Section 4: Blood and Immune Flashcards
Average person has ___L of blood
5L
_____L circulates through a person’s heart every 24 hours
14,000L
Large vs small vessels
Large vessels: High volume, low flow
Small vessels: Low volume, high flow
Vast network of small capillaries require…
Quite high pressures to force blood through
Muscular arteries and valves provide…
Pressurised directional flow from lungs to tissues and organs
Blood pressure ensures…
Even and efficient flow through small capillaries
Low enough to prevent capillary leakage but high enough to avoid coagulation
Why does blood move rapidly
Blood moves rapidly through tissues and small capillaries to ensure muscles and other organs are completely bathed in an oxygenated environment because tissue needs oxygen
Parts of heart
Right and left ventricles
Right and left atrium
Heart and lungs
Pulmonary artery extends from right ventricle to lungs where unoxygenated blood is bathed in oxygen and breathed in through the lungs
The blood is then drawn back in through the left atrium via the pulmonary vein
Left ventricle pumps blood out through the aorta and arterial system to tissues and organs
What is the source of haemopoietic stem cells
Bone marrow
Where do blood cells arise from
Tissue that resides inside bone
Is bone a small or large user of oxygen
Small
Is muscle a small or large user of oxygen
Small
Is the brain a small or large user of oxygen
Large/major
Generates lots of heat - hair prevents heat loss from skull, which is generated by the brain using/burning oxygen
Is the kidney a small or large user of oxygen
Large/heavy user
Filters blood
Liver - blood
Liver is a major recipient of blood via GI and spleen
How is blood divided across the body
Dependent on need
Pressure of arterial blood
Quite high, since arteries are muscular capillaries (thick muscular walls) so when left atrium pumps, those walls expand to carry pressure from the heart
How is pressure measured
Systolic and diastolic pressure
Normal blood pressure
Normal blood pressure is 120/80 (120mm of mercury = mercury of stigma monitor is 120mm high)
Systolic pressure
When blood is at full compression, i.e. left ventricle is squeezed at its tightest and arteries are expanded at their greatest
Diastolic pressure
Heart at complete rest
Too high and too low blood pressure
Too high: arteries are not expanding and contracting effectively, e.g. hardened or blocked (due to disease)
Too low: don’t have enough blood pumping through veins and arteries to supply tissues
Valves
Part of venous system
Prevent backflow because there is no pressure in the venous system - blood is draining back to the right ventricle that is not under as much pressure as the arterial system, i.e. ensures blood is always flowing in one direction
Major components of blood
Cells Proteins Lipids Electrolytes Vitamins, hormones Glucose
Blood - types of cells
RBC - carry haemoglobin
Erythroid
Myeloid (all white cells)
Lymphoid (B cells and T cells)
Haemoglobin contains…
Contains iron which carries oxygen
B lymphocytes come from … and provide …
Come from bone marrow and provide antibodies
Blood - types proteins
Albumin - most abundant Fibrinogen Haemoglobin Immunoglobins Complement Coagulation factors Electrolytes
Blood - lipids
When you eat a fatty meal, those lipids are taken up and transported through blood by lipoproteins
Blood - lipids: lipoproteins
Signal susceptibility to diff forms of coronary heart disease
LDL (low density lipoprotein) - common, ‘bad’ lipoprotein
HDL and VLDL - ‘good’ lipoprotein’
Lipoproteins are based on…
Density; the only way to isolate them is to centrifuge them at high speed
Since they have low density, they will float to the top of a centrifuged tube of blood
Blood - electrolytes
Salts and minerals maintain isotonicity
Deviation from the norm of the normal ions will result in significant illness
K+ is most tightly regulated - regulates many cellular functions
Blood - vitamins, hormones
Transported to various organs by blood
Blood - glucose
Major source of carbon and energy (6C source used by muscle through glycolysis)
Blood separation: centrifugation
One of the simplest forms of separation of blood
Plasma, buffy coat, RBC
Test for health by looking at no. of red cells
- not enough red cells –> anemic
- too many red cells –> blood becomes viscous
- remove plasma and test
Centrifugation: Plasma
55% of blood V
Blood that still has fibrinogen in it that hasn’t been clotted
Centrifugation: Buffy coat
Layer of white cells
Types of blood cells
Erythrocytes: ~5-6 million / ml
Leukocytes: ~10,000 / ml
Platelets: ~400,000 / ml
Blood cells - erythrocytes
Anucleated
Form flat disc
Purpose: carry oxygen to tissue
Major protein: haemoglobin
Blood cells - leukocytes
Immune defense
Neutrophil: responds immediately to microbial challenge
Nucleated
Blood cells - platelets
Coagulation and tissue repair
Important for releasing factors which regulate homeostatic mechanism of tissue repair
Multiple myeloma
Mature B cell malignancy - produces an antibody in a very high amount
Person develops a monoclonal antibody in their blood
Shows on electrophoresis
Patients start to urinate the antibodies
Why don’t you typically see myeloma cells in blood
They typically reside in bone / bone marrow
Since walls of capillaries are very weak…
They leak easily, so osmotic walls on either side of the capillary vessels must be balanced carefully (albumin)
First cells to arrive at site of infection
Neutrophils
Driven to migrate from the capillaries to site of infection by activation of complement
Coagulation factors - haemophilia
Haemophilia’s result from a missing component
Factor VIII deficiency most common form of haemophilia
What are blood cells
Cells that circulate in blood
Origin of blood cells
A single multi-potent stem cell that resides in bone marrow
Multi-potential haemopoietic stem cell
Rare
Has capacity to differentiate into any mature haemopoietic stem cells that populate the body
High conc in umbilical cord
CD34 antigen - can isolate cells relatively easily
Multi-potential haemopoietic stem cell - CD34 antigen technique
Monoclonal antibody usually has fluorescent tag / magnetic bead attached to it –> added to patient’s blood
Hold magnet to side of tube so all CD34 cells bind to the side of the tube where the magnet is and wash away all other cells (leukemic cells) –> relatively purified pop of CD34 cells
Treat patient with high dose of radiation which destroys white cells and leukemic cells
Transplant back the isolated CD34 cells
Treat patient with other factors
Innate immune system contains…
Basophil
Neutrophil
Eosinophil
Monocyte (–> macrophages)
Adaptive immune system contain…
Small lymphocyte –> T lymphocyte and B lymphocyte
B lymphocyte –> plasma cell
Factors that drive haematopoiesis
GM-CSF: Granulocyte macrophage colony-stimulating factor
EPO: Erythropoietin
G-CSF: Granulocyte colony-stimulating factor
Receptors on myeloid progenitor cells bind to…
GM-CSF, and stimulates these cells to differentiate into myeloid cells
What is used to re-populate white cells in leukemia patients following radio-ablation
GM-CSF and G-CSF
Oxygen being transported to tissue allows…
Oxidative phosphorylation –> generate high energy ATP
Membrane of alveoli - thickness
Very thin - allows oxygen to diffuse across it
Blood colour
Pulmonary blood: bright red
Venous blood: dark red
Where is oxygen picked up
Lung alveolus
Haemoglobin - lobes
4 lobes within haemoglobin
Each lobe has a heme molecule, which contains 4 Ns which complex the Fe2+ –> allows oxygen to bind and dissociate under pressures that are normally found at atmospheric pressure / sea level –> oxygen readily associates with Fe2+
Atmospheric pressure / sea level pressure
160mm of mercury
Haemoglobin and oxygen molecules
4 molecules of oxygen per haemoglobin molecule
Haemoglobin - association and dissociation
Oxygen dissociates because partial pressure of oxygen has reduced significantly to ~10-40mm of mercury
While haemoglobin is there, it picks up CO2 (bi-product of respiration)
When it gets out to the lungs, the partial pressure of CO2 reduces and dissociates
What molecules might displace O2
CO and cyanide
Pathways used to activate complement
Classical, lectin, alternative activation - cascades
Complement - classical pathway
C1 –> C4 and C2 –> C3 (major component)
- Antibody binds to antigen on surface of microbe
- C1 complement binds to antibody
- C1 complement creates enzymes that cleaves C4 and C2 complements. C4 is cleaved into C4a + C4b. C2 is cleaved into C2a + C2b
- C4b and C2b collectively forms C3 convertase
- C3 convertase cleaves C3 complement into C3a + C3b. C3b is added onto existing C3 convertase to form C5 convertase. C3a leaves as an anaphylatoxin.
Complement - C5a, C4a, C3a
Anaphylatoxins, which are sensed by the macrophage
Convertase complex
Where proteins bind irreversibly to surface (covalent)
Attract macrophages and neutrophils to site of infection
Macrophages won’t recognise a bacteria until…
It’s coated with complement proteins
Complement - lytic pores
Can insert into some types of bacteria which then immediately kill the bacteria
Formed at end stage of complement
Membrane attack complex (MAC)
Coagulation pathways
Intrinsic pathway (contact) Common Extrinsic pathway (tissue damage)
All are cascades
Coagulation pathway - enzyme thrombin
Exists as an inactive enzyme until activated by factor X
Coagulation pathway - enzyme plasminogen
Converted to active plasmin and cleaves the clot (thrombolysis)
Important for people who have had a thrombosis
Releases clot and saves patient from severe tissue damage
3 areas of immune response
Anatomical and physiological barriers
Innate immunity
Adaptive immunity
Anatomical and physiological barriers - examples
Intact skin - staforius
Cilary clearance
Low stomach pH - most pathogens don’t survive
Lysozyme in tears and saliva - good at disrupting surface of bacteria
Innate immunity - sub-types
Cellular component:
Myeloid lineage gives rise to WBC involved in cellular component
Humoral component:
Soluble proteins in blood designed to opsonise microorganisms as soon as they contact them
Innate immunity - examples
Cellular component:
Neutrophils and macrophages
Humoral component:
Complement
Lectin binding proteins that activate complement - recognises you need carbohydrates found on surface of bacteria –> anti-microbial peptides (e.g. guts, saliva) bind to surface of bacteria
Innate immunity vs adaptive immunity
Innate immunity doesn’t change / strengthen over time
Adaptive immunity strengthens / adapts the longer you are exposed to an antigen
Immune response - tolerance
In both innate and adaptive response, immune systems are said to be tolerant to self
If tolerance is broken, often leads to autoimmune disease
Main types of pathogens
Viruses
Bacteria, yeast, fungi
Protozoa and other parasites
Innate immunity provides…
Our first-line / immediate response to pathogen invasion
Innate immunity is highly developed with which 3 interlinked processes
Complement (C’)
Myeloid cells and phagocytosis (neutrophils and macrophages)
Pattern Recognition Receptors (PRR)
Innate immunity has no…
Memory
3 main types of pathogens require…
3 different defense strategies
Viruses
Intracellular pathogens
Defense relies on cellular immunity - must be able to distinguish infected from normal cells
Replicate on their own - carry genes that provide for enzymes that allow them to replicate once they use host machinery
Most viral infections onset within 24-48 hours and take 10 days from start to finish
Bacteria
Mostly extracellular pathogens
Defense primarily mediated by innate mechanisms and phagocytosis
Slightly higher order of pathogens
Protozoa and parasites
Complex multicellular organisms
Large - too big to be engulfed by phagocytic cells
Require direct killing by chemical mediators released by specialist myeloid cells (basophil, eosinophil, mast cell) - filled with cytotoxic chemicals released by degranulation (e.g. histamine)
Main bacteria distinguished by Gram stain
Gram positive bacteria
Gram negative bacteria
Gram positive bacteria
Thick peptidoglycan cell wall as defense –> lights up with Gram stain
Requires phagocytosis and aren’t killed directly by complement
Gram negative bacteria
Thin peptidoglycan layer surrounded by outer membrane –> doesn’t light up with Gram stain
Can often be lysed directly by complement membrane attack complex
Peptidoglycan wall
Present in most bacteria
Mechanism by which antibiotics work
Bacteria and antibiotics
Bacteria are able to develop resistance to antibiotics
Neutrophil extravasation
Ability of neutrophils to identify site of infection by recognising endothelial cells on inner wall of capillary that’s closest to infection
Whole process takes only minutes from first point of tissue injury
Neutrophil extravasation - steps
- Activation - chemokines from inflammation activates local endothelial cells lining an adjacent capillary wall
- Tethering - neutrophil tethers to inside capillary wall. Mediated by selectins unregulated on endothelial cells and sLe^x
- Adhesion - strong binding between neutrophil integrins and ICAM-1 on endothelium. Neutrophil immobilises and flattens
- Diapadesis - neutrophil squeezes between endothelial cells into interstitial space
- Chemotaxis - neutrophil migrates along a chemokine gradient to site of infection
sLe^x
Sialyl Lewis X
A carbohydrate antigen on neutrophils
Neutrophils phagocytosis of opsonised S aureus - colour
Neutrophils are phagocytosing opsonised S. aureus made green by a fluorescent dye
Neutrophils phagocytosis of opsonised S aureus - steps
Chemoattractants (e.g. C5a) are released that radiate away from bacteria and sensed by leading edge of neutrophil
Neutrophils migrate up chemoattractant gradient - polymerising actin filaments at leading edge and de-polymerising filaments at trailing edge
Neutrophils - complement
Neutrophils have receptors that bind deposited complement proteins, mainly C3b on the surface
Complement receptors
CR1, CR2, CR3, CR4
Myeloid cell receptors that bind activated complement components deposited on bacteria
CR1 is the main neutrophil receptor and binds to C3b
Cross-linking of surface CRs initiates phagocytosis
FcR (antibody) mediates phagocytosis - steps
- Antibody (IgM and IgG) bind to bacterial antigens
- Exposes antibody Fc region
- Neutrophil FcR binds multivalent Fc
- Activates phagocytosis
Phagocytosis - steps
- Ingestion - bacterium is captured by receptors, membrane invaginates into phagosome
- Fusion - phagosome and lysosome fuse to form a phagolysosome
- Acidification - phagolysosome acidifies with H+ pumped in
- Digestion - acidification activates protease and stimulates production of superoxides which kill bacteria (e.g. H2O2 and HOCl)
- Exocytosis - expulsion of digested microbe
Molecular pattern recognition
Innate mechanism
Pattern recognition receptors (PRR) bind complex molecules that are unique to microbes
Molecular pattern recognition - TLR
Toll-like receptors
Activation through TLR stimulates strong innate response through an important inflammation pathway
Pathogen Associated Molecular Patterns (PAMPs)
Molecules unique to microbes recognised by PRRS
Structurally complex
Evolutionarily stable
Stimulate the ‘power’ switch for adaptive response
LPS
Lipopolysaccharide
Membrane component of all gram negative bacteria
Type of PAMP
A ‘pyrogen’ - causes fever and inflammation when injected into bloodstream by binding and cross-linking of TLR4, stimulating an inflammatory response
TLR4 and LPS
TLR4 is the receptor for LPS
Septic shock
Release of LPS by Gram negative bacterial infections leads to life threatening septic shock
B cells and T cells have ability to…
Rearrange a genetic locus to form 2 important molecules; cell surface receptor on surface of T cells and other starts off as cell surface receptor but then becomes a soluble antibody molecule
In B cells, it’s called immunoglobulin locus
In T cells, it’s called T cell receptor locus
Immature B cell receptor
Has a receptor on surface, designed to recognise antigens
Adaptive immunity
Fundamental feature of all higher organisms
Genes that regulate adaptive immune response are identical in all species
Relies on randomly produced antigen receptors
Memory - secondary response stronger and more rapid than primary response
Affinity of B cells towards antigen increases with time and persistence of antigen
Adaptive immunity - vaccinations
When vaccinated, we generate an immune response which lasts almost our lifetime
Memory B cells are primed to recognise the toxin whenever we encounter it
Adaptive immunity - repertoire of B and T lymphocytes
Born with large repertoire of B and T lymphocytes
Each lymphocyte represents a diff antigen specificity randomly produced by rearrangement of genes coding for the antigen receptors
Adaptive immunity relies on…
Phenomenon of gene arrangement or recombination - the only genetic locus capable of this
Affinity
The ability of molecules to recognise and bind tightly to antigens
Immune repertoire developed before birth - problem and solution
Don’t know what antigens the individual will be exposed to throughout lifetime
Develops as many possible combinations of antigen-binding molecules as possible
Antigen - lymph nodes
Antigen is taken up into lymph nodes and develop germinal centres and lymphoid follicles
Starts to produce lots of progeny that have similar but developing antibody molecules
Features of transposon
Enzyme called transposase (does cutting and insertion)
Recognition sequences at end of transposal element
Transposase - working in ‘trans’
Enzyme can work on bits of genes without affecting its own location
RAG1 and RAG2
Transposases Recombination Activation Genes If not present, no immune system will develop Have a single exon Responsible for rearrangement Only active in B and T lymphocytes
Recognition sequences (RS)
Base pair sequences found at ends of any gene segment that rearranges
Substrate for RAG1 and RAG2 directed recombination
Immunoglobulin (Ig) protein fold
Common fold
Repeated Ig domains form antibodies
Loops: where antigen binding site and rearrangement occurs
Loops at ends of strands are not constrained so can vary amino acid sequences without affecting stability of fold (but not all Ig domains vary amino acid sequences)
Immunoglobulin (Ig) protein domain fold - β sheets
Ig protein domain fold is called a β-barrel of ~110 amino acids
Two stable anti-parallel β-pleated sheets joined in middle by a disulphide bond
When is there affinity
When the sum of attractive molecular forces at two surfaces exceed the repulsive forces
The higher the affinity, the fewer molecules it takes per unit volume to associate and dissociate slowly
Avidity
Results from multiple affinity contacts
Strength of binding can be orders of magnitude higher than individual affinities (avidity > sum of all affinities)
When an antibody binds to its antigen, it comes and stays together for a certain length of time
Affinity - molar concentrations
Number of molecules needed of antibody and antigen to give 50% bound as an antigen-antibody complex
Affinity - dissociation constant
How long it takes before the complex dissociates away from equilibrium after you remove the reactants
Equilibrium constant
The conc of antibody and antigen where 50% remains bound to the complex
Ig effector functions and associated Ig molecules
Activates complement: IgG, IgM
Secreted at mucosal surfaces: IgA
Placental transfer - foietal immunity: IgG
High affinity receptor on mast cells: IgE
Membrane bound form: IgM, IgD
Ig molecule gene name / classes (greek)
IgG: γ - most abundant IgM: µ IgA: α IgD: δ IgE: ε - least abundant Gene codes for heavy chain
B cells - Ig genes
Use µ gene first, resulting in a membrane bound IgM molecule - this is the B cell antigen receptor
After activation, B cell switches to using a heavy chain gene, typically γ
Complementarity
An antibody can form complementarity to virtually anything because the potential amino acid diversity at the antigen binding site is vast
Complementarity - affinity
Affinity arises when sum of attractive forces exceeds sum of repulsive molecular forces
Somatic hypermutation
Occurs in germinal centres in lymph nodes
Start off with large repertoire
A few of those B cells will be involved in recognition of pathogen, which go to lymph nodes and undergo affinity maturation
2-3 months after immunisation boosting, you now have memory cells which have a higher affinity antibody against that specific pathogen –> strong protection
Complementarity Determining Regions (CDR)
Amino acid variation is found in 3 discrete regions called CDR (1, 2, 3)
3 loops that connect strands in 1st domains of H and L chains
3 loops from V(H) and V(L) juxtapose in folded protein to form a roughly rectangular surface of ~800-1000 Å2
Loops form within beta strands
Where amino acid diversity occurs
2 identical antigen binding sites
Recombination in Ig locus
Germ-line genes segmented into clusters: Variable (V), Diversity (D), Joining (J) and Constant (C) regions
Light chain locus has no D segments
D to J, then V to D
Intervening DNA is lost
Recombination in Ig locus - precision
Joining is very imprecise, so base pairs are changed during repair
Leads to huge variation at VDJ joining
Most important mechanism for generating diversity in B and T cell repertoires at birth
Recombination in Ig locus - which sections code for CDR 1, 2 and 3
CDR 1 and 2: V segment
CDR 3: VDJ region
For every B cell…
There is one antigen specificity
Clonal selection theory
When a baby, have naive repertoire of B cells
Encounter an antigen –> one of those B cells have a receptor which responds weakly to antigen –> undergoes mitosis
Migrates to a lymph node and produce progeny
Within those progeny, since that gene is undergoing somatic hypermutation, some of those B cells will have slightly higher affinity for antigen
Over time, end up with lots of B cells with higher affinity designed for that specific antigen than original clonal B cell
Results in higher affinity and reactive B cells that sit in lymphoid tissue for lifetime, waiting for next time you’re exposed to that antigen
When that happens, memory B cells rapidly produce plasma cells
Lymph node follicle
Where clonal selection takes place
Occurs more often in exposed areas
Contains germinal centres
Contains T cells
Lymph node follicles - T cells
Generate B cell progeny which develop higher and higher affinity
Eventually, some of those become high affinity memory cells that provide protection
How cells become T cells
Haemopoietic lymphoid precursors migrate from bone marrow to thymus and mature into T lymphocytes (part of cellular adaptive response)
T lymphocytes expression of co-factors
When they reach the thymus, they express co-factors CD4 and CD8 (referred to as double positive immature thymocytes)
This is the only time T cell expresses both receptors together
T cells - learning in thymus
Go through a process of education in thymus, learning what ‘self’ looks like (MHC molecules expressed in thymic tissue)
T lymphocytes - co-factors
CD4 helper T cell (~80% in blood)
CD8 cytotoxic T cell (~20% in blood)
T lymphocytes - CD4 sub-factors
Treg - regulates immune response
Th1 - drive cellular response (when pathogen demands a cellular immune response)
Th2 - predominate when antibodies or humoral/B cell response required. regulates B cell response
Th17 - control of inflammatory response
Thymus
Primary lymphoid organ Gland Part of lymphoid tissue Largest (most active) just before/after birth and shrinks with age Sits at top of pericardium above heart
T cells - survival in thymus
Only a small percentage of T cells survive thymus as mature T cells
Most die from neglect, either:
- didn’t recognise correct antigen to allow them to undergo mitosis - negative selection
- have recognised right ‘self’ antigen, but responded too strongly –> actively killed by apoptosis - positive selection
MHC
Major Histocompatibility Complex
Polymorphic genes (diff in almost everyone except identical twins)
Genetic locus that regulates histocompatibility
Control tissue transplantation
Detect intracellular pathogens i.e. viruses and kills the cell
Code for HLA on cell surface
T cells recognise
- infected cells to either kill them or provide help to other cells that reduce ability for virus to replicate
- antibodies (MHC) that actively present viral antigens to them
Tissue / bone marrow transplantation - MHC match
Tissue transplanted from donor to recipient is as closely matched in MHC as possible, otherwise immune system of recipient will recognise donor cells as foreign and kill them - tissue rejection (learned)
What is ‘nonself’
Anything that changes MHC
e.g. viruses, bacteria
HLA
Human Leukocyte Antigens
Human version of MHC
Expressed on most cells
Present peptide antigens to T cells
Highly polymorphic
6 diff molecules expressed on human cells
Genes that encode for HLA molecules are diff across individuals
CTL
Cytotoxic T cells React to own cells when there's a change in MHC class I molecules, i.e. when they express a neo-antigen picked up from inside the cell (viral or altered self-antigen) Have ability to generate perforins / pore-forming molecules that are inserted into target cells to kill them - effective for anti-tumour activity
Production of congenic mice
A skin transplanted to B
Backcross AxB with B
Repeated >20 times
AxB(20) is congenic with original B strain
Congenic mice experiment
Infect mice with LCMV (brain virus)
Isolate lymphocytes from spleens after 1 week
Mix lymphocytes from strain A or B with virus infected epithelial cells from either A or B
Result: cytotoxic T lymphocytes only killed LCMV infected cells from same strain
Viral immunity antigens
Self: antigen(s) encoded by MHC
Non-self: antigen(s) encoded by virus
T cell receptor and MHC on target cell
T cell receptor recognises MHC on target cell which was changed because a new antigen was inserted
T cell receptor molecule makes physical contact
T cell receptor
Membrane bound Ig-like molecule on T lymphocytes
H2
Antigens on mouse cells
Types of HLA
Class I: A, B, C
Class II: DR, DP, DQ
Serve diff roles
How many diff antigens expressed on surface of cells?
12
MHC class I HLA - A, B, C
Polymorphic region - alpha helices that sit along top of sheet are polymorphic - amino acid sequence vary between individuals
Peptide comes from virus
As MHC is produced on ribosome and moves into Golgi, it picks up peptides, folds, and is transported to cell surface where it remains for a length of time
β2M (microglobulin) - purpose
Holds molecule in right conformation
MHC class II HLA - DR, DP, DQ
2 chains - α and β
Polymorphic groove - allows peptide to bind to it
Peptide longer than in class I as it comes from a diff source (typically from antigens taken from extracellular pathogens)
MHC restriction =
MHC + peptide
CD4 and CD8 - differences
CD4: helper T cells recognise antigens in MHC class II CD8: cytotoxic T cells recognise antigens in MHC class I
CD4 and CD8 - similarities
Accessory molecules that physically associate with TcR
Have intracellular tyrosine kinases associated with their cytoplasmic tails that initiate T cell signalling through phosphorylation
Crucial to immune activation
CD4+ helper - function
HELP
Interacts with surface of macrophage
Huge production of cytokines and proliferation of T cells which form basis of T cell help
CD8+ cytotoxic (CTL) function
KILLING
Recognises surface of infected cell
Complex forms, T cell become activated
Killing by introduction of perforins and granzyme that punch holes in target cell membrane and destroy cell viability
Roles of MHC class I and II - summary
MHC class I: Peptide source - intracellular Pathogen - viruses Responding T cells - CD8 Effector function - cytotoxic Capture short amino acid peptides generated by viral replication inside cell and presents them to cytotoxic cells
MHC class II: Peptide source - extracellular Pathogen - bacteria Responding T cells - CD4 Effector function - help
MHC polymorphism is restricted to…
The protein domains that form the peptide groove
MHC polymorphism - co-dominance
An individual expresses both maternal and paternal genes 2 x 3 MHC class I and 2 x 3 MHC class II molecules Total of 12 polymorphic molecules expressed in cells
Haplotype polymorphisms
Different variations of MHC
Anchor amino acids
Where amino acid side chains point down into MHC molecules and anchor them
Why is MHC so polymorphic
Diff in diff countries - haplotypes evolved to provide defense against particular pathogens those communities are likely to face
Polymorphism designed to create a broad capacity to provide protection for species as a whole
MHC polymorphism - major consequences
Tissue transplantation is difficult except identical twins - requires careful matching and immunosuppressive drugs
MHC polymorphisms strongly linked to many autoimmune diseases
When is tissue typing more/less important
Not as important in heart and lung tissue transport
Important if transplanting cells involved in immune recognition and defense (i.e. bone marrow)
Heterologous bone marrow transplant
Relies on patients that are as closely matched as possible (unlike analogous bone marrow transplant)
Most susceptibilities to a disease are of…
A strong MHC component
Major types of type I allergies (examples)
Asthma (1/6 NZers)
Allergic rhinitis (seasonal hay fever)
Skin (dermatitis) eczema, urticaria (hives)
Insect allergies (house dust mite, bee stings)
Animal dander
Drugs (penicillin)
Large food proteins (gluten, peanut)
Nickel (metal induced contact dermatitis)
Anaphylaxis - may involve other organs
Classifications of hypersensitivity
Type I:
Atopic allergy
IgE mediated
Immediate
Type II:
Complement mediated
Medium
Type III: Serum sickness Less common Immune complexes Medium
Type IV:
Delayed type (DTH) - involves adaptive immune response
Slow response
Mast cells
Innate immune cells in the myeloid lineage that reside in skin
Provide protection against complex organisms that can’t be engulfed by phagocytosis
Type I (atopic allergy) - mast cells
Have Fc receptor on surface called Fc epsilon - receptor for IgE class of antibodies Pre-coated with IgE that has been primed against a particular allergen; usually occurs early on in birth
What does IgE regulate
Low conc, but regulates atopic allergies
Type I hypersensitivity - pollen
Recognised and attaches to IgE
Lots of IgE on surface of mast cell –> cross-linking since pollen grains are large
Mast cells activate and degranulate, releasing chemicals, e.g. histamine, leukotrienes, prostaglandin, free radicals and substance P which work together to destroy the pollen
Type I hypersensitivity can cause…
Smooth muscles to constrict
Blood vessels to constrict
Mucous glands to produce mucous/release fluid –> swelling
Platelets attracted to side –> platelet aggregation of clotting
Sensory nerve ending stimulation –> pain
Recruitment of other innate cells which also have granules released at sites of inflammation
Type I hypersensitivity - CD4
Th1 or Th2
What regulates type 1 hypersensitivity
Histamine
Type II hypersensitivity
Involves FcR, complement and neutrophils
Antibody present in new-born baby that reacts to a protein on the surface of their RBCs basement membrane –> induces response by attracting neutrophils and complement
Neutrophil tries to digest membrane of RBC, resulting in lysis of RBCs
Causes hemolytic anemia AKA rhesus
Rhesus anemia - parents must be…
Mother must be rhesus negative and father must be rhesus positive
Gene is dominant –> always expressed
How does rhesus anemia affect babies
RhD from 1st baby may pass through placenta to mother, causing the mother to have B cells producing anti-RhD in her blood
First born not as affected
Anti-RhD passes onto next (2nd) baby through placenta, so when second baby is born, it succumbs to hemolytic anemia because antibody induces type II hypersensitivity
Rhesus - treatment
Test mother and father
If mother negative and father positive, mother will receive an antibody which kills any RBCs that might have been transferred into her blood from the first baby –> prevents development of rhesus in following children
Treatment of allergy by desensitisation - success rate
Works in approx 50% of patients
Treatment of allergy by desensitisation - method
Skin scratch test to identify allergens
Increasing dose of allergen injected every week for 12-24 weeks
Drives B cells to produce IgG rather than IgE against the allergen
Goal is to have IgG bind to allergen before it binds to IgE
Note: IgG is more abundant than IgE
Monoclonal antibodies - where was it first tested/observed
Spleens of mice to see if genes were changing over time, particularly if there were somatic mutations occurring in CR1, 2, and 3 that led to the higher affinity antibodies
Define monoclonal
Single specificity and single binding affinity - can be highly specific and used for a range of purposes
What is typically used when making monoclonal antibodies
Bacteria
Used to only use mouse genes, but now can use human genes –> humanised –> not rejected by body
Making monoclonal antibodies - PEG
PEG fuses membranes of cells together; mix splenocytes and mouse myeloma line in correct ratio to get ~1:1 fusion between 1 B cell and 1 myeloma cell –> produces hybridoma
Monoclonal antibodies - hybridoma
Hybrid between B cell and myeloma cell
Often still produces antibody the B cell was producing
Making monoclonal antibodies - selection
Add a selective chemical that kills off myeloma cells that haven’t fused and put them on a plate
By 2 weeks, there are colonies of hybridomas that have grown out of a single cell and produce monoclonal antibodies
Take the hybridomas and put them in wells, then take supernatant (containing antibodies) and put on top of antigen that was coated to other microtiter plate
Add colour agent to show where there is an antibody
Monoclonal antibodies - advantages
Highly specific for intended target, so no ‘off-target’ effects
Can be tailor-made with the right affinity
Humanised so stay in blood stream for months
No adverse reactions or toxicity to antibody
Can be modified to be bi-specific for great potency
Monoclonal antibodies - disadvantages
Expensive to develop and make commercially
Side effects of functions can be serious
What is serum
Plasma without the clotting factors
Haematocrit
A measure of the percentage of whole blood occupied by erythrocytes
The intrinsic and extrinsic pathways of blood clotting are identical after formation of _____
Prothrombin
Major function of RBCs
Gas transport
Blood functions
Transport heat
Protect against infectious disease
Transport nutrients
Regulate blood pH
Which plasma protein plays a role in blood clotting?
Fibrinogen
Which blood cell releases granules that intensify the inflammatory response and promote hypersensitivity (allergic) reactions?
Basophils
An acute allergic response can lead to…
Anaphylactic shock
Inability of the immune system to protect the body from a pathogen causes…
Immunodeficiency
Function of C3 component of complement
Forms part of a convertase on the bacteria and is recognized by neutrophils through the receptor CR1
An inflammatory response that occurs immediately upon exposure to an antigen is likely to be mediated by…
IgE and mast cells
Coagulation - Many parasites and other microbes that rely on blood flow produce…
Powerful anti-coagulants that typically target the thrombin step
Virulence factors
A protein produced by many microbes that inhibits the complement cascade
Which cell is most associated with innate immunity
Myeloid
CD34 antigen marker defines…
A small pop of cells giving rise to all blood cells
Thrombin cleaves….
Fibrinogen to fibrin during coagulation
What is a possible cause of cherry red blood
Poisoning
What is unique to the adaptive immune system
Immunoglobulins
What are essential components of gene recombination in Ig and TcR loci
RAG and recognition sequences
Fc receptors activate phagocytosis upon….
Cross-linking
Germ-line gene rearrangement involves…
imprecise joining of V, D and J segments, which are the most important part
Viral immunity requires…
Dual recognition of self MHC and viral antigen (T cells)
Maritoux skin test for tuberculosis - type
Type IV - delayed hypersensitivity response
Affinity maturation
Takes place within germinal centre of lymph node follicles
Results from somatic hypermutation of rearranged Ig gene progressively producing higher affinity antibodies after B cell encounters the antigen
Enzyme Factor Xa
Cleaves prothrombin to active thrombin
Cb3 forms a principle component of..
Irreversibly bound convertase complex on bacterial surfaces
What is the likely first event to happen when bacteria enters the skin
Complement proteins react and opsonise the bacteria
TLR binds to…
A range of molecules unique to viruses and bacteria
FcR
Myeloid cell receptors that bind antibodies coating a microbe, NOT to bacterial surfaces
Which step leads to greatest amino acid diversity in CDR3 loop and TcR V domains
Random editing of base pairs prior to joining D to J and V to D segments
Fcε (epsilon) receptor triggers…
Mast cell degranulation when bound to IgE and allergen
type I allergy
Monoclonal antibodies have a single specificity towards … and are produced by…
1 epitope
Hybridoma
Anaphylatoxins
Potent activators of neutrophil phagocytosis
Chemoattractants that recruit neutrophils to site of infection
Small polypeptides cleaved from complement C3, C4 and C5
What components are common to both intrinsic and extrinsic pathway of coagulation
Factor X (10) and prothrombin
Opsonisation - basic definition
Irreversible coating of bacteria by complement proteins
Ig domain
A highly stable protein fold that allows amino acid variability in loops connecting its β-strands
AIDS
Caused by depleting CD4 T-lymphocytes, preventing essential cytokine help to the immune system
T-lymphocytes - class I and class II
CD4 T cells recognise antigen presented by MHC class II CD8 T cells recognise antigen presented by MHC class I
Antibodies can…
Agglutinate and precipitate antigen
Neutralise antigen
Enhance phagocytosis
Clonal selection of B-cells - antigens
Antigens are responsible for determining which cells eventually become cloned
