Lecture 7: Inflammation and Mediators

What is the major hallmark of the innate immune response

Inflammatory response.

Does the inflammatory response vary depending on the type of injury

No. It is identical

Is inflammation a normal part of the immune response

Yes. It is designed to restore immune homeostasis by brining injured tissue back to normal

2 types of inflammatory response

Acute - short term response to an insult to the body

Chronic - ongoing inflammation

What can chronic inflammation be caused by

Foreign bodies, persistent infection, autoimmune disease

Can chronic inflammation be cured

No but it can be treated with anti-inflammatory agents such as aspirin and steroids

Steps of Acute Inflammation

The trigger

The Endothelial Gate

The Neutrophil Avalanche

The trigger

Sensing of PAMPs/DAMPs by tissue-resident macrophages and mast cells via TLRs. Does not have to be initiated by pathogens that cause infection. Injuries release damaged cellular contents at local sites. This triggers mobilization of phagocytic cells and innate immune cells to damaged area.

Purpose of innate immune cells moving towards damaged tissue

Clear cellular debris

Set the stage for wound repair

Mobilization is in part due to trans endothelial migration of leukocytes

The endothelial gate

How immediate early mediators (histamine, bradykinin) alter vascular permeability and upregulate endothelial P-selectin and E-selectin

The neutrophil Avalanche

influx of neutrophils after endothelial gate

Trigger mediators

Histamines, Leukotrienes, and prostaglandins

Histamines

Increased blood flow and capillary permeability, allowing plasma leakage to for interstitial fluid leading to swelling.

When are DAMPs

Immediately following tissue injury. Released by stressed cells undergoing necrosis. They act as endogenous danger signals.

What do DAMPs promote

Positive feedback, they exacerbate the inflammatory response. They release H2O2 from wounded endothelial cells. Trigger the release of chemokines (IL-8) and cytokines like IL-1, TNFa, and leukotrienes. Result in the recruitment of additional neutrophils from circulation.

Endothelial Gate

The process of early arriving neutrophils being activated. They promote further positive feedback. Locally produced chemokines and cytokines induce increased expression of endothelial cell adhesion molecules (ECAMs) and their ligands.

Result of endothelial gate

Vascular permeability

Leukocyte endothelial adherence

Rolling

Trans endothelial migration or extravasation to local tissue

General Steps of Leukocyte Extravasation

Tethering

Rolling

Activation

Adhesion

Crawling

Transmigration

Tethering

Is the initial capture of fast moving leukocytes from blood flow.

Occurs when endothelial P-selectin and E-selectin bind to Leukocytes PSGL-1.

Transient selectin-ligand interactions occur, slowing the circulating leukocytes.

Week bonds form and rapidly break under shear stress.

Defect in PSGL1

Contributes to impaired leukocyte trafficking

Rolling

Slow leukocyte movement along the vessel wall

Selectins work to slow leukocytes by binding to Sialyl-LewisX carbohydrates on leukocytes

Repeated formation and disruption of selectin-mediated bonds causes rolling

Allows the leukocytes to sample inflammatory signals on the endothelium

Activation

Chemokines displayed on endothelial surface (CXCL8/IL-8, CCL2) signal the leukocyte to stop by binding to leukocyte’s GPCRs.

Intracellular signaling trigger inside out integrin activation where leukocyte integrins converge from a low to high affinity state

Firm Adhesion

High integrity integrins bind to the endothelial adhesion molecules

The leukocyte becomes firmly attached despite blood flow

Defects in adhesion

Patients develop recurring bacterial infections and poor wound healing

Leukocyte Receptors involved in Adhesion

LFA-1

MAC-1

VLA-4

Endothelial ligands involved in Adhesion

ICAM-1

VCAM-1

Crawling (diapedesis)

Leukocytes migrate along the endothelial surface using the integrins.

Searching for an optimal site to exit the vessel

Cells locate endothelial junctions suitable for tissue entry

Crawling improves efficiency and minimizes endothelial damage

Transmigration

Leukocytes squeeze between endothelial cells (paracellular) or go through cells (transcellular). To exit the bloodstream and enter the tissue.

Uses adhesion molecules (PECAM-1, JAM, CD99, MMPs) at endothelial junction to help guide leukocyte through

To cross the basement membrane leukocytes follow chemokine gradients.

Excessive transmigration

Contributes to chronic inflammation

What are the big 3 of cytokines

TNF-a - the alarm cytokine

IL-1B - the fever cytokine

IL-6 - notify the liver

These cells coordinate the acute inflammatory response

Major sources of the big 3

Macrophages (primary source)

Dendritic Cells

Monocytes

Endothelial Cells

Fibroblasts (during chronic inflammation)

TNF alpha local effects

Activate endothelium

Increase expression of E-selectin, ICAM-1, and VCAM-1

Promotes leukocyte recruitment

Increases vascular permeability

TNF alpha Systemic Effects

Fever

Acute-phase protein production

Cachexia during chronic disease

Septic shock when produced excessively

Local effects of IL-1B

Endothelial activation

Enhanced leukocyte recruitment

Increased adhesion molecule expression

Systemic effects of IL-1B

Fever through hypothalamic prostaglandin production

Acute-phase response

Bone and cartilage destruction in chronic inflammatory disease

Local effects of IL-6

Supports leukocyte activation

Promotes B-cell differentiation

Systemic effects of IL-6

Stimulates hepatic production of CRP, Fibrinogen, serum amyloid A

Increases ESR indirectly through fibrinogen

Contributes to fever

Is the driver of acute phase protein production

How do the big three work together at the site of infection

After pathogen recognition and macrophage activation, the big three are released. This leads to endothelial activation, vasodilation, adhesion molecule expression, recruitment of neutrophils, and monocytes.

NFkB Key function

Detects danger signals

Translates extracellular signals into gene expression changes

Coordinates inflammatory responses

Promotes host defense against pathogens

Links innate and adaptive immunity

Major outcomes of NFkB activation

Cytokine production

Chemokine production

Leukocyte recruitment

Cell Survival

Antimicrobial Response

What is NFkB

A transcription factor that serves as a central regulator of innate and adaptive immune responses

It transforms a resting cell into an inflammatory effector cell

Is one of the major reasons why inflamed blood vessels become “sticky” to leukocytes

Upstream activators of NFkB

PRRs (TLRs)

Cytokines (TNFa/IL-1B)

DAMPs

Antigen receptor signaling (BCRS/TCRS)

All lead to inflammatory gene expression

How is NFkB normally kept

Inactive as a result of being bound to IkBs (inhibitory protein)

It can not enter the nucleus

Purpose of IkB

Inhibits NFkB

Prevents unnecessary inflammation

Stops inflammatory gene transcription

NFkB activation pathway

Inflammatory stimulus binds receptor

Activation of intracellular signaling complexes

Activation of IKK (IkB kinase)

IkB phosphorylation

IkB is ubiquinated and degrade

NFkB is released

NFkB enters nucleus

Inflammatory genes are transcribed

Genes controlled by NFkB

Cytokines: TNFa, IL-1b, IL-6, IL-12

Chemokines: CXCL8 (IL-8) CCL2, CCL5

Adhesion molecules: ICAM-1, VCAM-1, E-selectin

COX, acute phase response genes

It promotes the expression of all of these genes leading to these molecules productions

Activated endothelial cells

Increase their expression of E-selectin, P-selectin, ICAM-1, VCAM-1, and CXCL8 (IL-8)

(selectins, adhesion molecules, and chemokines)

How does NFkB serve as a bridge between innate and adaptive immunity

Helps initiate the innate immune response that produces signals which in turn activate the adaptive immune response. Ex: activates macrophages that release cytokines, activate DCs that present to B cells, leads to T cell activation

Rheumatoid Arthritis

Persistent production of TNFa, IL-1B, IL-6 leading to joint inflammation and destruction

IBD

Excess cytokines production in the GI tract leads to chronic intestinal inflammation

Psoriasis

Keratinocyte and immune cell NFkB activation leads to epidermal hyperproliferation

Cancer

Chronic NFkB signaling may promote cancer cell survival

How do croticosteroids impact NFkB

They increase IkB production while suppressing NKkB transcriptional activity

Reduce cytokine production —→ reduce inflammation

TNF inhibitors

Block upstream TNF signaling that reduces NFkB activation

JAK inhibitors

Reduce Inflammatory cytokine signaling leading to indirect reduction in NFkB driven inflammation

Pain, redness, and heat

Is explained by elevated cellular metabolism that increases vasodilation allowing increased blood flow. Increased expression of adhesion molecules on endothelial cells promotes leukocyte binding.

Swelling

Separation of previously tightly joined endothelial cells leads to extravasation of fluids from blood vessels into surrounding tissue.

Cells of inflammation

Most cells are PMNs

30-60 minutes response

PMNs accumulate

Phagocytize intruder or damaged tissues

Release lysosomal enzymes to attempt destruction

4-5 hour response

If cause persists, macrophage and lymphocytes invade. Macrophages process antigens and start adaptive response (TCRS —→ cytokines —→ BCRS—→ antibodies)

5-7 day response

Antibodies produced are detected as serum antibodies and humoral immune defense occurs

What are kinins, what are they responsible for, and how are they degraded

Are potent nerve stimulators. they act directly on local smooth muscle and cause muscle contractions. They are responsible for pain and itching. They are rapidly inactivated after proteolytic activation.

Coagulation pathway

Activated following kinin induced damage to blood vessels. Leads to plasma enzyme activation that stars cascade. Results in a physical clot or thrombus that prevents microorganisms from entering the blood stream .

Acute phase response

Occurs within minutes as a result of innate immune response to PAMPs and releases the Big Three.

Acute phase proteins

Function as soluble PRRs. Increase WBC production. Increase synthesis of hydrocortisone and ACTH.

CRP

Can bind to microorganism membranes and activate the complement cascade leading to cell lysis and enhanced phagocytosis. Commonly measured along with ESR.

ESR mechanism

During inflammation inflammatory cytokines stimulate the liver to produce acute phase proteins. These proteins reduce RBC repulsion, promote their aggregation (rouleaux formations), increase RBC mass and settling rate. ESR increases as a result. ESR increase can also be seen in anemia and pregnancy

Is CRP and ESR specific

No. They just detect or monitor significant inflammation in patients with suspected infections. CRP tells you what is happening now, ESR tells you what has been happenings.

Fever

One of the most common manifestation. Caused by endotoxins from bacteria. Monocytes and macrophages come into contact with the endotoxins and release endogenous pyrogens like IL-1 and interferons.

IL-1 and fever

IL-1 increases leading to hypothalamic induction to raise body temperature

Progression to chronic Inflammation

The cellular shift

T cell coordination

Tissue Consequences

The cellular shift

The transition from neutrophil dominated infiltrate to a mononuclear infiltrate (T cells, macrophages, plasma cells)

T-cell coordination

How chronic cytokine environments (ILs 6,12,23 +TNFa+ IL1B) polarize naive t-cells into pathogenic Th17 or Th1 cells turning innate response into a chronic adaptive response cycle.

tissue consequences

granuloma formation, continuous macrophage activation, and fibroblast driven fibrosis

Acute Inflammation

Dominated by neutrophils

Hours to days

Rapid recruitment of chemokines

Primary functions: phagocytosis, degranulation, ROS protection, pathogen control

Chronic inflammation

Days to months

Dominated by mononuclear cells (macrophages, T-cells, plasma cells)

Macrophages

Central effector cells of chornic inflammation

Phagocytosis debris and dead neutrophils

Produce the big three

orchestrate tissue repair and remodeling

T-lymphocytes

Recognize specific antigens

Produce cytokines that regulate immune response

Activate macrophages and coordinate adaptive immunity

Plasma cells

Terminally differentiated B cells

Secrete antigen specific antibodies

Support long term memory

Granuloma Formation

Organized collections of activated macrophages that wall off persistent pathogens or foreign material.

Fibrosis

Chronic cytokine production activates fibroblasts. TGF-B stimulates collagen and Extracellular matric deposition that can impair normal organ function.