Download Path Lect 18 Outline - Chronic Granulomatous Disease

Path Lect 18 Outline - Chronic Granulomatous Disease
Objectives
1. Define immunodeficiencies and discuss their classification into primary and secondary, specific and
nonspecific.
2. Discuss causes, pathogenesis, and clinical manifestations of neutropenia.
3. Discuss causes, pathogenesis, pathology, clinical manifestations, laboratory findings of chronic
granulomatous disease.
4. Discuss causes, pathogenesis, pathology, clinical manifestations, laboratory findings of leukocyte
adhesion deficiency.
5. Discuss causes, pathogenesis, pathology, clinical manifestations, laboratory findings of
myeloperoxidase deficiency.
Basically looking at Disorders that illicit an abnormal inflammatory response
Immunodeficiency Disorders
• A state in which the immune system’s ability to fight infections or malignancies is deficient. – abnormal
inflammatory response
• Classification: Two types of immunodeficiency disorders
– Primary
• Specific (lymphocyte defects) – adaptive immune system involves lymphocytes, specific
receptors, specific things that will activate the: e.x T-cell deficiencies, B-cell
deficiencies, immunoglobulin deficiencies)
• Nonspecific (phagocyte defects) – innate immune system nonspecific mediator activates
the phagocytes they response: ex. neutropenia, chronic granulomatous disease, leukocyte
adhesion deficiency, myeloperoxidase deficiency)
• Neutrophils - Main cell of nonspecific phagocyte and innate immune system
– Secondary (acquired deficiency)
• Example: AIDS
• Clinical features: Common to all immunodeficiency disorders include frequent or unusual infections
which may or may not respond well to antibiotic therapy.
Neutropenia – have issues with innate immune response
 Definition: A reduction in the absolute number of circulating neutrophils below 1,500/mm3.
• Normal WBC count is 5,000-10,000/mm3.
• Neutrophils account for 55-70% of the total WBC count (an absolute number of 2,500-8,000/mm3).
 Severity of neutropenia:
• Mild neutropenia (absolute number of neutrophils – 1,000-1,500 cells/mm3) – might not have
problems
• Moderate neutropenia (absolute number of neutrophils – 500-1,000 cells/mm3)
• Severe neutropenia (absolute number of neutrophils < 500 cells/mm3) – any infectious agent that
enters body will cause sever problems
• Agranulocytosis (absolute number of neutrophils < 100 cells/mm3) – infections = medical
emergency
 Clinical manifestations:
• Increased risk of bacterial infections - have issues with innate immune response
• Risk of bacterial infections is proportional to both the severity and duration of the neutropenia.
 Etiology:
• Acquired (most common)
• Hereditary
Acquired Neutropenia
• Causes: Drugs(chemotherapy) and chemicals
• Morphology: Depression of hematopoiesis in the bone marrow (bone marrow hypoplasia) – from an
inherited defect or radiation, or toxic effect from a drug
• Pathogenesis:
• Direct inhibition of myelopoiesis in most cases. -Myelopoiesis – differentiation of stem cells into
different components of your blood
• Antibody production that destroy circulating granulocytes. – if they think they are foreign –
autoimmune disorder
• Clinical manifestations: Neutropenia can go undetected, but is generally discovered when a patient has
developed infections. Frequent infections or infections in unusual areas
• Involved bacteria: Endogenous flora.
• S. aureus in skin infections
• E. coli and Pseudomonas in GI infections
• Mixed aerobes and anaerobes in oral infections.
Hereditary Neutropenia (three types)
1. AUTOIMMUNE NEUTROPENIA OF INFANCY (ANI)
• Characterized by increased destruction of neutrophils as a result of antineutrophil antibodies(from
mom) against human neutrophil antigen 1(on baby). - i.e. mom’s Ab will attempt to destroy baby’s
neutrophils
• Pathogenesis: Antineutrophil antibodies mediate peripheral destruction of the neutrophils.
• Etiology: The cause of antibody production is unknown.
• Epidemiology: Incidence is 1 in 1 million population. The average age at diagnosis is 8 months.
• Clinical manifestations:
• Recurrent mild infections in 80% of patients, such as otitis media, upper respiratory infections.
• Severe infections in 20% of patients such as pneumonia, meningitis, sepsis.
• Outcome: Usually a benign condition, lasts 6-24 months, then antibodies disappear - fetus recovers
2. SEVERE CONGENITAL NEUTROPENIA (SCN)
• Definition: A rare disorder with severe neutropenia present at birth.
• Genetics: Mutations of the ELA2 gene (encoding neutrophil elastase) for most cases. Makes
Neutrophils that have a shorter lifespan than normal.
• Morphology: Bone marrow reveals hyperplasia of neutrophil precursors(b/c inflammatory stimulus
continue to release stimuli, it’s not being eliminated b/c neutrophils don’t live long enough to mature
and go fight stimulus) but these cells cannot form enough mature neutrophils.
• Clinical manifestations: Recurrent fever and life threatening infections (stomatitis, cellulitis,
pneumonia, diarrhea, meningitis, peritonitis) caused by S. aureus, E. coli and Pseusdomonas.
• Laboratory studies: Severe persistent neutropenia.
3. Chronic Granulomatous Disease
• Definition: Chronic granulomatous disease (CGD) is a rare disorder characterized by absent or reduced
function of the respiratory burst in neutrophils (produces oxygen free radicals, important for
intracellular bacterial destruction).
• Cause. CGD represents a group of genetic disorders.
– caused by genetic defects in each of 6 components of the enzyme NADPH oxidase complex,
which catalyzes the respiratory burst in neutrophils.
• Activation of the NADPH oxidase requires several proteins normally present in the
cytosol of resting neutrophil, especially p47phox and p67phox .
• They translocate to the membrane of the phagocytic vacuole and help assemble the active
complex.
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Once the active complex is assembled, an electron transport system operates and
generates superoxide in the phagocytic vacuole.
Genetic forms of CGD.
• About 65% of cases, CGD occurs as an X-linked disease (i.e. males are affected, female carriers are
healthy) caused by a mutation in the gp91phox gene (these are the genes that make the NADPH
oxidase complex – they don’t produce protein products)
• About 25% of cases are inherited in an autosomal recessive manner and are caused by defects in the
p47phox gene. (need both copies to cause defect)
• About 5% of cases are inherited as an autosomal recessive manner and are caused by defects in
p67phox gene.
• About 1% of cases are inherited as an autosomal recessive manner and are caused by defects in
p22phox gene.
Clinical Manifestations
– Recurrent infections.
– Most children with CGD present in the first year of life with recurrent bacterial and fungal
infections. Infections can affect every organ system:
– Pneumonia is the most frequent infection, occurring in 80% of patients.
• Aspergillus is the most frequent isolate.
– Lymphadenopathy with suppurative lymphadenitis is present in 60% of cases.
• Supperation – vigorous inflammatory stimulus which recruits a lot neutrophils to the site
– lots of pus w/in the enlarged lymphnodes
• The cervical chain of lymph nodes is the most frequently involved. S. aureus is the most
common pathogen.
– Hepatic abscesses occur in 50% of patients.
• Abscesses – pocket of cell debris + neutrophils with inflammatory response around them
• Most of them are caused by S. aureus.
– Osteomyelitis occurs in 25% of patients.
• Ostenmyelitis – bone marrow cavity inflammation
• Serratia marcescens is the most common pathogen (a gram (-) from GI tract).
– Failure to thrive: Poor physical growth of any cause.
Pathogenesis: Catalase-positive pathogens.
• The most frequently encountered
• pathogens are S. aureus, Aspergillus spp, enteric Gram-negative bacteria (Pseudomonas, Serratia
marcescens and Salmonella spp), and Burkholderia cepacia.
• These organisms are catalase-positive,
• i.e. they produce catalase, which breaks down H2O2 into water and oxygen gas, to allow the
organism to better resist intracellular killing by H2O2.
• By contrast, patients with CGD are not susceptible to catalase-negative bacteria such as
Streptococcus and Pneumococcus.
Granulomas
• Another hallmark of the disease is the
presence of granulomas caused by
chronic granulomatous response to the
pathogen.
• In CGD patients, the response to
conventional bacteria is dominated by
mononuclear phagocytes, i.e.,
macrophages, which normally would
participate in the later stages of the
inflammatory response and whose
bacteria-directed killing ability is less
than that of neutrophils. – so you get a
less than stellar response i.e. abscess
Prognosis.
• In the 1960s, only 20% of children
survived beyond age of 5.
• As modern prophylaxis and treatment
strategies have been implemented, the mortality of patients with CGD in the United states is 2-5% per
year.
• Prophylaxis with antibiotics and antifungal agents has reduced the frequency of infections.
Laboratory tests.
• CBC shows leukocytosis; there is no neutropenia.
• Leukocyte adhesion molecules are present. these
are required for extravasation,
• Levels of IgG and complement =normal. these act
as opsonins and are required for efficient
phagocytosis, are as well.
• All of the players are here they just can’t perform
the oxidative burst needed to eliminate pathogens
Nitroblue tetrazolium test (NBT test).
• The standard assay for phagocytic oxidase activity.
• This colorless compound is reduced to blue
formazan by the activity of the NADPH oxidase
enzyme system.
• A drop of blood is placed on a microscope slide
coated with an activating agent, such as
lipopolysaccharide or phorbol ester – stimulate Neutrophil to produce oxidative burst
• Phagocytes adhering to the slide are activated by the lipopolysaccharide and upon incubation with NBT
develop blue inclusions.
• The number of NBT-positive cells(cells that picked up the blue) is scored under a microscope.
• No Blue = no active Neutrophils
Chemiluminescence assay.
• Luminescence produced by the direct transformation of chemical energy into electromagnetic energy
(light).
• The formation of oxygen radicals by neutrophils (respiratory burst) is detected in the
chemiluminescence apparatus by providing a substrate (the reagent luminol) that generates light when
excited by oxygen radicals.
• The light output can be detected by a photomultiplier tube and a signal generated that is proportional to
the degree of luminol excitation.
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The blue trace shows the response of normal neutrophils.
The red trace shows the response of deficient neutrophils from
CGD patients.
Blue is good for both lests
Principles of treatment.
• Early and aggressive treatment of infections, and
antimicrobial prophylaxis are the cornerstones of current
therapy for CGD.
• Bone marrow transplantation can cure CGD.
• However, this approach is fraught with clinically
significant morbidity and the risk of developing
potentially fatal severe graft-versus-host disease.
Leukocyte Adhesion Deficiency Syndromes
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Leukocyte adhesion deficiency (LAD) syndromes result from failures of leukocytes to participate in
inflammatory reactions because of missing surface adhesion molecules.
Two types: LAD I and LAD II.
LAD II neutrophils have a defect in rolling, implying impairment in selectin-mediated events.
LAD I neutrophils have a defect in tight adhesion, consistent with absent β2-integrin function.
Pathogenesis.
• The migration of leukocytes from the bloodstream to the tissue during inflammation is defective
• First step: Rolling. Selectins are a family of cell adhesion molecules - Transmembrane glycoproteins.
• In quiescent endothelial cells selectins are stored cytoplasmic granules called Weibel-Palade
bodies.
• Selectin is translocated to the plasma membrane where it functions as a receptor for monocytes
and neutrophils.
• Endothelial selectins are expressed after endothelial activation and bind to the leukocytes through
corresponding selectin-glycoprotein ligands.
• Selectin glycoprotein ligands are constitutively(always) expressed on all leukocytes and can bind to
selectin on the endothelium,
•
Binding of selectins to their corresponding ligands (Sialyl Lewis-X – LAD II) on the neutrophil
surface causes the leukocytes to roll on the endothelium of the venular wall.
• Selectins have a braking action that slows neutrophil transit.
 Second step: Adhesion.
• Binding to selectins tethers the leukocytes and exposes them to local chemokines.
• This activates β2-integrins, which are constitutively present on plasma membranes of leukocytes.
• Integrin activation will increase their affinity and avidity through clustering of integrin
glycoproteins.
• Integrins are recognized by ligands on the endothelial cells.
• Integrin activation is followed by firm adhesion i.e. sticking, after which transmigration occurs.
• Deficient β2-integrins expression on neutrophils limits the adhesive event and restricts neutrophil
movement from blood vessels.
Then diapadesis
Leukocyte Adhesion Deficiency I – Adhesion issues
• Definition: LAD I is an autosomal recessive disorder caused by mutations in the β2-integrin family.
• Epidemiology: LAD I has been reported in fewer than 400 individuals.
• Approximately equal numbers of males and females are affected.
• Most patients present within the first several months of life.
• Etiology: Mutations in the gene that codes for the β-chain (i.e. CD18) of β2-integrins mapped to
chromosome 21q22.3, (q is long part of chromosome, p – short portion) resulting in the production of a
truncated protein(don’t function properly).
• Two phenotypes of LAD I: “severe deficiency” and “moderate deficiency”.
• The severity of infections appear to be directly related to the degree of β2-integrin deficiency.
• Severe deficiency subtype
– Patients exhibit less than 1% of normal surface expression of β2-integrin.
– These patients succumb to infections within the first year of life.
• Moderate deficiency subtype.
– Patients exhibit 2-30% of normal surface expression of β2-integrin.
– Have fewer serious life-threatening infections.
– May survive into adulthood.
 Clinical manifestations.
• recurrent bacterial infections primarily localized to skin and mucosal surfaces.
– The most frequent infections include labial cellulitis, otitis media, upper and lower
respiratory tract infections, bowel infections, infections of the perirectal area.
– With tooth eruption, gingivitis and periodontitis develop.
• Sites of infections progressively enlarge and they may lead to septicemia.
• The most frequently encountered bacteria are S. aureus, Streptococcus spp, Escherichia coli, Proteus
mirabilis, Enterococcus spp., and P. aeruginosa.
• Delayed umbilical cord separation.
– LAD I may be diagnosed prior to the onset of infections when delayed umbilical cord
separation is observed in the absence of infection (beyond the normal range of 3-45 days,
median 10 days).
– Separation of the cord that occurs after 3 weeks of life is considered abnormal.
• The mean age of diagnosis is 1.5 months.
 Laboratory studies.
• Marked peripheral blood leukocytosis (5-20 times the normal values – remember this is from
inflamitory stimuli that is not being eliminated). WBC counts of 40,000 to 100,000 cells/mm3
(reference range: 5,000-10,000) are common.
• Neutrophils fail to mobilize. – can’t go into interstial space and eliminate pathogen
•
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Flow cytometry is used to assess the presence of the β2-integrins on leukocytes with the use of
integrin monoclonal antibodies.
Pathology
• The infections are indolent and necrotic.
• Serosanguinous exudate is present but not pus. b/c have no neutrophil activity in the infected area
• Biopsies of infected tissues demonstrate inflammations devoid of neutrophils, in contrast with
marked peripheral blood leukocytosis.
Principles of treatment
• The only cure for patients with severe LAD-I is hematopoietic stem cell transplantation or bone
marrow transplantation.
Leukocyte Adhesion Deficiency II - rolling
• Definition: A hereditary disease that results from a generalized defect in fucose production causing the
absence of Sialyl Lewis-X component of the selectin glycoprotein ligands.
• Etiology: Autosomal recessive genetic mutation in the transporter gene, on chromosome 11.
• The genetic mutation leads to the failure to convert mannose to fucose.
• The gene product transports fucose from the cytoplasm into the Golgi apparatus – to make Sialyl
Lewis-X
• Epidemiology: LAD II is an extremely rare condition (only 6 patients reported).
• Pathogenesis: Rolling, the first step in neutrophil recruitment to site of inflammation and is mediated by
the binding of the selectins to the carbohydrate structure Sialyl Lewis-X. LAD II results in an impaired
emigrations of leukocytes from the blood vessels to the site of infection. (b/c have no Sialyl Lewis-X)
• Clinical manifestations
• No delay in cord separation is observed.
• Patients suffer from early life from recurrent episodes of bacterial infections, mainly pneumonia,
periodontitis, otitis media and cellulitis.
• Infections are not life-threatening and are usually treated in the outpatient clinic.
• Overall infections are comparable to the moderate LAD I.
 Laboratory studies - flow cytometry
• During the infections, neutrophilia occurs.
• Diagnosis is confirmed by the analysis of peripheral blood leuykocytes by flow cytometry using a
monoclonal antibody to determine Sialyl Lewis-X expression.
Myeloperoxidase(MPO) Deficiency
 Definition
• Genetic disorder featuring lack of myeloperoxidase activity.
• An enzyme in granulocytic vacuoles, especially neutrophils – that help kill micro-organisms
• The defect is associated with higher susceptibility to infection, and higher incidence of neoplasm.
 Genetics
• Myeloperoxidase deficiency is an autosomal recessive disease.
• Most patients with MPO deficiency are compound heterozygotes, which means that they have a
different mutation on each allele of the MPO gene, one from each parent.
 Epidemiology
• MPO deficiency occurs 1 in 2,000-4,000 individuals in the general population.
 Pathogenesis
• MPO is a lysosomal enzyme found in the azurophilic granules of polymorphonuclear leukocytes and
lysosomes of monocytes.
• Monocytes contain about a third of the MPO contained in neutrophils.
• This enzyme catalyzes the formation of highly potent antimicrobial agents by reacting with
H2O2 and Cl¯ to form HOCl (hypochlorous acid). Hypochlorous acid is 50 times more potent in
microbial killing than H2O2.
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MPO deficient neutrophils are able to phagocytose, but their capacity to kill bacteria and fungi is
impaired to varying degrees.
Clinical manifestations
• MPO is a relatively benign immunodeficiency, and most patients with MPO deficiency are
asymptomatic.
• It is believed that alternative mechanisms could be efficiency in neutrophils lacking MPO.
• Severe infections occur in less than 5% of patients(these are usually fungal infections, particularly
Candida species), observed predominantly in MPO-deficient patients who also have diabetes
mellitus. Patients without diabetes mellitus rarely have problems.
• Patients with MPO deficiency show a high incidence of malignant tumors.
• Neutrophils mediate tumor cell cytotoxicity (neutrophil MPO system plays a key role in the
tumor surveillance of the host. As a consequence it may be hypothesized that this enzymatic
dysfunction underlies the unexpectedly high incidence of malignant diseases in MPO-deficiency
individuals).
• Benign autoimmune – but cancerous
Laboratory studies
• The presence or absence of myeloperoxidase
can be determined using numerous techniques
such as histochemical staining,
immunohistochemistry, and flow cytometry.
Myeloperoxidase stain