Lecture 2 - Cells of the immune system: review and updates

What is the innate immune response?

The immune response which is present from birth

• Non-specific response

What is the acquired immune response?

The immune response which is acquired through exposure to antigens

• Specific response

Which three inflammatory cytokines form part of the acute phase inflammatory response?

• IL-1β

• IL-6

• TNF-α

What is the first step of inflammation?

Tissue damage causes release of vasoactive and chemotactic factors

These trigger local increase in blood flow and capillary permeability 

What is the second step of inflammation?

(After release of vasoactive and chemotactic factors)

Permeable capillaries allow an influx of fluid and cells.

What is the third step of inflammation?

(After influx of fluid)

Phagocytes migrate to site of inflammation (chemotaxis)

What is the fourth step of inflammation?

(After phagocyte chemotaxis)

Phagocytes and antibacterial exudate destroy bacteria

What is the final step of inflammation?

(After bacteria destruction by phagocytes)

Resolution of inflammation 

What effect do acute phase cytokines have on the liver?

Acute phase proteins

• Activation of compliment opsonization

What effect do acute phase cytokines have on the bone marrow endothelium?

Neutrophil mobilization from bone marrow

• Phagocytosis

What effect do acute phase cytokines have on the hypothalamus?

Increased body temperature

• Decreased viral and bacterial replication

• Increased antigen processing

• Increased specific immune response

What effect do acute phase cytokines have on fat/muscle?

Protein and energy mobilization to allow increased body temperature

• Decreased viral and bacterial replication

• Increased antigen processing

• Increased specific immune response

What effect do acute phase cytokines have on dendritic cells?

TNF-α stimulates DC migration to lymph nodes and maturation

• Initiates adaptive immune response

What does CD mean?

(Example: CD3, CD4, etc)

Cluster of designation 

What does CyTOF mean?

Cytometry by time-of-flight

What does CyTOF use instead of fluorescence to label antibodies?

Different metal isotopes/elements 

• As this gives each antibody a different weight 

What is an advantage of CyTOF over standard flow cytometry?

Allows many antibodies to be used without the need for compensation (Up to 36 differently labelled antibodies)

What is PCA and what does it allow for?

Principle Component Analysis (PCA)

Allows for clusters of cells with similar antibody combinations to be plotted in 2D

• As up to 36 different antibodies can be used in CyTOF, plotting in 2D is easier to visualise than 36D

What is the software, ViSNE used for in mass cytometry?

Used to identify and visualise clusters so specific cells antibody staining can be seen.

• Similar to PCA

How do neutrophils destroy pathogens?

Ingest pathogens and place into phagosomes

• Oxygen species, cathepsins and lysozymes then destroy them

Also release extracellular proteins (DNA complexes) that trap/destory

What are neutrophil NETs?

Neutrophil Extracellular Traps (NETs)

• Large, web-like structures which are assembled on decondensed chromatin 

• Trap, neutralise and kill bacteria

What do mast cells release?

Toxic contents such as histamines

What is the role of eosinophils?

Release contents and damage large extracellular parasites 

• E.g. worms

What do NK cells express to allow them to kill infected cells during innate immune response?

Express members of a large family of innate antigen receptors 

How does altered MHC class I allow NK to directly kill a cell?

MHC class I is usually recognised by inhibitory receptors which provide a negative signal

Missing or altered MHC class I cannot provide the negative signal so NK cell is triggered by activating receptors

How does the TRAIL receptor allow NK cells to directly kill cells?

NK cells express the TNF family ligand TRAIL on cell surface

• Binds and activates DR4 and DR5 on cell target

• DR4/5 signal activates caspase 8, inducing apoptosis

How does Antibody Dependent Cell Cytotoxicity allow NK cells to directly kill a cell?

Antibody binds antigen on surface of target cell

• Fc receptors on NK cell then bind these antibodies

• Crosslinking of Fc receptors and antibodies signals the NK cell to kill the target cell

• Cell dies by apoptosis

How can NK cells indirectly kill pathogens via Macrophages?

Macrophages with phagocytosed pathogens release IL-12

• IL-12 binds to receptor on NK cells causing secretion of IFN-γ

• IFN-γ binds to receptors on Macrophages, improving killing

NK cells require which cytokine for development?

IL-15

Innate lymphoid cells (ILCs) originate from the same common lymphoid progenitor as T/B cells.

What supresses T/B development?

Id2 supresses development into T/B cells

• DNA-binding protein inhibitor

Innate lymphoid cells (ILCs) express lymphoid markers. What aspect of T/B cells do they NOT express?

ILCs do NOT express RAG-mediated recombined antigen receptors

True or False.

Like T-lymphocytes, Innate Lymphoid Cells (ILCs) also express TCRs.

False.

ILCs don’t express a TCR.

What are the 5 subtypes of Innate Lymphoid Cells (ILCs)?

• Natural Killer (NK)

• ILC1

• ILC2

• ILC3

• LTi

ILC1 express which transcription factor?

T-bet

ILC1 have similar cytokine profiles to which cells?

CD4 Th1 cells and NK cells

ILC1 cells are induced to produce IFN-γ by which cytokines?

IL-12 and IL-18

ILC2 express which transcription factor?

Gata3

ILC2 produce which cytokines?

IL-4, IL-5 and IL-13 (Th2 associated cytokines)

• Also produce IL-9

ILC2 are induced by which cytokines?

IL-25 and IL-33

ILC2 are crucial for the formation of secondary lymph nodes and Peyer’s patches.

True or False?

False.

LTi are crucial for formation of secondary lymph nodes

ILC2 are involved in expulsion of large extracellular parasites and mucus/collagen production

ILC3 express which transcription factor?

RORγt

ILC3 produce which cytokines?

IL-17 and/or IL-22

Lymphoid tissue inducer (LTi) cells express which transcription factor?

RORγt (just like ILC3)

Why are dendritic cells (DC) short-lived?

Short-lived to prevent prolonging of immune responses.

• Therefore not identified until much later than other immune cells

What is the role of Dendritic cells?

To take up, process and present antigens to T-cells

• Migrate from peripheral sites to lymph nodes to initiate an immune response 

  • Via activation of naïve T-cells

Which immune cell expresses CD11c, MHC class II and co-stimulatory molecules CD80/86 (B7.1/2)

Dendritic cells

What are Lin- (Lineage negative) cells?

Cells which do not express certain typical immune cell lineage markers

How many subsets of DCs have been identified in humans?

6 subsets have been identified 

Which type of DC is the most predominant?

DC2

A substantial number of lymphocytes return to where during circulation? 

The bone marrow

What are the two types of B-cells?

B1 B-cells and B2 B-cells

Which type of B-cells are dominant in the peritoneal and pleural cavities?

B1 B-cells

Which type of B-cells develop from the foetal liver

B1 B-cells

B2 B-cells secrete BCR rapidly as a natural (innate) antibody in the absence of T-cells

True or False?

False.

B1 B-cells are the ones to secrete their BCR as an innate-like antibody 

Which type of B-cell’s BCR has reduced diversity and is reactive against repetitive structures associated with pathogen antigens?

B1 B-cells

Which B-cell has high BCR diversity which is capable of recognising small individual epitopes on antigens?

B2 B-cells

Following antigen recognition, B2 B-cell BCRs internalise the antigen for processing.

What are the two ways the antigen is then presented?

MHC Class II (to CD4+ T-cells)

CD1a (to iNK T cells)

Following presentation of antigens to a T-cell, what happens to the B2 B-cell?

Undergoes somatic hypermutation and isotype switching of their BCR within germinal centres 

• Differentiation of B2 B-cells into plasma and memory cells

How does secretion of BCRs by B-cells help combat pathogens?

Neutralise toxins/pathogens

Render toxins insoluble

• Toxins are then cleared by Fc receptors and complement receptors

What are T-cells derived from?

Bone-marrow hematopoietic stem cells

• Via Common Lymphoid Progenitor (CLP)

What two combinations of TCR genes form the T-cell surface TCR?

TCR α/β or γ/δ genes

After leaving the bone marrow, where do T-cells migrate to?

Migrate from the bone marrow to the thymus

Once T-cells arrive in the thymus, what takes place?

Processed fragments of antigens complexed with MHC molecules bind to TCRs

This leads to signals that determines a T-cells fate 

• Positive and negative selection

What three fates do T-cells in the thymus have?

• NK T-cells (different to NK cells)

• γ/δ T-cells

• α/β T-cells

NK T-cells develop from which precursors?

The same CD4⁺/CD8⁺ double positive precursors

• Same as conventional MHC-restricted TCRα/β T-cells

What transcription factor do NK T-cells express?

The transcription factor PLZF

NK T-cells express TCR α/β which recognises what?

CD1d 

• Non-polymorphic MHC class I-like molecule

• Presents glycolipids

Also express markers seen on NK cells

What are invariant NK T-cells?

Expressed a very limited repertoire of TCRα and a restricted pool of TCRβ chains

What are diverse NK T-cells?

Express a more diverse TCRα/β

γ/δ T-cells develop from which precursors?

The same CD4^-/CD8^- and Double-Negative 3 precursors as conventional MHC-restricted TCRα/β T-cells

• BUT also express re-arranged γ/δ TCR genes

What transcription factor do γ/δ T-cells express?

SOX13

What is unique about γ/δ T-cell TCR diversity?

γ/δ T-cell TCR diversity is limited.

• Many tissues have almost monoclonal γ/δ T-cell populations

The 3D structure of γ/δ TCR is more akin to BCR than α/β TCR. This allows for what?

Allows for antigen recognition to not be limited to peptide-MHC complexes

Some γ/δ TCRs can recognise MHC-I-like molecules such as….

• CD1d

• MICA, RAE1 and ULBP

  • Expressed on ‘stressed’ cells

Which is faster γ/δ T-cell response or α/β T-cell response?

Due to location, number and antigen specificity, the kinetics of γ/δ T-cell responses are faster

• This allows the γ/δ T-cell response to be similar to an innate response

What is the differentiation from DP cells to SP T-cells also called?

Positive selection

What is a DP T-cell?

Double positive T-cell

• Is both CD4+ and CD8+

After leaving the thymus, what causes naive T-cells to further differentiate?

Antigen/MHC encounters on APCs

• This causes differentiation of naive CD4+ or CD8+ T-cells to effector/memory cells

What are some specialised CD4+ effector T-cells which ensure a pathogen specific immune response?

• Th1, Th2, Th17

• iTreg

• Tfh, Tfr

What does nTreg and iTreg mean?

Natural regulatory T-cells and Induced regulatory T-cells

What is the function of nTreg, iTreg and Tfr cells?

Ensure immune responses are controlled.

• These are regulatory cells