LECTURE 5 (cells of innate immunity) Flashcards
What are the properties of Neutrophils?
- Circulating phagocytes
- Short lived
- Rapid response + not prolonged defense
What are the properties of Monocytes/Macrophages?
- Circulate in blood + become macrophages in the tissues
- Provide a prolonged defense
- Produce cytokines that initiate and regulate inflammation
- Phagocytose pathogens
- Clear dead tissue and initiate tissue repair
What are the two pathways that Macrophages will develop?
CLASSICAL M1
- Induced by innate immunity (TLRs & IFN-y)
- Phagocytosis & initiate inflammatory response
ALTERNATIVE M2
- Induced by IL-4 & IL-13
- Tissue repair & control of inflammation
What are the properties of Dendritic cells?
- Found in all tissues
- Antigen processing and presentation
- Initiate inflammatory response & stimulate adaptive response
What are the properties of Mast cells?
- Found on skin & mucosa
- 2 pathways for activation: innate TLRs and antibody-dependent (IGE)
What are the properties of Natural killer cells?
- Found in blood & periphery
- Functions by direct lysis of cells & secretion of IFN-y
What is the pathway from Macrophages initiating itself?
Macrophages + Dendritic cells -> IL-12 -> Activate NK cells -> IFN-y -> Induce M1 macrophage pathway
What is the Complement system?
A set of interacting proteins released into the blood after production in the liver that act together as zymogens activating one another in cascade fashion after initiation from a variety of stimuli
What are the three major steps of the inflammatory response?
1) Recruitment of inflammatory cells and Anaphylatoxins
2) Opsonisation of pathogens
(uses opsonins to tag foreign pathogens for elimination)
3) Killing of pathogens
(membrane attack complex -> put holes in membrane)
What are the three pathways for the Inflammatory response?
- Alternative (innate)
- Lectin-binding/Mannose-binding pathway (innate)
- Classic (active)
What is the function of the inflammatory response?
- Mediate inflammation
- Enhance phagocytosis by opsonisation
- Cause lysis of particles by membrane pore formation
How is the Mannose-binding pathway initiated?
When mannose-binding lectin binds to carbohydrates on the pathogen
How is the Alternative pathway initiated?
Simple attraction of the early factors to the surfaces of microbes
(bacterial polysaccharides + lipopolysaccharide of the cell envelope of gram-negative bacteria both serve as potent-initiating stimuli)
What is Hereditary Angioedema?
PATHOPHYSIOLOGY:
- Autosomal dominant or de novo mutation
- C1 inhibitor deficiency/dysfunction
- Excessive bradykinin -> fluid extravasation into skin & mucosal tissues
SYMPTOMS:
- Cutaneous swelling
- Colicky abdominal pain, vomiting, diarrhoea
- Laryngospasm + airway obstruction
DIAGNOSIS:
- decreased C4 level
- decreased C1 inhibitor protein or function
TREATMENT:
- C1 inhibitor concentrate
- Bradykinin antagonist
- Kallikrein inhibitor
What is the first thing that happens in the acute inflammatory response?
Activation of the vascular endothelium in the breached epithelial barrier -> Cytokines + other inflammatory mediators released due to tissue damage induce expression of selectin-type adhesion molecules on the endothelial cells -> Neutrophils first to arrive
What are the steps of Extravasation of phagocytes?
1) Rolling phagocytes attach loosely to the endothelium by LOW AFFINITY, SELECTIN-CARBOHYDRATE INTERACTIONS -> E-selectin molecules on endothelium bind to MUCIN-LIKE ADHESION MOLECULES on the phagocyte membrane and bind briefly -> Force of blood flow into area causes cell to detach and reattach repeatedly -> rolls along endothelial surface until stronger binding forces can be found
2) Activation by CHEMO-ATTRACTANTS, Chemokines released in area during inflammation (e.g IL-8, complement split product C5a & N-formyl peptides) bind to receptors on phagocyte surface and trigger a G-PROTEIN-MEDIATED ACTIVATING SIGNAL -> Signal induces a CONFORMATIONAL CHANGE in integrin molecules increasing their affinity for IMMUNOGLOBIN-SUPERFAMILY ADHESION MOLECULES on endothelium
3) Arrest and adhesions interaction between interns and Ig-superfamily cellular adhesion molecules mediate tight binding of phagocyte to endothelial cell + phagocyte movement through extracellular matrix
4) TRANSENDOTHELIAL MIGRATION (phagocyte extends pseudopodia through the vessel wall and extravasates into the tissues)
What happens to Neutrophils whilst in tissues?
- Express increased levels of receptors for chemoattractants
- Exhibit chemotaxis migrating up a concentration gradient towards the attractant
- Release chemoattractive factors that call in other phagocytes
What is Leukocyte Adhesion Deficiency?
A rare autosomal recessive disease in which there is an absence of CD18, the common B2 chain of a number of integrin molecules -> migration of leukocytes us integrin-mediated cell adhesion -> patient suffer from an inability of their leukocytes to undergo adhesion-dependent migration into sites of inflammation
PATHOPHYSIOLOGY:
- Defect in CD18-containing integrins
- Impaired leukocyte adhesion & endothelial transmigration
SYMPTOMS:
- Skin & mucosal infections without pus formation
- Impaired wound healing
- Delayed umbilical cord separation
- Recurrent chronic bacterial infections
LAB FINDINGS:
- Leukocytosis & Neutrophilia
Where is the origin of Chemokines (IL-8)?
- Tissue mast cells
- Platelets
- Neutrophils
- Monocytes
- Macrophages
- Eosinophils
- Basophils
- Lymphocytes
Where is the origin of Complement split product C5a?
Classical or alternative pathways
Where is the origin of Leukotriene B4?
Membrane phospholipids of macrophages, monocytes & mast cells -> Arachidonic acid cascade -> Lipoxygenase pathway
Where is the origin of Formyl Methionyl peptides?
Released from microorganisms
What is Phagocytosis?
When phagocytes ingest and digest particulate debris such as microorganisms, host cellular debris ad activated clotting factors
What does Phagocytosis involve?
- Extension of pseudopodia to engulf attached material
- Fusion of the pseudopodia to trap the material in a phagosome
- Fusion of the phagosome with a lysosome to create a phagolysosome
- Digestion
- Exocytosis of digested contents
What is the function of Neutrophils during phagocytosis?
- Release granule contents in which neutrophils die -> forms pus
- Extrude nuclear contents, histones & neutrophil extracellular traps (NETs) -> Trap + kill pathogens and may damage tissues when enzymes + ROS get released into tissue
Which Anaphylatoxins are there?
- C3a
- C4a
- C5a
Which chemoattractants are there?
- IL-8
- C5a
- Leukotriene B4
- Kallikrein
What are the membrane receptors that both macrophages and neutrophils have for antibody and certain complement components?
- Antibody = IgG
- Certain complement components = C3b
What is Opsonisation?
When an antigen is coated with opsonins, adherence and phagocytosis is enhanced
[opsonisation = the means by which phagocytosis is enhanced]
Describe what happens in Intracellular killing
During phagocytosis, “respiratory burst” activates a membrane-bound oxidase that generates oxygen metabolites which are toxic to ingested microorganisms -> two oxygen-dependent mechanisms of intracellular digestion are activated as a result (NADPH oxidase + Myeloperoxidase)
Describe what NADPH oxidase does in Intracellular phagocytosis
NADPH oxidase reduces oxygen to SUPEROXIDE ANION which generates HYDROXYL RADICALS and HYDROGEN PEROXIDE which are microbicidal
Describe what Myeloperoxidase does in Intracellular phagocytosis
Myeloperoxidase in the lysosomes acts on HYDROGEN PEROXIDE and CHLORIDE IONS to produce HYPOCHLORITE which is microbicidal -> additionally, NITRIC OXIDE SYNTHASE converted arginine to nitric oxide which as antimicrobial properties
Which oxygen-independent degradative materials do the lysosomal contents of phagocytes contain?
- Lysozyme
[digests bacterial cell walls by cleaving peptidoglycan] - Defensins
[form channels in bacterial cell membranes] - Lactoferrin
[chelates iron] - Hydrolytic enzymes
During the acute inflammatory response, which pro-inflammatory cytokines are produced?
- IL-1
- IL-6
- TNF-alpha
Explanation: These cytokines have systemic effects on the tissues, including fever, production of acute phase proteins and leukocytosis
What are the effects of IL-1, IL-6 and TNF-alpha?
- IL-1, IL-6 & TNF-alpha = hypothalamus -> prostaglandins -> fever
- IL-1, IL-6 & TNF-alpha = Liver -> production of acute-phase proteins (e.g CRP, mannose-binding protein and complement components)
- IL-1 & TNF-alpha = bone marrow -> leukocytosis
What is Chronic Granulomatous Disease?
An X-linked recessive deficiency in the production of one of several units of NADPH oxidase which eliminates the phagocyte’s ability to produce many critical oxygen-dependent intracellular metabolites (O2-, -OH, O2 and H2O2). However, the two other intracellular killing mechanisms remain in tact.
PATHOGENESIS
- Impaired respiratory burst & decreased reactive oxygen species -> inhibition of phagocytic intracellular killing
- Patient infected with Catalase (-ve) organism -> bacteria generate their own H2O2 -> used as a substrate for myeloperoxidase and bacterium is killed
- Patient infected with Catalase (+ve) organism -> bacteria break down H2O2 -> host cells have no H2O2 to use + myeloperoxidase has no substrate -> RECURRENT INFECTIONS
CLINICAL MANIFESTATIONS
- Recurrent infections with catalase +ve bacteria + fungi
- Lungs, skin, liver, lymph node involvement
- Diffuse granulomas
Failure of phagocytic cells to generate oxygen radicals are detected by which tests?
- Nitroblue tetrazolium (NBT) reduction test
[normal = formazan +ve (purple-blue), abnormal = formazan -ve (yellow)] - Neutrophil oxidative index (NOI)
- Dihydrorhodamine test
What is the diagnosis and treatment for Chronic Granulomatous disease (GCD)?
DIAGNOSIS
- Measurement of neutrophil superoxide production (DHR flow cytometry + NBT testing)
TREATMENT
- Prophylaxis: TMP-SMX, Itraconazole, Interferon gamma
- Active infection: culture-based, antimicrobial therapy
- Hematopoetic cell transplant is curative
What are the 2 major mechanisms for dealing with viral infections?
- IFN-a/b
- Natural killer cells
What are Interferons?
A family of eukaryotic cell proteins classified according to the cell of origin. IFN-a and IFN-b are produced by a variety of virus-infected cells.
What are the functions of IFN-a and IFN-b?
- Act on target cells to inhibit viral replication (not the virus)
- Not virus-specific (inhibits viral protein synthesis)
- Activation of an RNA endonuclease -> digests viral RNA
- Phosphorylation of protein kinase -> activates elF2 -> inhibits viral protein synthesis
What are the functions of interferons collectively?
- Increase in the expression of class I and II MHC molecules and augment NK cell activity
- Increase the efficiency of presentation of antigens to both cytotoxic and helper cell populations
What is the importance of Interferon-a?
- Antiviral activity
- Used in treatment of hepatitis B and C infections
- Used to treat hairy B-cell leukaemia, Chronic myelogeous leukaemia and Capos sarcoma
What is the importance of Interferon-b?
Used to treat multiple sclerosis -> longer periods of remission + reduced severity of relapses
What is the importance of Interferon-y?
Used to treat Chronic granulomatous disease -> potent inducer of macrophage activation + inflammatory responses -> reverse the CGD patient’s inability to generate toxic oxygen metabolites inside phagocytic cells
What are the side effects of IFN therapy?
- Headache
- Fever
- Chills
- Fatigue
Side effects can be managed with ACETAMINOPHEN
What are the properties of Natural killer cells?
- Kill virally infected cells and tumour cells
- Increased in the presence of Interferons a and b and IL-12
- Share a common early progenitor with T cells but do not develop in the thymus
- Do not express antigen-specific receptors or CD3
- Activity doesn’t generate immunologic memory
What are the markers used clinically to enumerate NK cells?
- CD16 (FcRg)
- CD56 (CAM)
What are the 2 categories of receptor that NK cells employ?
- Killer activating receptor (KAR)
- Killer inhibitory receptor (KIR)
Explanation: If only KARs are engaged, the target cells will be killed. If both the KIRs and KARs are ligated, the target cell lives.
What is the major KAR expressed by NK cells?
NKG2D
What are MIC proteins?
Stress proteins that are expressed only when cells are infected or undergoing transformation
[Upon binding of KAR to a MIC protein -> NK cells become cytotoxic -> resulting in death of the target cell]
How do the KIRs function?
- Activate PROTEIN TYROSINE PHOSPHATASES which inhibit intracellular signalling and activation by removing tyrosine residues from various signalling molecules
- Bind to HLA-E (specialised type of MHC class I antigens) -> HLA-E bind to HLA-A, B and C -> during viral infections/transformed cells, the amount of class I HLA expression may be decreased, preventing leader sequences from binding to HLA-E -> decrease expression of HLA-E and make cells susceptible to NK mediated killing
What happens when NK cells are activated through the FcR (CD16)?
Only one signal is required because the antibody signals that there is an active infection -> occur through a mechanism called antibody-dependent cell-mediated cytotoxicity (ADCC)
