Enteroviruses as causative agents in type 1 diabetes: loose ends or lost cause?
Noel G Morgan & Sarah J Richardson
Institute of Biomedical & Clinical Science
University of Exeter Medical School
RILD Building, Barrack Road
Exeter EX2 5DW, UK
Correspondence:
n.g.morgan@exeter.ac.uk (+44-(0)1392-408300)
s.richardson@exter.ac.uk (+44-(0)1392-408225)
Keywords
Coxsackievirus; type 1 diabetes; persistent infection; pathogen recognition receptors; islets of
Langerhans
1
Abstract
Considerable evidence implies that an enteroviral infection may accelerate or precipitate type 1
diabetes (T1D) in some individuals. However, causality is not proven. We present and critically
assess evidence suggesting that islet β-cells can become infected with enterovirus, and argue that
this may result in one of several consequences. Occasionally, a fully lytic infection may arise and
this culminates in fulminant diabetes. Alternatively, an atypical persistent infection develops
which can be either benign or promote islet autoimmunity. We propose a model in which the
“strength” of the β-cell response to the establishment of a persistent enteroviral infection
determines the final disease outcome.
2
Linking enteroviruses to T1D
Enteroviruses are a genus of positive-sense, single stranded, non-enveloped, RNA viruses that
circulate widely and can cause a spectrum of mild respiratory and gastrointestinal (GI) symptoms
as well as more severe conditions such as myocarditis or poliomyelitis. In addition, some
enteroviruses, notably those belonging to the Coxsackievirus B species, have been implicated in
the development of T1D in humans, implying that an initial (mild) infection might have significant,
long-term, consequences in some individuals.
The cumulative evidence implying a firm association between enteroviral infection and the
development of T1D in humans is substantial but, unlike the situation in mouse models [1], this
evidence base remains largely correlative. Unequivocal confirmation of enteroviral infection
within the pancreatic islets has been hard to obtain and the majority of data have been derived
from epidemiological studies which show that markers of enteroviral infection occur commonly in
the circulation of patients at diagnosis of T1D [2-11]. One pertinent recent example comes from a
study of Italian families where enteroviral genome was detected at high frequency in the blood of
newly diagnosed T1D patients (79%) as well as in their parents (63%) and siblings (60%) but in only
1 of 29 (3%) relevant controls [8]. These data imply very firmly that enteroviral infections may
contribute to the onset of disease in some patients but the data also show that such infections are
not the sole determinants. This is not surprising, given that enteroviruses circulate widely in the
population whereas the incidence of T1D is much lower, even in populations where it is most
prevalent.
Clearly, any model which attempts to implicate enteroviruses in the aetiology of diabetes must
account for the fact that the vast majority of people who become infected do not go on to develop
diabetes. This is illustrated forcibly by studies among Cubans who were exposed to an epidemic of
echovirus early in the 21st century [12]. Within this group, large numbers of patients
seroconverted to islet auto-antibody positivity following enteroviral infection, but the prevalence
of overt T1D remained low. Thus, not all infected individuals went on to develop disease. This
situation is reminiscent of the paralysis associated with poliomyelitis which occurs in only a
minority of infected individuals [13]. One contributory factor in the case of T1D may be the
enteroviral serotype involved since data from the Finnish population implies that serotypes which
are closely related phylogenetically, have a differing potential to promote T1D [3, 6].
3
Autoimmunity and enteroviruses; what are the connections?
Is there a role for molecular mimicry?
The currently accepted wisdom considers that human T1D is mediated by a process of
autoimmunity in which pancreatic β-cells are destroyed in a highly selective manner [14].
Therefore, any attempt to relate autoimmunity and β-cell loss to enteroviral infection must
provide a plausible mechanism by which these processes are linked. One of the most obvious
possibilities for the development of autoimmunity is that some form of molecular mimicry occurs
in which the immune response to a viral antigen triggers the generation of antibodies which
serendipitously cross-react with sequences present in islet proteins. Support for this concept has
been presented since a conserved sequence within the enterovirus 2C protein is also present in
glutamate decarboxylase (GAD); a principal autoantigen in T1D [15, 16]. However, while this
epitope relationship may be true, it is not clear why this should serve as the trigger for islet
autoimmunity. Moreover, it is likely that the presence of circulating autoantibodies directed
against GAD is a marker of disease progression rather than of pathogenic significance. Thus, the
association between enteroviral peptides and islet autoantibody generation has been questioned
[17-21]. We conclude that, although the molecular mimicry mechanism has an attractive
symmetry, this is superficial and does not provide a firm basis to explain the mechanisms of β-cell
loss.
Migration of enteroviruses to islets
A second possibility is that enteroviruses may spread from their initial sites of infection to β-cells
and thereby provoke an (auto)immune response more directly. There is firm evidence that
enteroviruses can establish an infection in cells of the GI tract in patients with T1D [22], and this is
likely to be the source of islet infection. Supporting evidence comes from reports of the isolation
of enterovirus from the pancreas of two patients with recent onset T1D [23, 24] (although, in one
of these, the provenance of the virus has been challenged [1]).
It is also clear that other pathways exist by which an infection may spread to more remote organs,
since it is well understood that acute enteroviral infection can lead to myocarditis under certain
circumstances [25-27]. It is likely that, in such cases, a blood-borne infection ensues and, in
4
support of this, evidence of early viraemia during acute enteroviral infection has been provided
[28]. This might, then, lead to the spread of infectious particles to any internal organ that is
permissive for the virus. To address this possibility, pancreases recovered from several children
who had died of acute myocarditis following proven enteroviral infection were examined and the
islets found to contain enteroviral antigens [25, 29] and RNA [30, 31]. This provides very strong
evidence that enteroviruses can spread from the circulation to the pancreatic islets, at least under
conditions when the viral load is extremely high.
It is a moot point whether such transit also occurs during episodes of mild enteroviral infection but
the establishment of productive infection in the pancreatic islets appears to be uncommon.
Among a cohort of British children who died of causes unrelated to enteroviral infection (or T1D)
only 6% (3 of 50) had evidence of enteroviral infection in their islet cells (as determined by
immunostaining for the enteroviral capsid protein VP1). Moreover, in those who did, the number
of positively stained cells was vanishingly low. This contrasts with data obtained from the
pancreases of children who died soon after developing T1D, where VP1 was detected in the islets
of more than 60% (44 of 72) [29] and, in a subset of these same cases, the presence of enterovirus
was confirmed by in situ hybridisation [30]. Additional smaller studies have reached similar
conclusions [30, 32, 33]. Such evidence implies that enteroviral infection is intimately related to
diabetes; and might be causative. Alternatively, it is also possible that the islets of patients who
have contracted T1D are simply more susceptible to enteroviral infection; perhaps because of
ongoing damage to the β-cells. We do not favour this latter hypothesis since histological analysis
of human pancreas suggests that the majority (>60%) of insulin-containing, VP1-immunopositive,
islets are not insulitic [34], implying that viral infection precedes the process of immune cell
recruitment and β-cell damage. Moreover, the fact that seroconversion to islet autoantibody
positivity occurs in many patients (including monozygotic siblings [35]) who have contracted an
enteroviral infection but not developed T1D (at least, at the time of analysis) [12, 35] implies that
infection of the islet cells must be an early event in the process. Nevertheless, gaining firm
information about the pathological changes occurring within the pancreas during the initiation of
T1D remains difficult and the continuing dearth of appropriate samples for study represents a
significant impediment to progress.
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Islet cell susceptibility to infection differs between cell types
That islet cells can become infected with enteroviruses under some circumstances is not in dispute
but begs a further important question; namely, which of the various endocrine cell types support
enteroviral replication in vivo? The majority of data indicate a clear selectivity for β-cells (as
judged by the cellular distribution of immunoreactive VP1) [29, 32, 33] and this is consistent with
the selectivity of cell loss seen in the disease. In this context, it is also important to note that the
exocrine pancreas is largely immunonegative for VP1 in most human T1D patients, although
occasional VP1 immunopositivity occurs in cells of the duct epithelium [29, 30]. This situation is
quite different from that seen in mouse models where enteroviral infection of the acinar tissue
within the exocrine pancreas is often widespread, suggesting that there are very different
mechanisms regulating pancreatic infectivity and replication in rodents and humans [36-38].
To clarify the infectivity of islet cells, recent studies have addressed the expression of two
potential enteroviral "receptor" proteins, Decay Accelerating Factor (DAF) and CoxsackieAdenovirus Receptor (CAR), in human pancreas. CAR is considered to be a tight-junctional
component, whereas the physiological role of DAF is less clear and it is reportedly absent from
pancreatic islets [30]. By contrast, CAR is expressed in islet cells [39, 40], and antibodies against
CAR can block enteroviral infection of islets incubated in vitro [30]. However, CAR exists in
multiple isoforms and the precise isoform distribution has not been mapped in islet cells; nor is it
clear exactly which isoforms preferentially mediate enteroviral entry [41]. Some workers have
found that CAR is present in the majority of human islet cells [42] whereas others, using different
antibodies, report that it is present in β- but not alpha-cells within islets [39]. Interestingly, in the
latter study, CAR was detectable on a unique sub-population of human alpha cells located outside
the islets and within the pancreatic parenchyma [39]. The origin and ultimate destination of these
"extra-islet" alpha cells is unclear and it is not known why they should express CAR. However, one
might speculate that the protein is present to assist in their coalescence into primordial islets, by
promoting tight junction formation with other, single, endocrine cells that are encountered within
the pancreas. Whatever the explanation, it has not yet been demonstrated that such CAR-positive
alpha cells can become infected with enterovirus in vivo, and the weight of evidence still points to
a primarily β-cell focussed infection mechanism.
6
Non-canonical viral replication
Importantly, there are aspects of the biology of enteroviruses which appear unusual when they
infect β-cells in vivo. Most notable is the fact that morphological examination of the islets of
patients with T1D does not reveal evidence of large-scale cell lysis. This contrasts with expectation
since enteroviruses normally subvert the cell's transcriptional and translational machinery to
generate newly synthesised capsids which are then released by a lytic mechanism [43]. These new
virions are able to infect neighbouring cells, leading to extensive tissue damage; but such damage
is not typically seen in human T1D. Despite this, it cannot be excluded that lysis of a small number
of islet cells may occur early in the disease process since any debris will have been cleared by the
time tissue is removed for analysis (which may be weeks, months or even years after disease
initiation). An equivalent situation has been described in the heart where an early episode of
cardiomyocyte lysis is followed by the establishment of a more sustained, non-lytic, infection [44].
The fact that extensive cell lysis is not seen in the pancreas of patients with T1D is unlikely to be
simply a property of the cells themselves since infection of isolated human islets maintained in
tissue culture, can readily lead to extensive damage [45-47]. Moreover, in a rare form of T1D that
has been described most frequently in Japanese populations (known as "fulminant" diabetes),
large-scale lysis of islet cells does occur in vivo, and this disease may well be mediated by an
acutely lytic enteroviral infection of the islet cells [48-50]. If so, it differs markedly from the
situation seen more typically in the islets of patients with T1D in Western populations.
Interestingly, in fulminant diabetes the destructive process is so intense that both β- and alpha cell
lysis occurs [51, 52]. This might imply that each cell type can support enteroviral infection in those
ethnic groups who are most susceptible to fulminant diabetes but this remains to be clarified. A
further possibility is that the net damage to β-cells is so intense in fulminant disease that adjacent
"bystander" cells undergo a rapid demise as a result of the large-scale release of β-cell hydrolases
within the islet milieu (Figure 1).
Given that dramatic cell lysis does not normally occur during β-cell enteroviral infection in more
typical (i.e. non-fulminant) T1D, this implies that the viral replication cycle cannot proceed
according to a canonical mechanism. Any such conclusion may seem radical but there is already
evidence that the biology of enteroviruses is altered when they infect the heart. In particular,
Chapman and colleagues have shown that infection of human cardiomyocytes with enteroviruses,
7
in vivo, results in the loss of genomic sequences from the 5' end of the RNA [44, 53, 54]. Such
"terminally deleted" viruses have much reduced propensity to replicate and they rarely form
functionally competent virions that can be released to infect neighbouring cells. As such, they
persist in the host cells in a form which replicates only very slowly over a prolonged period. This
scenario has clear parallels with the situation found in the islets of patients with T1D, where only
occasional VP1 immunopositive β-cells can be detected and wholesale cell lysis is not apparent.
The presence of slowly replicating (genomically modified?) virus in a tiny proportion of human islet
cells may also contribute to the difficulties encountered in detecting viral RNA by RT-PCR in
pancreatic extracts, by comparison with isolated islets acutely infected in vitro, where abundant
quantities of fully replicative virus are present in each infected cell [55].
Defective trans-differentiation of infected pancreatic duct cells
Additional data emerging from studies of persistent enteroviral infection in cultured cell models
has also pointed to another important issue, which might shed further light on the processes
leading to net β-cell loss in human T1D. Hober and colleagues have established a persistent
enteroviral infection in a line of pancreatic ductal cells in tissue culture, and reported that this
leads to a reduced propensity for these cells to undergo differentiation [56]. This is important since
one mechanism by which β-cells might be replenished in humans occurs by a process of transdifferentiation of pancreatic duct cells [56, 57]. Thus, if the establishment of a persistent
enteroviral infection in the duct cells impedes this process, then β-cell replacement might be
compromised, further contributing to their overall demise under conditions when they are being
depleted by autoimmune-mediated mechanisms. This is important since, as emphasised earlier,
VP1 immunopositivity is sometimes detected in pancreatic ducts in patients with T1D, which
would be consistent with this model.
Alternative mechanisms of persistent enteroviral infection
The mechanism by which enteroviral persistence might be maintained within islet cells remains an
important issue and there are reports that enteroviruses can exist in an atypical double-stranded
RNA genomic form as a means to promote persistence [58-60] (Figure 1). This may seem odd,
given that the genome is normally a molecule of positive single stranded RNA and the fact that
dsRNA is not usually tolerated in cells. Normally, the presence of dsRNA is carefully monitored by
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various cellular surveillance systems which are designed to respond to its appearance by mounting
a vigorous anti-viral response [27]. Human islet cells are equipped with several intracellular
pattern recognition receptors [39, 47, 49, 61-63] (Figure 1), some of which are activated
exclusively by dsRNA and respond by initiating signalling cascades that promote type-I and type-III
interferon generation [64]. Frequently, the presence of longer length (i.e. enteroviral) dsRNA is
sensed most effectively by an RNA helicase enzyme, Mda5 [65] and this protein is present in
human islets. However, in islets, Mda5 is not expressed uniformly among the different endocrine
cells. Rather, it appears to be most abundant in alpha cells while β-cells are relatively deficient [39,
63, 66]. Thus, conceivably, the establishment of a persistent β-cell infection might be favoured by
this relative deficiency. Interestingly, Mda5 expression is increased in the residual β-cells of
patients with T1D [39, 66] which could be indicative of an enhanced response to viral infection
(Figure 1). This suggests, in turn, that the normally low activity of Mda5 in β-cells may be
important as a means to mitigate the development of autoimmunity.
Level of response to infection might determine final outcome
Following on from the evidence outlined above, it seems plausible that the magnitude of the antiviral response occurring at the level of β-cells during an enteroviral infection, could be critical to
determination of the final outcome. In this model, the better outcome arises when β-cells fail to
respond vigorously to enteroviral infection since, under these circumstances, they can continue to
survive and function. In support of this, it was shown that islet cells in 15-20% of adults without
diabetes, express VP1 (albeit at very low frequency) [29, 32]. This suggests that, if the cells allow
the virus to persist (perhaps in a slowly replicating form) without developing a robust response,
then a new equilibrium is established in which both the virus and the infected β-cell can co-exist.
If, however, this equilibrium is disturbed, such that the cell strenuously attempts to eliminate the
virus (e.g. by increasing the expression and activity of Mda5), this might then, counter-intuitively,
serve as the trigger that provokes autoimmunity and the progression towards T1D. In support of
this, certain polymorphic variants of Mda5 with reduced helicase activity are known to confer a
measure of protection against the development of T1D [67-70] and we individuals carrying these
protective variants are likely to respond only weakly to β-cell enteroviral infection.
Further evidence that a heightened β-cell response to enteroviral infection might mediate
autoimmunity comes from an analysis of the expression of MHC class I antigens on the islet cells of
9
patients and controls. Islet cell hyper-expression of MHC class I is a hallmark of T1D [71-73] and is
likely to occur as a result of increased interferon secretion during enteroviral infection [74, 75].
Importantly, islet cell hyper-expression of MHC class I is not seen in control individuals, even
among those whose islets stain positively for VP1. This supports the concept that weakly (or non-)
responding subjects are less likely to develop T1D.
This hypothesis inevitably raises additional questions, including whether it is the presence of
virally infected β-cells per se which triggers autoimmunity and then frank diabetes, or whether
more subtle mechanisms are operative. So far, the answers to these questions remain elusive, but
a recent study of the specificity of islet infiltrating T-cells in patients with T1D has shown that at
least a proportion of these are directed against islet antigens [76]. It is unclear whether a different
population may also be reactive against viral antigens. Nevertheless, the fact that some infiltrating
T-cells are reactive against islet proteins implies that, if viral infection underlies these processes,
then the virus may promote altered presentation of normally "silent" islet protein epitopes as the
primary means to elicit an immune response. Alternatively, (or additionally) the characteristic islet
cell hyper-expression of MHC class I noted above, may contribute to the enhanced presentation of
typical β-cell antigens thereby provoking an autoimmune response. Whichever mechanism is
operative, the virus can be considered to act as a Trojan horse, slipping quietly inside the normal
cellular defences but then, once recognised, eliciting an anti-viral response which alters the
intracellular environment to promote autoimmunity.
The role of interferons and protein kinase R (PKR)
It follows from this discussion, that the presence of enterovirus within the β-cells of patients with
T1D should be accompanied by a signature anti-viral response. In particular, it would be
anticipated that interferons type-I and -III, should be generated locally within the islet milieu
(Figure 1). This would then lead to the production of a characteristic interferon response in the
cells surrounding those carrying the persistent infection. One feature of this would be the hyperexpression of MHC class-I [71-73] (Figure 1) and direct evidence that this correlates with
interferon-alpha secretion in the islets of patients with recent-onset T1D has been provided [74].
Furthermore, a clear interferon signature has also been observed in the islets of the non-obese
diabetic (NOD) mouse early in life, although this is apparently not required for development of
10
T1D [77]. In addition, an interferon response has also been demonstrated in the blood of human
subjects who subsequently developed islet autoantibodies [78, 79] although this may reflect
interferon production outside the islets.
Although such evidence is persuasive, it must also be accepted that there are still many gaps to be
filled before the hypothesis that an islet viral response signature can be detected commonly in
human T1D, is considered watertight. In particular, as noted above in relation to MHC class-I
hyper-expression, the release of interferons should also promote the expression of additional
"interferon-response genes" (ISGs). Searching for this in human islet cells has not proved
straightforward. Thus, while interferon responses (and generation of ISGs) can be readily
demonstrated when human islets are isolated and exposed to enteroviruses in vitro [47, 61, 80,
81], detection of such gene products in the islets of patients with T1D has proved more elusive.
For example, one of the most prominent ISGs is "ISG15", a protein which becomes covalently
attached to other newly-synthesised viral proteins in virally infected cells, thereby targeting them
for degradation [82]. Surprisingly, ISG15 is readily detected in all human islets irrespective of the
presence or absence of enteroviruses, and it is present in delta- rather than β-cells [39]. The
reasons for this are unclear and the factors that drive expression of elevated ISG15 levels in delta
cells are unknown. However, the data imply that ISG15 is not expressed in response to viral
infection in these cells and, more significantly, there is little evidence that it is induced in β-cells, in
T1D. This seems surprising if the islets are bathed in interferons as a response to local viral
infection. Conceivably, this discrepancy might be explained by variations in the production of,
and/or the precise downstream response to, individual interferon isoforms which mediate
differences in receptor signalling [83].
A further unexpected observation relates to the up-regulation of a second anti-viral response
gene, PKR. It was shown that the small number of islet cells which are immunopositive for
enteroviral VP1 in human pancreas (and which, therefore, appear to be - or to have been infected with replicating virus), also express very high levels of PKR [32]. This provides very strong
support for the concept that β-cell VP1-immunoreactivity reflects an underlying enteroviral
infection, since PKR is a member of the family of PRRs known to be induced by these viruses [81,
84, 85], and whose primary role is to signal a state of cellular stress in which protein translation is
arrested (Figure 1). This then leads to the selective degradation of certain labile anti-apoptotic
11
proteins (such as Mcl-1 [32]) within the virally-infected cells, thereby increasing their vulnerability
to pro-apoptotic stimuli. The unexpected feature of the islet PKR response is that it occurs in such
a restricted manner. Rather than finding a marked up-regulation of this enzyme in all islet cells in
the vicinity of those which express VP1 (as might be expected if the release of a soluble factor such
as an interferon was involved), the most dramatic increase occurs uniquely in that subset of β-cells
which also express VP1. Indeed, this relationship is so strong that, under many circumstances,
immunodetection of PKR can be used, with confidence, as surrogate for VP1 in human pancreas.
Thus, it must be concluded that this component of the "anti-viral" response is mediated mainly by
an intracellular mechanism operating within each single infected cell, rather than by an
intercellular (soluble) mediator which acts on all neighbouring cells. How this happens is unclear.
Concluding remarks and future perspectives
In conclusion, there is growing evidence that enteroviral infection plays a direct role in promoting
islet autoimmunity in some patients with T1D and a recent meta-analysis of the cumulative
epidemiological evidence confirms a very clear association [11]. There is also strong evidence that
β-cells can become infected in vivo and can sustain an enteroviral infection. However, whether
(and, if so, how) such infections persist is unclear. The current state of play is summarised in
Figures 1 & 2 and it is evident that many loose ends must still be tied before causality is confirmed.
Nevertheless, all is not yet lost and the prospect that an enteroviral vaccination strategy might
ultimately be effective in, at least, some individuals who are predisposed to T1D, justifies a
continued effort.
Acknowledgements
We are pleased to acknowledge financial support from the European Union's Seventh Framework
Programme PEVNET [FP7/2007-2013] under grant agreement number 261441. Additional support
was from a Diabetes Research Wellness Foundation Non-Clinical Research Fellowship and, since
2014, a JDRF Career Development Award (5-CDA-2014-221-A-N) to S.J.R. The research was also
performed with the support of the Network for Pancreatic Organ Donors with Diabetes (nPOD), a
collaborative type 1 diabetes research project sponsored by the Juvenile Diabetes Research
12
Foundation International (JDRF) and with a JDRF research grant awarded to the nPOD-V
consortium. Organ Procurement Organizations (OPO) partnering with nPOD to provide research
resources are listed at www.jdrfnpod.org/our-partners.php.
We are grateful to Mark Russell and Pia Leete for stimulating discussions and assistance with the
preparation of images.
13
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Glossary:
Coxsackievirus: a virus that belongs to the family Picornaviridae and the genus Enterovirus of
nonenveloped, linear, positive-sense ssRNA viruses.
Coxsackie-Adenovirus Receptor (CAR): a type I membrane receptor that is expressed in tissues
such as heart, brain, and in epithelial and endothial cells. It may function as a cell adhesion
molecule at tight junctions through its interactions with extracellular matrix glycoproteins.
17
Decay Accelerating Factor (DAF): is a membrane glycoprotein that regulates the complement
system on the cell surface. DAF is used as a receptor by some coxsackieviruses and enteroviruses.
Enterovirus: is a member of the Picornaviridae family, and the genus Enterovirus that includes a
diverse group of small RNA viruses characterized by a single positive-strand genomic RNA.
Enteroviruses, which include polioviruses, echoviruses and Coxsackie viruses, are often found in
the respiratory secretions and stool of infected persons and have been associated with several
human and mammalian diseases.
Fulminant diabetes: a form of insulin-dependent diabetes which has been seen most commonly in
Japanese populations and occurs by virtue of an acute, large-scale, destruction of islet cells,
possibly mediated by a lytic enteroviral infection of the islet cells.
Insulitis: is a term defining the influx of immune cells around the periphery or within the islets of
Langerhans in the pancreas. It is associated with the elaboration of pro-inflammatory cytokines
that play a role in mediating β-cell death.
Interferons: are signaling proteins synthesised and released by host cells in response to the
presence of pathogens, such as viruses. Release of interferons from infected cells induces
protective responses in neighbouring cells, thereby reducing the likelihood of the spread of
infection.
ISG15: is a protein induced by the release of interferons that becomes covalently linked to newly
synthesised proteins (including viral proteins in infected cells) targeting them for degradation. The
process of “ISG15ylation” may also inhibit the assembly of functional viral capsids in virally
infected cells.
Islet autoantibodies: are antibodies directed against one or more of an individual’s own islet cell
proteins. These are often produced as an early response to islet damage during the development
of type 1 diabetes.
18
Glutamate decarboxylase (GAD): is the principal islet autoantigen in T1D. It is an enzyme with a
molecular mass of approximately 64kDa which is expressed in pancreatic β-cells and serves as an
autoantigen in many patients with T1D. Production of autoantibodies to GAD can also be detected
frequently in the circulation in close relatives of patients with T1D who do not, themselves,
necessarily go on to develop the disease.
Mda5: is a pattern recognition receptor encoded by the IFIH1 gene, which has been associated
with a pre-disposition to type 1 diabetes by genome wide association studies. Mda5 is a helicase
enzyme which binds dsRNA to initiate antiviral responses in cells.
Myocarditis: defines the inflammation of heart muscle, frequently associated with an enteroviral
infection. In young children with an acute enteroviral infection, myocarditis can be fatal.
Non-obese diabetic (NOD) mouse: is an animal model of type 1 diabetes, used very widely as a
surrogate for the human disease. NOD mice exhibit a susceptibility to spontaneous development
of T1D, usually with a marked female bias. Beta cells are selectively destroyed by a process of
insulitis in NOD mice although the extent of insulitis is much greater than that typically seen in
humans with T1D.
Pattern recognition receptors (PRRs): are a class of molecules which detect and respond to
sequences common to many forms of pathogens.
Protein kinase R (PKR): is a protein kinase and a member of one class of PRRs which becomes
activated in response to the presence of dsRNA in cells (and, possibly, also by other stressors,
including endoplasmic reticulum stress). Its primary substrate is a eukaryotic initiation factor,
eIF2α, which, when phosphorylated by PKR, attenuates the rate of protein synthesis in cells.
19
Figure legends
Figure 1. Schematic representation of the sequence of events which derive from infection of
pancreatic β-cells with either a fully replicative enterovirus (left; red circles) or a replication
deficient enterovirus (right; black circles).
The sequence of events arising from infection of islets is summarised in each case via a flow
diagram with the key components highlighted in sequentially numbered circles and described in
the accompanying box.
Figure 2. Schematic representation of the possible sequence of events that might ensue following
enteroviral infection in humans
An initial respiratory or GI tract infection could spread to the pancreas and may then either be
resolved, lead to acute islet cells loss and fulminant diabetes (especially in Japanese populations)
or establish a persistent infection within β-cells. The cells may then either respond only modestly
leading to long-term persistence and, conceivably, no adverse consequences or, alternatively, they
may respond strongly in an attempt to resolve the infection. In the latter case we propose that the
relative “strength” of the response could be critical to the development of autoimmunity and T1D.
20