Relationship between enterovirus-and-diabetes.pdf

BY ROMISSAA
ALY ESMAIL
ASSISTANT LECTURER OF ORAL MEDICINE,
PERIODONTOLOGY, DIAGNOSIS AND
DENTAL RADIOLOGY (AL-AZHAR
UNIVERSITY)

• Contents
• Introduction
• Enteroviruses in the blood and gut in T1DM
• The hygiene hypothesis
• Enterovirus, βeta cells and innate immunity
• Type I interferon (IFN) secreted by plasmacytoid dendritic cells (pDCs) and
pathogenesis in type 1 diabetes
• Enterovirus and adaptive immunity
• Enterovirus and host genes
• Viral-induced pathogenesis of T1DM
• Antiviral Vaccines and Therapeutics for T1D Prevention

Type 1 diabetes mellitus
(T1DM) is a common disease in
which insulin-producing
pancreatic β cells are impaired
by immune-mediated
mechanisms that develop as a
result of a complex interaction
of genetic and environmental
factors.

A role of environmental
factors in the development of
T1DM is suggested by the
continuously increasing
incidence of T1DM across the
world and differences in
incidence of the disease in
different geographic regions.
(REF. 2)
Jaïdane,
H.
(2008).

For example, the neighboring
populations of Finland and Russian
Karelia display a significant difference in
the incidence of T1DM, even though
their genetic profiles are similar. (REF. 3)

Environmental
changes are
believed to affect
the penetrance of
T1DM
susceptibility
alleles.
Key genes contributing to
T1DM disease susceptibility
are located in the human
leukocyte antigen (HlA) class
ii locus on chromosome 6,
and other loci associated with
T1DM have been identified in
the human genome.

Although rubella virus, mumps
virus, cytomegalovirus,
retroviruses and rotaviruses
have all been linked to the
development of T1DM,
evidence most strongly
implicates the involvement of
enteroviruses, especially
coxsackievirus B.
Viruses are candidates as
infectious risk factors in
the induction of strong
cellular immune
responses and in the
infection and damage of β
cells. Consequently, they
can cause local
inflammation of the
pancreas.

Data from initial
epidemiological studies
suggested a possible link
between enteroviruses,
in particular
coxsackievirus B4, and
T1DM. (REF. 2)
Jaïdane, H. & Hober,
(2008).

• Enteroviruses are frequently encountered human
pathogens with a wide variety of pathologic effect.
• they are small nonenveloped viruses (30 nm).
• The icosahedric capsid is made up of 60
capsomers; one capsomer comprises four
structural proteins
(vP1, vP2, vP3 and vp4).

Figure 1 Enterovirus structure and replication
Hober, D. & Sauter, P. (2010) Pathogenesis of type 1 diabetes mellitus: interplay between
enterovirus and host
Nat. Rev. Endocrinol. doi:10.1038/nrendo.2010.27

Anti-enterovirus
antibodies, as well as
enterovirus RNA, are
more frequently found
in the blood of patients
newly diagnosed with
T1DM than in healthy
individuals . (REF. 2,14–
21)

An association between the
presence of coxsackievirus B
RNA and elevated levels of
interferon α (IFN-α) has been
found in the blood of patients at
various stages of T1DM, but not
in individuals without the
disease. (REF. 22)
Chehadeh, w. et al.
(2000)

• Boys born of mothers positive for enterovirus-
specific IGM antibodies have a fivefold increased
risk of developing T1DM as compared with boys
born of mothers negative for these antibodies.

• Enteroviruses have been found in patients with
the typical autoimmune T1DM subtype, and a role
of enteroviruses is suspected in a fulminant non
autoimmune T1DM subtype that accounts for
20% of all cases of T1DM reported in Japan (REF.
25).
•

Prospective studies suggest that
the development of pancreatic
autoimmunity is concomitant with
an enteroviral infection. (REF.
2,27–29)
The prevalences of both enteroviral
RNA and enteroviral-specific
antibodies are greater in children with
islet autoimmunity than those
without. (REF. 14)

The environmental
Determinants of Diabetes in the
Young (TEDDY) is a large-scale
study that aims to improve
understanding of the causes of
T1DM from birth to disease
onset, and to highlight the role
of environmental factors in
triggering T1DM.

In the TEDDY studies, enterovirus infections screened by
nested reverse transcription PCR for enterovirus RNA
and/or anti-enterovirus IGM have been detected during
pregnancy in mothers whose children went on to
develop T1DM before the age of 15 years.(REF.
24,30,31)
Elfving, M. et al. (2008).

IFN-α is considered a marker of virus
infection.
the expression of IFN-α by β cells of
patients with T1DM (detectable by
immunohistochemistry and by an
increase in IFN-α mrna expression)
has been reported, (REF. 32,33)
which could be a sign of the presence
of a virus in these cells.

Enteroviruses, especially
coxsackievirus B4, have
been isolated from the
pancreas of patients with
T1DM. (REF. 34,35)
Viral particles and the
enterovirus VP1 capsid
protein have been found in
β cells in pancreatic tissue
from three of six organ
donors with T1DM, but not
in the pancreatic tissue of
26 organ donors without
T1DM. (REF. 34)

IN postmortem pancreatic tissue(without
inflammatory changes in the islets) in a child who
tested positive for islet-cell auto antibodies,
enteroviral VP1 could be detected, which is in
agreement with a possible role of enteroviruses in
the early stages of the pathogenic process leading
to T1DM. (REF. 38)

A role Entero viruses in the
T1DM is supported by
hygiene hypothesis is a
theory that explains why
T1DM incidence is
paradoxically higher in
industrialized countries
where the sanitary
conditions are optimal and
where enterovirus infections
are less prevalent in the
population.

According to the hygiene
hypothesis, viral
infections during
childhood would protect
individuals from the risk
of developing T1DM or
would delay disease
onset.
Thus, an inverse
correlation would exist
between T1DM incidence
in a given area and the
frequency of enterovirus
infections within the
population. (REF. 42)

Figure 2 Crosstalk between viral infection, genetic background and early education of the immune
system
Beeck, A. O. d. & Eizirik, D. L. (2016) Viral infections in type 1 diabetes mellitus — why the β cells?
Nat. Rev. Neurol. doi:10.1038/nrendo.2016.30

Figure 2 | Crosstalk between viral
infection, genetic background and
early education of the immune
system. Genetic background and
early immune education, either alone
or in combination, define the
individual's capacity to modulate the
cell autonomous response upon viral
infection.
These diverse responses to viral
infection can lead to different outcomes,
including excessive β-cell loss (with or
without viral persistence) and triggering
of an autoimmune response.
Repeated viral infection might accelerate the ongoing
autoimmune assault against β cells, culminating in
clinical disease. AS, alternative splicing; ER,
endoplasmic reticulum; T1DM, type 1 diabetes
mellitus.

Enterovirus, the immune
system andT1DM

• Various serotypes of human
enterovirus B frequently encountered in
the environment (coxsackievirus B1, B2,
B3, B4 and B5, and echovirus 6, 7, 11,
25, 29) can infect and damage human β
cells in vitro.

Enteroviruses, by the
double-stranded
RNA(dsRNA) generated
during their replication ,
activate the production
of IFN-α and IFN-β.
Genes that encode
molecules of
the(MHC) are then
induced by IFN.

Coxsackievirus B4 activates the
expression of MHC molecules in
human fetal islet cell cultures.
Coxsackievirus B induces
cytokines, chemokines and
intercellular adhesion
molecule 1 in human islets
in vitro, which can activate
mononuclear cells in the
islets that may have a role
in the pathogenesis of
T1DM.

Figure 1 Regulation of key antiviral responses in pancreatic β cells
Beeck, A. O. d. & Eizirik, D. L. (2016) Viral infections in type 1 diabetes mellitus — why the β cells?
Nat. Rev. Neurol. doi:10.1038/nrendo.2016.30

Figure 1 | Regulation of key antiviral responses in pancreatic β
cells. Following infection, replicating coxsackievirus subtype B
(CVB) produce cytosolic double-stranded RNA (dsRNA), a
nonphysiological form of mRNA recognized by the cytoplasmic
receptor MDA5. Binding of MDA5 to the dsRNA activates the
transcription factors NF-κB, IRFs and STATs, triggering production of
type I interferons and chemokines, thus contributing to local
inflammation (insulitis).
Type I interferons (IFN-α and IFN-β), type II interferon (IFN-γ) and
the cytokines TNF and IL-1β contribute to β-cell destruction among
genetically susceptible individuals. Type I interferons bound to the
IFN-α/β receptor (IFNAR) signal via TYK2 and JAK1 and induce
activation of STATs and expression of interferon-stimulated genes
(ISGs) with antiviral properties. Proinflammatory cytokines
promote the activation of JNK1, which induces the intrinsic
(mitochondrial) apoptotic pathway through the proapoptotic
protein BIM and its phosphorylated form (P-BIM).

PTPN2 modulates β-cell death induced
by interferons by regulating activation of
P-BIM via JNK1. BIM and/or JNK1 are
downregulated by BACH2, GLIS3 and
CTSH.
PTPN2 also functions as a negative
regulator of the STAT signalling pathway,
whereas USP18 exerts negative feedback
on interferon-induced STAT signalling
and mitochondrial apoptotic pathways in
β cells.

Candidate-gene regulated
factors implicated in type 1
diabetes mellitus are framed
in red and the consequences
of their modulated
expression and/or activity
on biological function or
type 1 diabetes mellitus risk
indicated.
Kinases and
phosphatases are
indicated by green
ovals and
transcription factors
by grey ovals.

• The detection of entero-viral protein in human β
cells is accompanied by over expression of classI
MHC molecules, which could be involved in the
activation of the autoimmune processes by
enterovirus infection.

• Furthermore, the infection of
mice with coxsackievirus B4 results
in autoimmune diabetes
accompanied by the replication of
the virus in β cells (REF. 50)
•
• Owing to the local inflammation
in the pancreas that results from
infection of β cells, enteroviruses
are strongly suspected as risk
factors for T1DM.

A pancreatic infiltrate,
called insulitis, which
reflects the immune
response to β cells, is
present at the
symptomatic onset of
T1DM.
Studies of the pancreas
in patients with T1DM
suggest that the disease
has remarkable
heterogeneity. (REF. 52)

However, the presence of
enteroviral protein (VP1) in
insulitis-free islets from a
child without T1DM who
was islet-cell-autoantibody-
positive open the debate
about the possible role of
viruses in the early stages of
the disease. (REF. 38)
The detection of
enteroviruses in patients
newly diagnosed with T1DM
could be seen as a final step
in an ongoing autoimmune
process of β-cell damage
.(REF. 53,54)

But Several groups have published data
which indicate that enteroviruses may
target β cells, which is in agreement
with the hypothesis that they may play
a role in the initial disturbances of β
cells.
β-cell death might precede immune-c
infiltration in the islets and may actua
contribute to immune infiltration.(RE
55)

Indeed, in human islets
infected with coxsackievirus
B5, expression of pattern-
recognition receptors (such
as toll-like receptor 3, RIG-1
and IFIH-1) increases.
these receptors help the
innate immune system
recognize microorganisms.

Neo-autoantigens could
be generated by
misfolding of proteins
(such as insulin) as a
result of β-cell stress.
β-cell death and stress
may initiate the
autoimmune attack that
precedes immune-cell
infiltration in the islets.

Figure : Viral infections or other
inflammatory stimuli (1) will activate APCs
(2) that secrete mediators that attract
lymphocytes to "rolling" and entry into the
pancreatic tissue (3). Some beta cells might
be directly destroyed via cytotoxicity (4) and
others by cytokines secreted from
lymphocytes (5) or activated APCs (6).
Courtesy of Urs Christen, La Jolla Institute
for Allergy and Immunology, San Diego, CA.

Figure : During a localized virus infection in
the pancreas. Cytokines, such as interleukin-
1β (IL-1β), IL-12, interferon-γ (IFNγ) and
tumour necrosis factor (TNF), secreted by
immune cells, such as macrophages,
dendritic cells (DCs), CD4+ and CD8+ T cells
and natural killer (NK) cells, could cause
direct damage (such as apoptosis) of β-cells
cells but could also induce self-defence
mechanisms;
for example, IFNγ induces indoleamine 2,3-
dioxygenase (IDO) expression by β-cells.
Chemokines such as CXC-chemokine ligand
10 (CXCL10), CC-chemokine ligand 2 (CCL2)
and CCL20 secreted by β-cells recruit
macrophages and other inflammatory cells
to the islet area.

Natural killer group 2, member D (NKG2D) expressed by NK
cells binds to retinoic acid early transcript 1 (RAE1) expressed
by β-cells in non-obese diabetic (NOD) mice, and is
associated with β-cell damage. NKp46 engagement by an
unknown ligand on β-cells causes degranulation of NK cells.
Negative co-stimulatory molecules, such as programmed cell death ligand 1
(PDL1) expressed by β-cells can modulate diabetogenic T cell attack.

Continue
Figure :
β-cell antigens (such as
peptides derived from
insulin or pro-insulin)
can be presented by
MHC class I molecules
and recognized by
diabetogenic CD8+ T
cells.
MHC class II molecules
expressed by antigen-
presenting cells such as
DCs can also present β-
cell antigens (peptides
such as WE14 from
chromogranin A, insulin
and pro-insulin) that
can induce islet
antigen-specific CD4+ T
cell expansion. . FASL-
expressing T cells can
mediate apoptosis
through interaction
with FAS expressed on
β-cells.

Figure : Dying tissue cells release
nucleic acids; these form large
complexes with antimicrobial
peptides, such as LL37 in humans
and cathelin-related antimicrobial
peptide (CAMP) in mice, and with
endogenous DNA-binding proteins,
such as high mobility group protein
B1 (HMGB1).
These DNA and RNA complexes
activate pDCs via Toll-like receptor 9
(TLR9) and TLR7, respectively, and
they induce the production of type I
IFN.

Fig. :In turn, type I IFN promotes T
cell activation, autoantibody
production by B cells and the release
of neutrophil extracellular traps
(NETs) that consist of DNA–
antimicrobial peptide complexes.
Autoreactive antibodies activate
neutrophils and form DNA-
containing immune complexes that
are preferentially endocytosed by
pDCs via Fc receptors.
Self-nucleic acids also activate
classical DCs (cDCs) and they promote
the release of inflammatory cytokines
and the priming of T cells that are
specific for self antigens in a process
that is also facilitated by type I IFN.

Enterovirus and adaptive immunity:
• Persistence and β-cell impairment.
• Bystander activation.
• Molecular mimicry.
• Tolerance.
• Enhancing Antibody.

Coxsackievirus B and other enteroviruses
are able to cause persistent infections
The viral persistence would
result from an inappropriate
immune response to
enteroviruses, as suggested by
a reduced activation and
cytokine production by Tcells
from patients with T1DM in
response to in vitro challenge
with coxsackievirus B4
antigens. (REF. 69 )

The detection of
enterovirus components
in gut mucosa of patients
with T1DM suggest that
the intestine could
provide a reservoir for
persistent enterovirus .
(REF. 23)
Interestingly, enterovirus
RNA has been detected
in pancreatic tissue of
patients newly
diagnosed with T1DM
and also in the tissue of
patients with long-
standing T1DM. (REF. 36)

•
• In addition, there is persistent
infection (through the
presentation of viral antigens
and self- antigens) might result
in the induction of an antiviral
and autoimmune response.

T1DM might rely on
cyclical processes,
also called
relapsing–remitting
(like those observed
in chronic viral
infections). (REF. 72)
In this process, an
autoimmune
response could be
raised against
neoautoantigens at
the same time as
cellular stress or
inflammation and
during each relapse.

JA Bluestone et al. Nature 464, 1293-1300 (2010)
doi:10.1038/nature08933
Immunologic history of type 1 diabetes.

Every β-cell damage event
would trigger the recognition of
other autoantigenic epitopes.
this process would
increase the
population of
neoantigens and
epitopes displayed,
and result in the
activation of
autoreactive t cells, a
process called epitope
spreading. (REF. 72,73)

Alocal infection of β cells and the inflammatory
reaction that accompanies it, which causes tissue
destruction and release of sequestered self-antigens
from the islets, can induce the recruitment of auto
reactive T lymphocytes directed against these
antigens but not against viral antigens. (REF. 77,78)

A molecular mimicry has been detected between pathogens
and autoantigens recognized by antibodies or T cells of
patients with autoimmune diseases. (REF. 76)
Induction or acceleration of autoimmunity could be
related to infections with pathogens that have
similar structures as autoantigens. (REF. 79,80)

well-known model of molecular
mimicry in T1DM involves the 2C
viral protease of enterovirus and
the islet auto antigen GaD65 that
share a common amino acid
sequence called PEVREK. (REF. 81)
However, the hypothesis
that this homology has a
role
in T1DM pathogenesis has
been challenged and is the
subject of controversy. (REF.
75,82)

A defect in tolerance towards β-cell antigens at the
peripheral level is based on anomalies of regulatory T
lymphocytes (T. REG).
T.REG cells, which are a specialized subset of T cells
whose functions include the suppression of auto-
reactive T cells, play a role in the progression of
T1DM. (REF. 90)
They maintain at least a partial tolerance to self-
antigens in the pancreas.

Whether viral infections have
effects on T.REG cells is not known;
however, the hypothesis cannot be
excluded because toll- like receptor
signaling, provoked
by viral infections, can inhibit the
suppressive activity of T.ReG cells
indirectly by causing inflammation,
(REF. 91)and possibly directly as
well

Thus, in addition to their possible role in the
activation of β-cell-reactive T cells, viruses can
reduce the capacity of T.REG cells to maintain
tolerance. So , whether enterovirus infections have
such an effect on T.REG cells should be investigated.

The ADE of coxsackievirus B4 infection of monocytes
involves CAR (coxsackie virus and adeno-virus
receptor), FcγRII and FcRrIII (receptors for the Fc
portion of the IgG molecule) at the cell surface and
results in the synthesis of IFN-α by these cells.

The virus-induced synthesis of
IFN-α (increased by specific
antibodies) is associated with the
development of a TH1-type
immune reaction, which can
contribute to the development of
autoimmune reactions .
The Antibodies directed against VP4 or
its peptides were more prevalent and at
higher levels in patients with T1DM than
in those without the disease. (REF.
105,107)

• The spreading of coxsackievirus B to β cells can,
consequently, be promoted by the ADE of the enterovirus
infection, which canfor that reason be implied in the early
steps of the immune response against these cells.
• The persistence of enteroviruses could be associated with
an increased production of enhancing antibodies.

30–40%, and the
risk that siblings of
patients with
T1DM have of
developing the
disease is 6.0%
versus only 0.3%
for the general
population.
The occurrence of
T1DM in
monozygotic twins
is
Susceptibility and
resistance to T1DM
are both affected
by genetic factors.

Several loci are
associated with the
disease, which suggests
that its pathogenesis is
polygenic.
Genes within the HlA
region that encode
antigen-presenting
molecules, especially
the DQ and DR loci, are
associated with the risk
of T1DM. (REF. 111)
In addition, some of
the loci are associated
with antiviral activities.

OAS1 gene
a single nucleotide polymorphism exists in the
OAS1 gene that is strongly associated with basal
activity of the gene product, 2'5'-oligoadenylate
synthetase 1 (OAS1) a key antiviral enzyme that
synthesizes oligoadenylates that activate a latent
RNase (RNaseL). this enzyme degrades
endogenous and viralRNA, and hence impedes viral
replication and promotes cell death.

How would the
antiviral enzyme
activity of
individuals with
variants of OAS1
increase their risk
of T1DM ?
One explanation is
that increased
enzyme activity,
induced by the
binding of
2'5'oas1 to
intracellular viral
dsRNA, directly
damages sensitive
β cells (by
RNaseL-mediated
degradation of
cellular RNA).

Another explanation is that
variants that confer a low risk
of T1DM produce one of the
isoforms of the protein (p48)
that has a proapoptotic
activity that is independent of
its synthetase activity,
which could trigger the
apoptosis of β cells infected
with enterovirus and hinder
viral spread to neighboring
cells.

IFIH1
• IFIH1 is a gene that has a role in innate
immunity by the recognition of
picornavirus RNA. (REF. 117)
• The Picornaviridae family includes the
enteroviruses, such as coxsackievirus B4,
which are potential environmental triggers
for T1DM.

• How does IFIH1 act to confer a risk of T1Dm?
• following to viral infections, the transcription
of the IFIH1 gene is up regulated.
• IFIH1 signaling activates IFN-regulator factors
and transcription factors, like nuclear factor κB,
and activates target genes such as those that
encode IFN-α and IFN-β and interferon-
inducible genes (Figure 2).

Figure 2 IFIH1-mediated β-cell response to infection with an enterovirus

• In response to enterovirus infections, individuals with
elevated IFIH1 levels may have an increased stimulating
capacity of dendritic cells and a elevated production of
pro-inflammatory cytokines, which may promote the
development of T1DM.
• Signal transduction pathways that involve
cytoplasmic IFIH1 considerably increase the ability of
human monocyte-derived dendritic cells to stimulate
proliferation of CD4+ T cells.

Figure : Regulation of
key antiviral responses
in pancreatic β cells
• Beeck, A. O. d. & Eizirik, D. L.
(2016) Viral infections in type 1
diabetes mellitus — why the β cells?
• Nat. Rev. Neurol.
doi:10.1038/nrendo.2016.30

Figure 1 | Regulation of key
antiviral responses in pancreatic
β cells.
Following infection, replicating coxsackievirus subtype B (CVB) produce cytosolic double-
stranded RNA (dsRNA), a nonphysiological form of mRNA recognized by the cytoplasmic
receptor MDA5. Binding of MDA5 to the dsRNA activates the transcription factors NF-κB,
IRFs and STATs, triggering production of type I interferons and chemokines, thus
contributing to local inflammation (insulitis). Type I interferons (IFN-α and IFN-β), type II
interferon (IFN-γ) and the cytokines TNF and IL-1β contribute to β-cell destruction among
genetically susceptible individuals. Type I interferons bound to the IFN-α/β receptor
(IFNAR) signal via TYK2 and JAK1 and induce activation of STATs and expression of
interferon-stimulated genes (ISGs) with antiviral properties.

Proinflammatory cytokines promote the activation
of JNK1, which induces the intrinsic (mitochondrial)
apoptotic pathway through the proapoptotic
protein BIM and its phosphorylated form (P-BIM).
PTPN2 modulates β-cell death induced by
interferons by regulating activation of P-BIM via
JNK1. BIM and/or JNK1 are downregulated by
BACH2, GLIS3 and CTSH.
PTPN2 also functions as a negative regulator
of the STAT signalling pathway, whereas
USP18 exerts negative feedback on
interferon-induced STAT signalling and
mitochondrial apoptotic pathways in β cells.
Candidate-gene regulated factors implicated
in type 1 diabetes mellitus are framed in red
and the consequences of their modulated
expression and
/or activity on biological function or type 1
diabetes mellitus risk indicated. Kinases and
phosphatases are indicated by green ovals
and transcription factors by grey ovals.

Viruses play a dual role in T1DM.
Devil or angel? viral infections can
protect individuals from the risk
of developing T1DM.
The results of studies performed
in animal models indicate that
enteroviral infections before
weaning are beneficial, whereas
infections after this period have
an opposite effect.

Together, the nature of the
virus, its tropism for β cells,
and the timing of the
infection have an important
role in T1DM occurrence.
The interplay between
enterovirusinfection, β cells,
the immune system (innate
and adaptive) and host genes
and the processes able to play
a part are summarized in
Figure 3.

Figure 3 Virus-induced pathogenesis of type 1 diabetes mellitus (T1DM)

Antiviral Vaccines and
Therapeutics for T1D
Prevention

what stages of islet
autoimmunity (IA) and
T1D development
these may prove
useful, indicating
current clinical trials
aimed at preventing
enterovirus infection
for prevention or
treatment of T1D.
Abbreviations: VLP,
virus‐like particles;
CVB, coxsackievirus

Antiviral drugs could offer
an option for
preventing/treating T1D by
eradicating infections by
diabetogenic EVs.
Currently, individuals
newly diagnosed with T1D
are being recruited into
the DiViD and Intervention
Trial in Norway (EU Clinical
Trials Register EudraCT No.
2015-003350-41).
This represents the first
randomised clinical trial with
antiviral drugs to test the
hypothesis that a six-month-long
treatment with pleconaril–
ribavirin combination can
eliminate persistent EV infection
in the pancreas

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