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X-linked Hyper IgM Syndrome

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Simone Sasse

on 4 December 2013

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Transcript of X-linked Hyper IgM Syndrome

X-linked Hyper IgM Syndrome
CD40 ligand
B-cell activation is a two-step process
1. Antigen binding
2. CD40/CD40L interaction between B cell and T helper cell
B Cell Development
Case Study: Dennis Fawcett
Antibody-mediated bacterial killing
What Ab can be produced by someone with a CD40L deficiency?
X-linked Hyper IgM syndrome
X-linked Hyper IgM Syndrome
Question 6
Dennis has an X-linked immunodeficiency and yet there is no informative family history. Is he a new mutation or is his mother a carrier of the defect in the CD40L gene? Can we test his sister to determine whether she is a carrier? Would it be useful to examine the T cells of Dennis' mother and sister for nonrandom X inactivation?
The case of Subject III-3
Overview of B Cell Development
Antigen-Independent Phase
Ig-gene rearrangement
IgM expression
negative selection (central tolerance)
Antigen-Dependent Phase
Antigen activation in SLO
Class switching
Affinity maturation
B Cell Activation Requires Two Signals
1. Antigen Binding
cross-linking of mIg
2. CD40/CD40 ligand interaction
T helper cell binds MHC Class II-peptide
T cells secrete activating cytokines recognized by B cells
cytokines deliver differentiation, proliferation, and survival signals to B cells
Sites of B Cell Activation
Secondary Lymphoid Organs
cortex: rich in B cells
paracortex: rich in T cells
entering Ag percolates, foci of activated B cells form
germinal centers develop; activated B and T cells
Germinal Centers
Three important events:
2. class switching
3. plasma and memory cell formation
1. affinity maturation
Class Switching
mediated by B cell interaction with T helper cell (CD40/CD40L necessary)
allows the variable heavy chain to associate with a constant region of any isotype
effector functions of antibodies secreted determined by isotype of heavy chain constant domain
cytokines determine the class produced
Class Switching
VDJ unit combines with any C gene segment
switch regions upstream of C segments

mediated by switch recombinase and activation-induced cytodine deaminase (AID)
in this example, end up with IgE, with an IgG intermediate
Ig Isotypes
Each human Ig has specialized functions and a unique distribution
determined by class of secreting plasma B cell, Fc portion mediates effector function
5 major types
What does that antibody do?
first antibody type developed during response
activate complement system
found in blood and lymph
IgG1: opsonization

mucosal immunity

sensitization of mast cells

transport across placenta
diffusion into extravascular sites
main Ig in mucosal secretions
immunity to parasites

The Case of Dennis Fawcett, cont.
14 days later: no Ab
to tetanus toxoid or typhoid O and H antigens
Dennis' anti-A and anti-B titers measured
Dennis has RBC of Type O. Why does he have Ab to A and B RBC?
FACS Analysis
of peripheral blood lymphocytes
11% react with antibody to CD19 (B cell marker)
DPT Booster Injection
and Typhoid Vaccine
diptheria toxoid, pertussis, tetanus toxoid
Bacteria in the intestine have Ag closely related to A and B Ag.
87% react with anti-CD3
(T cell marker)
2% react with anti-CD56
(NK cell marker)
All of CD19 cells have
surface IgM and IgD,
none with IgA or IgG
normal results, except...
Activated T cells do not bind soluble CD40
Dennis is treated with intravenous gamma globulin, one injection 600 mg/kg body weight per month
Dennis' brother and sister do not have the disease. No family history.
a bag of blood plasma, which contains gamma globulin
gamma globulins are a class of globulins, identified by serum protein electrophoresis
significant portion are Igs
Dennis remained free of infection!
IgM in response to T-cell independent Ag
no other Ab isotypes

cannot make any Ab to T-cell dependent Ag: vulnerable to many bacteria
T-cell independent Ag
Ag binding to mIg provides both signals for B cell activation
no T cell help required
TI-1:PAMP activation through TLR provides signal 2
TI-2: cross-links mIg and CD21 (via complement C3d)
Question 2
Question 2. Normal mice are resistant to Pneumocystis carinii. SCID mice, which have no T or B cells but normal macrophages and monocytes, are susceptible to this infection. In normal mice, Pneumocystis carinii are taken up and destroyed by macrophages. Macrophages express CD40. When SCID mice are reconstituted with normal T cells they acquire resistance to Pneumocystis infection. This can be abrogated by antibodies to the CD40 ligand. What do these experiments tell us about this infection in Dennis?
activation of macrophages by T cells via CD40/CD40L
histopathology of the lung shows P. carinii infection
Failure to develop leukocytosis
Why are CD40 ligand-deficient males unable to develop leukocytosis?
leukocytosis: rise in WBC count during inflammation
MHC/TCR interaction induces expression of CD40L on T helper cells
CD40/CD40L interaction provides second signal for B-cell activation
CD40/CD40L interaction induces cytokine production in T cell
CD40/CD40L interaction induces cytokine expression
An example: production of IgE
CD40/CD40L interaction, along with TCR/MHC interaction, induces production of IL4
IL-4 induces class switching from IgM to IgE (and IgG1)
CD40/CD40L engagement also leads to upregulation of B7 expression on B cell. Increases co-stimulation of B and T cells and further upregulation of IL-4 synthesis.
CD40 activation leads to isotype switching
CD40 crosslinkage activates protein tyrosine kinases such as Lyn and Syk, creating a signaling pathway leading to production of transcription factors
Together with cytokine regulation, results in class switching recombination, transcription of mRNA, and IgE synthesis
Promotes antigen phagocytosis by macrophages and neutrophils
activated macrophages release GM-CSF: granulocyte-macrophage cell-stimulating factor, which normally activates neutrophils and causes them to divide
develops into neutropenia (neutrophil deficiency)
neutrophils are usually the first responders to a site of inflammation
they phagocytose bacteria and release antimicrobial granules
Precipitation of antibody-antigen products. Product can be flushed from the system before bacteria attacks cells
most abundant WBC
neutropenia can lead to severe sores and blisters in mouth and throat
infested with many bacteria, neutrophils normally protect oral mucosa
give patients GM-CSF to account for lack of macrophage secretion
Antibody is bound to surface of target cell and Fc receptor of NK cells
Allows attacking cell to recognize and destroy antigen
Question 4
Most IgM is in the blood and less than 30% of IgM molecules get into the extravascular fluid. On the other hand, over 50% of IgG molecules are in the extravascular space. Furthermore we have 30 to 50 times more IgG molecules than IgM molecules in our body. Why are IgG antibodies more important in protection against pyogenic bacteria?
Dennis Fawcett
pyogenic bacteria are found in extravascular areas and must be opsonized by IgG molecules to aid with clearance
5 years old
Severe acute infection of ethmoid sinuses
Recurrent since he was 1
Had pneumonia from infection with
Pneumocystis carinii
when 3 years old
treated successfully with antibiotics
Group A Beta-hemolytic streptococci cultured from nose and throat
Neisseria meningitidis
White Blood Cell Count
Doctors expect high WBC count
Count is normal: 4200/microliter
26% neutrophils (low)
56% lymphocytes (normal)
28% monocytes (elevated)
Staphylococcus aureus
various pus-producing bacteria:
leading cause of hospital-acquired infections
skin infections, respiratory disease, and food poisoning
reside in back of nose and throat, may spread to meninges (type B outbreak on campus)
Hyper IgM
nonrandom X inactivation at the cell level is characteristic of female carriers, indicative of a defective X chromosome
What is X inactivation?
dosage compensation for 2 Xs in females and 1 X in males
females inactivate 1 X early in development
generally a random event, with females expressing a mosaic of maternal and paternal X chromosomes
seen phenotypically in female calico cats with both orange and black genes: random inactivation
Thomas, Caroline, et al. "Correction of X-Linked Hyper-IgM Syndrome by Allogenic Bone Marrow Transplantation," New England Journal of Medicine
Pedigree of a Family with X-Linked Hyper IgM Syndrome
2 maternal uncles had died at ages 6 months and 2 years of protracted diarrhea
Complement is Antibody-mediated
inability to counter cryptosporidium infection
what's going on in this pedigree?
The classical pathway of the complement system is antibody-dependent
First, antibody (IgM or IgG) binds to antigen
C1 complex binds to two antibody Fc sites

Dennis was given monthly injections of gamma globulin. What's another possible treatment?
bone marrow transplant
Successfully treated with IV antibiotics.

Serum tested for antibodies to streptolysin O, an antigen secreted by streptococci.

No antibodies to streptococcal antigen found.
Flow cytometric analysis: soluble CD40 protein
normal individual: two pop of cells, CD40 and non-CD40 binding
patient: CD40 fluorescence coincides with nonspecific control (no CD40 binding)
T cells activated (marked by CD25 presence)
Serum Igs
IgG: 25mg/dl (very low)
IgA: undetectable
IgM: 210mg/dl (elevated)
Poorly organized structures with absence of secondary follicles and germinal centers
Lymph node biopsy
Subject III-3 received a bone marrow transplant from his sister.
Normal lymph node
HXIM lymph node
intravenous immune globulin
bone marrow transplantation
ability to create Ab to vaccines
Microsatellite Typing before and after Bone Marrow Transplantation of the the patient. Samples also displayed from the patient's mother, father, and his donor sister.
Pyogenic Bacteria
Pyogenic (pus-forming) bacteria are resistant to phagocytosis unless opsonized with antibody and complement.
Why? These bacteria are gram-positive, more resistant to destruction
Dennis' ethmoiditis caused by Streptococcus pyogenes, a pyogenic bacterium
allele 2 associated with the CD40 mutation (10 bp deletion)
Subject III-3 changed blood type for O- to A+ (blood type of his sister)
Opportunistic Infections
Dennis is susceptible to pyogenic and opportunistic infections
Opportunistic infections: bacteria, viruses, fungi that normally reside in our bodies cause infections when cell-mediated immunity is defective.
Dennis' pneumonia was caused by Pneumocysitis carinii
Common opportunistic infection in AIDS and cancer patients

Dennis' anti-A titer was 1:3200 and anti-B titer was 1:800, both very elevated. Anti-A and anti-B Ab IgM class only.
Question 1
Dennis' B cells expressed IgD as well as IgM on their surface. Why did he not have any difficulty in isotype switching from IgM to IgD?
Antibody-mediated complement system leads to the creation of MAC and lysis of bacteria
Question 3
Why did Dennis make antibodies to blood group A and B antigens but not to tetanus toxoid, typhoid O and H, and streptolysin antigens?
Intestinal bacteria have an Ag closely related to A and B Ags. Dennis creates a cross-reactive Ab for A and B Ags.
Dennis' body has no antigens comparable to tetanus, typhoid, and streptolysin antigens. He cannot create them naturally because of his CD40L malfunction.
Mature, naive B cells express both IgM and IgD. The role of IgD is unknown, but it is necessary for Ag interaction.
There is no switch region between the constant M and D regions. No recombination is necessary for IgD transcription because recombination occurs between switch regions.
Coat pathogens in antibodies such that they are not a threat
Chemical reaction releases oxidative material , kills bacterial cell
Question 5
Newborns have difficulty in transcription of the CD40 ligand gene. Does this help to explain the susceptibility of newborns to pyogenic infections?
Yes. Without CD40L, newborns cannot create many of the necessary antibodies for opsonization
They have, however, inherited IgG from their mother, which confers passive immunity during early life
Cyclosporin A, a drug widely used for immunosuppression in graft recipients, also inhibits the transcription of the CD40L gene. What does this imply for patients taking this drug?
Graft recipients taking Cyclosporin A will also have inhibited class switching and will be unable to make all isotypes upon new infection.
Thank you!
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