T1D vaccine

Question 1

What is insulitis?

Answer 1

The infiltration of the islets of Langerhaans by lymphocytes. Leads to beta cell destruction and formation of pancreatic lesions

Question 2

Which immune cells are present within the infiltrate during insulitis?

Answer 2

CD45+ cells (common to all leukocytes)

CD4+ T helper cells

CD8+ cytotoxic T cells

CD20+ B cells

CD68+ macrophages

Question 3

What is the pattern of lymphocyte concentration during T1D progression?

Answer 3

As more beta cells are killed and less insulin is produced, more lymphocytes infiltrate the pancreas. CD8+ cytotoxic T cells and CD20+ B cells are present in the greatest amount. Once all the beta cells are killed off, all the immune cells leave.

Question 4

How does the enterovirus infect beta cells?

Answer 4

Insulin secretory granules within human beta cells express the Coxsackie and Adenovirus Receptor (CAR-SIV) During exocytosis, CAR-SIV binds enteroviral particles and the particles are then taken in by endocytosis

Question 5

How do beta cells become primed for response to viral infection?

Answer 5

Virus enters systemic circulation Production of IFN Beta cells upregulate production of antiviral proteins

Question 6

What are the possible beta cell responses to a systemic enteroviral infection?

Answer 6

1. Effective beta cell antiviral defence. Beta cells not infected. Infection cleared from systemic circulation
2. Acute enteroviral infection of beta cells
3. Persistent enteroviral infection of beta cells

Question 7

What occurs during an acute enteroviral infection of beta cells?

Answer 7

The hosts antiviral response is impaired. The enterovirus establishes a lytic infection. There is rapid synthesis of large numbers of viral particles. Following cell lysis, viral and beta cell specific antigens are released, triggering activation of islet autoreactive immune cells. There is some beta cell destruction but eventually the virus is cleared and this stops.

Question 8

What are the markers of an acute enteroviral beta cell infection?

Answer 8

Excess of positive strand viral RNA over negative strand RNA.

High levels of VP1 due to viral replication.

Question 9

What occurs during a persistent enteroviral infection of beta cells?

Answer 9

The hosts antiviral response is partially successful. There are deletions in the 5’ UTR of the viral genome leading to problems with replication. The virus produces equal amounts of +ve and -ve RNA strands, forming dsRNA. Presence of dsRNA activates pathogen recognition receptors such as Mda5 leading to enhanced IFN production. IFN upregulates MHC I on beta cell surface leading to insulitis and autoimmune destruction of beta cells

Question 10

What are the markers of a persistent enteroviral infection?

Answer 10

The virus produces equal amounts of +ve and -ve viral RNA, so dsRNA forms.

Low levels of VP-1 due to less viral replication

Question 11

How is genetic predisposition thought to lead to T1D?

Answer 11

Many SNPs are associated with T1D. SNPs are found in genes relating to

• increased IFN response to systemic infection
• increased activity of Mda5
• increased expression of MHC I

Meaning that a persistent infection and autoimmune response is more likely

Question 12

What form is the genome of human enteroviruses and what are the implications of this?

Answer 12

+ve ssRNA Can act as mRNA and be directly translated into a protein, unlike -ve ssRNA that must first be converted into +ve RNA by RNA-depenent-RNA-polymerase

Question 13

Why is T1D thought to be triggered by an environmental trigger rather than genetic?

Answer 13

The rate at which incidences of T1D are increasing is faster that what can be explained by genetics

Question 14

What is the evidence linking viral infection to T1D?

Answer 14

Detection of VP-1, Mda5, RIG-1, PKR and dsRNA in beta cells of diabetic patients Absence of Mcl-1 in beta cells of diabetic patients

Question 15

Why does detection of VP-1 in beta cells indicate viral infection?

Answer 15

VP-1 is a protein encoded by enteroviral RNA which is involved in formation of the viral capsid

Question 16

Why does detection of Mda5 and RIG-1 in beta cells indicate viral infection?

Answer 16

Mda5 and RIG-1 induce expression of IFNs (e.g. IFNalpha) in response to detection of viral dsRNA. Mda5 normally expressed only in alpha cells

Question 17

Why does detection of PKR in beta cells indicate viral infection?

Answer 17

PKR is involved in translational arrest in virally infected cells (turning of protein synthesis). Works by phosphorylating a factor in the protein synthesis pathway called eIF2alpha.

Question 18

Why does absence of Mcl-1 in beta cells indicate viral infection?

Answer 18

Mcl-1 is a labile protein (exists for short amount of time). PKR activity removes all labile proteins.

Question 19

What are the implications of Mcl-1 absence in beta cells?

Answer 19

Mcl-1 is an anti apoptotic protein. Absence makes the cell more sensitive to pro apoptotic stimuli. Possible explanation for beta cell destruction

Question 20

Why does detection of dsRNA in beta cells indicate viral infection?

Answer 20

Enterovirus ssRNA produces dsRNA as it undergoes replication

Question 21

Describe the genome replication process of +ve ssRNA viruses

Answer 21

The viral +ve ssRNA is directly translated into proteins involved in genome replicaiton, such as RNA dependent RNA polymerase.

The +ve ssRNA is then recruited to viral replication complexes.

Formation of dsRNA intermediates.

Replication takes place in membrane invaginations to avoid dsRNA detection.

Question 22

Which type of enterovirus is freqently associated with T1D?

Answer 22

Group B Coxsackie viruses (CVB)