There is a new strain of bacteria that is causing low grade fevers, lower respiratory tract infection and
wheezy bronchitis in children, elderly and the immune compromised. This bacterium is transmitted from
person to person through air droplets, and later travels to the digestive causing inflammation and
diarrhea. Once isolated, you find out this bacterium is gram-negative, coccobacillary, with a capsule
that is heavily made up of non-immunogenic polysaccharides, meaning it is very difficult to elicit an
immune response against this capsule.
The bacterium’s protective polysaccharide capsule is very similar to H. influenzae.
You name the bug Haemophilus sovegae, but call it H. Soveg for short.
1. In your first attempt at creating the vaccine, you decide to target the vaccine against the bacterium’s
protective polysaccharide capsule. You isolate a polysaccharide molecule with highly repetitive
structures, and inject it into mice. While you got a response to your antigen, it was mostly low affinity
IgM antibodies. No matter how many times you boost, you are unable to generate the high affinity
IgG antibodies necessary for an effective vaccine. Explain why.
A polysaccharide is considered a T-independent type 2 antigen, because it has a highly
repetitive structure and is a non-protein antigen. For the most part, both TI-1 and TI-2 will only
elicit an IgM response. Be sure you can distinguish a Type 2 TI from a Type 1 TI antigens.
2. What might be the best strategy for eliciting an immune response to this antigen and what principle
from lecture would explain why this may work?
For this question you need to think about linked antigen recognition. Because
polysaccharides are poor inducers of T-dependent B cell responses, you would need to
conjugate it to a large protein. The idea of linked antigen recognition is that B cells will
recognize the polysaccharide, but take in the entire polysaccharide-protein complex, and
present peptides of the protein on MHCII receptors. At the same time, DC’s are able to pick
up and present the large protein to T cells. T cells activated against peptides of the large
protein are now capable of activating (CD40 ligand, cytokines) any B cell presenting peptides
of the protein.
This allows B cells that are capable of recognizing the polysaccharide, with stimulatory
signals (CD40L, cytokines) to induce class switching and undergo affinity maturation.
3. You have finally created a vaccine that is able to elicit a B cell response that results in the generation
of IgG antibodies. Yay! However, the titer of antibodies is very low, and you would like to boost this
titer by injecting the mice with an adjuvant. What could you inject the mice with in order to boost
their ability to create more, higher affinity IgG antibodies? Also consider what would lead to an
increase in memory B cells.
Think about what pushes B cells to become class-switched, affinity matured memory B cells.
CD40L:CD40 interaction, types of differentiated T cells that express cytokines that drive IgG
production in B cells, etc.
4. You then test your new vaccine on mice that have T cells deficient in CXCR5. You notice a decrease
in antibody titer and class switching. Explain why.
After a T cell has been activated by a DC, it needs to upregulate CXCR5 in order to meet B
cells at the border of the T and B cell zones. Without CXCR5, T cells would not be able to
provide CD40L and cytokine stimulation to B cells.
5. In what ways is a B cell ‘primary focus’ different from a germinal center?
A primary focus is important for quickly generating antibodies during a primary response.
Activated B cells can migrate to form a primary focus, where they continue to proliferate, and
differentiate into plasmablasts. Plasmablasts do not progress through a germinal center, and
are therefore usually of lower affinity than cells that leave the germinal center, and are usually
IgM+. Plasma cells can result from the germinal center and become antibody-producing cells
that are class-switched and affinity matured.
Some activated B cells, together with their activating T cell, can migrate into a lymphoid follicle,
where they continue to proliferate and form a germinal center. Within the germinal center B cells
can undergo affinity maturation and class-switching. By cycling between the light and dark
zones, B cells are able to undergo somatic hypermutation, test their affinity to antigens
presented by follicular DCs, and be selected based on affinity. There T cells can provide survival
signals to B cells with high affinity receptors, and they can become memory B cells or
plasmablasts.
6. You find out that H. Soveg has very close similarity to a common gut microbe (commensal), however
H. Soveg causes inflammation within the intestines, but the commensal microbe does not.
Coincidence Detection! Commensal bacteria might activate/stimulate PRR receptors on
the surface of epithelial however pathogens contain virulence factors, which are factors
that allow them to penetrate into epithelial cells or lamina propria thereby setting off the
host’s intracellular PRRs. Inflammation, in this case, is dependent upon stimulating both
sets of PRRs (extracellular and intracellular). Commensal bacteria typically do not invade
the gut cells or lamina propria.
a. Explain why this H. Soveg is able to generate an inflammatory response.
b. How does the human body protect itself from generating inappropriate inflammatory
responses to commensal bacterial?
c. Give an example.
You find the H. Soveg also makes a toxin! What are two ways that vaccines can help defend against
this toxin, besides eliminating the bug itself?
This question was a little misworded, it should have been worded as “what are two mechanisms
that antibodies can remove toxins.” Vaccines can be generated against toxins to provide
neutralizing antibodies. The example from lecture was mucosally associated IgA, see slides
below. IgA has to mechanisms for removing toxins: 1) IgA within the lumen can bind to toxin
leading to its removal 2) can bind to toxins within the lamina propria and transport across
epithelial cells into the lumen for secretion.
7. What are the unique features about mucosal tissue and what purpose do they serve?
Most important ones:
1) Intraepithelial Lymphoid Cells – Mostly CD8 T cells, they have two purposes – Kill
epithelial cells infected with virus and/or cells that display distress signals (NKG2d,
CD8alpha:alpha homodimers)
2) Microfold cells (M cells) – Highly endocytic, allows for sampling of the lumen, transports
material via transcytosis.
3) Peyer’s patches and other secondary lymphoid tissues (MALT, GALT, NALT, etc)
8. In thinking about causes of autoimmunity, explain why and how each person could develop
autoimmunity?
a. A patient has a deficiency in Foxp3.
A patient without Foxp3 would be unable to generate any type of regulatory T cells. It is the
transcription factor that drives the regulatory T cell (Treg) phenotype. Tregs are important in
preventing autoimmune responses.
b. In a patient that has an HLA allele strongly associated with autoimmunity.
Do not forget HLA are the genes that make up MHC molecules. The question is indirectly asking
how do mutations within MHC molecules lead to propensity to develop autoimmunity. If an MHC
molecular has a propensity to induce autoimmunity, this is suggesting that something about
this MHC will lead to activation of Autoreactive T cells.
c. In a patient that is deficient in AIRE.
AIRE is a transcription factor responsible for expressing tissue specific antigens within the
thymus. This allows developing T cells to be negatively selected against proteins that are
typically expressed in other tissues.
9. What are the key differences between Central and Peripheral tolerances?
Central Tolerance stems from regulatory T cells (Tregs) that are created in the thymus. During
negative selection, autoreactive T cells can be deleted, or can be induced to become regulatory
T cells. Think of central = “center” of T cell development = thymus.
Peripheral Tolerance occurs outside of the primary lymphoid organs (bone marrow and
thymus). This can include the induction of regulatory T cells (iTregs) from mature naïve T cells,
induction of anergy in autoreactive T cells, etc.
10. A patient is infected with a virus that expresses a protein that has 90% homology with insulin. You
later find this patient how now developed Type 1 Diabetes, characterized by auto reactive CD8 T
cells to insulin. What are some reasons for why this person developed T1D?
In this case, the immune system will mount a significant response to the viral protein. Since this
protein has 90% homology with insulin, any cell that recognizes the viral protein is highly likely
to be able to recognize insulin as well. CD8+ T cells that are generated that can recognize both
the viral protein and insulin will destroy the insulin-producing cells and cause T1D. Note: since
insulin is a self-antigen, many of the lymphocytes that can respond to viral protein and insulin
will likely be deleted. Selection is not perfect, however, so some may escape and allow this to
response to occur, since this is occurring in an inflammatory environment (viral infection) that
can likely overcome many of the inhibitory efforts of Tregs and the like.
11. List all of the autoimmune diseases from lecture that are either driven by auto-antibodies, auto-Tcells, or both. Also briefly describe its mechanism of action.
Autoimmune
Disease
Systemic Lupus
Erythematosus
(SLE)
Hemolytic
Anemia
Immune Effector
Autoantibodies
(Form immune
complexes with
target)
Autoantibodies
(Induce complement
activation,
phagocytosis of
target)
Reactivity/Tar
get
Chromatin
(Histones and
DNA)
Erythrocytes
(RBCs)
Mechanism
Tolerance
Affected
Pathological Outcome
Lodging of immune
complexes in kidney
glomeruli
B cell
Kidney inflammation
(nephritis); proteinuria
Kidney dysfunction
Deposition of immune
complexes in blood vessels;
complement activation
B cell
Skin inflammation (rash)
Lysis of erythrocytes
B cell
Anemia
Graves’ Disease
Autoantibodies
(Activation of target)
Thyroid
stimulating
hormone
receptor (TSHR)
Consitutive activation of
TSHR
B cell
Hyperthyroidism;
dysregulated metabolism
Myasthenia
gravis
Autoantibodies
(Inhibition of target)
Acetylcholine
receptor
Inhibition of acetylcholine
signaling
B cell
Peripheral muscle
weakness
Type-1 diabetes
Autoreactive CD8+ T
cells
Beta islet cells
(produce insulin)
Rheumatoid
arthritis
Autoreactive CD4+ T
cells
Synovial joint
antigens
Multiple
sclerosis
(MS)
Autoreactive Th1,
Th17 CD4+ T cells
Myelin sheath
protein
Lysis of beta cells in
pancreas
Production of MMP/RANKL
by synovial fibroblasts;
recruitment and activation
of osteoclasts
Demyelination of neurons
T cell
Loss of insulin production;
dysregulated blood
glucose metabolism
T cell
Degradation of bone and
cartilage in joints
T cell
Neurodegeneration;
paralysis
441 Lecture #18 Slide 75 of 28
Savan
11/09/2016
12. Design an experiment that would provide direct evidence of B-cell mediated autoimmunity? T
cell mediated autoimmunity?
The answers for this are taken directly from Autoimmunity #2 section slides #14 and #17.
13. A fetus is a type of allogeneic graft, why is it not rejected?
The most natural transplant: Fetus, which is
an allograft and not rejected!
•
•
•
•
Fetus contain paternal allogeneic MHC and
minor antigens
Mother does not reject fetus even when she
has had multiple pregnancies in which the
same alloantigens are expressed
Breakdowns in maternal-fetal tolerance can
lead to spontaneous abortion and other
complications such as preeclampsia
Placental tissue barrier
– Trophoblast (outer layer of the
placenta) does not express classical
MHC class-I and class-II
– A non classical MHC, HLA G , is
expressed on the trophoblast. this binds
to inhibitory receptors on NK cells and
prevent NK mediated killing.
The most natural transplant: Fetus, which is
an allograft and not rejected!
•
Local immunosuppressive response in the
mother
– Expression of indoleamine 2,3
dioxygenase (IDO) is enhanced at the
placental interface. IDO catabolizes
tryptophan making it unavailable. T
cells deprived of tryptophan become
anergic. Administration of an inhibitor
of IDO into pregnant mice caused rapid
rejection of allogenic but not syngenic
fetuses.
– Placental environment leads to
induction of regulatory T cells specific
for paternal antigens
14. What is the difference between the major and minor histocompatibility antigens?
Minor-H antigens can be derived from any polymorphic cellular protein
15. How does your body become tolerized to their own MHC molecules?
Think about negative selection within the thymus. Antigen presenting cells that present self
peptides (mTECs), will also be presenting process MHC molecules. Therefore, whatever MHC
molecules the mTECs may harbor are presented as self peptides during negative selection, and
T cells that bind too tightly will be eliminated.