Inflammation, inflammasomes, PhD course

PhD Course 2023

Immunology:
Inflammation,
inflammasomes

Attila Gacser

Questions to Consider

•How can the immune system detect invasion by pathogens and alert the

immune system- immunological triage?

•How the inflammation is regulated, and how intracellular PRR’s are

involved in the inflammatory responses?

•Can inflammation cause cell death and how this process is involved in host

immune response?

•What are the role of inflammasomes?

Thought Experiment

•Which limb of the immune system would you rather give up?

•Innate

•Adaptive

Poll

Which arm of the immune system is more important?

• Innate

• Adaptive

Far More Rapid Progression of Bacterial

Infection in Mice Lacking the Innate Immune
Response Than in Mice Lacking the Adaptive

Immune Response

The Innate Immune System Uses Multiple

Fixed Barriers to Prevent Infection

The Innate Immune Response is the

Layered Front Line of the Host Defense

System Using Pattern Recognition

Definition of Inflammation

• A normally beneficial host response to foreign challenge or

tissue injury that leads ultimately to the restoration of tissue
structure and function.

• Normally is self-limiting while prolonged inflammation can

lead to disease

Initiation of an Inflammatory Response

• Causes: Response to bacterial replication (PAMP’s)

• Response to tissue damage (DAMP’s)

• Response to macrophage mediator release

Poll

What are the goals of the Inflammatory Response?

• Prevent the initial establishment of infection

• Prevent the spread of infection from the invasion site.

• Recruit effector cells for assistance.

• Alert and mobilize B cells and T cells.

Goals of the Inflammatory

Response

• Prevent the initial establishment of

infection

• Prevent spread of infection from

invasion site

• Recruit effector cells for assistance

• Alert and mobilize B cells and T cells

Infection is Initiated By Bacterial
Adherence and Penetration of the

Epithelial Barrier

Persistent Local Infection of Tissues

Induces Adaptive Immunity

The innate immune system uses numerous “sensors” to

detect problems

One of the best understood is the Toll-like receptor (TLR)

system

TLRs are expressed on the cell surface or in endosomes &

monitor the environment for pathogens

Toll- An Important Immune Molecule In Flies

• Toll, discovered in 1996, is a

drosophila developmental gene

• When this gene was knocked out, the

fruit flies succumbed to massive

fungal infection

• This indicated Toll encoded an

important immune molecule

Bruce Beutler: TNFα

Jules Hoffman: TLR’s

Ralph Steinman: dendritikus sejtek

Charles A. Janeway (1943-2003)

Toll-like receptors (TLR)Structure-Function

Toll-like Receptors Recognize Unique Pathogen-

associated Molecular Patterns (PAMP)

Gram positive bacteria

ssRNA (HIV)

Gram
negative
bacteria

Lipopolysaccharide Binds to CD14 Which Interacts

With TLR-4 Inducing Translocation of NFkB

MyD88 TRIF Mal MyD88

TRIF

MyD88 MyD88

MD2

TLR2/1 or 2/6 TLR3 TLR4 TLR5 TLR7/8/9

Mal

NFkB

NFkB IRF3

NFkB IRF3

NFkB

TNFa
IL6
IL8

TNFa IFNb
IL6 RANTES
IL8

TNFa IFNb
IL6

RANTES

IL8

TNFa
IL6
IL8

IRAKs

TRAF6

RIP1

TBK1

TBK1

IRAKs

TRAF6

IRAKs

TRAF6

IRAKs

TRAF6

NFkB

TNFa
IL6
IL8

IRF5/7

IFNa

TRAM

Cytosolic sensors

One subset = NOD-like receptors (NLR)

23 genes in humans

34 genes in mice

All have Nucleotide Oligomerization

Domains (NOD)

All have LRR domains (in this aspect like

intracellular TLRs)

Pathogen recognition receptors and inflammatory signaling pathways

• Toll-like receptors (TLRs) senses pathogen associated molecular patterns at cells surface and
endosomal compartment.

• Nod-like receptors (NLRs) are cytosolic sensors for pathogen and danger associated molecular
patterns.

http://www.invivogen.com/

NOD-like receptors

LRR

C-terminal

N-terminal

effector

NOD (NACHT)

NOD: nucleotide-binding oligomerization domain
CARD: caspase activation and recruitment domain
PYD: pyrin domain

Casp1

Casp1

Casp1

Anatomy of an Inflammasome

NLRC4

AIM2

NLRP3

Inflammasomes

When NLR portion recognizes PAMP (or a DAMP in the case of NLRP3) the
inflammasome oligomerizes with an adapator protein and capsapse 1. Once
Caspase-1 become apart of the inflammasome it is activated and capable of
cleaving cytokines and inducing cell death.

Nod-Like Receptor (NLR)
•NLRP3

•NLRC4

•AIM2

Caspase 1
Enzyme capable of
cleaving pro-forms of
cytokines (IL-1b, IL-18)

Often

times

we

use

the

name

NLRP3

interchangeably

with

inflammasome

by

accident.

However, the NLRs are actually

components of the inflammasome.

“Adaptor” protein

Inflammasome Components
1. NLR (NLRP3, NLRC4, AIM2)
2. Adaptor protein
3. Caspase 1

Steps that lead to upregulation of NLRs

+

PM

MyD88

MyD88

MyD88

TLR5

TLR4

TLR2

Nucleus
NLRC4

AIM2

NLRP3

1. TLRs detect pathogen

2. Upregulate gene expression of NLRs
(NLRP3, NLRC4, and Aim2)

3. Increased NLR proteins
and Pro-IL1b in cytoplasm

NLRs

pro-IL-1β

pro-IL1β

Transcription

Note: Pro-IL-18 is
NOT upregulated in
this manner. It is
constitutively
(always) expressed.

TLR signaling cascade leads to gene
upregulation

PM

NLRC4

AIM2

NLRP3

4. Presence of NLRs in cytoplasm awaiting either PAMP or DAMP stimulus

Detect and activated
by PAMP stimuli ONLY

Detects and is activated by BOTH PAMP and

DAMP stimuli

PAMP stimuli

Damage Associated Molecular Patterns (DAMPs) –
Non-PAMP Stimuli

PM

b. Pore formation

PM

a. Pore-forming toxins
c. Crystals
•Asbestos

•Silica

+

Remember, broadly there are two types of stimuli that can activate the

inflammasomes.

Detection, Inflammasome assembly, and activation of Caspase 1

PM

NLRC4

AIM2

NLRP3

5. NLRS detect activating Stimuli (PAMPs and/or DAMPs)

Flagellin

DNA

PAMP/DAMPs

Representative of a DAMP

Other Inflammasome Components

Float around the cytoplasm until recruited by

NLRs during inflammasome assembly.

Casp1

Casp1

Casp1

Casp1

6. Inflammasome assembly and activation
“activation” means all of the components are
assembled and Caspase 1 is active and able to
cleave other proteins.

Detection, Inflammasome assembly, and activation of Caspase 1

PM

NLRC4

AIM2

NLRP3

Flagellin

DNA

PAMP/DAMPs

Representative of a DAMP

Casp1

Casp1

PM

NLRC4

AIM2

NLRP3

7. Activated Caspase 1 can now cleave Pro-IL1b and IL-18 into active forms.

Flagellin

DNA

PAMP/DAMPs

Representative of
a DAMP

Casp1

pro-IL1β

pro-IL18

IL1β

IL18

Proteolysis

PYROPTOSIS

Active forms
•These cytokines send
inflammatory signals to
other cells

•Mediate inflammation and
fever

Detection, Inflammasome assembly, and activation of Caspase 1

Inflammatory cell death
•Elimination of
pathogen niche

•Antigen exposure

•Cytokine release

gasdermin clevage

Summary of major inflammasome pathways

NLRP1b

inflammasome

NLRP3

inflammasome

NLRC4

inflammasome

AIM2

inflammasome

Pro IL-1β
Pro IL-18

Pro IL-1β
Pro IL-18

Active caspase-1

Molecular Cell 25, 713–724, March 9, 2007

animation

http://www.adipogen.com/

Inflammasome activators

pro-IL-1β
IL-1β

gout

atherosclerosis

arthritis

diabetes

ekzema

inflammatory
bowel disease

Inflammasomes: multiprotein complexes that trigger the maturation of IL-1β and IL-18 in

myeloid cells

high IL-1β level

Inflammasome activation

Heritable disorders associated with inflammasome activation

Disease

Clinical features
Gene

mutated

Anakinra
response

Familial cold
autoinflammatory
syndrome (FCAS)

Fever, arthralgia,
cold-induced urticaria
NALP3

yes

Muckle-Wells
syndrome
(MWS)

Fever, arthralgia,
urticaria,
sensorineural
deafness,
amyloidosis

NALP3

yes

Chronic infantile
neurological
cutaneous and
articular syndrome
(CINCA, NOMID)

Fever, severe
arthralgia,
urticaria, neurological
problems, severe
amyloidosis

NALP3

yes

Familial
Mediterranean
fever (FMF)

Fever, peritonitis,
pleuritis,
amyloidosis

Pyrin

partial

Adapted from: The inflammasomes: guardians of the body. Martinon et al., Annu Rev Immunol, 2009.