HC2: Introduction to the immune system

Narcolepsy

Narcolepsy is a neurological disorder linked to HLA and autoimmunity. When a narcolepsy-patient is excited, his muscles relax and he passes out.

Defense mechanisms

The human body is challenged by many different types of pathogens, which trigger different immune responses. The body is made of a very firm physical and chemical barrier, which prevents pathogens from invading and taking over. The basic defense mechanisms of the body are organized in 3 layers:

1. Physical barriers
2. Innate immune system
3. Adaptive immune system

Physical barrier

The physical barrier prevents bacteria from entering the body. This is mostly done by commensal flora, specific bacteria which protect the body. Epithelia form a tough impenetrable barrier which lines the outer surface and inner cavities of the body, for example:

• Respiratory tract: various airway epithelial functions collectively form a major host defense
  • Cilia ensure that pathogens are moved upwards
• Skin: contains a stratum corneum which makes it even harder for pathogens to enter
• Gastro-intestinal tracts: contain goblet cells → mucus production
  • Goblet cells are large white cells
  • Mucus contains antibacterial enzymes
• Urine tracts

The human body itself also possesses many bacteria, which aren't damaging but protect the body from intruders. They ensure that other bacteria don't colonize.

Innate immune system

The innate immune system delays pathogenic replication and spreading until the adaptive immune system can take over. Without the innate immune system, spreading of the pathogen can't slow down. Without the adaptive immune system, the pathogen can't die completely.

In case of an injury, the physical barrier is damaged → intruders can easily enter the body. As a result, the innate immune system is switched on. The innate immune system is pre-programmed → it is activated quickly, a few hours after the pathogen has entered. The innate immune system blocks about 95% of the pathogen-attacks. It isn't a changeable system → it can only process certain pathogens with a limited amount of receptors and proteins.

Leukocytes

Leukocytes are white blood cells. Blood usually contains 4,5-10⁹leukocytes/L. The morphology and normal distribution of leukocytes in blood has certain properties:

• Eosinophils (5)
  • Color depends on liquid used
  • 1-6% of leukocytes
• Monocytes (2)
  • Relatively large
  • 2-10% of leukocytes
  • Coffee-bean shaped big nuclei
  • Produced daily
  • Can stay in tissue for months-years
• Neutrophils (3,4)
  • Produced continuously by the bone marrow
  • 40-75% of leukocytes
  • Only survive for 1-2 days in tissue once they have left the bloodstream
  • Segmented nuclei
• Lymphocytes (1)
  • Can stay in tissue for months
  • 20-50% of leukocytes
  • Relatively small volume of cytoplasm
  • Relatively large nucleus
• Basophils (6)
  • Play an important role in allergic responses
  • <1% of leukocytes 

Origin of immune cells:

White blood cells can be created in 3 different ways. All leukocytes originate from hematopoietic stem cells, which reside in the bone marrow:

• Lymphoid line: hematopoietic stem cell → common lymphoid

HC4: Pathology of normal immune response

Techniques to visualize diseased tissue

Pathology always starts with tissue:

• Biopsies
• Resections
• Autopsies

3 techniques can be combined to make a final diagnosis:

• Immunohistochemical techniques
  • Possible to make a diagnosis in an hour
  • Tissue is frozen
• Light microscopy
  • Most important
  • A tissue specimen goes into a paraffine block to make very thin slices → the slices are stained
    • Glomeruli of a healthy kidney have silver staining
    • In case of chronic inflammation, blue dots are present → lymfocytes
• Electron microscopy
  • Used to look for details
    • For example podocytes

Myocardial infarction:

Normal light microscopy of the heart shows that all cells have a nucleus (the heart is a muscle). Many blood vessels and few lymphocytes are present. A myocardial infarction is an ischemic injury, usually caused by a thrombus in a coronary artery:

1. Cell death: necrosis of myocytes
   • Hours after the incident
   • Cardiomyocytes lose their nucleus
   • Neutrophils are the first leukocytes to arrive to clean up the necrotic tissue
2. Inflammatory reaction with neutrophilic granulocytes
   • Days after the incident
   • Ischemic tissue is infiltrated massively by cells with lobulated nuclei → neutrophilic granulocytes
3. Fibroblast proliferation: remodeling of connective tissue through collagen disposition
   • Weeks after the incident
   • Fibrotic tissue is starting to replace the dead cells
4. Scar formation
   • Months after the incident

This is a general mechanism that can be applied in all tissues → the same process can happen in inflamed skin or lung tissue.

Histomorphology of inflammatory cells and their markers

Inflammatory cells have a distinct histomorphology:

• Granulocytes
  • Indicate an acute infection or inflammatory process
  • Polymorphonuclear cells
    • Neutrophils
      • Most frequent
    • Eosinophils
      • Less frequent
    • Basophils
      • Very rare
  • Marker: MPO
• Lymphocytes
  • Are associated with chronic processes
  • Dark nuclei
  • Small cytoplasm
  • Mononuclear cells
  • Markers
    • CD3: entire population
    • CD8: cytotoxic T-cells
    • CD4: T-helper cells
• Plasma cells
  • Produce antibodies
  • Pericentric nucleus
  • B-cells
  • Marker: CD20
• Macrophages
  • Large cells
  • Present to clean up the mess → digest all sorts of things
  • May turn into multinucleated cells
  • Marker: CD68
• Other cells
  • Dendritic cells
  • Fibroblasts
  • Histiocytes

Visualizing what one cannot directly see

Some questions cannot be answered by "just" looking at light microscopy:

• How can B-cells be distinguished from T-cells?
• Is the complement system involved?
• Are these large cells indeed macrophages or are they tumor-cells?
• Is there a SARS-CoV2 related protein in this cell?
• Are immune complexes involved?

Immunohistochemistry:

Immunohistochemistry is a technique which makes it possible to visualize proteins in tissue. Proteins are stained with a color. This is done by picking an antibody that fits nicely into the protein and then adding color to it.

This technique can be used to visualize B- and T-cells in

HC6: Mechanisms of adaptive immunity

T-cell activation

There are several types of lymphocytes:

• CD8 T-cell
  • Mass destructors → kill cells
• B-cell
  • Production of antibodies
• Regulatory T-cell
  • Dampen everything
• CD4 T-cell
  • Mostly helper-cells → aid other cells in fighting pathogens

There are 2 types of MHC molecules, which are distinguished by their peptides being produced in 2 different cellular compartments:

• MHC class I: peptides derived from protein synthesis by the presenting cell → the antigen is produced by the cell itself
  • An exception is cross-presentation of endocytosed proteins on MHC class I molecules
  • Can be expressed by all nucleated cells
  • CD8 T-cells bind to the a3domain of MHC class I
    • The cell is infected and needs to be cleared
• MHC class II: peptides derived from endocytosed proteins → the antigen is taken up from the surroundings
  • These proteins are extracellular
  • Can be expressed by all antigen-presenting cells
    • Dendritic cells always present MHC class II
  • CD4 T-cells bind to the b2domain of MHC class II
    • The cell itself isn't infected and doesn't need to be cleared

How do T-cells get activated?

Activation of the adaptive immune system starts when T-cells are activated. T-cells get activated by antigen presenting cells. Antigen presenting cells are part of the innate immune system:

1. Dendritic cells, macrophages or neutrophils engulf pathogens such as bacteria and viruses and digest these to obtain antigens
   • The bacteria is engulfed by a phagosome, which fuses with a lysosome that degrades the bacteria into smaller particles
     • Lysosomes have enzymes and a very low pH
2. The cells present the foreign antigens with MHC molecules on their surface
3. A naive T-cell binds transiently to the antigen presenting cell it meets
   • The TCR screens the peptide through the MHC-complex
4. The TCR delivers an antigen-specific signal
   • T-cell-antigen presenting cell interaction is stabilized for days
5. A second signal is required to trigger activation of the naive T-cell → co-stimulation
   • Co-stimulation plays an important role → the combination of an antigen-specific signal and a co-stimulatory signal is required to activate a naive T-cell
   • The co-signal is derived from an alarm signal from the dendritic cell
     • A B7 unit is expressed → IL-2 production
       • If there isn't any B7 expression, the T-cell won't be activated
       • A B7 unit has a CD80 or CD86 receptor, which will connect to CD28 on the naive T-cell
6. Activated T-cells switch on the expression of various genes, including interleukin-2 (IL-2)
   • IL-2 drives proliferation and differentiation of activated naive T-cells
     • Naive T-cells express the low affinity IL-2 receptor, activated cells express the high-affinity receptor
     • Immunosuppressive drugs act by suppressing IL-2 production or action → T-cell responses are dampened
       • For example cyclosporin, tacrolimus and rapamycin

HC8: B-cell development and antibodies

SARS-CoV2

The SARS-CoV2 (COVID-19) virus can bind to the ACE2 receptor molecule. SARS-CoV2 has a spike protein, the S1 domain of this protein can bind to the ACE2 receptor in the respiratory tract and intestines. When the virus arrives, the following happens:

1. Multiple spike proteins bind to different types of ACE2 molecules
2. The virus has a small genome of only 30.000 base pairs → it needs human cells to proliferate
3. Depending on the antibodies available, the virus is blocked a little
4. If the epitopes on the spike proteins aren't blocked, the virus invades the human cells
5. If everything goes well, antibodies that are fully blocking destroy the virus after a few days

Diversity

Every lymphocyte is different, because during development T-cells and B-cells preform rearrangements of immunoglobulin genes. Different parts of V-, J- and D-regions are coupled. This process can reach a diversity of 2 x 10⁶. The diversity can grow even more at the junction site where V, D and J are coupled → junctional diversity. Here, there is an enormous deletion and inversion of nucleotides which increases the diversion up to 10¹²-10¹⁴.

There are several molecular processes in precursor B-cells and peripheral B-cells:

1. The process of rearrangement takes place in the bone marrow
2. Immature B-cells go into the periphery
3. Immature B-cells arrive in the lymph node via arteries and exit the blood stream
4. The immature B-cells arrive in the germinal center
5. The immature B-cells come into contact with an antigen
6. Under support of T-cells, proliferation, somatic hypermutation and class switch recombination of the B-cells takes place → plasma cells and memory B-cells are created
   • This process takes a few days
7. Antibodies are created

Somatic hypermutation:

Somatic hypermutation takes place in the gene segments that coat particles for the variable domains:

• The heavy chain's VDJ-exon
• The light chain's VJ-exon

There are 3 contact spots per antibody chain → complementary determining regions (CDRs). These are the spots where the mutation takes place during the germinal center response. Different rounds of mutation ensure that the antibody can fit into the epitope:

• There is an increased diversity
• The affinity for the antigen is increased

Class switch recombination:

Class switch recombination is critical to change the effective function of an antibody. These functions take place in the constant domains of antibodies, for instance whether an antibody is brought easily onto the epithelial layer. IgA plays a very important role in this process.

CSR exclusively takes place in immunoglobulin genes. A portion of the heavy chain locus is removed from the chromosome, and the gene segments surrounding the deleted portion are rejoined to retain a functional antibody gene that produces antibodies of a different isotype. This can occur multiple times and does not happen randomly → antibody genes are produced to create antibodies according to the B-cells that are most busy at the moment.

Immune monitoring

In total, there are more than

HC10: Repair mechanism

Apoptosis

Apoptosis is a pathway of cell death that is induced by a tightly regulated suicide program. This can be:

• Physiologic: pre-programmed
• Pathologic: associated with necrosis

Morphology:

Apoptosis has the following characteristics:

• Decrease in cell size
  • Shrinkage
• Increase in chromatin concentration
  • Hyperchromasia: the nucleus becomes small and dark
  • Pyknosis → karyohexis → karyolysis
    • Pyknosis = shrinkage
    • Karyohexis/karyolysis = nuclear fragmentation
• Increase in membrane blebs

This is a very organized process → parts are recycled for the body to reuse. Apoptosis always results in phagocytosis, which is done by macrophages.

Biochemistry:

During apoptosis, the following processes take place:

1. Protein digestion
   • This is done by a family of caspases that ensure that the pre-programmed suicide happens in an organized way
     • A family of enzymes
2. DNA-breakdown
3. Phagocytic recognition

There are several subcellular responses to caspase activation:

• Lysosomal response → auto-digestion
• Smooth endoplasmic reticulum (SER) activation
• Mitochondrial swelling
• Cytoskeleton breakdown
  • Thin filaments
    • Actin, myosin
  • Microtubules
  • Intermediate filaments
    • Keratin, desmin, vimentin, neurofilaments, glial filaments

Necrosis

Necrosis isn't programmed and is much faster than apoptosis. It occurs when there is an immediate irreversible, progressive injury. There isn't any time for a nicely orchestrated process:

• A lot of fluid leaves the cell
• Neutrophils are attracted to the cell → inflammatory response
• Macrophages travel to the cell after a while

Necrosis versus apoptosis:

It is important to differentiate between necrosis and apoptosis:

• Apoptosis: normal death/replacements
  • Shrinkage → reduced cell size
  • Fragmented nucleus
  • Intact plasma membrane
  • Cellular content may be intact
  • No adjacent inflammation
  • Often physiologic
• Necrosis: premature death due to causes
  • Swelling → enlarged cell size
  • Karyolysed nucleus
  • Disrupted plasma membrane
  • Enzymatic digestion of cellular content
  • Frequent adjacent inflammation
  • Invariably pathologic

Clinical perspectives

Graft versus Host disease:

The pathway of apoptosis is activated in graft versus host disease. This is cytotoxic T-lymphocyte-mediated apoptosis

Dysregulated apoptosis:

Apoptosis is also activated in cancer. P53 can signal that there is too much injury in DNA and apoptosis is triggered. If P53 is mutated, the apoptosis-process is disregulated and damaged DNA will not result in apoptosis → cancer will progress.

Endometrial cancer:

In a case of endometrial cancer, grade 1 and stage I, the following is visible:

• Complex hyperplasia → atypical proliferation of epithelial cells with partly solid growth
• Squamous metaplasia → the tumor should be classified as endometrioid
• Large areas with necrosis in the center of the atypical proliferation
• Hypertrophic myometrial smooth muscle cells in the surrounding myometrium
• Atrophic ovaries
• Focal tubal metaplasia in the cervix

Ischemia and hypoxia:

Ischemia and hypoxia both cause cell injury. In both cases, there is a reduced oxygen availability. Ischemia is more damaging because there isn't any blood supply at all, which also will lead to nutrients deficiency.

Intracellular accumulations

Due to metabolic scenarios of cellular stress

HC12: Introduction to infectious diseases

The earth's history

Bacteria form an essential part in the development of higher organisms:

1. 4 billion years ago: the first bacteria (prokaryotes)
2. 2,5 billion years ago: the first 1-cellular organisms (eukaryotes)
3. 1,5 billion years ago: the first multicellular eukaryotes
4. 0,5 billion years ago: the first fish

If the age of the earth was 1 year, bacteria would've come in February and humans only would have existed in the last minute of December 31st.

Determination of bacteria

Bacteria can be classified based on the structure of the cell wall

• No cell wall → mycoplasma
• Gram-negative → escherichia coli
• Gram-positive → streptococcus pyogenes
• Acid-fast → mycobacterium tuberculosis

Gram negative and positive bacteria can be distinguished with gram stains:

• Gram-negative
  • Curved → vibrio campylobacter
  • Rods → escherichia and salmonella
  • Cocci → neisseria
    • Spherical
• Gram-positive
  • Rods
    • Spore forming → clostridium
    • Non spore forming
  • Cocci
    • Groups → staphylococci
    • Chains → streptococci

Interaction

Virulence factors describe what the bacterium does with the host (the human). Defense mechanisms describe how the host reacts to the bacterium.

S. pyogenes:

Streptococcus pyogenes is a gram-positive bacteria. They are called extracellular bacteria → cannot survive in a cell. Virulence factors of S. pyogenes are:

• Structural → leads to acute inflammation
  • Peptidoglycan
  • Lipoteichoic acid
• Products
• Characteristics → acute inflammation

S. pyogenes can enter the body when the barrier function is impaired, for example when the granulocyte function doesn't work due to alcoholism. Defense mechanisms of S. pyogenes are made up of the innate immune system.

Laboratory tests

There are 2 blood tests which can be used to test for inflammation:

• C-reactive protein (CRP)
  • Acute phase protein
    • There are lots of cytokines → liver responds by exciting lots of proteins (also CRP) → acute phase response
  • Rises within hours
  • Disappears in days
• Erythrocyte sedimentation rate (ESR)
  • Measure of the amount of proteins in the blood
    • When there are a lot of proteins, they glue together as "stacks"
      • Many proteins can do this
  • Dependent on the concentration of plasma proteins versus red blood cells
    • Fibrinogen
    • Immunoglobulins
  • Nonspecific
  • Rises and disappears more slowly

Both are blood tests that aren't specific for a specific bacterium. CRP is an acute marker, ESR is a chronic marker.

Entamoeba histolytica

Entamoeba histolytica is a human pathogen. It lives in the bowel, where it may make a person ill. It is excreted with the feces in 2 forms:

• Cyst form: survives in the environment → bacteria in this form can be transmitted to other humans

Trophozoites: doesn't survive in the environment → dies within seconds/minutes

HC14: Viruses

Viral diseases

Viral infections are much more common than bacterial infections. Viruses are a major cause of human disease:

• Common cold: rhinoviruses, coronaviruses
• The flu: influenza
• Stomach flu: norovirus
• Fever blister: herpes simplex virus
• Meningitis or poliomyelitis: enterovirus
• HIV-AIDS
• Congenital CMV, rubella
• Et cetera

A virus is a small infectious agent that replicates only inside the host cell. It is a package containing either DNA or RNA (not both), but it is not a cell → it doesn't have organelles and fission. Surface components determine attachment to cell types.

Two important facts about viruses:

• Al viruses are obligate parasites that can only replicate in a cell
• All viruses are parasites of the host protein synthesis machinery → they must make mRNA that can be translated by host ribosomes
  • Viruses use the host cell to make proteins from their mRNA

Viruses are classified by comparing morphology and replication cycles. This provides common basis for naming and allows common clinical approach.

Viral structure

There are very large and very small viruses. Viruses with a larger genome are more complex.

A virus can be naked or enveloped:

• Naked virus
  • Capsid
  • Nucleic acid
• Enveloped virus
  • Envelope
    • A lipid bilayer containing spikes
    • Surrounds the capsid
  • Capsid
  • Nucleic acid

Terminology:

A few important terms that are used to determine the structure of a virus:

• Nucleic acid: DNA or RNA genome
• Capsid: protein structure surrounding the nucleic acid
• Nucleocapsid: capsid + nucleic acid
• Capsomere: subunit of the capsid

Capsid:

Capsid viruses have a symmetrical arrangement:

• Helical
• Icosahedral: 20 triangular seats

Capsid functions are:

• Packaging of viral parts
• Protection of nucleic acids
• Transport of nucleic acids from cell to cell
• Provision of specificity for attachment to the cell
  • Spike proteins

Envelope:

The basic structure of an enveloped virus is as follows:

• Center with DNA or RNA
• A protein coat (capsid)
• A lipid membrane envelope

Capsid versus enveloped viruses:

Capsid and enveloped viruses differ from each other in multiple ways:

• Naked capsid virus
  • More resistant to environmental conditions
    • Drying
  • More difficult to kill with detergent
  • Sensitive to:
    • Chlorine and iodine
  • Enters the host cell by virus-induced endocytosis
  • Exits the host cell by cell lysis
• Enveloped virus
  • More susceptible to environmental conditions
  • Easier to kill with lipophilic detergentia than non-enveloped viruses
  • Sensitive to:
    • Chlorine and iodine
    • Ethanol or propanol 70-95%
    • Chlorhexidine 0,05-0,5%
    • Ammonium, phenol
  • Enters the host cell by membrane fusion or endocytosis
  • Exits the host cell by budding

DNA and RNA

A virus is either a DNA or an RNA virus. It never can have DNA and RNA simultaneously.

DNA viruses:

General properties of DNA viruses are:

• Resemble host cell DNA
• Processing occurs within the host cell nucleus
• Relatively stable within the cell → can persist easily
• Usually have

HC16: Invaders

Pathogenesis

All invaders can make people ill. They are micro-organisms which interact with a host. Humans are full of micro-organisms, which usually don't do anything.

There are 10¹⁴bacteria in the body. Some of these take place and colonize the bowel and mouth, forming protection from other bacteria. Others are only carried and don't cause any harm, or reside in the body in latent form.

Disease symptoms are mainly caused by the human immune host response → it takes 2 to tango. There are only a few micro-organisms which actually cause disease without there being an immune response.

Virulence factors

A virulence factor is any component, product or characteristic that contributes to the ability of a micro-organism to cause disease:

• Structural components
  • Survival of the adverse environment
  • Attachment to human structure
  • Entry into the human body
  • Induction of an inflammatory response
• Products
  • Toxins
  • Enzymes
• Tricks
  • Kill a cell
  • Decreasing the normal immune response
  • Evading the normal immune response
  • Latency

These factors help the virus to:

• Get inside
  • Adhesion proteins or glycoproteins
  • Endocytosis induced by micro-organisms
  • Fimbriae
  • Hyphae
  • (Lipo)techoic acid
• Do harm
  • Enzymes destructing tissue
  • Hyphae
  • Lipopolysaccharide (LPS)
  • Lipoteichoic acid
  • Peptidoglycan
  • Toxin that activated adenylate cyclase
  • Toxin acting as a superantigen
  • Toxin inhibiting the release of neurotransmitters
  • Toxin with cytotoxic activity
• Turn off/hide from the defender
  • Antigenic shifts
  • Antigenic variation
  • Biofilm
  • Protein inhibiting opsonization
  • Intracellular survival
  • Capsule
  • Molecular mimicry
  • Ability to remain latently in the host

It isn't necessary to memorize these factors.

Streptococcus pyogenes

Streptococcus pyogenes is a primary pathogen. It acts via a transient carrier in the throat.

Diagnostics show that it is a:

• Catalase test → gram positive cocci
• Culture plate → Group A b-hemolytic cocci

Symptoms:

Symptoms of an infection with streptococcus pyogenes are:

• Local or disseminated infections
  • Tonsillitis, otitis media
  • Impetigo, erysipelas
    • Erysipelas are skin infections
  • Childbed fever, sepsis
• Exotoxin-mediated syndromes
  • Scarlet fever
  • Streptococcal toxic shock syndrome
• Immunological effects
  • Acute rheumatic fever
  • Acute glomerulo-nephritis

Adhesion:

S. pyogenes has little hairs on its cell wall that touch the epithelial cell. The cell wall of s. pyogenes contains:

• Peptidoglycan
  • S. pyogenes is a gram positive bacteria
• Lipoteichoid acid
• Potrusions connecting to components on the human cell
  • Without these, adhesion wouldn't be possible → the bacteria would be gone after swallowing

Virulence factors:

Several other virulence factors pay an important role:

• Streptolysin → a toxin which makes pores in the cell
  • Belongs to a class of pore-forming, membrane-active, exotoxins
    • After formation of the pore, the epithelial cell dies
  • Causes the lysis of red blood cells that is visible on a culture plate → b-hemolysis
• Streptokinase → lysis of clot
  • In case blood clotting is involved → the bacteria can go anywhere
    • Blood

HC18: Immune deficiencies and infection risk

Components of host defense

There are 6 components of host defense:

• Immunoglobulins
• T-cells
• Complement system
• Granulocytes
• Macrophages
• NK-cells

Impaired barrier function

There are many causes of an impaired barrier function:

• Literally
  • Wounds
  • Insect bites
  • Penetration of the skin by catheters
  • Skin/mucosal toxicity by cytostatic drugs
• Functionally
  • Urinary catheter, incomplete emptying of the bladder
  • Lack of gastric acid
  • Lack of tears
    • M. Sjörgen, an auto-immune disease
  • Disturbance of normal airway cleaning
    • Intubation, abnormal mucus, ciliary function, coughing, COPD
• Impaired colonization resistance
  • All protecting bacteria are removed
    • Can be caused by antibiotics

Microorganisms:

Various microorganisms can cause an impaired barrier function:

• Literal: the microorganisms that already are on the skin or mucosal membranes
  • Primary pathogens
    • Staphylococcus aureus → skin or nose
      • The most common cause of wound and catheter infection
    • Streptococcus pyogenes → skin or throat
  • Commensal opportunistic pathogens
    • Staphylococcus epidermidis → bowel
      • Cannot cause wound infections, but can cause catheter infections
    • Escherichia coli → bowel
    • Bacteroides fragilis → bowel
    • Clostridium sp. → bowel
    • Streptococcal sp. → throat or vagina
    • Candida albicans → throat or vagina
• Functional
  • Urinary catheter, incomplete emptying of the bladder → escherichia coli
  • Lack of gastric acid → salmonella species, vibrio cholera
  • Lack of tears → haemophilus influenzae, streptococcus pneumoniae
  • Disturbance of normal airway cleaning → streptococcus pneumoniae

Candida infection:

If all protecting bacteria are removed due to impaired colonization resistance, the candida will remain, multiply and cause local diseases, for example thrush ("spruw"). This can be caused by using antibiotics. It also is very common among HIV-patients.

Clostridium difficile:

Clostridium difficile is a gram-positive, anaerobic, rod-shaped bacteria that also survives in cases of impaired colonial resistance. It cannot be wiped out by antibiotics and isn't carried by everyone. When it starts to multiply, it causes pseudomembranous enterocolitis. A pseudomembrane consists of mucus and numerous granulocytes. This can cause the whole bowel to become necrotic.

A solution for such an infection is fecal microbiotica transplantation:

1. A healthy person donates their feces
2. The feces are washed and made into a fluid
3. The feces are transmitted to the bowel via a catheter
4. New and healthy microbiota are infused

This works in 80-90% of the cases.

Complement deficiency

Causes of complement deficiency are mainly genetic:

• Classical pathway
• Alternative pathway
• MB lectin pathway
• Terminal pathway (MAC)

One pathway can comprise for the falling out of the other. However, one is very essential to function correctly:

• Terminal pathway → forms the membrane attack complex (MAC)
  • This can cause MAC Neisseria meningitis and Neisseria gonorrhoea infections

Hypogammaglobulinemia

In case of hypogammaglobulinemia, there is an immunoglobulin deficiency. Causes are:

• Congenital
  • X-linked a-gammaglobulinemia
    • The B-cells are lacking
  • As part of SCID

HC20: Diagnostics of infectious diseases

Gram-staining

The Gram-stain test was developed by Hans Christian Gram (1853-1938). It works as follows:

1. Fixation of the bacteria on a glass slide
2. Crystal violet staining → slides through the cell wall
3. Iodine treatment → forms crystal complexes
4. Decolorization → the crystal complexes disappear from the gram-negative bacteria
5. Counter stain safranin

Streptococcus pneumoniae:

A 70-year-old woman who smokes has an increased cough with sputum. Gram-stains are made and show Gram-positive duplex cocci. She is infected by the streptococcus pneumoniae bacteria.

Ziehl-Neelsen

Ziehl-Neelsen staining is used to color acid-fast bacteria, for instance mycobacterium tuberculosis. These bacteria have a lipid in their layer which can specifically be colored to identify it as an acid-fast bacterium.

Other colorization methods

Some other colorization methods are:

• Auramine: can also stain acid-fact bacteria
  • Has a higher sensitivity but a lower specificity than Ziehl-Neelsen
• Giemsa stain: used to diagnose plasmodium falciparum in a thin blood smear
  • Plasmodium falciparum causes malaria

Rapid antigen tests

Rapid antigen tests can be useful because they are indeed rapid, however they have a very low sensitivity → the practical value is limited. SARS-CoV-2 immunochromatographic tests, for instance, have a sensitivity of 30-80%. There is a specific antibody on a piece of paper, and it is tested whether an antigen attaches to it. Immunochromatographic tests are based on pregnancy tests.

Culture of microorganisms

There is methodology based on culture of microorganisms. This is usually used for research of bacteria and viruses, but these days hardly for diagnostics. Culturing has certain characteristics:

• Time-dependent
• Specimen quality is important
• Further analysis is required
  • Morphology
  • Metabolism
  • Antigens
• Some bacteria and many viruses cannot be cultured

In contrary to bacteria, viruses can only be cultured on living cells.

Staphylococcus aureus endocarditis:

A 21-year-old man has fever, chills and a heart murmer. He has suffered from intravenous drug abuse. The doctor decides to take a blood culture:

1. The blood culture is taken and put in a bottle
2. The blood culture is left to grow
3. The culture is taken out of the bottle and put on agar
4. Gram staining and determination

This process takes a while.

Afterwards, further analysis is done:

• Rapid simple tests
  1. Catalase test: the bacteria is a gram-positive staphylococcus
  2. Coagulase test: positive → the bacteria is staphylococcus aureus
• Biochemical properties
• MALDI-TOF
• Antibiotic susceptibility testing

MALDI-TOF mass spectrometry

MALDI-TOF (Matrix-Assisted Laser Desorption/Ionization Time of Flight) is a mass spectrometry test. Several steps are followed:

1. A colony of an unknown organism is selected
2. The organism is prepared onto a MALDI target plate
3. Different organisms spend a different time in the tubes
4. The MALDI-TOF profile spectrum is generated
5. The data is interpreted
6. The species are identified

PCR

PCR is a main diagnostic method these days. Cycles of heating and cooling are used to multiply DNA → PCR is a form of nucleic acid amplification:

1. Target DNA strands are selected

HC extra: Mycobacterial infections (tuberculosis)

Mycobacterium tuberculosis epidemiology

Mycobacterium tuberculosis is characterized by:

• Droplet nuclei
• Alveolar macrophage
• Intracellular survival
• Spread to lymph nodes
• Hematogenous spread
• T-cell immunity → latent stage
• Reactivation

In regions such as southern Africa, 50% of patients with TB have HIV. HIV affects the CD4 cells, which increases the chance of getting TB enormously.

Pathogen and pathogenesis

Mycobacterium tuberculosis can have 6 different clinical manifestations:

• Latent
  • Many patients don't have symptoms but are carriers
• Subclinical
  • Cannot only be transmitted during forceful coughs
• Limited disease
  • Pneumonia in 1 lung lobe
• Extensive disease
  • Pneumonia in both lungs
• Multi-organ
  • Spread by macrophages migrating everywhere in the body
  • For example masses in the abdomen
• Miliary TBC
  • Very rare
  • Granulomas all over the body
    • The granulomas are visible as speckles
    • The necrosis breaks through to the vessel wall → the bacteria are spread via the blood stream

Diagnosis and treatment

Testing:

TB can be diagnosed clinically or via diagnostic tests:

• Clinical
  • Complaints
    • Cough for longer than 2 weeks
    • Fever, night sweats, weight loss
    • Depends on organ involvement
  • Physical examination
• Diagnostic tests
  • Radiography: never specific for TBC
  • Histology: unusual
    • Granulomas
    • Central necrosis
      • Looks like cottage cheese
    • Multinucleate giant cells
  • Microbiological tests: essential for a correct diagnosis
    • Acid fast staining
      • For example Ziehl-Neelsen
      • The cell wall has a high content of complex lipids → mycolic acids
      • Acid fast bacteria stay pink, non-acid fast bacteria become blue after decoloring
    • PCR
    • Culture
      • Takes 2 weeks to 2 months until the bacteria have grown
    • Antibiotic susceptibility testing
    • Genotyping
  • Indirect
    • Tuberculin skin test
      • A small amount of antigens is injected → the T-cell response is measured
      • Positive mantoux test → the T-cells respond in the skin
      • These tests are not specific for TBC
    • Interferon gamma release assay

1. A small amount of blood is put in 1 positive and 1 negative control tube
2. Antigens specific for mycobacterium tuberculosis are put in the tube
3. If the cells have produced the cytokine interferon g, the person has been exposed to mycobacterium tuberculosis in the past or present

Therapy:

The treatment of TBC depends on whether it is active or latent. Multiple drugs are prescribed → a patient has to take at least 2 drugs at the same time:

• Active TBC → treatment has to be a combination of:

• Isoniazide
  • For 6-9 months
• Rifampicin
  • For 6-9 months
• Pyrazinamide
  • For 2 months
• Ethambutol
  • For 2-9 months

• Latent TBC → it is acceptable to treat with 1 or 2 drugs:
  • Isoniazide
  • Rifampicin
  • Isoniazide and rifampicin

It isn't necessary to remember the names of

HC23: Principles of antibiotic pharmacotherapy

Goal of drug treatment

The main goal of drug treatment is to achieve a drug concentration within the body that is therapeutically relevant and appropriate. Knowledge about pharmacokinetics is relevant to determine the appropriate dose and to maintain drug concentrations within the therapeutic index. The most prescribed antibiotic is amoxycillin.

To describe the route of a drug, the ADME aspects are used:

• Absorption
• Distribution
• Metabolism
• Excretion

The processes of metabolism and excretion together form elimination.

Absorption

Absorption consists of the passage of a drug from its site of administration into the circulation. There are 3 main routes of administration for antibiotics:

• Oral
• Epithelial surfaces
• Intravenous injection

Transport:

During absorption, drug molecules are transported across the cell membranes. This can happen via:

• Passive transport → diffusion
• Active transport → carrier-mediated transport using energy (ATP)

Which form is used, depends on the physicochemical properties of the drug molecule:

• Molecular size
  • Drugs with a size smaller than 1000 dalton can be diffused over membranes
    • Bigger drugs need to be injected intravenously
• Relative lipid solubility
  • Drugs with a high lipid solubility can cross the membrane by diffusion
  • Hydrophilic drugs cross the membrane via carriers

Concentration-time curve:

Absorption can be shown in a concentration-time curve. 2 relevant parameters are:

• Cmax: the maximal plasma concentration
• tmax: time when the concentration is maximal

The therapeutic range of a drug lies between the MEC (minimal effective concentration) and MTC (maximal toxic concentration). A drug with a wide therapeutic range is preferable. Penicillins, for instance, are antibiotics with a very wide therapeutic range, while gentamycins have a narrow therapeutic range.

The concentration can be influenced by:

• The dose
• The dose frequency

Case:

A 21-year-old patient is treated with ciprofloxacin (chinolon) because of a severe and complicated cystitis. Ciprofloxacin should not be taken in combination with dairy products, like milk or yoghurt, because:

• It forms a complex with calcium, magnesium, zinc and iron
  • The drug will be too large to be transported across the cell membrane
• The absorption reduces
• The effectiveness reduces

Factors:

Several factors can influence absorption:

• Food intake
• Gastrointestinal motility
• pH at absorption site
• Drug-drug interaction

Bioavailability:

The bioavailability (F) is the fraction of an administered dose that reaches the systemic circulation as an intact drug. This typically is less than 100%, due to:

• Poor absorption
• Metabolic degradation → first pass effect
  • If the drug is in the portal vein, it has to pass the liver before it reaches the systemic circulation

The bioavailability of oral drugs is less than 100%, while the bioavailability of intravenous drugs is 100%.

Distribution

It is relevant to know the distribution of a drug to determine whether a drug is able to reach the site of infection and to determine the effective dose.

Fluid:

Once administered, drugs are distributed over different body fluid

HC25: Epidemiology

Definition of epidemiology

Epidemiology is the study of the occurrence and determinants of illnesses and their spread in the population. It is about the effects between:

• Environment
• Host
• Microorganism

Incidence, prevalence and attack rate

The 3 most important terms in epidemiology are:

• Incidence: the part of the population which develops the disease in a certain time period
  • Number of new cases (n)/(population number (N) x time period)
• Prevalence: the part of the population which has the disease at a certain time point
  • Number of existent cases (n)/population number (N)
• Attack rate: the percentage of "attacked" people
  • Number of people affected/population number (N)

Case:

The following data is available:

• 10 beds, 10 days
• 5 surgical site infections
• 24 patients
• 95 patient days

The following calculations can be done:

• The prevalence on day 3 is: 1/9 = 11%
• The prevalence on day 10 is: 2/8 = 25%
• The incidence is 5/95 patient days
  • 100 patient days is 1 patient staying in the hospital for 100 days, or 4 patients staying for 25 days
• The attack rate is 5/24 = 21%

Endemic, epidemic and pandemic

3 important terms that describe to what extent a disease has spread are:

• Endemic: the disease occurs continuously in a certain part of the population, but doesn't spread any further
  • For example malaria in sub-Saharan Africa
• Epidemic: the disease occurs more frequently than normal and there are more patients than expected
  • For example Ebola in 2015/2016
• Pandemic: an epidemic on worldwide level
  • Can be caused by a DNA-shift → a part of the genome of the pathogen changes

Filoviruses

Currently the Ebola virus is causing an epidemic. In Africa, confirmed cases of Ebola HF have been reported in many countries.

Structure:

Marburg and Ebola are filoviruses, which have a distinguishing structure:

• Negative-stranded RNA virus
• Envelope
• Threadlike structure
• Very broad cell tropism → can infect nearly every cell in the body

Transmission:

Fruitbats form the normal reservoir of Ebola. The hosts are reindeers and monkeys → these animals are infected, but the virus normally stays inside the rainforest. People living around the rainforest can get infest by eating meat of these animals. In conclusion, filoviruses can be transmitted in multiple ways:

• Primary transmission: contact with fruitbats/infected mammals
  • Bushmeat
• Interhuman transmission: contact between body secretions
  • Sweat, mucosae, bloodstream, non-intact skin, aerosols (limited)
• Nosocomial transmission: inadequate sterilization of materials
  • One of the biggest outbreaks of Ebola was caused by hospital needles not being properly sterilized

Incubation period:

The incubation period is the moment of infection up to the moment of the first symptoms. It is important to know the incubation period to determine how long a patient has to be isolated. In this period, the patient has a clinical disease.

Both Ebola and Marburg

HC extra: COVID-19

COVID-19 epidemiology

The start of a pandemic:

COVID-19 originates from Wuhan, in the province Hubei in China. Over 11 million people live there.

Numbers:

Important numbers to know are:

• Incidence: new number of cases/population
• Case fatality rate: number of deceased/total number infected
• R0reproductive number: average number of new cases infected by 1 infected person

In case of COVID-19, almost all numbers aren't correct:

• Only what is tested is known
  • What isn't tested, isn't known
  • Misclassification
• There is always delay
  • Reports are never up to date
  • Disease in specific subgroups can be underreported
• There are always aberrations
  • R0represents the average number, but not the real patterns of transmission

Transmission:

The transmission chain of COVID-19 consists of:

• Reservoir: bats
• Source: humans
• Host: humans

Transmission happens via different routes:

• Droplet-borne route: droplets cannot travel very far because they sink to the ground due to gravity
• Short-range airborne route: transmitted by aerosols
• Long-range airborne route: transmitted by aerosols
• Fornite route: via surface contact or contact between people

Clinical aspects

Clinical course:

From January to May 2020, there were 1,320,488 laboratory-confirmed COVID-19 cases, of which:

• 14% were hospitalized
• 2% were admitted to the intensive care unit
• 5% died

These percentages are only based on the confirmed COVID-19 cases → aren't realistic.

Symptoms of COVID-19 are:

• Cough (50%)
• Fever (43%)
• Myalgia (36%)
• Headache (34%)
• Dyspnea (29%)
• Sore throat (20%)
• Diarrhea (19%)
• Nausea/vomiting (12%)
• Loss of smell or taste, abdominal pain, rhinorrhea (10% each)

Complications of COVID-19 can be:

• Pulmonary embolism
• Acute respiratory distress syndrome
  • Possibly due to a cytokine-storm → hyper-inflammatory state

Diagnosis:

COVID-19 can be diagnosed with molecular techniques → the SARS-CoV2 viral load is highest in respiratory samples. For instance, in case of PCR, the sensitivity is the highest in lower airway samples.

COVID-19 diagnosis can also be done using serology. This is useful when the patient has had the virus for a while → antibodies can be detected around day 10.

Lastly, diagnosis of COVID-19 can be done using radiology. COVID-19 has a specific pattern on a CT-scan. A score is given to the images → a CO-RAD score of 4 or 5 indicates that there is a COVID-19 infection and a CO-RAD score of 1 or 2 indicates that an infection by this virus is unlikely.

Treatment:

There are 3 treatment options to treat COVID-19:

• Supportive care
• Anti-viral
  • Remdesivir
  • Convalescent plasma
• Anti-inflammatory
  • Corticosteroids
  • Dexamethasone
  • Anti-IL6 (tocilizumab)
  • Anti-IL1 (anakinra)
  • Bradykinine-inhibitor (Icatibant)
  • Anti-CD147 (meplazumab)
  • JAK-inhibitor (ruxolitinib)

Remdesivir is an anti-viral way to treat COVID-19. It was originally designed for the Hepatitis C virus, and later used for Ebola and Marburg. It is a ribonucleotide analogue inhibitor of viral RNA polymerase. It gives a faster clinical improvement but no difference in mortality. Usually it is prescribed 10 days after the symptoms start, so it may be more

HC28: Pathology of allergy

Classification

Classification is the act of forming a distribution into groups, classes, orders and families according to some common relations or attributes. A classification system is used to:

• Enhanced the quality of communication
  • Especially between experts involved in the field
• Provide a logical structure for categorization
  • For epidemiologic, prognostic or interventional studies
• Assist in the management of individual patients
  • To this extent, categories should be mutually exclusive and predictive of the subsequent behavior of the disease

Conjunctivitis

Conjunctivitis is a type I hypersensitivity reaction. In case of direct hypersensitivity reactions, there is a fast immunological reaction in a sensitive individual, often called an allergy.

Morphology:

Histological images of conjunctivitis are different from most images of type I hypersensitivity reactions:

• Eosinophils are strongly present
• IgE is hard to find in a histological image
• Mast cells are present, but are hard to distinguish

Symptoms:

Conjunctivitis is an ocular allergy caused by pollen release spores. Symptoms are:

• Itching
• Watery discharge
• Redness

The upper ENT may also be involved.

Therapy:

Therapy consists of:

• Antihistamines
• Mast cell stabilizers
• Non-steroidal anti-inflammatory agents

Anti-GBM nephritis

Anti-GBM nephritis is an example of Type II hypersensitivity. GBM stands for glomerular basal membrane. Type II hypersensitivity reactions are mediated by antibodies directed to antigens on the cell-surface in the extracellular matrix. It is caused by cell destruction, an inflammatory reaction or intervention with the normal function. Deposited IgG-antibodies activate the complement system → inflammatory reaction.

Process:

In case of anti-GBM nephritis, the following happens:

1. Anti-GBM antibodies bind to the basal membrane of capillaries
2. Leukocytes (such as neutrophils) are activated and attack the vessel walls → the basal membrane breaks
3. The vessel walls become infected → glomerulonephritis and alveolar capillaritis
   • Capillary loops are destroyed
4. Inflammatory mediators come into Bowman’s space → lining epithelium starts to proliferate → crescent formation
   • Histological images of glomerulonephritis show a crescent

The IgG antibodies work against the alpha-3 chain of collagen type IV, a complement of GBM. The alpha-3 chain is part of the basement membrane in the kidney and the lung. This causes leukocytes to attack the membrane with antigens. Anti-GBM nephritis typically occurs in the kidneys and lungs.

Goodpasture syndrome:

About 50% of anti-GBM nephritis cases occurs without the lungs being involved. In case of Goodpasture-syndrome, both kidneys and lungs are involved. This usually is paired with hemorrhagia (bleeding). The alveolar walls are inflamed and pneumocytes are clearly visible.

Smokers:

Glomerulonephritis typically occurs in smokers → smoking destroys the alveolar epithelia → antibodies travel from the lungs to the kidneys. This can be diagnosed with serology for anti-GBM antibodies.

Fibrinoid necrosis

Fibrinoid necrosis is a type III hypersensitivity reaction. An important difference between type II and III hypersensitivity is that in case of type III the antibodies bind to solubles.

Morphology:

Histological images of fibrinoid necrosis show:

• Fibrine
• Necrosis
• Inflammation

Process:

Fibrinoid

HC30: Pathology of autoimmunity

Autoimmune diseases and hypersensitivity

Hypersensitivity reactions and autoimmune diseases correlate with each other:

• Hypersensitivity reactions
  • Mechanisms show that the immune system overreacts to certain stimuli
  • Are present in many autoimmune diseases
    • Type I hypersensitivity forms an exception
• Autoimmune diseases
  • Have a hypersensitivity reaction as part of their pathogenesis

Autoimmunity and infection

Patients with a variety of auto-immune diseases report that an infectious disease seems to precede the development of their auto-immune disease. In these cases, an infection can be:

• A trigger
  • For the production of antibodies that cross-react with autoantigens
• An enhancer
  • For activating the immune system by activating inflammatory cells
• A second, third of fourth hit in a complex pathogenesis

Examples are:

• Many patients with IgA nephropathy report flu-like episodes with gastrointestinal and upper airway involvement before the occurrence of hematuria
• Patients with ANCA-associated vasculitis often report that their initial symptoms consisted of upper airway disease with sinusitis

Koch’s postulates:

The association of specific microorganisms with diseases came about as a consequence of the work of the German physician Robert Koch. He formulated a set of criteria that could be used to identify the pathogen responsible for a specific disease. These criteria came to be known as Koch’s postulates:

• The microorganism must be found in abundance in all organisms suffering from the disease, but should not be found in healthy organisms
• It must be possible to identify microorganism from a diseased organism
• The microorganism should then cause disease upon introduction into a healthy organism
• The microorganism should be identified as being identical to the original specific causative agent
• If detection of the microorganism predates disease and/or if its quantity correlated with disease severity a causal relationship becomes more likely

In case of autoimmunity, the microorganism is replaced by a factor such as an antibody. The criteria are a little less strict than for infectious diseases → for instance, the factor must be present in most organisms.

ANCA-associated vasculitis:

Anti-neutrophil cytoplasmic autoantibodies (ANCA) pass these criteria. The antibodies are directed against components in granules of neutrophils, for example against proteinase-3, myeloperoxidase and elastase. Most patients with systemic vasculitis have high values of these antibodies, while healthy patients hardly have any of them.

Other immune mediated diseases are:

• Rheumatoid arthritis
• Sjögren’s disease
• Systemic sclerosis
• Idiopathic interstitial fibrosis of the lung

In some patients, antibodies can be present long before they have symptoms or the disease is diagnosed.

Rheumatoid Arthritis

Rheumatoid Arthritis is a systemic, chronic inflammatory disease, affecting many tissues but principally the joints → hand-knuckles start to swell. A typical patient is a 40-year-old woman. It is relatively frequent in the population, but there are many treatment options.

Histology:

Histological images show a proliferating synovitis that frequently destroys the cartilage or bone, with disabling arthritis. There are villous projections of granulation tissue with inflammatory infiltrate and proliferation of synovial lining

HC32: Vasculitis

Vasculitis

Vasculitis is a hypersensitivity reaction. It is a systemic disease. There is an inflamed blood vessel and fibrinoid necrosis caused by white blood cells that have been stimulated by ANCA. Every blood vessel can be damaged and eventually lead to death.

Peri-arteritis nodosa

In the 19^th century, many descriptions of similar patients appeared. The general name given to their symptoms was periarteritis/polyarteritis nodosa. There was an inflammation of the blood vessels → arteritis, which was arranged in a nodular fashion → nodosa. Both similarities and variations were noticed:

• Similarities
  • Illness of the vessels
  • Rapid course
  • Death
• Variations
  • Organ involvement
  • Differences in histology

This mainly occurred in combination with kidney disease and progressive muscle weakness. Several hypotheses on the etiology of vasculitis were made based on environmental factors, viruses and genetics.

Subtypes

Henoch Schonlein purpura:

Henoch Schonlein purpura is a subtype of vasculitis mainly affecting the skin and gut, but other organs may be affected as well. Patients are mainly children and elderly. The disease is characterized by:

• Abdominal pain
• Purpura on the legs
  • Purple point bleedings

Henoch Schonlein can also affect the kidneys → deposition of IgA in the glomeruli of the kidneys. It is possibly related to IgA nephropathy, which is the most common immune mediated disease in the western world. Therefore, IgA vasculitis is a suggested alternative name for Henoch Schonlein purpura. In other subtypes of vasculitis, no glomerular disposition is visible → the ANCA-test is negative.

Churg-Strauss syndrome:

In 1951, Jacob Churg and Lotte Strauss published on 13 patients with vasculitis who had prominent lung involvement and were previously known with asthma. This typical combination of the lung with eosinophilia became known the Churg-Strauss Syndrome.

Churg-Strauss syndrome is a vasculitis typically involving the lungs with an eosinophilic infiltrate, in patients previously known with asthma. Both asthma and in this case also vasculitis are strongly related to eosinophilic granulocytes. The disease is also known as EGPA (eosinophilic GPA).

Granulomatosis with polyangiitis:

Granulomatosis with polyangitis (GPA) is type of vasculitis that used to be known as Wegener’s granulomatosis. It is a combination of:

• Upper airway vasculitis
• Renal involvement
• Histologically proven presence of granulomas

Typical for people with GPA is the “saddle nose”. This is chronic and is caused by vasculitis destroying the cartilage in the nose → there is no tissue left in the middle of the nose. This may be associated with staphylococcus aureus infections.

Anti-neutrophilic cytoplasmatic auto-antibodies

In case of systemic ANCA-associated vasculitis, anti-neutrophilic cytoplasmic auto-antibodies (ANCA) are detected. These antibodies are a form of IgG and are directed against components in the primary granules of neutrophils, for example:

• Proteinase-3
• Myeloperoxidase
• Elastase
• Other components

ANCA-test:

The ANCA-test is used a lot for diagnostics. It is a type of indirect immunofluorescence:

1. Granulocytes (neutrophils) from healthy donors are put on a glass slide
2. Patient serum is put on the slide
3. ANCA directed towards neutrophils binds to the neutrophils

HC35: Infections and autoimmunity

Overlap

Infection, inflammation and autoimmune diseases can overlap, but this isn’t necessary:

• In case of rheumatoid arthritis, there is inflammation but no infection
• In case of cholera and tetanus, there is infection but no inflammation
• In case of myasthenia gravis, there is autoimmunity but no inflammation

From infection to autoimmunity:

Infection can lead to autoimmunity. An infection induces a normal immune response, but the immune molecules or cells react with self-antigens which are similar to those of the pathogens → cross-reactivity. This can be caused by some microorganisms having qualities that resemble human qualities → normally the body doesn’t react to self-antigens due to positive and negative selection. This is called tolerance, which is expressed by the regulatory T-cells.

Acute rheumatic fever

Case:

A 9-year-old boy of Turkish descent acutely falls ill:

• Fever
• Arthritis of right knee, later of both knees
• Heart murmur
• High inflammation
  • ESR: 140 mm/hour
  • C-reactive protein: 160
    • Normally this is <3
• ECG: prolonged PR interval
• Echocardiography: mitral valve abnormalities

The diagnosis is acute rheumatic fever (ARF).

Clinical presentation:

ARF is an infection and an autoimmune reaction. It is a result of throat infection, but not of skin infection, by streptococcus pyogenes. Immunological effects of this bacteria are:

• Acute rheumatic fever
  • An autoimmune response
• Acute glomerulonephritis
  • A type III hypersensitivity reaction

Not the bacteria, but the immune response causes symptoms. ARF is a syndrome → a combination of symptoms and signs:

• Heart: carditis
• Joints: arthritis
• Neurological: chorea
• Skin: nodules and/or erythema marginatum

Epidemiology:

ARF mainly occurs in developing countries. Recurrent ARF may lead to rheumatic heart disease (RHD):

• Incidence of ARF: 0,5 million/year → 300.000 develop RHD
• Prevalence of RHD: 15 million
• Mortality of RHD: 233.000/year

ARF mainly occurs in children of young age, while RHD is more common among elderly. Approximately 3-6% of the population is susceptible.

Pathogenesis:

Pathogenesis is made up out of:

• Molecular mimicry: auto-immune hypersensitivity
  1. Type II reaction: initial damage → exposes self-antigens
     • IgG formed against streptococcus pyogenes reacts with an M-protein (and possibly group A carbohydrates) on the bacterium
       • M-proteins prevent opsonization
         • Have 3 parts: the response is directed against middle components which have rheumatoid epitopes
  2. The complex cross reacts with:
     • Myosin
     • Heart sarcolemma
     • Synovium
     • Articular cartilage
  3. Subsequent type IV response: Th1 cellular responses
     • The myocardium contains Aschoff bodies
• Genetic predisposition

The prognosis of the first episode is quite good → 27% has no abnormalities after 1 year. Therapy consists of:

• Anti-inflammatory drugs
• Slow-releasing penicillin: to prevent re-infection of streptococcus pyogenes

Jones criteria:

To diagnose ARF, the Jones criteria are used:

• 2 major or 1 major and 2 minor manifestations must be present, plus evidence of antecedent group A streptococcus infection
  • Chorea and indolent carditis do not require evidence of antecedent group

HC37+38: Pharmacology: immunosuppression

What is immunosuppression?

Immunosuppressive drugs are drugs that lower the body’s normal immune response → they interfere with the immune system homeostasis. They are a variety of drugs that prevent the production of antibodies. Commonly, they are used to prevent rejection of the recipient’s body of an organ transplanted from a donor.

Immunosuppression is needed in case of:

• Autoimmune disease
• Transplantation

Examples of such cases are acute Graft versus Host disease and systemic lupus erythematosus. Glucocorticosteroids are examples of immunosuppressive drugs.

History

The history of immunosuppression is a successful story:

1. 1954: the first successful renal transplant
   • Identical twin donor without immunosuppression
2. 1959: the first successful allograft
   • Non-identical twin
   • Sublethal body irradiation
3. 1962: the first successful allograft with an unrelated donor
   • Azathioprine
   • > 1 year survival
4. 1963: reversal of rejection with steroids
5. 1967: the first heart transplant
   • Died of rejection in several days

Immunosuppressive theory

In reality, transplantation trades 1 set of problems for the other. Immunosuppressive drugs shift balances → new problems arise. Transplantation-patients have a multi-drug therapy of drugs with a low therapeutic index which very susceptible to side effects. Multidrug regimens allow for lower doses of each drug → minimize toxicity while providing adequate immunosuppression. They work at different signals/pathways of immune activation.

Acquiring the desired effect is always paired with toxic effects. Every drug has a certain toxicity:

• NIT: non-immune toxicity
  • Give side effects like nephrotoxicity
    • Side effects that aren’t immune related but organ related
  • A drug doesn’t affect primary action mechanisms
• ISE: immunosuppressive effect
  • Effective level
  • Risk of infections is higher
• ID: immunodeficiency
  • Infections, hospitalization

NIT and ID can overlap:

• Positive effects have a trade-off
• There already is toxicity at low levels

For this reason, drug concentrations have to be measured continuously → TDM (therapeutic drug monitoring) is necessary. Blood samples are continuously taken to measure the drug concentration.

Goals of immunosuppression

Goals of immunosuppression are:

• Providing adequate immunosuppression
• Minimizing adverse effects
• Treating adverse effects and chronic, drug-related problems
• Screening for drug-related complications

Current drugs are good for preventing acute rejections, but not chronic antibody mediated rejections → there is a recent improvement in short-term outcomes but less improvement in long-term outcomes. Rejections are often chronic.

The focus is on different combinations:

• Reducing doses of each individual drug to improve outcomes
• Combining of multiple, low-dose drugs is more effective than 1 single drug in its optimal dose

Achieving immunosuppression:

Immunosuppression can be achieved by blocking the immune system. The adaptive immune system can be targeted at multiple sites:

• Belatacept: blocks direct interaction between antigen presenting cells and T-cells
• Cyclosporin and tacrolimus: inhibit T-cell proliferation and block downstream signaling → less IL-2 production
• Rapamycin and MMF/azathioprine → inhibit proliferation of activated T-cells

Classification

Immunosuppressants can be classified in 3 groups:

• Inhibitors of cytokine production