Kyudai Oral Bioscience&
OBT Research Center
Joint International Symposium 2019
PROGRAM & ABSTRACTS

March 2 – 3, 2019
Lecture Room A/B, Faculty of Dental Science,
Kyushu University

K OB

2019

■ Date:March 2-3, 2019
■ Venue:

Lecture Room A/B, Faculty of Dental Science,
Kyushu University, Fukuoka, Japan■ Organization

Kyudai Oral Bioscience
Oral Health・Brain Health・Total Health Research Center■ Contact

Eijiro Jimi
Oral Health・Brain Health・Total Health Research Center
Laboratory of Molecular and Cellular Biochemistry,
Faculty of Dental Science, Kyushu University,
Fukuoka 812-8582, Japan
Tel & Fax: +81 92-642-6332
E-mail: ejimi@dent.kyushu-u.ac.jp

KOB & OBT Joint International Symposium 2019

PROGRAMKOB & OBT Joint International Symposium 2019

Session 1
Oral Health and Brain Health

Chaired by Dr. Zhou Wu

Critical roles of Cathepsin B in linkage
between periodontitis and neuroinflammation
Junjun Ni 1, Zhou Wu 1,2
1

Department of Aging Science and Pharmacology, 2OBT Research Center
Faculty of Dental Science, Kyushu University.

Neuroinflammation, inflammation of the brain, is strongly implicated in Alzheimer’s
disease (AD), which can be enhanced by systemic inflammation. The concept suggests a
possible link between periodontitis and AD because periodontitis is an oral chronic
infection that elicits systemic inflammatory responses. There is now growing clinicalevidence that chronic periodontitis is closely linked to the initiation and progression of
AD. Chronic periodontitis can be a significant source of covert systemic inflammation
within the general population, because a high percentage of adults are suffering fromperiodontitis, and the prevalence of periodontitis increases with age. In the present talk,we will introduce our recently works on clarifying the mechanisms linking between
periodontitis and AD, thereby helping to understand how does oral infection impact on
brain.
We have recently found that Cathepsin (Cat) B plays a critical role in inducing ADlike phenotypes, including microglia-mediated neuroinflammation, intracellular Aβ
accumulation in neurons and impairment of the learning and memory functions in the
middle-aged mice following chronic systemic exposure to LPS from P.gingivalis(PgLPS). The microglia CatB- dependent in PgLPS-mediated intracellular Aβ
accumulation in neuron. We have clarified the novel mechanisms of CatB involving in
microglia-mediated neuroinflammation. Firstly, CatB involves in chronically NF-kB
activation in microglia,
result in prolonging microglia-mediated neuroinflammation, because CA-074Me, thespecific CatB inhibitor, inhibits the autophagic IkBa degradation. Secondly, CatB
involves in microglia aging, because leakage of CatB in microglia during aging are
responsible for mitochondria-derived reactive oxygen species (ROS) generation result in
proinflammatory mediator production. The CatB-dependent microglia aging promotion
are important for periodontitis inducing AD-like phenotypes, because the senescent-type
microglia respond in an exaggerated manner to systemic inflammation.
In conclusion, the novel mechanisms of CatB in involving microglia-mediatedneuroinflammation are dependent on mitochondria-derived ROS generation and NF-kB
activation. We propose that CatB may as a therapeutic target for preventing periodontitisassociated cognitive decline in AD.

The mechanism of periodontitis link to Alzheimer’s disease:gingipains involvement
Yicong Liu
Zhe Jiang University, China
Alzheimer’s disease (AD) is the world’s most common neurodegenerative disease. Althoughclinical evidence has already showed the positive link between chronic periodontitis and cognitive
decline in AD, the precise mechanism underlying the relationship remains unclear.
Porphyromonas gingivalis (P.g), the keystone pathogen in chronic periodontitis, has been identified
in the brain of AD patients, and the level of the toxic cysteine proteinases from P.g called gingipains
has been found to correlate with tau and ubiquitin pathology. We hypothesized that Gingipains may
activate microglia, brain-resident immune cells, directly resulting in neuroinflammation. Gingipains
comprises Rgp and Kgp, and both Rgp and Kgp are mediated host cell responses and the subsequent
intracellular signaling in the infected cells. In the present talk, I will introduce our recently works forclarifying the potential effects of Rgp and Kgp on microglia activation. We provide the first evidence
that Rgp and Kgp cooperatively contribute to the P. g-induced cell migration and expression ofproinflammatory mediators through the activation of PAR2. The subsequent activation of PI3K/Akt andmitogen-activated protein kinase/ERK kinase/ERK pathways contributes to cell migration andinflammatory response of microglia.
It is known that female not only have a higher risk of developing AD, but also showed significantly
age-related faster decline than male. Finally, I will introduce our ongoing works on exploring genderdifference of microglia-mediated neuroinflammation during P.g infection. We found that the antiinflammatory mediators in the P.g infected microglia, including TGFb1, Arginase-1 and IL-10 weresignificantly lower in the P.g infected microglia from female aged mice brain than that of from maleaged ones, and the estrogen receptor (ER)-a agonist increased the Arginase-1 expression in the P.g
infected primary microglia from the aged female mice brain.
In conclusion, gingipains may be key mediators of periodontitis link to AD, gingipains inhibitors
could be valuable for treating periodontitis-related neurodegeneration in AD. And ER-a agonist could
be beneficial for reducing the risk of AD in old female individuals by suppressing the microgliamediated neuroinflammation.

Spontaneous Activity-Dependent Discrete Circuit Formation in theDeveloping Olfactory Bulb
Satoshi Fujimoto
Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan

In the mouse olfactory system, odor information detected by ~1000 types of olfactory
receptors is represented by the glomerular map on the olfactory bulb. Mitral cells, which
are second-order neurons, extend multiple dendrites to multiple glomeruli in the earlystages of development, but dendrites are pruned as development progresses and finallyconnect one primary dendrite to one glomerulus. However, it remains unknown how thisdiscrete connectivity is established. Here we found that dendrite pruning was perturbed
by silencing neuronal activity in mitral cells, while the pruning occurred normally underthe blockade of activity derived from olfactory sensory neurons. Spontaneous activity ofthe mitral cell was directly observed by two photon calcium imaging of awake neonatesin vivo and even isolated olfactory bulbs in vitro. We also found that the pattern of activity
becomes decorrelated during development. Pharmacological experiments revealed thatspontaneous activity is mediated by glutamatergic synaptic transmission and gap junction.
Dendrite pruning was suppressed in the mitral cells lacking NMDA receptors. Thus, thespontaneous activity generated in the olfactory bulb establishes the discrete olfactory
circuit.

Exploring Potential Therapies for Alzheimer’s DiseaseZhenzhen Quan1, Hong Qing1
Beijing Institute of Technology, China

Alzheimer’s disease (AD) is a progressive neurodegenerative disease which affects atleast 47 million people worldwide as per World Alzheimer Report of 2016. Currently,
there are no specific drugs or therapies that could cure AD effectively. Revealing themechanism and exploring effective therapy for ND are the hotspots for current research.
My topic is mainly based on two parts: 1) NSC transplantation on AD study. We initiallyapproved that NSC transplantation significantly improves the ability of short-term
memory and certainly ameliorates the ability of long-term memory in tau/tta mice; wealso found that NSC transplantation could reduce the number of neurofibrillary tangles inhippocampal CA1 region; the transplanted NSC could differentiate into neurons andmicroglia. These indicated NSC transplantation has the potential to improve the ability oflearning and memory in AD model mice which is possibly due to its ability of integrating
into the impaired neural circuits and eliminating the neurofibrillary tangles; 2)
Application of optogenetics on AD study. Previous studies have indicated the importantrole of CA3 on learning and memory, our recent studies found that, manipulation ofhippocampal CA3 firing via luminopsins significantly modulates spatial and episodic
short-term memory, especially working memory. Regarding this, we further found thatmanipulation of CA3 by optogenetics could ameliorate the cognitive dysfunctions of AD
model mice through improvement of the synapse plasticity of CA3 circuits.

KOB & OBT Joint International Symposium 2019

Session 2
Oral Health

Chaired by Dr. Masafumi Moriyama

The effect of systemically-administered Mesenchymal stem cells for
peri-implant Mucosa
Ikiru Atsuta1*, Yasunori Ayukawa1, Nobuyuki Ueda1, Takayoshi
Yamaza2, Yuri Matsuura1, Ryosuke Kondo1, Kiyoshi Koyano1
1Section of Implant and Rehabilitative Dentistry, Division of Oral Rehabilitation,
Faculty of Dental Science, Kyushu University, Fukuoka, Japan2
Department of Molecular Cell and Oral Anatomy, Faculty of Dental Science, Kyushu
University, Fukuoka, Japan

Dental implant therapy is one of the most important prosthodontic options for edentulous
patients. Dental implants based on the concept of “osseointegration”, a term explaining
the fixation of a titanium implant in the bone, have shown dramatic continual
improvement. However, the peri-implant tissue is always exposed to the possibility of
inflammation because the titanium body penetrates into the oral mucosa. Actually, theperi-implant tissue has many close similarities to normal gingiva, but many reports havenoted that the ability of the peri-implant epithelium to seal against the oral environment
is poor. Therefore, the improvement of local defense in mucosa is strongly expected toenable successful outcomes in oral implant patients.
In this study, we have attracted attention to mesenchymal stem cells (MSCs), because
these cells generate a great deal of interest in many clinical settings, including immunomodulation and tissue engineering, where studies have demonstrated the feasibility oftransplanted MSCs becoming a new mode of cellular therapy. However, the relationshipbetween MSC-based therapy and PIE-implant interface sealing is not well understood.
The hypothesis of the present study was that systemic MSCs accumulate around theimplant in the early stage and promote PIE formation and soft tissue attachment to theimplant surface.

A Novel disease discovered and established in 21th Century:
IgG4-related disease-Mechanistic insights from both clinical and immunologic
understanding of this condition Takashi Maehara1, Hamid Mattoo2, Vinay Mahajan2, Emanuel DellaTorre3, Cory Perugino4, Masafumi Moriyama1,5, Miho Ohta1, Naoki
Kaneko2, Noriko Ishiguro1, Akira Chinju1, Ryusuke Munemura1, John
Stone4, Shiv Pillai2, Seiji Nakamura1.
1

Section of Oral and Maxillofacial Oncology, Division of Maxillofacial Diagnostic andSurgical Sciences, Faculty of Dental Science, Kyushu University, Fukuoka, Japan
2
Ragon Institute of MGH, MIT, and Harvard, Massachusetts General Hospital,
Harvard Medical School, Boston, MA, USA
3
Universita Vita-Saluta San Raffaele, IRCCS San Raffaele Scientific Institute, Milan, Italy
4
Division of Rheumatology, Allergy and Immunology, Massachusetts General Hospital,
Harvard Medical School, Boston, Massachusetts, USA
5
Faculty of Dental Science, OBT Research Center, Kyushu University, Fukuoka, Japan

Objective: IgG4-related disease (IgG4-RD) is a chronic, systemic inflammatory
condition with T cells and polarized class-switching to IgG4 secreting B cells withinsevere tissue fibrosis. Important mechanistic insights in this disease have been derived
from studies of B-cell depletion therapy. Actually CD4+T cells are the most abundant
cells within tissue lesions, so disease-specific CD4+T cells could provide help to specific
B cells during T-dependent immune responses. We thus investigated disease-related
CD4+T cells in this disease on International IgG4-RD collaboration research.
Methods: We used flow cytometry to identify CD4+ effector/memory T cells in 101
patients with IgG4-RD. Next-generation sequencing of the T cell receptor β chain gene
was used to identify clonal expansions of effector CD4+T cells. We use transcriptomes ofisolating CD4+ cytotoxic T lymphocytes (CD4+CTLs). Furthermore, by using IL-4
capture assay, we use transcriptomes of isolating IL-4 secreting Tfh cells based on theirfunction. Tissue infiltration by these T cells was examined using quantitative multi-color
imaging in IgG4-RD subjects compared with healthy controls and other diseases.
Results: CD4+ effector/memory T cells were expanded in IgG4-RD. Next-generation
sequencing revealed prominent clonal expansions of CD4+CTLs and Tfh cells in subjects
with IgG4-RD. Clonally-expanded CD4+CTLs that expressed pro-fibrotic cytokine. IL-4
expressing Tfh cells are surprisingly sparse in human secondary lymphoid organs (SLOs).
Notably, most Tfh cells in IgG4-RD expressed IL-4. IL-4+Tfh cells can be detected incell-cell contact with IgG4+B cells, some of these B cells express AID in SLOs and TLOs.
The proportion of IL-4+Tfh cells in IgG4-RD tissues correlates tightly with tissue IgG4
plasma cell numbers and plasma IgG4 levels in patients but not with the total plasma
levels of other isotypes. The numbers of CD4+CTLs declined during clinical responses to
B cells depletion therapy, suggesting a role of these cells in the pathogenesis of thisdisease.
Conclusions: IgG4-RD is prominently linked to the tissue infiltration of clonallyexpanded CD4+CTLs and IL-4+Tfh cells. Especially 1) reactivation of CD4+ CTLs might
require antigen presentation by activated B cells in tissue sites, 2) disease-related Tfh
subpopulation in SLOs and TLOs that is linked to IgG4 class switching.

PTHrP-PPR autocrine regulation of mesenchymal progenitor cell fateorchestrates tooth eruption
Akira Takahashi1,2, Yoichiro Ogino1, Kiyoshi Koyano1, Noriaki Ono2,
Wanida Ono2
1

Division of Oral Rehabilitation, Faculty of Dental Science, Kyushu University
Department of Orthodontics and Pediatric Dentistry, University of Michigan,
School of Dentistry2

Formation of functional skeletal tissues requires highly organized steps of mesenchymal
progenitor cell differentiation. The dental follicle (DF), a sac-like membranous tissue
surrounding the developing tooth bud, harbors mesenchymal progenitor cells for various
differentiated cells constituting the tooth root-bone interface, and coordinates tooth
eruption in a manner dependent on signaling by parathyroid hormone-related peptide(PTHrP) and its receptor, PTH/PTHrP receptor (PPR). In humans, primary failure of tooth
eruption, a rare autosomal dominant disorder that exclusively affects tooth eruption, ischaracterized by a cessation of tooth eruption before emergence despite an unobstructed
eruption path. PFE is caused by loss-of-function mutations in PPR. However, the identityof mesenchymal progenitor cells in the DF and how they are regulated by PTHrP-PPR
signaling remain unknown. Here we show that the PTHrP-PPR autocrine signal maintains
physiological cell fates of DF mesenchymal progenitor cells to establish the functional
periodontal attachment apparatus and orchestrates tooth eruption. A single cell RNA-seq
analysis revealed cellular heterogeneity of PTHrP+ cells, wherein PTHrP+ DF
subpopulations abundantly expressed PPR. Cell lineage analysis using tamoxifeninducible PTHrP-creER mice revealed that PTHrP+ DF cells differentiated into
cementoblasts on the acellular cementum, periodontal ligament (PDL) cells and alveolar
cryptal bone osteoblasts during tooth root formation. PPR-deficiency induced a cell fateshift of PTHrP+ DF mesenchymal progenitor cells to non-physiological cementoblast-likecells precociously forming the cellular cementum on the root surface associated withupregulation of Mef2c and matrix proteins, resulting in loss of the proper periodontal
attachment apparatus and primary failure of tooth eruption closely resembling human
genetic conditions caused by PPR mutations. These findings reveal a unique mechanism
whereby proper cell fates of mesenchymal progenitor cells are tightly maintained by anautocrine system mediated by PTHrP-PPR signaling to achieve functional formation ofskeletal tissues.

KOB & OBT Joint International Symposium 2019

Special Lecture

Chaired by Prof. Eijiro Jimi

From Kyushu to Okayama: My research history
Ryusuke Yoshida
Department of Oral Physiology, Graduate School of Medicine, Dentistry andPharmaceutical Sciences, Okayama University

After I obtained a Ph.D. in Science from Kobe University in 2002, my research career inthe taste field started as a post-doctoral fellow at the professor Ninomiya’s laboratory in
Kyushu University. At that time, my main research theme was to analyze molecular andphysiological properties of taste receptor cells. To do this, a predecessor and I developed
the experimental setup for recording taste cell responses, which is able to apply taste
stimuli restricted to the apical side with keeping the structure of taste buds. After
recording taste responses, the taste cell was collected from a taste bud and subjected with
RT-PCR to analyze molecular expression. I had a hard time to establish this system; I
spent about 3 years for the first publication of a paper in this field. This technique wasgreatly improved by using transgenic mice expressing fluorescent protein in a subset of
taste bud cells and laser scanning microscopy. This technique has further advantages; weare able to measure transmitter release from taste cells and also able to analyze the effectof endocrine factors on taste cell responses since apical and basolateral side of a taste bud
is completely segregated in this recording system. Now, famous researchers in taste fieldof USA comment on this technique as “Technical tour de force”. In 2016, I was selected
as a member of OBT research center. This would be a very important step to get current
position in Okayama University. In addition, many members in Kyushu University kindly
supported for this personnel affair. From October in 2018, I moved to Okayama
University as a professor in Department of Oral Physiology. Although I still have not get
deeply into my research works because of many other works, problems in man power,
etc., now I ready to take a new step in Okayama University. I hope that this talk will be
as reference for and encourage young researchers.

KOB & OBT Joint International Symposium 2019

Session 3
UGM Summer Course Report

Chaired by Prof. Fusanori Nishimura

KOB & OBT Joint International Symposium 2019

Session 4
Graduate Student’s Session

Chaired by

TLR8+CD68+monocyte/macrophage contribute to the pathogenesis of
Sjögren’s syndrome via TNF-α secretion.
Mizuki Sakamoto, Masafumi Moriyama, and Seiji Nakamura
1

OBT Research Center, 2Section of Oral and Maxillofacial Oncology, Division of
Maxillofacial Diagnostic and Surgical Sciences, Faculty of Dental Science, Kyushu
University, Fukuoka, Japan
Objectives
Sjögren’s syndrome (SS) is an autoimmune disease characterized by lymphocytic
infiltration into the salivary and lacrimal gland with the concomitant destruction ofglandular tissue. Recent studies indicate that Toll like receptors (TLRs) play animportant role in the initiation of autoimmune diseases including rheumatoid arthritis
and systemic lupus erythematosus. In this study, we thus addressed to identify thedisease-associated TLR related molecules in SS, and then examined both expression andfunction of those molecules.
Methods
Gene expression was analyzed by using DNA microarray in labial salivary glands from
3 patients with typical SS and 3 healthy controls (HC). DNA microarray analysis was
performed in the two groups to screen for TLR family (TLR1-TLR10). The candidate
TLRs were validated by real-time PCR and immunohistochemical staining in SS(n=19), and HC (n=10). Moreover, the productivity of TNF-α in human monocyte cellline (U937) was examined by ELISA. Finally, we established human Tlr8
transgenic/mouse Tlr8 knock-out (huTlr8 Tg/mTlr8 KO) mice and confirmed theirphenotypes in each organ (submandibular gland, lung, kidney, pancreas, and liver).
Results
DNA microarray and PCR validated the up-regulation of TLR8 in SS compared with
HC. Immunofluorescence double staining confirmed that the expression of TLR8
almost merged with that of CD68, known as a monocyte/macrophage marker.
Moreover, TNF-α productivity of U937 was significantly increased by over-expressionof TLR8 in comparison with those of untreated U937 and TLR8 KO U937. In vivo,
huTlr8 Tg/mTlr8 KO mice showed strong infiltration of inflammatory cells insubmandibular glands and lungs compared with Wild-type mice.
Conclusion
TLR8 is considered to recognize self-RNA as well as viral RNA and then accelerates
autoimmune responses via production of inflammatory cytokines, such as TNF-α.
Therefore, our current data suggest that TLR8+CD68+ monocyte/macrophages might
promote the initiation of SS via TNF-α secretion.

Characterization of disrupted microbiota on the tongue dorsum
observed in frail elderly adults
Mikari Asakawa1, Toru Takeshita1,2, Shinya Kageyama1, Kenji
Takeuchi1,2, Michiko Furuta1, Shino Suma1, Yoshihisa Yamashita11Section of Preventive and Public Health Dentistry, Division of Oral Health, Growthand Development and 2OBT Research Center, Faculty of Dental Science, Kyushu
University, Fukuoka, Japan.

Tongue microbiota is a primary source of numerous microorganisms constantly ingested
with saliva, and the overgrowth of non-commensal bacteria due to the disruption of the
tongue microbiota would threaten the health of elderly adults with swallowing
impairment. This study investigated tongue microbiota composition of 325
institutionalized frail elderly adults aged ≥70 years by 16S rRNA gene ampliconsequencing and explored disrupted tongue microbiota in the elderly adults.
The tongue microbiota of most individuals was constituted by well-known predominantoral bacterial taxa such as Streptococcus salivarius, Rothia mucilaginosa, and Veillonella
atypica, whereas Streptococcus agalactiae, Corynebacterium simulans, and Actinomyces
radicidentis deposited as non-oral bacterial taxa in the expanded Human Oral
Microbiome Database (eHOMD 16S rRNA RefSeq version 15.1) occupied ≥10% in the
microbiota of the 24 individuals. These individuals demonstrated significantly poorer
physical function and nutritional condition than the individuals with the normal
microbiota.
These results suggest that tongue coating could be a reservoir of unexpected bacteria in
the frail elderly adults, especially those with declined physical function or nutritional
condition.

SPOCK-1 is a novel epithelial mesenchymal transition (EMT) inducer
in calcium channel blocker-induced gingival overgrowth.
Rehab Alshargabi
Department of Periodontology, Division of Oral Rehabilitation, Faculty of Dental
Science, Kyushu University, Fukuoka, Japan.
【Background】
Drug-induced gingival overgrowth (DIGO) is a side effect associated with Ca+2 channel
blockers such as Nifedipine. The exact mechanism by which those drugs causes
overgrowth is not fully understood. However, previous studies suggested that both EMT
and reduced catheptin-L activity could contribute to the development of DIGO. Moreover,elongated rete pegs are typical feature of DIGO. SPOCK-1, a multi-domain secreted
proteoglycan, has been reported to induce EMT in several cancer cells as well as to inhibit
cathepsin-L activity. In addition, SPOCK-1 belongs to SPARC family, some of which arehighly expressed in basement membrane zone. Therefore, we hypothesized that SPOCK1 may associate with the pathology of DIGO.
【Methods】
Calcium channel blocker-induced gingival overgrowth (CCBGO) and non-overgrowthsamples were collected from patients undergoing periodontal surgery in the
Periodontology Section at Kyushu University Hospital. SPOCK-1 protein level waschecked by western blotting and IHC staining. Spock-1 transgenic (Tg) mice weregenerated. Spock-1 Tg mice and wild-type (WT) littermates were sacrificed at 8 weeks
after birth. The expression of TGF-β1, E-Cadherin, Vimentin, Mmp2, Mmp9, Slug were
examined by western blotting and IHC staining following morphological observation.
【Results】
SPOCK-1 protein level was significantly up-regulated in CCBGO samples comparedwith control samples. Morphological observation of Tg mice revealed more thickened
gingival tissues at the molar region compared with WT mice. Histologically, moreelongated rete pegs and accelerated collagen accumulation were observed. E-cadherin
protein expression was significantly down-regulated while TGF-β1, Vimentin, Mmp2,
Mmp9, Slug expressions were up-regulated.
【Conclusion】
The results strongly indicate the importance of SPOCK-1 as an inducer of the EMT
process in the pathophysiology of DIGO.

Effects of TNF-a on Senescent human dental pulp cells
Aoi Nozu¹, Sayuri Hamano¹,², Atsushi Tomokiyo³, Daigaku Hasegawa³,
Shinichiro Yoshida³, Hideki Sugii³,
4
Hiromi Mitarai , Keita Ipposhi¹,³, Naohisa Wada4, Hidefumi Maeda¹,³
1

Department of Endodontology and Operative Dentistry,2 Division of Oral
Rehabilitation, Faculty of Dental Science, Kyushu
3 University, OBT Research Center,
Faculty of Dental Science,
KyushuUniversity,
Division of Endodontology, Kyushu
4
University Hospital, Division of General Dentistry, Kyushu University HospitalObjectives: Aging is characterized by chronic, low-grade inflammation, and thisphenomenon has been termed as inflammaging. It has been reported that the concentrationof tumor necrosis factor-a (TNF-a) is higher in the blood of aged people compared withthat of young people. Therefore, the aim of this study was to investigate how TNF-aaffected pulp cells during aging.
Methods: Human dental pulp cells (HDPCs) were cultured until reaching the plateau of
their growth, and the cells were isolated at actively (yHDPCs) or inactively (sHDPCs)
proliferating stages, whose gene expression of senescence-related factors, TNF-a, and its
receptor, TNFR1 was examined by quantitative PCR. In addition, on odontoblastic
differentiation of yHDPCs and sHDPCs cultured in control culture medium (CM) or CM
containing 2mM CaCl2 as differentiation medium (DM) with or without TNF-a was
examined by quantitative PCR and alizarin red staining. Furthermore, the contribution of
the signaling via TNFR1 to mineralization of sHDPCs was examined by transfection of
TNFR1 siRNA.
Results: sHDPCs expressed senescence-related molecules while yHDPCs did not. The
expression level of TNF-a in sHDPCs was significantly higher than that in yHDPCs.
When these HDPCs were cultured in DM, the mineralization activity of sHDPCs was
lower compared with that of yHDPCs. However, the administration of TNF-a to thisculture medium altered these responses: yHDPCs showed downregulated mineralization
per a cell, while sHDPCs exhibited a significant increase. Furthermore, the expression of
TNFR1 in sHDPCs was significantly upregulated compared with that in yHDPCs.
Downregulation of TNFR1 expression led to decreased mineralization of TNF-a-treatedsHDPCs.
Conclusions: These results suggested that sHDPCs preserved the odontoblastic
differentiation capacity, and TNF-a promoted odontoblastic differentiation of sHDPCs
through increased expression of TNFR1.

KOB & OBT Joint International Symposium 2019

Session 5
Oral Health & Total Health

Chaired by Dr. Shinsuke Mizutani

Oral function and decline in cognitive function
Masahiro Yamaguchi1), Shinsuke Mizutani2,3),
Rui Egashira1), Toru Naito1)
1) Section of Geriatric Dentistry, Department of General Dentistry, Fukuoka Dental College, Fukuoka, Japan
2) OBT Research Center, Faculty of Dental Science, Kyushu University, Fukuoka, Japan
3) Section of Geriatric Dentistry and Perioperative Medicine in Dentistry,
Division of Maxillofacial Diagnostic and Surgical Sciences,
Faculty of Dental Science, Kyushu University, Fukuoka, Japan

In Japan, about 4.62 million elderly people aged 65 years or older are thought to be
developing dementia. A condition called mild cognitive impairment (MCI) occurs in the
preliminary stage of dementia, and if left untreated, about 40% of those with MCI willdevelop dementia within 5 years; however, dementia onset may be delayed by appropriate
treatment. Therefore, it is important to notice MCI at an early stage and take effective
countermeasures. For patients who do not have a family doctor, dentists who havefrequent contact with the elderly who visit for regular dental checkups could easily
become involved in the early detection of MCI. Recent findings have suggested that oral
conditions such as periodontal disease or oral function are related to a decline in cognitive
function. For example, epidemiological studies have reported that severe periodontitis
and the impaired ability to pronounce /pa/ are related to MCI in Japanese people. However,few studies have examined oral condition or function in detail. Therefore, we examinedpresent teeth, periodontal inflammation, degree of periodontal disease, masticatory
function, swallowing function, and tongue pressure in dental outpatients. We used theJapanese version of the Montreal Cognitive Assessment and the Mini-Mental State
Examination as cognitive function tests. In addition, we measured serum apolipoprotein
A1, complement protein C3, and transthyretin as biomarkers for MCI. The results showedthat higher age and low tongue pressure were related to a decline in cognitive function.
In this presentation, we introduce studies on the relationship between oral
condition/function and MCI. We also discuss our study results, which suggest an
association between low tongue pressure and a decline in cognitive function.

Necroptosis of Adipocytes Induced by Osteocalcin
Takahito Otani1, Miho Matsuda2, Akiko Mizokami3, Norio Kitagawa1,
Hiroshi Takeuchi4, Eijiro Jimi2,3, Tetsuichiro Inai1 and Masato Hirata5
1

Division of Functional Structure, Department of Morphological Biology, Fukuoka
Dental College
2
Laboratory of Molecular and Cellular Biochemistry, Faculty of Dental Science,
Kyushu University
3
OBT Research Center, Faculty of Dental Science, Kyushu University
4
Division of Applied Pharmacology, Kyushu Dental University
5
School of Dental Medicine, Fukuoka Dental College

Osteocalcin, one of non-collagenous bone matrix proteins, is known to regulate themetabolism of glucose and energy in the uncarboxylated form designated as GluOC. We
examined the effects of GluOC on adipocytes and adipose tissue of mice, and thenrevealed the signaling pathway on which GluOC increased the expressions of adiponectin,
an adipokine regulating the metabolism of glucose and lipids, and peroxisome
proliferator-activated receptor-γ (PPARγ), a master regulator of adipogenesis, via its
putative receptor GPRC6A. Furthermore, we also found that GluOC at the concentrationof over 20 ng/ml inversely abolished the expression of adiponectin, and that themechanism was induced by the programmed necrosis (necroptosis) in 3T3-L1 adipocytes.
The adipocyte bound by GluOC via GPRC6A increased the expression of FoxO1, atranscription factor regulating cell death, via cAMP-PKA-CREB signaling axis together
with activation of p300, a transcriptional coactivator of CREB followed by the expressionof FasL, a member of tumor necrosis factor superfamily. FasL expressed in the plasma
membrane triggered Fas activation followed by the phosphorylation and trimerization of
mixed lineage kinase domain-like protein (MLKL), a major regulator of necroptosis.
Finally, calcium influx, reactive oxygen species (ROS) production, lipid peroxidation andmitochondrial fragmentation occurred in the downstream of MLKL activation leading tonecroptosis. Although high-dose GluOC decreased the cell number of adipocytes, it was
maximally about by 30% even after 96 h, probably because direct cell-cell contact was
required for necroptosis via Fas/FasL. We have here revealed the signaling pathway on
which high-dose GluOC induced necroptosis in 3T3-L1 adipocytes.