Immunology in Endocrinology

Dr. Jim Johnson, Ph.D. Assistant Professor, Dep t. of C ellular and Physiological Sciences and Dep t. of Surgery,
University of British C olumb ia 5358 Life Sciences Building Lab W eb site: www.diab etes.ub c.ca
Office: (604) 822-7187 E-mail: jimjohn@interchange.ub c.ca
Immunology in Endocrinology

Outline and Objectives:

1) Immunology basics - self versus non-self

2) Immunology basics - specific cell types
3) Immunology basics - T-cell activation
4) Immunology in Endocrinology - Other diseases
5) Immunology in Endocrinology - Type 1 diabetes
6) Immunology in Endocrinology - Type 2 diabetes
Immunology basics - Self versus non-self

‘Immunity’ refers to the global ability of the host to resist the

predation of microbes that would otherwise destroy it. Immunity
has many facets, but the greatest dichotomy separates adaptive
immunity (‘acquired immunity’) from innate immunity (‘natural
immunity’ or ‘innate resistance’).

A key element of immunity is the recognition of ‘self’ and of ‘non-

self’. Errors in this recognition lead to autoimmune diseases,
likely type 1 diabetes.

A complex array of cell types control this process.

Immunology basics - Innate vs. Adaptive Immunity

The immune system is typically divided into two categories--innate and

adaptive--although these distinctions are not mutually exclusive.

Innate immunity
Innate immunity refers to nonspecific defense mechanisms that come into
play immediately or within hours of an antigen's appearance in the body.
These mechanisms include physical barriers such as skin, chemicals in the
blood, and immune system cells that attack foreign cells in the body. The
innate immune response is activated by chemical properties of the antigen.

Adaptive immunity
Adaptive immunity refers to antigen-specific immune response. The adaptive
immune response is more complex than the innate. The antigen first must be
processed and recognized. Once an antigen has been recognized, the
adaptive immune system creates an army of immune cells specifically
designed to attack that antigen. Adaptive immunity also includes a "memory"
that makes future responses against a specific antigen more efficient
Immunology basics - Cellular vs. Humoral Immunity
Questions or discussion
Where are the ‘weak links’ in the immune system?

?
Immunology basics - Specific cell types
Immunology basics - Hematopoietic stem cells
Questions or discussion
What is the proof, in the lab or in the clinic, that these cells are truly stem cells?

?
Immune cell types
Immune cell types

Red blood cell

Monocyte
Neutrophil

Platelet
Immune cell types - B cells
B lymphocytes (from Bursa of Fabricius)

These cells are major antibody producers.

•Plasma B cells are actively producing
antibodies.
•Memory B cells respond quickly following
a second exposure to a specific antigen
Immune cell types - T-cells

T-lymphocytes - mature in the thymus

These cells are a major mediators of cell-mediated immunity.

T-lymphocytes - T-cell receptor
T-lymphocytes talk with antigen-present cells using the T-cell receptor.

The signal from the T cell complex is enhanced by

simultaneous binding of the MHC molecules by a co-receptor.

The co-receptor on helper T cells is CD4, which exclusively

binds the class II MHC.

The co-receptor on cytotoxic T cells is CD8, which is specific

for class I MHC.

The co-receptor not only ensures the specificity of the TCR for
the correctly-presented antigen but also allows prolonged
engagement between the antigen presenting cell and the T cell
and recruits essential molecules (e.g., LCK) inside the cell that
are involved in the signaling of that activated T lymphocyte.
T-lymphocytes - T-cell activation
T-lymphocytes - T-cell activation signalling pathways
T-cell activation - The role of Ca2+ signals
A cytosolic Ca2+ signal is an essential early event in T cell activation.
T-cell activation - The immunological synapse
T-cells ‘lock’ onto antigen presenting cells via a ‘synapse’.
T-cell activation - The immunological synapse
T-cells ‘lock’ onto antigen presenting cells via a ‘synapse’.

T-cell activation signals proceed

the formation of the ‘synapse’.
T-cells - Immunosuppressant drugs block T-cell activation

Islet transplant drugs:

Tacrolimus (FK506) - blocks calcineurin

Sirolimus (rapamycin) - blocks mTOR
Daclizumab - antibody against IL-2R

T-cell activation - Shape changing
T-cell activation - signals controlling cytoskeleton reorganization
Questions or discussion
Any questions on T-cell activation. What types of signal coding are involved?

?
Immune cell types - Subsets of T-cells
Helper T cells - Once activated, they divide rapidly and secrete small proteins called cytokines
that regulate or "help" the immune response.
Cytotoxic T cells - Destroy virally infected cells and tumor cells, and are also implicated in
transplant rejection. These cells are also known as CD8+ T cells, since they express the CD8
glycoprotein at their surface. Through interaction with helper T cells, these cells can be
transformed into regulatory T cells.
Memory T cells - A subset of antigen-specific T cells that persist after an infection. They quickly
expand to large numbers of effector T cells upon re-exposure to their cognate antigen, thus
providing the immune system with "memory" against past infections. Memory T cells comprise two
subtypes: central memory T cells (TCM cells) and effector memory T cells (TEM cells). Memory
cells may be either CD4+ or CD8+.
Regulatory T cells - (a.k.a. suppressor T cells). These are crucial for the maintenance of
immunological tolerance. Their major role is to shut down T cell-mediated immunity toward the end
of an immune reaction and to suppress auto-reactive T cells that escaped the process of negative
selection in the thymus. Naturally occurring Treg cells can be distinguished from other T cells by
the presence of an intracellular molecule called FoxP3. Mutations of the FOXP3 gene can prevent
regulatory T cell development, causing fatal autoimmune disease.
Natural Killer T cells - bridges the adaptive immune system with the innate immune system. NKT
cells recognize glycolipid antigen presented by a molecule called CD1d, rather than MHC. Once
activated, these cells initiate both cytokine production and release of cell killing molecules.

T cells - possess a distinct TCR on their surface. 5% of all T cells.
Autoaggressive T cells - characterized by CD40 expression, which is typically is associated with
antigen-presenting cells. Th40 cells are expanded in autoimmune subjects.
Regulatory T cells - Help suppress responses when appropriate
This subset of T cells is identified by the expression of CD4, CD25 and FoxP3.
Regulatory T cells - Help suppress responses when appropriate

Mechanism of bystander suppression. Tregs that are activated by DCs in an

antigen-specific fashion release IL-10. Once released, IL-10 inhibits immune
reactions not only against the initial antigen (dot) but also against other
antigens (square, triangle) in a nonspecific fashion.
Questions or discussion
Do antigen-specific Tregs have potential therapeutic value?

?
Immune cell types - Natural killer cells

Natural killer cells

These cytotoxic cells are a major

component of the innate immune
system.

They release granules of perforin

and granzyme that cause cells to
undergo apoptosis.
Immune cell types - Macrophages
Macrophage (greek for big eaters)

These cells roam tissues looking for ‘foreign

cells’ or dead cells.

They also display antigens via MHC class II.

Immune cell types - Neutrophils
Neutrophils are granulocytes, filled with neutrally-staining granules which
contain enzymes that help the cell to kill and digest microorganisms it has
engulfed. The neutrophil has a lifespan of about 3 days.

Non-segmented neutrophils are younger.

Immune cell types - Neutrophil targeting and tissue invasion
Immune cell types - Neutrophils communicate with other cells
Through cell–cell contact and secreted
products, neutrophils recruit and
activate monocytes, dendritic cells
(DCs) and lymphocytes, and products
of monocytes, macrophages and T
cells activate neutrophils. Tissue
macrophages ingesting apoptotic
neutrophils produce less interleukin-
23 (IL-23). IL-23 triggers T cells in
secondary lymphoid tissues to
produce IL-17. IL-17 triggers stromal
cells in the bone marrow to produce
granulocyte colony-stimulating factor
(G-CSF). G-CSF promotes
proliferation of neutrophil precursors
and release of neutrophils into the
circulation (see text for references).
BLyS, tumour-necrosis factor-related
ligand B-lymphocyte stimulator;
CXCL12, CXC-chemokine ligand 12;
IFN, interferon-; TNF, tumour-
necrosis factor.
Neutrophils - Release serine proteases with specific targets
Neutrophils - A movie
Questions or discussion
What dangers are there for people who are ‘neutropaenic’?
What could cause this?

?
Autoimmune diseases - A partial list
•Type 1 diabetes (possibly also type 2 diabetes)
•Graves’ diseases
•Hashimoto’s diseases
•Systemic Lupus Erythematosus (SLE)
•Multiple sclerosis
•Rheumatoid arthritis
Autoimmune diseases - Graves’ and Hashimoto’s diseases

Graves' disease is a thyroid-specific autoimmune Chronic thyroiditis (Hashimoto's disease) is a

disorder in which the body makes antibodies to
the thyroid-stimulating hormone receptor (TSHR),
leading to hyperthyroidism, or an abnormally
strong release of hormones from the thyroid gland.
slowly developing persistent inflammation of
the thyroid which frequently leads to
hypothyroidism, a decreased function of the
thyroid gland. Middle-aged women are most
commonly affected.
Autoimmune diseases - Systemic lupus erythematosus

Systemic Lupus Erythematosus (SLE) is a chronic, usually life-long, potentially fatal

autoimmune disease characterized by unpredictable exacerbations and remissions with
variable clinical manifestations. In Lupus SLE there is a high probability for clinical
involvement of the joints, skin, kidney, brain, lung, heart, serosa and gastrointestinal tract.
African-Americans and Asians are disproportionately affected. There is a female to male sex
bias of 8:1.
Autoimmune diseases - Multiple sclerosis

Multiple sclerosis is a central nervous system disorder marked by decreased

nerve function with initial inflammation of the protective myelin nerve covering
and eventual scarring. Symptoms and severity of symptoms vary widely and may
progress into episodes of crisis alternating with episodes of remission.
Autoimmune diseases - Rheumatoid arthritis

Rheumatoid arthritis is a systemic autoimmune disease which initially attacks the

synovium, a connective tissue membrane that lines the cavity between joints and
secretes a lubricating fluid.
Questions or discussion
What do autoimmune diseases have in common? What does this tell us?

?
Autoimmune diseases - Type 1 diabetes
Type 1 diabetes is one of the most common and most expensive autoimmune
diseases.

Frequent urination

Excessive thirst

Unexplained weight loss

Extreme hunger

Sudden vision changes

Tingling or numbness in hands or feet

Feeling very tired much of the time

Very dry skin

Sores that are slow to heal

More infections than usual

Nausea, vomiting, and stomach pains
Type 1 diabetes - Diagnosis
Type 1 diabetes - A model autoimmune disease

Type 1 diabetes:

1) is associated with marked infiltration

of T-lymphocytes.
2) can be transferred from one mouse to
another by auto-reactive T-cells.
3) can be impeded (in mice) by
treatments that suppress the immune
system

Insulin / CD3+ T-cells

Type 1 diabetes - a model autoimmune disease
Type 1 diabetes - Natural history of disease progression
Type 1 diabetes - Genetic susceptibility IDDM 1 - HLA genes

•In humans, the 3.6-Mb (3 600 000 base pairs) MHC region on chromosome 6 contains 140
genes. About half have known immunological functions.
•MHC class II molecules on the surface of antigen-presenting cells display a range of peptides
for recognition by the T-cell receptors of CD4+ T helper cells.
•Thus, MHC class II molecules are central to effective adaptive immune responses, but
conversely, genetic and epidemiological data have implicated these molecules in the
pathogenesis of autoimmune diseases.
•The strength of the associations between particular MHC class II alleles and disease render
them the main genetic risk factors for autoimmune disorders such as type 1 diabetes.
Type 1 diabetes - genetic susceptibility IDDM 1 - HLA genes

MHC Class 1 MHC Class 2

Type 1 diabetes - genetic susceptibility IDDM 1 - HLA genes
Type 1 diabetes - genetic susceptibility IDDM 2 - insulin gene
The IDDM2 locus contributes about 10% of the type 1 diabetes risk that is synergistic with HLA.

The IDDM2 locus contains a sequence of repeated DNA called a variable number tandem repeats (VNTRs).
There are three classes of VNTR in the insulin gene:
• Class I has alleles that range from 26 to 63 repeat units - common in Caucasians (70% of alleles), confers risk.
• Class II has alleles that average around 80 repeat units - rare.
• Class III has alleles ranging from 141 to 209 repeat units - protective.

The presence of at least one class III allele is associated with a 3-fold reduction in the risk of type 1 diabetes,
compared with common I/I homozygote genotype. Because the VNTR occurs in a non-coding region, its
influence on diabetes risk cannot be attributed to an alteration of the protein sequence. The VNTR probably
affects the transcription of the insulin gene. Class III alleles are associated with 15-30% lower INS mRNA in the
pancreas, but higher INS mRNA in the thymus.

The thymus helps train the developing immune system by deleting autoreactive T cells. Because the longer
VNTRs cause more insulin to be produced in the thymus, the detection and deletion of autoreactive T cells may
be more efficient. This improved immune tolerance to insulin would lessen the risk of a future onset of type 1
diabetes caused by anti-insulin antibodies.

Chromosome 11
Questions or discussion
Is there an alternative hypothesis for the effects of the insulin VNTR?

?
Type 1 diabetes - Human susceptibility genes IDDM 3-18
Type 1 diabetes - Genetically susceptible NOD mice
Mouse diabetes genes are denoted with lower case (i.e. iddm1-27).
Human
NOD mouse
Type 1 diabetes - Hundreds of cures for NOD mice
Questions or discussion
Do any of these work in humans? What does that tell us?

?
Type 1 diabetes - Autoantibodies
Most, if not all, are normally found inside beta-cells.
Type 1 diabetes - Autoantibodies to a zinc transporter, ZnT8
Slc30A8 is also a type 2 diabetes susceptibility gene… interesting…
Type 1 diabetes - Insulin is an early autoantigen in humans
Antibodies to insulin are usually the earliest to be detected in high-risk infants.
Type 1 diabetes - Insulin is a ‘primary’ autoantigen in NOD mice
Type 1 diabetes - Triggers?

Examples:
• Viruses
• Cow’s milk (insulin)
Type 1 diabetes - Mechanisms of
-cell death

Pancreatic beta-cells are specifically targeted in type 1 diabetes.

Type 1 diabetes - Mechanisms of
-cell death

Cytokines play an important role in

beta-cell death in type 1 diabetes.
Type 1 diabetes - Mechanisms of
-cell death
Type 1 diabetes - Mechanisms of
-cell death
Type 1 diabetes - Mechanisms of
-cell death
Questions or discussion
Do any of these work in humans? What does that tell us?

?
Type 2 diabetes - Possible role for the immune system?

Ramlo-Halsted et al.
Type 2 diabetes - Possible role for the immune system?
Type 2 diabetes - Possible role for the immune system?

http://www.hsph.harvard.edu/GSH-LAB/tnf-ins.html
Type 2 diabetes - Resident macrophages are found in islets

Islet Macrophage
Type 2 diabetes - Macrophages are abundant in T2D islets
Questions or discussion
If type 2 diabetes is affected by systematic inflammation, how can we use
this information?