƄƑƖƏ ŰƈƇ

Yasutaka Ueda
Tomoaki Ueda
Figure 1.
Novel target antigen structures that we have identified for CAR-T cell therapy against multiple
myeloma.

Shinsuke Kusakabe
Akihisa Hino

We are focusing on cellular immunotherapy, especiallychimeric antigen receptor (CAR)-T cell therapy for cancers. cells
specifically recognize and activate cancer cells using the cancerspecific antigen recognition domain derived from mAbs.
Activated CAR-T cells kill tumor cells and also proliferate
extensively. CD19 CAR-T cells showed surprisingly high efficacy
against acute lymphoblastic leukemia and malignant lymphoma.
We discovered that the active conformer of an integrin 7 could
serve as a specific therapeutic target for multiple myeloma (MM),an incurable hematologic cancer characterized by theaccumulation of neoplastic plasma cells in the bone marrow(BM). The clinical trial of the CAR T-cell targeting the active
conformation of the integrin 7 for MM is now underway. We
screened more than 10,000 mAb clones raised against MM cells
and identified R8H283 as an mAb that bound to MM cells but not
to normal hematopoietic or non-hematopoietic cells. R8H283
specifically recognized CD98hc. R8H283 did not react with
CD98hc monomer, but bound to CD98hc forming heterodimers
with the light chains, which are amino acid transporters. MM cells
expressed abundant CD98 heterodimers to take up amino acids
for constitutive immunoglobulin production. Although CD98
heterodimers were also expressed in normal leukocytes, R8H283
did not react with them. Normal leukocytes expressed CD98hc
glycoforms different from those expressed in MM cells, which
may account for the lack of R8H283 reactivity in normal
leukocytes. R8H283 exerted significant anti-MM effects withoutharming normal hematopoietic cells.

■ Visiting Scientist

1■ Support Staff

3

Development of CAR-T cell therapy targeting antigen
structures formed as a result of post-translational
events in various cancers
We have applied the same strategy to different types of cancer.
For hematological cancers such as acute myeloid leukemia (AML),
the only hurdle to developing CAR-T cell therapy is the lack of asuitable cancer-specific cell surface antigen. We have alreadygenerated a large number of mAbs that react with AML cells andhave identified those that specifically react with AML cells. We
have also applied the same strategy to various types of solid
tumors in collaboration with various cancer treatment
departments at Osaka University Hospital. We recently
demonstrated that our screening method also works inglioblastoma.

Development of CAR NK cells
A major problem with CAR T cells is the high cost of producing
them from autologous T cells for each patient. Since NK cells do
not cause GVHD even when derived from allogeneic donors, CAR
NK cells derived from a single donor can be used for multiple
allogeneic recipients. The efficacy of cord blood-derived CD19
CAR NK cells has recently been demonstrated. We generate cord
blood CAR-NK cells using our original CARs.

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Figure 2.
CAR-NK cells can be used for allogenic recipients.

Recent Publications
1. Fukushima K, et al. Clostridium butyricum MIYAIRI 588 contributes tothe maintenance of intestinal microbiota diversity early after
haematopoietic cell transplantation. Bone Marrow Transplant. (2024).

4. Hino A, et al. Prolonged gut microbial alterations in post-transplantsurvivors of allogeneic haematopoietic stem cell transplantation. Br J
Haematol. (2022).

2. Nakagawa T, et al. Identification of glioblastoma-specific antigens
expressed in patient-derived tumor cells as candidate targets forchimeric antigen receptor T cell therapy. Neurooncol Adv. 5:vdac177(2023).

5. Hasegawa K, et al. Selective targeting of multiple myeloma cells witha monoclonal antibody recognizing the ubiquitous protein CD98
heavy chain. Sci Trans Med. 14:eaax7706 (2022).

3. Uchihara Y, et al. DNA damage promotes HLA class I presentation bystimulating a pioneer round of translation-associated antigen
production. Mol Cell. 82:2557-2570 e2557 (2022).

54

55

Immunology

Microbiology and Immunology

Nobuhiko Kamada, PhD■ Professor

Nobuhiko Kamada■ Associate Professor

Sho Kitamoto■ Assistant Professor

Kohei Sugihara

Hiroko Kitamoto

Our team has been studying the role of microbiota in thepathogenesis of gastrointestinal diseases, such as inflammatory
bowel disease (IBD) and colorectal cancer (CRC). It has beenreported that certain pathogenic members of commensal
bacteria (namely ‘pathobionts’) are enriched under disease
conditions and contribute to disease pathogenesis. However, theprecise mechanisms by which such pathobionts thrive in disease
conditions and trigger and/or exacerbate disease remain
incompletely understood.
We demonstrated that L-serine metabolism plays a vital role inthe survival of some pathobionts, including adherent–invasive
Escherichia coli, a pathobiont associated with IBD. AIEC utilizes
L-serine supplied by dietary protein intake in the gut lumen.
Therefore, deprivation of dietary L-serine can suppress the
expansion of AIEC in the gut. However, certain commensal
bacteria can help AIEC overcome the dietary nutrient limitation.
The presence of mucolytic bacteria allows AIEC to grow evenunder the restriction of dietary L-serine. Mucolytic bacteria, suchas Akkermansia muciniphila, degrade the mucus layer andfacilitate the encroachment of AIEC to the epithelial niche. In theepithelial niche, AIEC acquires L-serine from the amino acid poolof the colonic epithelium, allowing them to grow even under
diet-derived L-serine restriction. Thus, the interaction between
pathobionts and mucolytic commensal bacteria helps
pathobionts switch their source of essential nutrients from thediet to host cells. In addition to AIEC for IBD, we are identifying

56■ Postdoctoral Fellow

2■ Research Assistant1■ Support Staff

1

pathobionts associated with various gastrointestinal diseases,
such as CRC and intestinal fibrosis. We aim to elucidate themechanisms by which these pathobionts adapt to diseasespecific environments. Moreover, we are identifying partner
bacteria that cooperate with pathobionts to promote diseaseprogression and suppressors that inhibit the expansion ofpathobionts. Also, we focus on the microbial and immune
connections between the oral and gut mucosae in thepathogenesis of gastrointestinal diseases. We have discoveredthat inflammation in the oral mucosa results in the outgrowth of
inflammatory oral pathobionts. Amassed oral pathobionts thennaturally translocate to the gut and contribute to thepathogenesis of gut diseases. In addition, inflammatory immune
cells arising during oral inflammation can also migrate to the gut.
Gut-migrated inflammatory T cells of oral origin are activated byectopically colonized oral pathobionts and contribute toinflammation in the gut mucosa. Klebsiella aerogenes is the most
predominant bacteria species found in the inflamed oral cavity of
periodontitis mice. We found that K. aerogenes employs chaperon
usher pili (CUP) 1 for colonization in the inflamed gut but not in
the oral mucosa. Thus, CUP1 serves as a key adhesion moleculefor site-specific adaptation of the oral pathobiont.
Note: The PI is jointly appointed at IFReC and the University ofMichigan (USA). These research projects were conducted ateither IFReC or the University of Michigan.

Figure.
Pathobionts promote various gastrointestinal diseases, including IBD, CRC, and intestinal fibrosis.
Some pathobionts are oral origins. Certain symbiotic bacteria, such as mucolytic bacteria, act as‘cooperator’ and help pathobionts acquire required nutrients. Some symbiotic bacteria may serve as‘suppressors.’ These suppressors may directly and indirectly inhibit the expansion of pathobionts andsubsequent disease progression.

Recent Publications
1. Guo Y, Kitamoto S, Caballero-Flores G, Kim Y, Watanabe D, Sugihara K,
Núñez G, Alteri CJ, Inohara N, Kamada N. Oral pathobiont Klebsiellachaperon usher pili provide site-specific adaptation for the inflamedgut mucosa. Gut Microbes. 16(1):2333463 (2024).
2. Sugihara K, Kitamoto S, Saraithong P, Nagao-Kitamoto H, Hppstal M,
McCarthy C, Rosevelt A, Muraleedharan CK, Gillilland III MG, Imai J,
Omi M, Bishu S, Kao JY, Alteri CJ, Barnich N, Schmidt TM, Nusrat A,
Inohara N, Golob JL, Kamada N. Mucolytic bacteria license pathobionts
to acquire host-derived nutrients during dietary nutrient restriction.
Cell Reports. 40(3):111093 (2022).
3. Kitamoto S, Nagao-Kitamoto H, Jiao Y, Gillilland III MG, Hayashi A, ImaiJ, Sugihara K, Miyoshi M, Brazil JC, Kuffa P, Hill BD, Rizvi SM, Wen F,
Bishu S, Inohara N, Eaton KA, Nusrat A, Lei YL, Giannobile WV, andKamada N. The intermucosal connection between the mouth and gut
in commensal pathobiont-driven colitis. Cell. 182(2):447-462 (2020).

4. Nagao-Kitamoto H, Leslie JL, Kitamoto S, Jin C, Thomsson KA, Gillilland
MG 3rd, Kuffa P, Goto Y, Jenq RR, Ishii C, Hirayama A, Seekatz AM,
Martens EC, Eaton KA, Kao JY, Fukuda S, Higgins PDR, Karlsson NG,
Young VB, Kamada N. Interleukin-22-mediated host glycosylation
prevents Clostridioides difficile infection by modulating the metabolic
activity of the gut microbiota. Nat Med. 26(4):608-617 (2020).
5. Kitamoto S, Alteri CJ, Rodrigues M, Nagao-Kitamoto H, Sugihara K,
Himpsl SD, Bazzi M, Miyoshi M, Nishioka T, Hayashi A, Morhardt TL,
Kuffa P, Grasberger H, El-Zaatari M, Bishu S, Ishii C, Hirayama A, Eaton
KA, Dogan B, Simpson KW, Inohara N, Mobley HLT, Kao JY, Fukuda S,
Barnich N, Kamada N. Dietary L-serine confers a competitive fitness
advantage to Enterobacteriaceae in the inflamed gut. Nat Microbiol.
5(1):116-125 (2020).

57

Immunology

Cutaneous Allergy and Host Defense

Yumi Matsuoka-Nakamura, MD/PhD

Our laboratory, established in November 2022, is on a mission
to unravel the complexities of inflammatory skin diseases such as
atopic dermatitis (AD) and acne vulgaris. We are also deeply
immersed in understanding how Methicillin-resistant
Staphylococcus aureus (MRSA) evolves multi-drug resistance andperpetuates persistent infections within hospital environments.
One of our pivotal research focus lies in exploring the dynamics
between the accessory gene regulator (Agr) quorum sensing (QS)
system of S. aureus and the host's innate immune response, bothat the level of the skin and systemic infections. QS, a mechanism
allowing bacteria to sense and respond to their population
density, plays a crucial role in their survival strategy by modulating
gene expression to optimize nutrient acquisition and competewith other bacteria for resources. Our investigations underscorethe significance of QS in understanding the role of pathogenic
bacteria in diseases, positioning it as a promising target for
innovative therapeutic approaches.
In an earlier study, we leveraged bacterial whole-genome
sequencing to examine S. aureus strains from the skin of Japanese
infants. Our findings indicated a higher propensity for AD
development in infants with S. aureus colonization on theircheeks. Notably, infants hosting S. aureus strains withspontaneous mutations in the agr QS system were more likely toremain healthy. This discovery points to a potential link between
S. aureus, QS functionality, and the early onset of AD.
This year, we've made significant strides by reporting theimpact of the skin microbiome and skincare interventions on AD
in a Japanese birth cohort study. Analyzing 177 infants, all58■ Professor

Yumi Matsuoka-Nakamura■ Assistant Professor

Yuriko Yamazaki■ Postdoctoral Fellow

1■ Support Staff

3

receiving skincare, we aimed to decipher the role of dysbiosis in
AD's infancy onset. By the age of one, we observed AD
development in 13 infants (7.3%), while three (1.7%) werediagnosed with a food allergy (FA), specifically to egg white (EW),with an overlap of one case also presenting AD. Furthermore, 61
infants (34.5%) without AD or FA showed sensitization to EW.
Interestingly, the EW-sensitized group was associated with ahigher use of moisturizer (TAM) and elevated serum total IgE
levels compared to their healthy counterparts. Conversely, theAD group utilized less TAM yet exhibited higher IgE levels thanhealthy infants. Our microbiome analysis did not reveal significant
diversity differences between healthy and AD groups, but it
linked early life bacterial population shifts to subsequent AD
development. Additionally, a negative correlation between TAM
use and Streptococcus sp. abundance, alongside a positive
correlation with Cutibacterium acnes, suggests that skincare
practices could beneficially alter microbial compositions,potentially reducing AD risk. These insights advocate for thepreventive potential of skincare interventions, including
moisturizer use during infancy, against AD and FA.
Currently, we are delving into how Agr-QS influences MRSA's
adaptation in hospital settings and its evolution towards
multidrug resistance. Given MRSA's global health challenge,
understanding its adaptive mechanisms in causing recurrent
disease is crucial. Our ongoing project investigates MRSA strainsfrom a neonatal intensive care unit (NICU) outbreak in a Japanesehospital, aiming to decode the bacterial attributes that facilitate
S. aureus's survival and persistence in hospital contexts.

Figure.
Overview of skin care intervention cohort study.
AD, Atopic dermatitis; HS, Healthy subject; TAM, Total amount of moisturizer used.

Recent Publications
1. Aoyama R, Nakagawa S, Ichikawa Y, Inohara N, Yamazaki Y, Ito T,
Sugihira T, Kono M, Akiyama M, Takahashi H, Takaya A, Ichikawa F,
Nakano T, Tanaka S, Koyano Y, Fujimoto M, Núñez G, Shimojo N,
Nakamura Y. Neonatal skin dysbiosis to infantile atopic dermatitis:
Mitigating effects of skin care. Allergy. (2024).
2. Yamazaki Y, Ito T, Tamai M, Nakagawa S, Nakamura Y. The role ofStaphylococcus aureus quorum sensing in cutaneous and systemic
infections. Inflamm Regen. 44(1):9 (2024).
3. Matsumoto M, Nakagawa S, Zhang L, Nakamura Y, Villaruz AE, Otto M,
Wolz C, Inohara N, Núñez G. Interaction between Staphylococcus Agrvirulence and neutrophils regulates pathogen expansion in the skin.
Cell Host Microbe. 29(6):930-940 (2021).

4. Miyachi H, Wakabayashi S, Sugihira T, Aoyama R, Saijo S, KoguchiYoshioka H, Fujimoto M, Núñez G, Matsue H, Nakamura Y. Keratinocyte
IL-36 Receptor/MyD88 signaling mediates Malassezia-induced IL-17dependent skin inflammation. J Infect Dis. 223(10):1753-1765 (2021).
5. Nakamura Y, Takahashi H, Takaya A, Inoue Y, Katayama Y, Kusuya Y,
Shoji T, Takada S, Nakagawa S, Oguma R, Saito N, Ozawa N, Nakano T,
Yamaide F, Dissanayake E, Suzuki S, Villaruz A, Varadarajan S,
Matsumoto M, Kobayashi T, Kono M, Sato Y, Akiyama M, Otto M,
Matsue H, Núñez G, Shimojo N. Staphylococcus Agr virulence is criticalfor epidermal colonization and associates with atopic dermatitis
development. Sci Transl Med. 12:551 (2020).

59

Single Molecule Imaging

Imaging

temporal precedence to get closer to identifying causal circuits.
We are also expanding this approach to develop partner rodent
studies, where we can get a much finer handle on circuitarchitecture.

Toshio Yanagida, PhD
Ben Seymour, MD/PhD■ Professor

The lab has also been involved in capacity building activities.
We have launched a fully-acredited course in neurotechnology

for chronic pain (https://cpnn.ac.uk/neurotech-course/) which
offers a series of 28 lectures and quizzes, in a free, online platform.
We have been also running a series of practical workshops, again
with UK-Japan collaboration build in. And lastly, we continue toactively pursue research that is diverse and inclusive, and inwhich the patient perspective plays a central role in the research
process.

Toshio Yanagida
Ben Seymour

How do humans and animals protect themselves so effectively
against injury and illness for their entire lives? The brain is criticalpart of the systems that guide defensive action and physiology.
This physiology involves two distinct components: behaviour
that allows you to avoid injury/infection in the first place: a key
part of which is the pain system that directs nocifensive actions(and also the closely related itch system). The second component
is the behaviour that allows you to recover from injury or infection
in a way that minimizes further harm. This system is best
considered a multi-system system that incorporates brain,
immune and endocrine axes, that work together to optimize
body defence and recovery. We propose that there is a central
homeostatic control circuit that governs this behaviour, and ourexperiments aim to discover where in the brain this is located,and how it controls physiology and behaviour. Discovering thiscircuit would open the door to a new generation of treatments ofpain, fatigue and mood disorders, and offer a fundamental new
approach to immunomodulation.
Our lab uses a combination of computational modelling,
imaging, and behavioural experiments to understand how thissystem works. We have been building increasingly sophisticatedmodels of defensive and recuperative behaviour, designed tocapture the core computations that the brain makes in the face of
threat and significant injury. We have discovered algorithms –that is, information processing systems – that can be shown to
optimize efficient behaviour (in terms of reward harvesting) in
dangerous environments (Mahajan et al, 2024). This allows, forexample, animals to forage safely for food when exploring
potentially dangerous new habitats or environments; and arises
from having multi-module value memory systems in the brain,

60

and employing a risk-sensitive decision-making. Overall, thisallows us to begin to build a picture of what is under control, andthis specificity should enable us to work backwards to identify
the proposed central control circuit involved in changes inhomeostasis (i.e. injury/illness allostasis).
Next, therefore, we have applied this to experiments probing
how behaviour changes after an injury, using various behaviouraltasks, and with concurrent neuroimaging. Using thiscomputational approach, we can set out predictions for how thehomeostatic priorities change, given optimal survival
assumptions (Seymour, Crook and Chen, 2023). We have now
applied this to experiments in humans with inflammatory arthritis(i.e. rheumatoid arthritis), and successfully shown that these
theoretical models predict how behaviour is shaped in patientswith inflammation and associated symptoms of fatigue and pain(see figure).
Using the computational signature alongside imaging data,
we can then look across the brain to identify key areas involved in
representation and control. However, as the brain is a complex
information processing network, we use graph theory models to
identify hubs of information flow. This approach has revealed akey role for the insula – an evolutionarily old brain regions closely
associated with interoception (sensing the internal body),
homeostasis and emotion; a region also closely connected to the
hypothalamus and other key homeostatic structures (Mancini etal, 2024).
The next step in our research is to try and prospectively trackthese processes using longitudinal studies, whereby we can use

Figure.
Identifying potential injury-homeostatic circuits in the brain.
a) we use functional brain imaging to study brain activity whilst patients with inflammatory arthritis perform various motivation anddecision-making tasks; b) computational analysis of behaviour yields a parameter profiles (computomic panels) that c) uniquely predict
symptoms of inflammation (pain and fatigue); and d) these can be mapped to the brain using hub centrality metrics that derive fromgraph theoretic analysis of brain information networks.

Recent Publications
1. Mahajan P, Tong S, Lee SW and Seymour B. Balancing safety andefficiency in human decision making. bioRxiv, pp.2024-01 (2024).

4. Onysk J, Gregory N, Whitefield M, Jain M, Turner G, Seymour B and
Mancini F. Statistical learning shapes pain perception and prediction
independently of external cues. eLife, 12 (2023).

2. Seymour B, Crook RJ and Chen ZS.. Post-injury pain and behaviour: acontrol theory perspective. Nat Rev Neurosci. 1-15 (2023).
3. Mancini F, Mahajan P, Guttesen AÁV, Onysk J, Scholtes I, Shenker N, Lee
M. and Seymour B. Enhanced behavioural and neural sensitivity topunishments in chronic pain and fatigue. bioRxiv, pp.2024-04 (2024).

61

Imaging

Immunology and Cell Biology

Masaru Ishii, MD/PhD■ Professor

Masaru Ishii■ Associate Professor

Yasuhito Yahara■ Assistant Professor

Yutaka Uchida
Kentaro Fujii■ Postdoctoral Fellow

3■ Research Assistant2■ Visiting Scientist

2■ Support Staff

2

Figure 1.
A dynamic cellular system inside the living body.

Crowds of people crossing streets, cars streaming along
roadways, and trains that come and go carrying passengers:
These are examples of ubiquitous, tightly organized humandynamics that support and vitalize societies. Similarly, in thehuman body, a variety of cells with different roles move andfunction according to their features and locations; collectively
they comprise complex biological systems. A typical example isthe immune system. Lymphocytes and macrophages migrate to
every region of the body and gather in specific environments to
exchange information and maintain normal immune responses.
Thus, organisms are shaped by organizational dynamics. These
dynamics are natural in life and do not occur following death.
Understanding the dynamic cellular society by elucidating how
cells move, or are moved, to maintain life is a fundamental themein the life sciences. However, researchers have only recently
succeeded in analyzing cellular movement. Recent advances in invivo fluorescence imaging technologies have enabled researchers
to look inside the bodies of live animals and analyze cellular andmolecular dynamics. In our laboratory, we have developed anovel multiphoton excitation microscopy technique for studying
the movement of immune cells in vivo using minimally invasive
observational and analytical methods that maintained
spatiotemporal information intact. Since we successfully
observed the interior of live bone tissue, we have energeticallypursued the analysis of cellular dynamics for various types of cells(Figure 1).
Especially as one of the most impacting achievements in thefiscal year of 2023, we demonstrated the spatiotemporal

62

heterogeneity of inflammatory responses in the liver. The liver isthe main gateway from the gut, and the unidirectional sinusoidal
flow from portal to central veins constitutes heterogenous zones,
including the periportal vein (PV) and pericentral vein zones;
however, the functional differences in the immune system ineach zone remain poorly understood. Here, intravital imaging
revealed that inflammatory responses were suppressed in PV
zones. The zone-specific single-cell transcriptomics detected an
immunosuppressive macrophage subset enriched in PV zones
that highly expresses IL-10 and Marco, a scavenger receptor
sequestering pro-inflammatory PAMPs/DAMPs and consequently
suppressing immune responses. Induction of the Marco+
immunosuppressive macrophages depended on gut microbiota,
and especially, a specific bacterial family, Odoribacteraceae, wasidentified to induce this macrophage subset via its postbiotic,
isoallo-lithocholic acid. Intestinal barrier leakage results ininflammation in PV zones, which was markedly augmented by
Marco-deficient conditions. Chronic liver inflammatory diseases
such as primary sclerosing cholangitis (PSC) and non-alcoholicsteatohepatitis (NASH) showed decreased Marco+ macrophages.
Functional ablation of Marco+ macrophages led to PSC-likeinflammatory phenotypes related to colitis and exacerbatedsteatosis in NASH in animal experimental models. Collectively,commensal bacteria induce Marco+ immunosuppressive
macrophages, consequently limiting excessive inflammation at
the gateway. Failure of this self-limiting system promotes hepatic
inflammatory disorders such as PSC and NASH (Figure 2).

Figure 2.
Periportal macrophages protect against commensal-driven liver inflammation.

Recent Publications
1. Miyamoto Y, Kikuta J, Matsui T, Hasegawa T, Fujii K, Okuzaki D, Liu Y-C,
Yoshioka T, Seno S, Motooka D, Uchida Y, Yamashita E, Kobayashi S,
Eguchi H, Morii E, Tryggvason K, Shichita T, Kayama H, Atarashi K,
Kunisawa J, Honda K, Takeda K, Ishii M. Periportal macrophages
protect against commensal-driven liver inflammation. Nature.
629:901–909 (2024).

3. Taniguchi S, Matsui T, Kimura K, Funaki S, Miyamoto Y, Uchida Y, Sudo
T, Kikuta J, Hara T, Motooka D, Liu Y-C, Okuzaki D, Morii E, Emoto N,
Shintani Y, Ishii M. In vivo induction of activin A-producing alveolar
macrophages supports the progression of lung cell carcinoma. Nat
Commun. 14(1):143 (2023).

2. Shimizu K, Kikuta J, Ohta Y, Uchida Y, Miyamoto Y, Morimoto A, Yari S,
Sato T, Kamakura T, Oshima K, Imai R, Liu Y-C, Okuzaki D, Hara T,
Motooka D, Emoto N, Inohara H, Ishii M. Single-cell transcriptomics ofhuman cholesteatoma identifies an activin A-producing
osteoclastogenic fibroblast subset inducing bone destruction. Nat
Commun. 14(1):4417 (2023).

63

Imaging

Chemical Imaging Technique
imaging. The study highlights the importance of NP elasticity inenhancing clearance from the body, thereby reducing potential
side effects associated with long-term accumulation. The ability
to track NP dynamics in vivo using 19F MRI offers a powerful tool

for real-time and longitudinal monitoring, which is crucial for theclinical translation of nanoparticle-based imaging agents andtherapeutics.

Kazuya Kikuchi, PhD■ Professor

Kazuya Kikuchi■ Associate Professor

Masafumi Minoshima■ Assistant Professor

Tomoya Yamamoto■ Postdoctoral Fellow

1■ Research Assistant1■ Support Staff

2

Figure 1.
(top) Schematic image of elastic polymer NPs containing PFCE prepared by polymerization on
micelle. (bottom) Size distribution of polymer-coated NPs using a nanoparticle analyzer and 19F NMR
spectrum of polymer-coated NPs containing PFCE (TFANa was used as an internal standard).

"Elastic Polymer Coated Nanoparticles with Fast
Clearance for 19F MR Imaging"
Magnetic resonance imaging (MRI) is a critical noninvasive tool
for molecular imaging, with 19F MRI being particularly
advantageous due to its high sensitivity and the 100% naturalabundance of 19F atoms. The low endogenous background signalof 19F in the body enables high-contrast imaging, making it ideal
for tracking 19F-containing probes in deep tissues and monitoring
enzyme activity in vivo. Our group has developed FLAME(FLuorine Accumulated silica nanoparticle for MRI Contrast
Enhancement), consisting of perfluorocarbon (PFC)-encapsulated
silica NPs. However, the efficacy of silica nanoparticles (NPs) as 19F
MRI probes has been hampered by their propensity to accumulate
in the liver, leading to prolonged retention and potential side
effects such as chronic inflammation and toxicity. To address thislimitation, elastic nanomaterials, including nanogels andpolymersomes, have emerged as attractive cargo for drugdelivery systems (DDSs). With advances in nanomaterial synthesis
and the characterization of stiffness, the role of NP elasticity inbiological systems has gained increased attention. Thus, we aim
to develop PFC-encapsulated elastic NPs to improve delivery
efficacy and reduce long-term accumulation.
The synthesis involved forming perfluoro-15-crown-5-ether(PFCE)-encapsulated micelles followed by surface-templated
reversible addition-fragmentation chain transfer (RAFT)
polymerization. This process used a trithiocarbonate-based chaintransfer agent (CTA) for controlled polymerization. The NPs had
an average size of 162 nm and a negative surface charge (−30.0mV), similar to silica NPs. We confirmed the deformability of theobtained NPs with high-speed atomic force microscopy (HS-64

AFM). They could shrink and return to their original shape underpressure without leakage of the core compound, unlike stiff silica
NPs that only slid on the substrate without significant
morphological changes. The 19F NMR spectrum showed a sharp
peak corresponding to the encapsulated PFCE, with thetransverse relaxation time (T2) of the encapsulated PFCE inpolymer NPs being slightly shorter than that of non-encapsulated
PFCE, indicating well-dispersed NPs in aqueous solution andsubstantial sensitivity in 19F NMR signal detection. The amountsof adsorbed serum proteins on the NP surfaces were quantitatively
analyzed. Notably, the amounts of protein adsorbed on thepolymer NPs were below the detection limit and significantly
lower compared to the silica NPs. This indicates their potential for
mitigating unwanted immune responses and improved behavior
in biological systems. influence the biodistribution of
nanomaterials.
We further visualized the biodistribution of elastic polymer NPs
using 19F MRI. The NPs were administered intravenously to amouse and monitored for three weeks. Acute toxicity was notobserved after injection. At 2 weeks after injection, the 19F MRI
signal intensity of the polymer NPs in the liver and spleen
gradually decreased over time. Additionally, we compared thebiodistribution of elastic polymer NPs with that of stiff silica NPs
using multi-color 19F MRI within a single mouse. The results
demonstrated that the polymer-coated NPs exhibited a gradualdecline in signal intensity post-administration, indicating
effective clearance from the liver or degradation over time. Thisbehavior contrasts with stiff silica NPs, which tend to accumulate
and persist in the liver.
The development of these elastic polymer-coated NPs
represents a significant advancement in the field of molecular

Figure 2.
1
H/19F MRI images of a mouse injected with polymer and silica NPs. Time course of 19F MRI signal
intensity changes in liver. Normalized by 19F MRI of polymer NPs on Day 1.

Recent Publications
1. Torii K, et al. No-wash Fluorogenic Labeling of Proteins for Reversible
Photoswitching in Live Cells. Chem Sci. 15:1393-1401 (2024).

4. Nishiura M, et al. Visualization of Multiple Localizations of GLUT4 byFluorescent Probes of PYP-tag with Designed Unnatural Warhead.
Chem Sci. 14:5925-5935 (2023).

2. Konishi Y, et al. Elastic Polymer-coated Nanoparticles with Fast
Clearance for 19F MR Imaging. Angew Chem Int. Ed. 62: e202308565(2023).
3. Minoshima M, et al. Development of a Versatile Protein Labeling Tool
for Live-Cell Imaging Using Fluorescent β-Lactamase Inhibitor. Angew
Chem Int. Ed. 62: e202301704 (2023).

65

Imaging

Immune Response Dynamics
treatment phenocopied COMMD3/8 complex deficiency,
suggesting that celastrol may target the COMMD3/8 complex in
the context of humoral immune responses and autoimmunity.

Kazuhiro Suzuki, MD/PhD■ Professor

Kazuhiro Suzuki■ Assistant Professor

Akiko Nakai
Taiichiro Shirai

Our research focus has been to discover novel mechanisms that
control lymphocyte migration and elucidate their physiologicaland pathological significance. Our previous studies showed that
inputs from adrenergic neurons to the β2-adrenergic receptor
expressed on lymphocytes enhance the responsiveness of specificchemokine receptors and inhibit lymphocyte exit from lymph
nodes (Nakai et al. J Exp Med. 2014). This mechanism was found to
generate diurnal variations in lymphocyte numbers in lymph
nodes and consequently the magnitude of adaptive immune
responses in phase with the circadian oscillation of adrenergic
neuron activity (Suzuki et al. J Exp Med. 2016). These studies
provided insights into the molecular basis of the interaction
between the nervous and immune systems.
In our efforts to elucidate the mechanism of the crosstalk
between the β2-adrenergic receptor and chemokine receptors,
we identified a protein complex consisting of copper metabolism
MURR1 domain-containing (COMMD) 3 and COMMD8 (COMMD3/8
complex) as a positive regulator of chemokine receptor signaling.
Our study demonstrated that the COMMD3/8 complex plays
important roles in the control of B cell migration and the inductionof humoral immune responses (Nakai et al. J Exp Med. 2019).
However, the contribution of the COMMD3/8 complex to thepathogenesis of immunological disorders was unclear.
Based on the important role of the COMMD3/8 complex inhumoral immune responses, we tested its involvement incollagen-induced arthritis, a B cell-dependent mouse model ofrheumatoid arthritis. COMMD3/8 complex deficiency induced at
the onset of arthritis inhibited disease progression. This was
accompanied by a reduced humoral immune response to66■ Postdoctoral Fellow

0■ Research Assistant2■ Visiting Scientist

0■ Support Staff

3

Since alanine substitution of C170 (C170A) on COMMD3
rendered the COMMD3/8 complex resistant to celastrol while
preserving the function of the protein complex, we generated amouse strain expressing COMMD3C170A from the endogenous
Commd3 locus and examined whether the effects of celastrol are
abolished in these mice. B cells isolated from COMMD3C170A mice
showed complete resistance to celastrol in chemotactic migration.
Humoral immune responses and collagen-induced arthritis in the
mutant mice were not suppressed by celastrol treatment. Thesefindings indicated that the COMMD3/8 complex is a major targetof celastrol (Shirai et al. Sci Immunol. 2023).
Our study suggests that the COMMD3/8 complex is involved in
the progression of antibody-mediated autoimmunity. However,the point of action of the COMMD3/8 complex in the pathogenesisof autoimmune diseases has not been clearly defined. Additionally,
since most of our knowledge about the COMMD3/8 complex has
been obtained in mice, the relevance of our findings to
autoimmunity in humans remains to be established. Futurestudies will overcome these limitations and realize thedevelopment of a novel therapy targeting the COMMD3/8
complex in immune disorders.

collagen. These findings indicated that the COMMD3/8 complex
contribute to the pathogenesis of rheumatoid arthritis (Shirai et al.
Sci Immunol. 2023).
Prompted by this finding, we performed a chemical screen to
identify inhibitors of the COMMD3/8 complex that could be used
for the treatment of autoimmune diseases. Since the function of
the COMMD3/8 complex depends on the association between
COMMD3 and COMMD8, we sought for compounds that disrupt
the physical interaction between the two COMMD proteins. Afterscreening of a chemical library that was relatively enriched innatural products, we identified celastrol as the most potent
compound. Celastrol is a bioactive molecule extracted from a
medicinal herb, Tripterygium wilfordii, and exhibits antiinflammatory properties. However, its mechanism of action had
been poorly understood. Celastrol disrupted the COMMD3/8
complex in living cells or in the purified form, indicating direct
action of celastrol on the COMMD3/8 complex. Using site-directedmutagenesis, molecular dynamics simulations (Figure) and liquid
chromatography-tandem mass spectrometry, we revealed thatcelastrol covalently binds to cysteine 170 (C170) on COMMD3 todissociate the COMMD3/8 complex (Shirai et al. Sci Immunol.
2023).
We then asked whether celastrol reproduces the functional
consequences caused by COMMD3/8 complex deficiency.
Celastrol inhibited chemotactic migration of B cells in vitro and invivo. Celastrol treatment suppressed antibody responses with
reduced production of germinal center B cells and plasma cells.
The progression of collagen-induced arthritis was blocked bycelastrol treatment started at the disease onset. Thus, celastrol

Figure.
Celastrol inhibits the COMMD3/8 complex and alleviates autoimmunity. (A) An in silico model ofthe interaction between celastrol and the COMMD3/8 complex. (B and C) Effects of celastrol on theseverity (B) and histopathology (C) of collagen-induced arthritis. Scale bar, 500 μm.

Recent Publications
1. Shirai T, Nakai A, Ando E, Fujimoto J, Leach S, Arimori T, Higo D, vanEerden FJ, Tulyeu J, Liu Y-C, Okuzaki D, Murayama MA, Miyata H,
Nunomura K, Lin B, Tani A, Kumanogoh A, Ikawa M, Wing JB, Standley
DM, Takagi J, Suzuki K. Celastrol suppresses humoral immune responses
and autoimmunity by targeting the COMMD3/8 complex. Sci Immunol.
8: eadc9324 (2023).

3. Suzuki K, Hayano Y, Nakai A, Furuta F and Noda M. Adrenergic control of
the adaptive immune response by diurnal lymphocyte recirculation
through lymph nodes. J Exp Med. 213:2567-2574 (2016).
4. Nakai A, Hayano Y, Furuta F, Noda M and Suzuki K. Control of lymphocyte
egress from lymph nodes through β2-adrenergic receptors. J Exp Med.
211:2583-2598 (2014).

2. Nakai A, Fujimoto J, Miyata H, Stumm R, Narazaki M, Schulz S, Baba Y,
Kumanogoh A and Suzuki K. The COMMD3/8 complex is a determinantof GRK6 specificity for chemoattractant receptors. J Exp Med. 216:16301647 (2019).

67

Imaging

Biophotonics

Nicholas Isaac Smith, PhD

The Biophotonics laboratory develops tools for label-free
analysis of single cells. Single-cell analysis is a popular target for avariety of research fields, usually pursued by labeling surface
markers, by introducing fluorescent dyes into the cell, or byinvasive, yet comprehensive, techniques such as single cell RNA
sequencing. In contrast, our tools are based on label-free opticalmethods, which aim to produce some of the same discriminatory
capability as the more invasive methods. Additionally, label-freemethods are based on endogenous contrasts of the cell, and canalso find novel features that can be used to discriminate between
cell phenotypes or cell states.
In the last year we were fortunate to complete several projects.
We have proposed that unlabeled Raman measurement of single
cells can have sufficient sensitivity to provide discrimination
which has previously needed labels or sequencing to achieve.
Now, we established more clear limits on exact levels of sensitivity
when used for phenotyping cells and established safe laser
power regimes where we demonstrate that no adverse effects oncells are occurring. While optical techniques usually fall into
categories of either imaging, or non-imaging single measurement(as in flow cytometry), we showed that cell phenotyping can bedone in imaging or non-imaging modes, or even partway
between the two extremes, where some amount of spatial
information is captured, increasing phenotype discrimination
accuracy compared to single point measurement, while still■ Associate Professor

Nicholas Isaac Smith■ Assistant Professor

Alison Hobro■ Support Staff

1

allowing for higher throughput than full imaging which istypically too slow for biologically relevant phenotyping. By
comparing 4 different possible modes of measurement, andassessing each in terms of the discrimination power, as well as
any induced change in the sample (denoted as differential
spectra in the figure at different power and exposure regimes),
we showed how to maximize the measured features of each cellwhile ensuring that no laser-induced changes are occurring.
We also used Raman imaging to elucidate both morphologicaland biochemical changes occurring in the liver in mouse fed witha high-fat diet, leading to the onset of NASH (a type of fatty liverdisease). While it is known that a high fat diet leads to aggregationof lipids in the liver, we used Raman imaging to assess how they
were distributed, and whether they were correlated with those in
the diet itself. We found that oleic acid dominated the lipid
buildup in the liver, as a direct result of the diet, while otherdietary components, such as sucrose and glucose were notdeposited. This type of unlabeled imaging using no a priori
information can be used for a 3-way quantitative comparison ofcomponents across a) diet, b) diseased tissues and c) healthy
tissues, giving weights for each component that denote theiractual strength. The possibility to identify emergent differences
also exists, and in this study we were able to observe lipid
peroxidation occurring in the NASH liver samples, as a distinct
component not present to the same degree in control samples.

Figure.
For single-cell Raman phenotyping, we created several techniques, including no spatial information for the fastest throughput, using full
spatial information for the greatest amount of data, and several hybrid techniques that blend the ability to read intracellular components
across regions in the cell while still allowing high throughput. The left four panels (a) show the different modalities, compared by theirability to classify MEF vs Raw264 cells (b). At higher throughputs, we can forgo imaging completely, and use only a single point illumination,
but the ability to characterize each still is still improved by spatially averaging the information across each individual cell. Spectra shownin (c) are differential spectra showing any changes induced across sequential measurements with the ideal result there being no significant
change induced by the measurement itself. The spatially averaged measurement system we developed is significantly better than single
point measurement for single-cell phenotyping. In (d) we see the full Raman imaging of NASH-affected liver, where the building of lipids,in this case primarily oleic acid occurs in holes in the distribution of proteins and other components. The identification of the components
requires no a priori information or labeling.

Recent Publications
1. Hobro AJ, et al. Imaging vs non-imaging Raman spectroscopy forhigh-throughput single-cell phenotyping. Analytical Chemistry.
(2024).

4. Sugiyama T, et al. Label-free Raman mapping of saturated andunsaturated fatty acid uptake, storage, and return toward baseline
levels in macrophages. Analyst. 146 (4):1268-1280 (2020).

2. Lelliott PM, et al. Cellular Adhesion Is a Controlling Factor in Neutrophil
Extracellular Trap Formation Induced by Anti-Neutrophil Cytoplasmic
Antibodies. ImmunoHorizons. 6 (2):170-183 (2022).

5. Pavillon N, Hobro AJ, Akira S and Smith NI. Noninvasive detection ofmacrophage activation with single-cell resolution through machinelearning. Proc Natl Acad Sci USA. 115(12):E2676-E2685 (2018).

3. Pavillon N, and Smith NI. Deriving accurate molecular indicators ofprotein synthesis through Raman-based sparse classification. Analyst.
146, pp. 3633-3641 (2021).

68

69

Systems Immunology

Bioinformatics

A. AIR Representation

B. AIR Cluster Antigen specificity

C. Disease Diagnosis Accuracy

1

HIV

NSCLC

AIH
Clonotype

MyImmune■ Professor

Daron Standley■ Associate Professor

Kazutaka Katoh

DT
TT
PT
NB

CRC

0.8

ROC AUC

Daron Standley, PhD

COVID-19

0.9

MS
T1D

0.7

0.6

0.5
0

100

200

Songling Li
Soyoung Park
■ Postdoctoral Fellow

1■ Research Assistant2■ Visiting Scientist

4

Molecular function from simulation of protein
dynamics

We have formulated the problem of disease diagnosis from
Adaptive Immune Receptors (AIRs) as a classification problem thatcan be solved by ML. The classifier, MyImmune, required thedevelopment of a more useful representation of AIRs than theconventional “clonotype” definition, that could group AIRs fromdifferent individuals (Fig. 1A). The resulting clusters were found tobe >95% specific in terms of targeted antigens (Fig. 1B). We thenvalidated MyImmune on three classes of disease: infectious(COVID-19, HIV), autoimmune (Autoimmune Hepitaitis, Type-1
diabetes) and cancer (non-small cell lung cancer, colorectal
cancer). MyImmune demonstrated significant improvement overseven alternative classifiers using the same inputs: mean ROC AUC
0.893 vs 0.777, respectively. Moreover, we observed linear
improvement on MyImmune’s performance with the size of the
training data (Fig. 1C). A surprising finding for cancer is that
healthy individuals harbor a reservoir of potential tumorinfiltrating lymphocytes that can be identified by MyImmune
feature importance.

Drawbridge model for T cell receptor activation. Despite thecritical importance of the T cells in immune responses, it is still
unclear how T cell receptor (TCR) binding to specific peptide-MHC(pMHC) complexes results T cell activation. Based on extensive
multiscale molecular dynamics simulations we formulated thedrawbridge model (Figure 2A) in which the TCR receptor releasesa brake on CD3 proteins upon pMHC binding. This model isconsistent with a wide range of experimental data, including
mutagenesis of the hinge and FG loop residues, as well as recent
cryo-EM analysis.
Mechanism of activity enhancing antibodies in SARS-CoV-2. SARSCoV2 infection enhancing antibodies, discovered in the Arase
Laboratory, exert their effect in an Fc-independent manner.
Together, we hypothesized that this mechanism involves
crosslinking two neighboring spike proteins. Long-range
molecular dynamics simulations on the Fugaku computer support
a model in which such crosslinking decouples the N-terminal
domain from the receptor-binding domain (RBD), allowing the
RBD to transition from the “down” to the “up” orientation, where
it can engage with the host entry receptor ACE2.

400

500

600

Figure 1.
MyImmune Disease Classification. A, B cell receptor (BCR) heavy chain networks were constructed for 10 COVID-19 patients; in these networks,
nodes are colored by donor, and edges represent clonotype or MyImmune similarity. Clonotype networks are typically small and do not connect
different donors. MyImmune networks are much larger and generally connect many donors. B, BCR networks derived from single-cell immune
profiling of diphtheria-tetanus-pertussis (DTP)-vaccinated donors were demonstrated by ELISA to exhibit 96% antigen purity, surpassing theapparent 82% purity achieved by assigning antigens to the same B cells using fluorescently labeled DTP antigen probes. C, The performance of
MyImmune on new indivuduals (i.e. not seen by the ML training) was benchmarked for six diseases in three categories using the Area Under the
Receiver Operating Characteristic Curves (ROC AUC). The MyImmune performance increased linearly with the number of donors up to a thresholdvalue of ~200, after which the classifier was nearly perfect.

A. drawbridge model

Classification of health/disease from Adaptive Immune
Receptors

300

Number of Donors

B. Infection-enhancing antibody model

Figure 2.
A, The drawbridge model for TCR triggering. The TCR elongates upon TCR-pMHC binding, resulting in an increased mobility of the CD3 proteins,which carry the ITAM signaling domains. B, Infection-enhancing antibodies crosslink neighboring spike proteins via their NTD domains, resulting indecoupling of the NTD and RBD domains, thereby catalyzing the RBD down to up transition, which is an important part of the infection process.

Recent Publications
1. Saputri DS, et al. Deciphering the antigen specificities of antibodies byclustering their complementarity determining region sequences.
mSystems 8: e0072223 (2023).

4. Kitagawa Y, Akizuki S, Ito Y, et al. Construction of a T cell receptor
signaling range for spontaneous development of autoimmune
disease. J Exp Med. 220 (2023).

2. Millius A, et al. Circadian ribosome profiling reveals a role for the
Period2 upstream open reading frame in sleep. Proc Natl Acad Sci U S
A 120: e2214636120 (2023).

5. Eerden FJ v, et al. TCR-pMHC complex formation triggers CD3
dynamics. eLife (2023).

3. Mitsui Y, et al. Expression of the readthrough transcript CiDRE inalveolar macrophages boosts SARS-CoV-2 susceptibility and promotes
COVID-19 severity. Immunity 56:1939-1954 e1912 (2023).

70

71

Bioinformatics

Statistical Immunology

Yukinori Okada, MD/PhD■ Professor

Yukinori Okada■ Assistant Professor

Yuya Shirai■ Research Assistant5■ Support Staff

2

RA- or SLE-like clusters were exclusively dominant, showing
immunological polarization between RA and SLE across AIRDs. By
adopting RA as a flagship disease, in depth clinical analysis
revealed that such patient clusters differentially defined clinicalheterogeneity in disease activity and treatment responses, such astreatment resistance in the RA patients with SLE-like immunophenotypes. Inborn human genetics represented by polygenic
risk score based on the RA case-control genome-wide association

study (GWAS) and within-case stratified GWAS were associatedwith patient characteristics, disease activity, and immunephenotypes such as dendritic cells for RA-interstitial lung disease.
Our study demonstrated a value of cohort-wide and cross-disease
immuno-phenotyping to elucidate clinically heterogenous
patient subtypes existing within the single disease in an immune
cell type-specific manner. (Tanaka H et al. Ann Rheum Dis 2024).

Figure 1.
Large-scale Single cell sequencing with human genome data of severe COVID-19 in Japanese.

Goal of our laboratory

Human genomes in non-humans from human body

Genetic backgrounds of individuals have substantial impacts
on risk of a wide range of immune-related diseases. Statisticalimmunology is a research field that evaluates causality of human
genetic variations on immune-related diseases, using statisticaland bioinformatics approaches. The goal of our laboratory is todevelop such methods and apply to the latest large-scale disease
genome and multi-layer omics data.

Sharrow human DNA included in faecal samples can result in asmall number of human reads in gut shotgun metagenomic
sequencing data. It has been unclear whether and how muchpersonal information can be quantitatively reconstructed fromsuch contaminated reads. Based on metagenome resources ofJapanese, we constructed the genomic approaches to reconstructpersonal information from the faecal metagenomes. Genetic sex
could be accurately predicted based on the sequencing depth ofsex chromosomes. Individuals could be re-identified from thematched genotype data based on human reads recovered fromthe faecal metagenomic data. The method enabled us to predictthe genetic ancestries of the samples. We then performed ultradeep shotgun metagenomic sequencing of the faecal samples,
which demonstrated that the genotypes of both common andrare variants could be reconstructed from faecal samples. This
included clinically relevant variants. Our study showed thathuman reads within gut metagenomic data is personal identifiable
information (Tomofuji Y et al. Nat Microbiol 2023).

Single cell sequencing with human genome data ofsevere COVID-19
Coronavirus disease 2019 (COVID-19) represents a serious
global public health issue. Toward next pandemic, immunologicalmechanisms of the dysfunctional immune response in severe
COVID-19 infection are elusive. We analyzed single-celltranscriptomes and T and B cell receptors (BCR) of ~900,000
peripheral blood mononuclear cells from 150 Japanese subjects
with COVID-19 as well as healthy controls along with hostgenome-wide genotype data (Figure 1). We found a series of
immunological features: (i) COVID-19 patients showed a low
fraction of nonclassical monocytes. (ii) Cell-cell communication
analysis inferred decreased cellular interactions involving
nonclassical monocytes in severe COVID-19. (iii) Clonal expansionsof BCR were evident in the COVID-19 patients rather than TCR. (iv)
Disease risk genes identified by COVID-19 genome-wide
association study showed cell type-specific expressions inmonocytes and dendritic cells. (v) Further, a COVID-19-associatedrisk variant at IFNAR2 had COVID-19-specific and monocytespecific expression quantitative trait loci effects (Edahiro R et al.
Nat Genet 2023).

72

Deconvoluting heterogeneity across autoimmune
rheumatic diseases
Deconvolution of immunological and clinical heterogeneity
across autoimmune rheumatic diseases (AIRDs) is of needs. We
conducted large-scale and cohort-wide immuno-phenotyping of
46 peripheral immune cells from >1,000 Japanese patients of 11
AIRDs. Multimodal clustering of immune-phenotypes deciphered
underlying disease-cell type network, providing novel immune
cell type specificity shared or distinct across AIRDs. Individual
patient-level clustering deconvoluted the AIRD patients into
several clusters with different immunological features. Of these,

Figure 2.
Deconvoluting network across peripheral immune cells and autoimmune rheumatic diseases.

Recent Publications
1. Yamamoto K, Sonehara K, Namba S, Konuma T, Masuko H, Miyawaki S,
BioBank Japan Project, Kamatani Y, Hizawa N, Ozono K, Yengo L,
Okada Y. Genetic footprints of assortative mating in the Japanese
population. Nat Hum Behav. 7:65-73 (2023).
2. Edahiro R, Shirai Y, Takeshima Y, Sakakibara S, Yamaguchi Y, Murakami
T, Morita T, Kato Y, Liu YC, Motooka D, Naito Y, Takuwa A, Sugihara F,
Tanaka K, Wing JB, Sonehara K, Tomofuji Y; Japan COVID-19 Task Force,
Namkoong H, Tanaka H, Lee H, Fukunaga K, Hirata H, Takeda Y, Okuzaki
D, Kumanogoh A, Okada Y. Single-cell analyses and host genetics
highlight the role of innate immune cells in COVID-19 severity. Nat
Genet. 55:753-767 (2023).

3. Tomofuji Y, et al. Reconstruction of the personal information fromhuman genome reads in gut metagenome sequencing data. Nat
Microbiol. 8:1079-1094 (2023).
4. Tomofuji Y, et al. Analysis of gut microbiome, host genetics, andplasma metabolites reveals gut microbiome-host interactions in theJapanese population. Cell Rep. 42:113324 (2023).
5. Tanaka H, Okada Y, Nakayamada S, Miyazaki Y, Sonehara K, Namba S,
Honda S, Shirai Y, Yamamoto K, Kubo S, Ikari K, Harigai M, Sonomoto K,
Tanaka Y. Extracting immunological and clinical heterogeneity acrossautoimmune
rheumatic
diseasesbycohort-wide
immunophenotyping. Ann Rheum Dis. 83:242-252 (2023).

73Quantitative Immunology■ Associate Professor

Our group applies computational and single cell genomics
techniques to understand the immune system. We develop
computational methods to analyze single cell data. We integrate
experimental data, including transcriptome, chromatin
accessibility, protein expression, immune repertoire, and spatialtranscriptomics, with publicly available information into networkmodels of immune regulation. We apply this framework to study
gene regulatory networks controlling immune cell development
and function.

Unit

Diego Diez

experiments opens the door to approaches that study the
immune system using mathematical modeling and machinelearning. Transcriptional regulatory networks are critical
determinants of cell identity and function. We use machinelearning to model immune transcriptional regulatory networks.
Using the expression level of the regulators as a proxy for theiractivities we apply these methods to study how transcriptional
networks change during immune cell differentiation and disease.

Applications to immunology
Development of computational methods
An important problem in single cell genomics is how to
combine different datasets while correcting for batch effects. A
key focus is on preserving the original cell population structure
while not introducing bias. We have developed Canek, a methodthat leverages a fuzzy logic framework that enables efficientbatch correction without bias. Another problem is theidentification of marker genes. In collaboration with Alexis
Vandenbon at Kyoto University, we have developed
singleCellHaystack, a method to identify differentially expressed
genes from multi-dimensional representations of single cell
genomics data.

Mathematical modeling
The large number of cells obtained in single cell genomics

Using single cell transcriptomics, protein expression, immune
repertoire, and chromatin accessibility we study thedifferentiation of NKT cells in the thymus and spleen of SKG and
WT mice. SKG mice have a mutation in ZAP70 that weakens TCR
signaling, resulting in a bias towards development of NKT1
compared to NKT2 in the wild type (BALB/c). This approachenables us to understand how changes in regulatory networks
during development effect NKT specification. In a clinical setting,
we apply single cell genomics to get insight into IgA nephritis
onset and therapies. We use transcriptome, protein expression
and immune repertoire of immune cells in PBMCs and tonsils
from IgAN patients before and after tonsillectomy and steroid
immunosuppression. We collaborate with other groups at IFReC
to study diverse aspects of immune responses in a basic andclinical setting.

Figure.
Single cell genomics identifies RNA and TCR features of differentiating NKT cells.

Recent Publications
1. Vandenbon A & Diez D. A universal tool for predicting differentially
active features in single-cell and spatial genomics data. Sci Rep.
13:11830 (2023).

4. Vandenbon A and Diez D. A clustering-independent method forfinding differentially expressed genes in single-cell transcriptome
data. Nat Commun. 11(1):4318 (2020).

2. Loza M, Teraguchi S, Standley DM & Diez D. Unbiased integration of
single cell transcriptome replicates. NAR Genom Bioinform. 4, lqac022(2022).

5. Teraguchi S, Saputri DS, Llamas-Covarrubias MA, Davila A, Diez D,
Nazlica SA, Rozewicki J, Ismanto HS, Wilamowski J, Xie J, Xu Z, LozaLopez MJ, van Eerden FJ, Li S, Standley DM. Methods for sequence andstructural analysis of B and T cell receptor repertoires. Comput Struct
Biotechnol J. 18:2000-2011 (2020).

3. Diez D, Morte B & Bernal J. Single-Cell Transcriptome Profiling of
Thyroid Hormone Effectors in the Human Fetal Neocortex: Expressionof SLCO1C1, DIO2, and THRB in Specific Cell Types. Thyroid. 31:15771588 (2021).

74

75

Events & Outreach Activities

Events & Outreach Activities

International Symposium on Microbiology and Immunology
The 13th International Symposium of IFReC
This symposium was co-organized with the Center for Infectious Disease Education
and Research (CiDER) at Osaka University, the project for "Self-referential immune
perception" (Grant-in-Aid for Transformative Research Areas, JSPS). Centering on basic
research achievements relating to microbiology and immunology, this symposium
consists of 12 lectures by world-leading scientists. This symposium was a significant
opportunity for the participants to share ideas and expertise for further developmentof microbiology, immunology, and life science.

■■ Date:February 9, 2024■■ Venue : Yuichi Yamamura Life Hall, Senri Life Science Center,
Suita-city, Osaka

Speaker (order of presentation)

78

Title

Kazuhiro Suzuki(IFReC, Osaka University)‘Targeting B cell migration in the pathogenesis of autoimmunity’

Hisako Kayama(IFReC, Osaka University)‘The role of microbial metabolites in development of ulcerative colitis’Ana Ivonne Vazquez Armendariz(LIMES-Institute, University of Bonn)‘Complex murine and human organoid systems to model pulmonary diseases’

Sho Yamasaki(IFReC, Osaka University)‘Immune response to host metabolites‘Joseph Craft(Yale School of Medicine)‘Tissue Adaptation and Tissue Damage in Autoimmunity’

Wataru Ise(CiDER, Osaka University)‘Regulation of plasma cell survival and migration to the survival niches forestablishing durable antibody response’

Laura McCoy(University College London)‘Preservation of memory B cell homeostasis in an individual producing broadly
neutralizing antibodies against HIV-1’

Raul Andino(University of California San Francisco)‘Studies on enterovirus particle structure and functions’Leyuan Ma(University of Pennsylvania)‘Boosting Chimeric Antigen Receptor T cell therapy via a synthetic vaccine’

Lynette Beattie(University of Melbourne)‘A dual role for gamma delta T cell-derived IL-4 in CD8 T cell priming’

Eric Huseby(University of Massachusetts)‘MHC-II polymorphisms regulate non-cognate negative selection to CD4 T cellorchestrators of type-1 diabetes’

Adrian Hayday(The Francis Crick Institute, London)‘Regulating tissue immunosurveillance by innate and adaptive uses of the T cell
receptor’79Events & Outreach Activities

The 2 nd Doherty Institute & partners –IFReC Immunology Symposium■■ Date:February 8, 2024■■ Venue : Taniguchi Memorial Hall, Osaka University

Finnish-Japanese Immunology Symposium
Emerging concepts in immunology - From basic
mechanisms to treating immune mediated diseases■■ Date:August 29-30, 2023■■ Venue : University of Turku, Finland

Speakers (order of presentation)

Speakers (order of presentation)

1. Akihiko Yoshimura (Keio University)

16. Kazuhiro Suzuki (IFReC)

2. Riitta Lahesmaa (University of Turku)

17. Sirpa Jalkanen (University of Turku)

3. Satu Mustjoki (University of Helsinki)

18. Mika Rämet (Tampere University)

4. Shimon Sakaguchi (Osaka University)
5. Akira Takeda (University of Turku)
6. Tomohiro Kurosaki (IFReC)
7. Hisashi Arase (IFReC)
8.

Speaker (order of presentation)

80

Title

Sammy Bedoui(UoM/Doherty Institute)‘T cell memory, metabolism and the microbiome’

Nobuhiko Kamada(IFReC)‘Decoding the Functional Impact of Gut Microbiota in Inflammatory Bowel’Andrew Brooks(UoM/Doherty Institute)‘Polymorphism in Killer cell immunoglobulin-like Receptor genes diversifies
Natural Killer Cell responses’

Yasutaka Okabe(IFReC)‘Stromal-immune cell interactions in non-classical lymphoid tissues’

Sidonia Eckle(UoM/Doherty Institute)‘Antigen recognition and associated immune functions of MAIT cells’

Daron M. Standley(IFReC)‘Medical diagnosis using adaptive immune receptor repertoires’

Axel Kallies(UoM/Doherty Institute)‘Cytotoxic CD8 T cells in chronic infection and cancer’

Atsushi Tanaka(IFReC)‘Regulation of TCR signaling molecules in Treg cells’

Cecilia Naucler (University of Turku and Karolinska
Institutet)

9. Shizuo Akira (IFReC)
10. Hiroshi Ohno (RIKEN)
11. Hisako Kayama (IFReC)
12. Kazuyo Moro (IFReC)
13. Olli Silvennoinen (University of Helsinki)
14. Hayato Takahashi (Keio University)
15. Laura Airas (University of Turku)

Joint workshop with ImmunoSensation at the
University of Bonn to promote joint research■■ Date:June 14-15, 2023■■ Venue : University of Bonn, Germany

Scott Mueller(UoM/Doherty Institute)‘Dissecting the tissue contexts of immunity’

Masahiro Yamamoto(IFReC)‘The novel VeDTR mouse system reveals roles of immune cells in anti-tumor
immunity’Jose Villadangos(UoM/Doherty Institute)‘Antigen-presenting cells and molecules, the initiators of adaptive immunity’

A joint workshop was held to promote collaboration between IFReC at Osaka University and ImmunoSensation at the University ofBonn. Ten PIs from IFReC and 16 PIs from ImmunoSensation participated in this workshop, the aim of which was to promote matchingfor further joint research in addition to existing ones. The researchers shared information about their research in advance, and gave
short talks at the workshop introducing their research, followed by individual interviews with potential collaborators. As a result, morethan ten new joint research projects were established. The workshop commenced with a presentation by Masahiko Hayashi, Directorof the JSPS Bonn Office where he introduced the competitive funding provided by the JSPS, which is available to support the joint
research.

81

Events & Outreach Activities

The 2 nd ImmunoSensation2 - IFReC International
School on Advanced Immunology
The 2nd International School on Advanced Immunology was held in Germany. Fifteen leading immunologists were invited as
lecturers, and 49 exceptional participants with remarkable research achievements were selected from a large number of worldwide
applicants. The high level of research by the participants and the active exchanges to promote interaction among them were wellreceived by both lecturers and participants.

■■ Date:September 17-21, 2023■■ Venue : Seehotel Maria Laach, Germany

Lecturers
Zeinab Abdullah (University of Bonn, Germany)
Julie Magarian Blander (Weill Cornell Medicine, USA)
Marco Colonna (Washington University School of Medicine, USA)James Chen (UT Southwestern Medical Center, USA)
Kate Fitzgerald (UMass Chan Medical School, USA)
Matthias Geyer (University of Bonn, Germany)
Masaru Ishii (Osaka University, Japan)
Raphaela T. Goldbach-Mansky (National Institutes of Health-NIAID/DIR, USA)
Naoki Hosen (Osaka University, Japan)
Kazuyo Moro (Osaka University, Japan)
Ann Marshak-Rothstein (UMass Chan Medical School, USA)
Joachim Schultze (University of Bonn, Germany)
Hans Stauss (UCL Institute of Immunity & Transplantation, UK)
Brigitta Stockinger (Francis Crick Institute, UK)
Kiyoshi Takeda (Osaka University, Japan)

82

83Events & Outreach ActivitiesNGS Expo 2023
The "NGS Expo", which started in 2022 is an academic event for users andpotential users of next generation sequencer, and organized by IFReC and RIMD,
Osaka University. About 800 people participated in this symposium including
online. The organizers incorporated "online salon", a communication tool
between participants, and nearly 50,000 page views were obtained.

■■ Date:November 15-16, 2023■■ Venue : Osaka International Convention Center (Grand Cube Osaka)

Colloquia and Seminars■■ IFReC Colloquia : Important events allowing IFReC researchers to gather together,and have been held every two months.

Date

Speakers
James Badger Wing (Human Single Cell Immunology)

May 25, 2023

Mohamad Alkadi & Daisuke Okuzaki (Human Immunology (Single Cell Genomics))
Nicholas Isaac Smith (Biophotonics)

July 20, 2023

September 28, 2023

November 8, 2023

January 25, 2024

March 21, 2024

Diego Diez (Quantitative Immunology Research Unit)
Burcu Temizoz, Takayuki Shibahara, and Ken Ishii (Vaccine Science)

Rei Watanabe & Manabu Fujimoto (Cutaneous Immunology)
Shunya Ikeda & Naoki Hosen (Cellular Immunotherapy)
Fumitaka Muramatsu & Nobuyuki Takakura (Signal Transduction)
Oluwaseun Fatoba & Toshihide Yamashita (Molecular Neuroscience)
Takuya Koike & Tomohiro Kurosaki (Lymphocyte Differentiation)
Taichi Nakatani & Takashi Nagasawa (Stem Cell Biology and Developmental Immunology)
David Calianese & Shigekazu Nagata (Biochemistry & Immunology)
Shimpei Kawamoto & Eiji Hara (Aging Biology)■■ IFReC Seminar & IFReC ImmunoSeminar*
A wide variety of scientists has been invited as speakers.
*Inviting world-prestigious immunologists mainly online.

Date

Speakers
Bernard Malissen*

April 12, 2023

May 16, 2023

May 24, 2023

July 21, 2023

(Group leader at Centre d'imuunologie de Marseille-Lumity/Founding-director of Center for
Immunophenomics, Marseille, France)
Keisuke Nagao(Senior Investigator, Cutaneous Leukocyte Biology Section, NIAMS, NIH, USA)

Arthur Weiss*(Ephraim P. Engleman Distinguished Professor/Professor of the Department of Medicine, UCSF, USA)
Yosuke Kumamoto(Assistant Professor, Department of Pathology, Immunology and Laboratory Medicine, Rutgers, theState University of New Jersey, USA)

Kai-Michael Toellner
August 8, 2023

September 19, 2023

September 29, 2023

November 2, 2023

January 22, 2024

February 19, 2024

84

(Professor, Adaptive Immunology, Institute of Immunology and Immunotherapy, University ofBirmingham, UK)

Olivier Lantz*(Director, the Clinical Immunology Laboratory at Institut Curie's Paris hospital, France)

Hiroki Kato
(Professor, Institute of Cardiovascular Immunology, Medical Faculty, University of Bonn, Germany)

Andreas Schlitzer(Professor, Quantitative Systems Biology, LIMES-Institute, University of Bonn, Germany)

Ruth A. Franklin(Assistant Professor, Stem Cell and Regenerative Biology, Harvard University, USA)

H. Suenaga, R. Ito, T. Takahashi, and M. Suzuki(PIs, Central Institute for Experimental Animals (CIEA), Japan)

85Events & Outreach Activities

Outreach Activities
In 2023, we have organized various outreach events by online and face-to-face. An interaction with the general public is a goodstimulus for researchers. Especially, an approaching to high school students and high school teachers is important for the future
development of science and technology.

Introduction of IFReC and WPI program for Super Science High School Students■■ July 15, 2023@Meijo University, Nagoya■■ December 27, 2023@Okazaki Conference Hall, Aichi

The 12th WPI Science Symposium■■ Date :November 23, 2023■■ Venue :Akira Suzuki Memorial Hall, Hokkaido University, Sapporo■■ Speakers : Hisashi Yamamoto (ICReDD), et al.

Life Science Seminar for High School Students■■ Date :August 7, 2023■■ Venue :Taniguchi Memorial Hall, RIMD and IFReC■■ Speakers : Yoko Fukushima (Osaka University School of Medicine), et al.

Online Seminars for High School Teachers■■ August 1, 2023: “A Molecule that controls winter depression and inflammation”

Science Café at the Nakanoshima Festival “The heart; present & the future”

Speakers : Taiichiro Shirai (WPI-IFReC) and Takashi Yoshimura (WPI-ITbM, Nagoya University)■■ March 27, 2024: “The forefront of genetic analysis”
Speakers : Daisuke Okuzaki (IFReC) and Daisuke Motooka (RIMD Osaka University)■■ Date :December 3, 2023■■ Venue :Lecture Hall A, Osaka University School of Medicine■■ March 28, 2024: “Mathematical approach to life phenomena”
Speakers : Takahiro Nemoto (WPI-PRIMe) and Kantaro Fujiwara (WPI-IRCN, University of Tokyo)■■ Speakers : Moyu Hasegawa and Kenji Miki (WPI-PRIMe, Osaka University)

86

87Immunology

SpringX Super School - Online Lecture series forprotecting our lives from infectious diseases –by CiDER and IFReC■■ Speakers :・Tokiko Watanabe :“A wide variety of virus research” (April 28, 2023)

・Kazuhiro Suzuki :“Regulation of Autoimmunity” (July. 28, 2023)

・Yasutaka Okabe :“Macrophages protect our health” (Oct. 27, 2023)

・Kiyoharu Fukushima : “Interstitial pneumonia/NTM diseases” (Jan. 26, 2024)

Japanese Language Class
Message from Ms. Tajima, an instructor of the class 2024.
“Hi. I’m Kaori Tajima. It’s been very fun to talk with you in the class. I can’t wait to see new members,
too. The class is focused on speaking. You are expected to talk about yourself, using the grammar and thevocabulary we learn in the class. I’m really looking forward to seeing you!”

Information

88

89

Information

Advanced Postdoc Program at IFReC

Major Awards

IFReC has been recruiting post-doctoral researchers for its Advanced
Postdoc Program. This program offers three-year employment andfunding (3 million JPY per year) for original research to promising
young researchers. Selected applicants have access to continually
upgraded state-of-the-art facilities at IFReC for their research, including
equipment for single-cell analysis.

Highly Cited Researchers (HCR) 2023 by Clarivate TM
Nobuhiko Kamada
Shimon Sakaguchi

Debrecen Award for Molecular
Medicine 2023

Karl Meyer Lectureship Award 2023
by Society for Glycobiology

Shimon Sakaguchi

Taroh Kinoshita"Rising Fellow" by Toyota Physicaland Chemical Research InstituteMochida Memorial Award 2023

Soyoung Park

Tomohiro Kurosaki

Original Support Programs for Young Researchers
To strengthen our international research network and our basis forinternational collaborative research, IFReC has established two kindsof financial support programs for researchers. 1) “IFReC Kishimoto
Foundation Fellowship,” which has been used to invite international
researchers to Osaka. 2) “Program for International Circulation of Young
Talented Researchers” for those who wish to participate in overseas
research activities. Since 2009, about 150 researchers have received
these grants.

Support for Paper Submission
This program aims to support the dissemination of research results by
young researchers of IFReC.

The Ishidate-Ueno Award 2023 by
Chugai Foundation

The 14 th Ikushi Prize by Japan Society
for the Promotion of Science

Yukinori Okada

Emi ItoGrant for Next Generation Principal Investigators
This program aims to foster the next generation of principal investigators at IFReC. In particular, challenging research thathas the potential to create a new field of study in immunology is selected. The grant helped to generate excellent researchachievements in 2023.

Osaka University Prize 2023
Kiyoharu Fukushima
Hiroko Kitamoto

90

91

Information

Common Facilities (IFReC, RIMD, Animal Resource Center)
IFReC and its parent institution, the Research Institute for Microbial Diseases (RIMD) are located on the same
site, constituting a large research complex. The complex contains the Core Instrumentation Facility, the Animal
Resource Center and the Network Administration Office, all of which are jointly operated by IFReC and RIMD. The
Core Instrumentation Facility is equipped with various highly advanced instruments and skilled technicians provide
in-house services to IFReC and RIMD researchers. The Animal Resource Center consists of three buildings for specificpathogen-free (SPF) animals and the live immuno-imaging facility. With a largecapacity animal-breeding facility in
IFReC, researchers are able to choose animal rooms suitable for their experiment purpose. Using these common
facilities, IFReC researchers are able to effectively and smoothly carry out their experiments to promote their worldleading research at IFReC.

Composition & Finance● Composition
Composition since the establishment of IFReC

as of Feb. 1, 2024

350
Researchers (JPN)

Researchers (Overseas)

Administrative and Support Staff

300
97

250

117

102

105

106

92

102

70

200

103

90
90

89
86
55

52

150

6
3

38

100

48

55

50

4

5

2

1

50

54

117

119

128

138

137

130

124

124

117

2011

2012

2013

2014

2015

2016

2017

2018

2019

43

44

51

54

47

54

43

42

24
16
8

53

42

110

129

142

162

151

141

2020

2021

2022

2023 2024

93
65

44

0
2008

2009

2010

Year

❶ IFReC Research Building❷ Integrated Life Science Building❸ Main Building, Research Institute for Microbial Diseases, RIMD

● Finance❹ South Building, Research Institute for Microbial Diseases, RIMD

Sources❺ Cutting-Edge Research Building for Infectious Diseases

Expenditure❻ Animal Resource Center for Infectious Diseases
From the
University
9.5 %Animal Resource Center for Infectious Diseases

Network Administration Office■ Specific pathogen-free (SPF) animal facility■ Provision and maintenance of network infrastructure:■ Sperm/ embryo freezing and preservation

LAN system and servers (web, mail, mailing lists, etc.)

Main Grant
34.1%

JPY
4,523,306,486r■ In vitro fertilization and embryo transplantation■ Intracytoplasmic sperm injection

Core Instrumentation Facility■ Basic and advanced instruments

Live immuno-imaging facility■ In-house service■ DNA sequencing, cell sorting, electron microscopy, mass

■ SPF animal experiment facility with 11.7T MRI, in vivo

imager & two-photon microscope.

92

Personnel
23.9 %

JPY
4,523,306,486

Donation
5.0%■ Transgenic and knock-out animals■ Genome editing in experimental animals

Management
Expenses Grant
19.1 %

Individual
Research
Projects
33.3 %

Grant-in-Aid
for Scientific
Research
9.2%

Contract Research
23.1 %

Equipment
11.2 %

Project
Activities
30.2 %

Travel 1.3 %spectrometry and next-generation sequencing analysis■ Radio isotope facility

93

Information

Selected Articles
Single-cell analyses and host genetics highlight the role of innate immune cells in COVID-19
severity.

Tumor-derived semaphorin 4A improves PD-1–blocking antibody efficacy by enhancing
CD8+ T-cell cytotoxicity and proliferation.

Edahiro R, Shirai Y, Takeshima Y, Sakakibara S, et al.

Naito Y, Koyama S, Masuhiro K, Hirai T, et al.

Nat Gen. 55:753-767 (2023).

Sci Adv. 9 (20): eade0718 (2023).

Mechanisms underpinning the dysfunctional immune
response in severe acute respiratory syndrome coronavirus
2 infection are elusive. Atsushi Kumanogoh (Graduate
School of Medicine, Osaka University/IFReC), Yukinori Okada(Graduate School of Medicine, Osaka University/IFReC/
RIKEN/Graduate School of Medicine, University of Tokyo) andthe research group found that CD14+CD16++ monocytes,
a rare subset among monocytes, are involved in COVID-19
severity. Further, COVID-19 severity-associated genes suchas IFNAR2, have specific function mainly in monocytes anddendritic cells.

Although immune checkpoint inhibitors have brought
major advances in cancer treatment, their effectiveness is still
far from satisfactory. We need to develop novel biomarkers
and therapies that strengthen their effectiveness. Atsushi
Kumanogoh (Graduate School of Medicine, Osaka University/
IFReC), and the research group shows that histologically
Semaphorin 4A (Sema4A)-positive non-small cell lung cancer(NSCLC) responded significantly better to anti-programmed
cell death 1 (PD-1) antibody than Sema4A-negative NSCLC.

Schematic of the role of Sema4A in the tumor microenvironment.
Sema4A expressed by tumor cells ameliorated anti-tumor function
and proliferation of CD8+ T cells by promoting mTORC1-S6K signaling
and polyamine synthesis.

Bacterial induction of B cell senescence promotes age-related changes in the gut microbiota.

94

PGL-III, a rare intermediate of Mycobacterium leprae phenolic glycolipid biosynthesis,
is a potent Mincle ligand.

Kawamoto S, Uemura K, Hori N, Takayasu L, et al.

Ishizuka S, van Dijk JHM, Kawakita T, Miyamoto Y, et al.

Nat Cell Biol. 25:865-876 (2023).

ACS Cent Sci. 9 (7):1388-1399 (2023).

We all experience aging, a phenomenon in which our
biological functions deteriorate. A research group led by
Eiji Hara (Aging Biology, IFReC) has revealed that longterm stimulation by gut microbiota causes the induction ofcellular senescence in immunoglobulin A (IgA)-producing
B cells, resulting in changes in the production and diversityof IgA, which in turn causes dysbiosis of gut microbiota.
This research clarifies the mechanism of changes in the gut
microbiota with aging. Furthermore, it could lead to thedevelopment of methods to improve age-related dysbiosis
of the gut microbiota, which is thought to be one of thecauses of various age-related diseases.

Although leprosy (Hansen's disease) is one of the oldest
known diseases, the pathogenicity of Mycobacterium
leprae (M. leprae) remains enigmatic. Indeed, the cell wallcomponents responsible for the immune response against
M. leprae are as yet largely unidentified. The research groupof Sho Yamasaki (Molecular Immunology, IFReC/RIMD/CiDER,
Osaka University) identified phenolic glycolipid-III (PGL-III) asa M. leprae-specific ligand for the immune receptor Mincle.

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Information

Single-cell transcriptomics of human cholesteatoma identifies an activin A-producing
osteoclastogenic fibroblast subset inducing bone destruction.

Circadian ribosome profiling reveals a role for the Period2 upstream open reading frame insleep.

Shimizu K, Kikuta J, Ohta Y, Uchida Y, et al.

Millius A, Yamada R, Fujishima H, Maeda K, et al.

Nature Commun. 14:4417 (2023).

Proc Natl Acad Sci U S A. 120 (40): e2214636120 (2023).

Cholesteatomas are made up of cysts or bumps in theear that consist of skin, collagen fibers, skin cells, fibroblasts,
keratin, and dead tissue. However, the exact mechanism
for the creation of cholesteatomas remains unknown. The
research group of Masaru Ishii (Graduate School of Medicine/
Graduate School of Frontier Biosciences/IFReC) revealed thecause of cholesteatomas, which may help in developing new
therapies for patients who are suffering from this disease.

Circadian rhythms, the internal biological clocks that
regulate our daily activities, are essential for maintaining
health and well-being. The research group of Arthur Millius(RIKEN/present: IFReC), Kazuhiko Maeda (Host Defense,
IFReC), Daron Standley (System Immunology, IFReC), andHiroki Ueda (RIKEN) showed how translation and posttranscriptional processes influence the body's internal clock
and its impact on sleep patterns.

Schematic of osteoclastogenesis induced by cholesteatoma fibroblasts
expressing activin A.

Disruption of the Per2 uORF disrupts the amplitude of circadianrhythms (left) and reduces sleep in mice (right).

IL-1β, PGE2, and TNF-α secreted from infiltrating CD45+ cells, particularly
macrophages, induced activin A-expressing pathogenic fibroblasts; theactivin A acted in conjunction with RANKL to promote ectopic
osteoclastogenesis. Construction of a T-cell receptor signaling range for
spontaneous development of autoimmune disease.

Opposing roles of RUBCN isoforms in autophagy and memory B cell generation.

STIM-mediated calcium influx regulates maintenance and selection of GC B cells.

Tsai C, Sakakibara Y, Kuan Y, Omori H, et al.

Yada Y, Matsumoto M, Inoue T, Baba A, et al.

Sci Signal. 16:803 (2023).

J Exp Med. 221 (1): e20222178 (2024).

Memory B cells depend on autophagy for their survival,
but the protein Rubicon is thought to hinder this process. Dr.
Chao-Yuan Tsai and the researchers of Osaka University havediscovered a shorter isoform of Rubicon called RUBCN100,
which enhances autophagy in B cells. Mice that lacked thelonger isoform, RUBCN130, produced more memory B cells
in a way that relied on autophagy. These findings provide
further insight into the role of Rubicon in autophagy.

Germinal centers (GCs) are specialized microenvironments
where antigen-specific B cells undergo antibody affinity
maturation and clonal expansion. However, the requirementsof BCR signaling in GC B cells remain poorly understood.
Yoshihiro Baba (Kyushu University), Tomohiro Kurosaki(Lymphocyte Differentiation, IFReC) and the research group
showed that stromal interacting molecule (STIM)-deficient B
cells have reduced B cell competitiveness compared to wildtype B cells during GC responses.
A model for the functional interaction between RUBCN100 and
RUBCN130 in regulating autophagy.

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Publications
Gp130-HIF1α axis-induced vascular damage is prevented by the short-term inhibition of
IL-6 receptor signaling.

1

Abbaoui A, Fatoba O, Yamashita T. Meningeal T cells function in
the central nervous system homeostasis and neurodegenerative
diseases. Frontiers in Cellular Neuroscience 17, 1181071 (2023).

19

Choi HS, Yum JH, Jeong Y, Lim S, Kumagai T, Cha HJ, Park S.
Fluorogenic bisazide cyanine probe as a highly efficient acrolein
detection tool for diagnosing triple negative breast cancer.
Sensors and Actuators B-Chemical 380, 133404 (2023).

2

Abe S, Asahi T, Hara T, Cui GW, Shimba A, Tani-ichi S, Yamada K,
Miyazaki K, Miyachi H, Kitano S, Nakamura N, Kikuta J, Vandenbon
A, Miyazaki M, Yamada R, Ohteki T, Ishii M, Sexl V, Nagasawa T, Ikuta
K. Hematopoietic cell-derived IL-15 supports NK cell developmentin scattered and clustered localization within the bone marrow.
Cell Reports 42, 113127 (2023).

20

Cui MY, Yamano K, Yamamoto K, Yamamoto-Imoto H, Minami S,
Yamamoto T, Matsui S, Kaminishi T, Shima T, Ogura M, Tsuchiya
M, Nishino K, Layden BT, Kato H, Ogawa H, Oki S, Okada Y, Isaka Y,
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3

Agata SN. Cloning of human Type I interferon cDNAs. Proceedingsof The Japan Academy Series B-Physical and Biological Sciences
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4

Akiyama Y, Sonehara K, Maeda D, Katoh H, Naito T, Yamamoto K,
Morisaki T, Ishikawa S, Ushiku T, Kume H, Homma Y, Okada Y, Biobank
Japan Project. Genome-wide association study identifies risk lociwithin the major histocompatibility complex region for Hunner-type
interstitial cystitis. Cell Reports Medicine 4, 101114 (2023).

21

Dangerfield EM, Ishizuka S, Kodar K, Yamasaki S, Timmer MSM,
Stocker BL. Chimeric NOD2 Mincle Agonists as Vaccine Adjuvants.
Journal of Medicinal Chemistry 67, 5373-5390 (2024).

22

De Vincentis A, Tavaglione F, Namba S, Kanai M, Okada Y, Kamatani
Y, Maurotti S, Pedone C, Incalzi RA, Valenti L, Romeo S, VespasianiGentilucci U. Poor accuracy and sustainability of the first-step
FIB4 EASL pathway for stratifying steatotic liver disease risk in thegeneral population. Alimentary Pharmacology & Therapeutics 59,
1402-1412 (2024).

5

Alshoubaki YK, Lu YZ, Legrand JMD, Karami R, Fossat M, Salimova E,
Julier Z, Martino MM. A superior extracellular matrix binding motif
to enhance the regenerative activity and safety of therapeuticproteins. Npj Regenerative Medicine 8, 25 (2023).

6

Aoyama R, Nakagawa S, Ichikawa Y, Inohara N, Yamazaki Y, Ito T,
Sugihira T, Kono M, Akiyama M, Takahashi H, Takaya A, Ichikawa
F, Nakano T, Tanaka S, Koyano Y, Fujimoto M, Núñez G, Shimojo N,
Nakamura Y. Neonatal skin dysbiosis to infantile atopic dermatitis:
Mitigating effects of skin care. Allergy (2024).

23

Dede EÇ, Gizer M, Korkusuz F, Bal Z, Ishiguro H, Yoshikawa H,
Kaito T, Korkusuz P. A pilot study: Nano-hydroxyapatite-PEG/
PLA containing low dose rhBMP2 stimulates proliferation andosteogenic differentiation of human bone marrow derived
mesenchymal stem cells. Jor Spine 6, e1258 (2023).

7

Arai M, Fukuda A, Morimoto R, Nakamura Y, Ci ZH, Sakaguchi
S. Protocol to evaluate cell lineage stability of mouse naturaland induced regulatory T cells using bisulfite sequencing. Star
Protocols 3, 101694 (2022).

24

Duperron MG, et al. Genomics of perivascular space burden
unravels early mechanisms of cerebral small vessel disease. Nature
Medicine (2023).

25

Edahiro R, et al. Single-cell analyses and host genetics highlightthe role of innate immune cells in COVID-19 severity. Nature
Genetics 55, 753 (2023).

8

Arai Y, Yamanaka I, Okamoto T, Isobe A, Nakai N, Kamimura N,
Suzuki T, Daidoji T, Ono T, Nakaya T, Matsumoto K, Okuzaki D,
Watanabe Y. Stimulation of interferon-b responses by aberrant
SARS-CoV-2 small viral RNAs acting as retinoic acid-inducible
gene-I agonists. Iscience 26, 105742 (2023).

26

9Arase N, Sasaoka Y, Narita J, Kiyohara E, Hashimoto K, Shinzaki S, Nojima
S, Takagi J, Fujimoto M. Anti-α6β4 integrin autoantibodies inhibit thebinding of laminins to α6β4 integrin in patients with pemphigoid
and affect the gastrointestinal tract. Journal of The European
Academy of Dermatology and Venereology 38, 404-412 (2024).

Ekemen S, Bilir E, Soultan HEA, Zafar S, Demir F, Tabandeh B,
Toprak S, Yapicier O, Coban C. The Programmed Cell Death Ligand
1 and Lipocalin 2 Expressions in Primary Breast Cancer and Their
Associations with Molecular Subtypes and Prognostic Factors.
Breast Cancer-Targets and Therapy 16, 1-13 (2024).

27

Emori C, Kodani M, Abbasi F, Mori M, Ikawa M. PABPN1L is required
for maternal mRNA degradation after meiosis resumption. Journalof Reproduction and Development 70, 10-17 (2024).

10

Azekawa S, et al. Serum KL-6 levels predict clinical outcomes andare associated with MUC1 polymorphism in Japanese patients with
COVID-19. Bmj Open Respiratory Research 10, e001625 (2023).

28

11

Bassler K, Schmidleithner L, Shakiba MH, Elmzzahi T, Koehne
M, Floess S, Scholz R, Ohkura N, Sadlon T, Klee K, Neubauer
A, Sakaguchi S, Barry SC, Huehn J, Bonaguro L, Ulas T, Beyer
M. Identification of the novel FOXP3-dependent Treg celltranscription factor MEOX1 by high-dimensional analysis ofhuman CD4+ T cells. Frontiers in Immunology 14, 1107397 (2023).

Endo T, Kobayashi K, Matsumura T, Emori C, Ozawa M, Kawamoto
S, Okuzaki D, Shimada K, Miyata H, Shimada K, Kodani M, IshikawaYamauchi Y, Motooka D, Hara E, Ikawa M. Multiple ageing effects
on testicular/epididymal germ cells lead to decreased malefertility in mice. Communications Biology 7, 16 (2024).

29

Fisch D, Pfleiderer MM, Anastasakou E, Mackie GM, Wendt F, Liu XY,
Clough B, Lara-Reyna S, Encheva V, Snijders AP, Bando H, Yamamoto
M, Beggs AD, Mercer J, Shenoy AR, Wollscheid B, Maslowski KM, Galej
WP, Frickel EM. PIM1 controls GBP1 activity to limit self-damage and to
guard against pathogen infection. Science 382, 67eadg2253 (2023).

12

Ben-Shalom N, Sandbank E, Abramovitz L, Hezroni H, Levine T,
Trachtenberg E, Fogel N, Mor M, Yefet R, Stoler-Barak L, Hagin D,
Nakai A, Noda M, Suzuki K, Shulman Z, Ben-Eliyahu S, Freund NT.
132-adrenergic signaling promotes higher-affinity B cells andantibodies. Brain Behavior and Immunity 113, 66-82 (2023).

30

Fukuda M, Fujita Y, Hino Y, Nakao M, Shirahige K, Yamashita T.
Inhibition of HDAC8 Reduces the Proliferation of Adult Neural
Stem Cells in the Subventricular Zone. International Journal of
Molecular Sciences 25, 2540 (2024).

13

Butler-Laporte G, et al. Exome-wide association study to identify
rare variants influencing COVID-19 outcomes: Results from the
Host Genetics Initiative. PLoS Genetics 18, e1010367 (2022).

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14Campos AI, Namba S, Lin SC, Nam K, Sidorenko J, Wang HW,
Kamatani Y, Wang LH, Lee S, Lin YF, Feng YCA, Okada Y, Visscher
PM, Yengo L. Boosting the power of genome-wide association
studies within and across ancestries by using polygenic scores.
Nature Genetics 55, 1769 (2023).

Fukushima K, Kudo H, Oka K, Hayashi A, Onizuka M, Kusakabe
S, Hino A, Takahashi M, Takeda K, Mori M, Ando K, Hosen N.
Clostridium butyricum MIYAIRI 588 contributes to the maintenanceof intestinal microbiota diversity early after haematopoietic celltransplantation. Bone Marrow Transplantation (2024).

32

Fukushima K , et al. MGIT-seq for the Identification of
Nontuberculous Mycobacteria and Drug Resistance: a Prospective
Study. Journal of Clinical Microbiology 61 (2023).

15

Cao SY, Liu YS, Gao XD, Kinoshita T, Fujita M. A lipid scramblase
TMEM41B is involved in the processing and transport of GPIanchored proteins. Journal of Biochemistry 174, 109-123 (2023).

33

Funakoshi K, Kato Y, Narazaki M, Kumanogoh A. Paradoxical
improvement of dyspnoea in relapsing polychondritis by
glucocorticoid tapering. Bmj Case Reports 17, e257861 (2024).

16

Chalalai T, et al. TRAF6 signaling in dendritic cells plays protectiverole against infectious colitis by limiting C. rodentium infection
through the induction of Th1 and Th17 responses. Biochemical
and Biophysical Research Communications 669, 103-112 (2023).

34

Furukawa A, Shuchi Y, Wang JQ, Guillen-Poza PA, Ishizuka S,
Kagoshima M, Ikeno R, Kumeta H, Yamasaki S, Matsumaru T, Saitoh
T, Maenaka K. Structural basis for plastic glycolipid recognition of
the C-type lectin Mincle. Structure 31, 1077 (2023).

17

Chen SW, et al. A genomic mutational constraint map using
variation in 76,156 human genomes. Nature 625 (2024).

35

18

Chiang MR, Shen WT, Huang PX, Wang KL, Weng WH, Chang
CW, Chiang WH, Liu YC, Chang SJ, Hu SH. Programmed T cells
infiltration into lung metastases with harnessing dendritic cells incancer immunotherapies by catalytic antigen-capture sponges.
Journal of Controlled Release 360, 260-273 (2023).

Ga H, Taguchi A, Honjoh H, Nishijima A, Eguchi S, Miyamoto Y,
Sone K, Mori M, Osuga Y. Prognosis of patients with endometrial
cancer or atypical endometrial hyperplasia after complete
remission with fertility-sparing therapy. Archives of Gynecology
and Obstetrics 308, 1629-1634 (2023).

36

Hashimoto R, Minoshima M, Kikuchi K. Rational Design ofHydroxylated Thiazole Orange Photocages for Green LightTriggered DNA Recombination. Chembiochem 25 (2024).

Kang S, Onishi S, Ling Z, Inoue H, et al.

Proc Natl Acad Sci U S A. 121(2): e2315898120 (2024).
Interleukin (IL)-6, an essential indicator of cytokine
release syndromes (CRS), regulates vascular homeostasis
and inflammation. Inhibition of IL-6 receptor (IL-6R)
signaling is beneficial for various CRS; however, it is limited
by adverse effects related to poor understanding of
mechanisms involved. Tadamitsu Kishimoto and Sujin Kang,and their research group discovered that hypoxia-inducible
factor (HIF1) α signaling is activated by IL-6R trans-signaling
in endothelial cells, which promotes vascular inflammatory
responses and endothelial permeability by glycolysis.

Giving medication a short half-life anti-IL-6 to patients ofSepsis, ARD, Burns, etc. is expected to suppress vasculardamage or secondary infection.

CGRP sensory neurons promote tissue healing via immune cells.
Lu YZ, Nayer B, Singh SK, et al.

Nature. (2024).
Nociceptive sensory neurons have a crucial role as
immunoregulators and exert both protective and harmful
effects depending on the context. However, how neuro–
immune interactions affect tissue repair and regeneration
following acute injury is unclear.
Collaborating with Shizuo Akira (Host Defense, IFReC), a
research team led by Mikaël Martino (Monash University)
demonstrated that the removal of a specific subtypeof sensory neurons containing the Nav1.8 ion channel
significantly impairs skin wound repair and muscle
regeneration following injury.

Graphical summary of the study by Mikaël Martino.

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He ZY, Tung NTC, Yahara Y, Makino H, Yasuda T, Seki S, Suzuki K,
Futakawa H, Kamei K, Kawaguchi Y. Association between serum
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Hirai T, et al. Sema6D forward signaling impairs T cell activation andproliferation in head and neck cancer. Jci Insight 9, e166349 (2024).
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Ihara F, Yamamoto M. The role of IFN-γ-mediated host immune
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Ikazaki T, Ishikawa E, Tamashima H, Akiyama H, Kimuro Y, Yoritate M,
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Ise W, Kurosaki T. Tissues of origin matter to plasma cell longevity.
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Ishigaki K, et al. Multi-ancestry genome-wide association analyses
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103

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