Innate Immune system. Overview of the immune system Immune system Innate (nonspecific) 1 st line of defense Cellular components Humoral components Adaptive.

Innate Immune system

Overview of the immune system

Cells of the immune system

Cellular componentsCell Mechanism

Neutrophils Phagocytosis and intracellular killingInflammation and tissue damage

Macrophages Phagocytosis and intracellular killingExtracellular killing of infected or altered self targetsTissue repairAntigen presentation for specific immune response

NK and LAK cells Killing of virus-infected and altered self targets

Eosinophils Killing of certain parasites

Innate Immunity Features

• Preformed: Rapid-Available on Short Notice

• No Memory:Not Enhanced by Prior Exposure:

• Broad Specificity• Dependent on species, strain, sex.

Innate vs. Adaptive Immunity• Innate Immunity• Pathogen recognized by

receptors encoded in the germline: pattern recognition receptors

• Receptors have broad specificity, i.e., recognize many related molecular structures called PAMPs (pathogen-associated molecular patterns)

• Immediate response

• No memory of prior exposure

• Adaptive Immunity• Pathogen recognized by receptors

generated randomly: B-cell (BCR) and T-cell (TCR) receptors for antigen

• Receptors have very narrow specificity; i.e., recognize a particular epitope after processing

• Slow (3 -5 days) response (because of the need for clones of responding cells to develop)

• Memory of prior exposure

Component of Innate Immunity

Innate Immune system

First line Second line1) Mechanical barriers A- cells 2) Chemical & biochemical inhibitors 1- Natural killer3) Normal flora 2- Phagocytes

B- Soluble factors C- Inflammatory barriers

First line1) Mechanical barriers - Intact skin - Mucous coat - Mucous secretion - Blinking reflex and tears - The hair at the nares - Coughing and sneezing reflex

First line2) Chemical & biochemical inhibitors - Sweet and sebaceous secretion - Hydrolytic enzymes in saliva - HCl of the stomach - Proteolytic enzyme in small intestine - Lysozyme in tears - Acidic pH in the adult vagina

Second lineA) cells1- Natural killer (NK)Definition: Large granular lymphocytes Innate cytotoxic lymphocytes

Source : Bon marrow precursors

Location : 10% or 15% of lymphocytes in peripheral blood 1% or 2% of lymphocytes in spleen

Tumor cells

Function : Cytotoxic for Viral infected cells Bacterial, fungal, parasitic infection

Responsible for antibody–dependent cell mediated cytotoxicity (ADCC)

Second line2- Phagocytes Specialized cells for capture, Ingestion and destruction of invading

microorganisms

* Polymorphoniclear leucocytes, mainly neutrophils: granulocytes circulate in blood

* Mononuclear cells (macrophages) - Monocytes in blood

- Histocytes in connective tissues

- Fixed reticuloendothelial cells in liver spleen, lymph nods, bon marrow

Second lineB- Soluble factors

1- Acute phase protein (Plasma protein, CRP=C reactive protein, Fibrin.)

2- Complement (proteins in serum, body fluids)3- Interferons (Proteins against viral infections)4- Properdin (Complement activation)5- Beta lysine (Antibacterial protein from Platelets)6- Lactoferrrin,Transferrin (Iron binding protein)7- Lactoperoxidase (Saliva & Milk)8- Lysozyme (Hydrolyze cell wall)9- Antibiotic peptides , 29-34a.a. and enriched with cys.

(Difensins (Neu, IL-1 and TNF-a is inducer), Cryptocidins (gut))

Complement system

Not Ab mediated

Com

plem

ent 1. Inactive

complement proteins are in constant circulation.

2. Complement proteins are activated by various mechanisms.

3. These are the consequences...

Phagocytosis

The engulfment, digestion, and subsequent processing of microorganisms by macrophages and neutrophils

1) Chemotaxis & attachment: a- Attraction by chemotactic substances (microbes, damaged tissues) b- Attachment by receptors on surfaces of phagocytes

Phagocytosis

2) Ingestion:

* Phagocyte pseudopodia surround organism forming phagosom * Opsinins and co-factors enhance phagocytosis * Fusion with phagocyte granules and release digestive, toxic contents

Phagocytosis

3- Killing (two microbicidal routes) a- Oxygen depended system (powerful microbicidal agents) Oxygen converted to superoxide, anion, hydrogen peroxide, activated oxygen and hydroxyl radicals. b- Oxygen-independent system (anaerobic conditions) Digestion and killing by lysozyme. Lactoferrin, low pH, cationic proteins and hydrolytic and proteolytic enzymes

C) Inflammatory Barriers* Tissue damage by a wound or by invading pathogen

* Inflammatory response: Tissue damageRelease of chemical mediators from Leukocytes (Histamine, fibrin, kinins, cytokines) Invading microbe Redness of tissue Tissue temperature Vasodilatation of capillaries Capillary permeability Influx of fluids Influx of phagocytes into tissues

Infla

mm

atio

n Inflammation gives rise to localized reddening, swelling,

increased temperatures, and pain.

The function of inflammation is to localize tissue damage, localize responses, and then to restore tissue function.

The action of localized leukocytes is augmented (i.e., enhanced) via the attraction of neutrophils and monocytes normally found in circulation.

Microbial materials such as LPS, flagellin (making up bacterial flagella), activated complement, and even bacterial DNA serve as indicators of infection which in turn activates the production of pro-inflammatory cytokines (immune-system activating chemicals).

In addition to the cell-to-cell interactions underlying inflammation, the inflammatory response involves localized increases in blood flow, leakage of blood vessels, and attraction of leukocytes from the blood.

Inflammation

Inflammation

Fev

er Inflammation is a great tool, unless it becomes

chronic or non-localized. Chronic inflammation typically has an underlying

cause (e.g., ongoing infection). Non-localized inflammatory responses gives rise to

body-wide vessel dilation and leakage, resulting in precipitous drops in blood pressure called Shock.

Endotoxin signals inflammatory responses and systemic infections with Gram-negative bacteria can give rise to a very dangerous condition known as Septic Shock.

Fever is the preferred systemic response to bacterial infection.

Fevers are elevated body temperatures induced either by pathogen molecules or by body molecules produced in response to pathogen molecules.

Fever results in temperatures that, ideally, inhibit microbes while enhancing body defenses.

Characteristics:- rapid- does not generate immunologic memory- dependent upon germline encoded receptors recognizing structures common to

many pathogens

Innate Immunity- First Line of Defense

InnateImmunity

Innate immune system recognize 10 3 molecular pattern whereas adaptive immune system recognize more than 10 7 antigens.

Recognition of Microbial Recognition of Microbial NonselfNonself

• Depends on recognition of “Pathogen Associated Molecular Patterns Depends on recognition of “Pathogen Associated Molecular Patterns (PAMPS)”(PAMPS)”– Gene products unique to microbes-Gene products unique to microbes-Molecular Molecular

SignaturesSignatures of microbial invaders of microbial invaders– Conserved among microbes of a given class-InvariantConserved among microbes of a given class-Invariant– Essential for microbial survivalEssential for microbial survival– LPS (Gram negative bacteria); Peptidoglycan (Gram LPS (Gram negative bacteria); Peptidoglycan (Gram

positive bacteria)positive bacteria)

Pattern Recognition Receptors (PRRs)

• Recognize PAMPsRecognize PAMPs• Receptors expressed on macrophages Receptors expressed on macrophages • Signal to induce inflammatory cytokines and activate host Signal to induce inflammatory cytokines and activate host

defense mechanismsdefense mechanismsExamplesExamples: :

Macrophage mannose receptorMacrophage mannose receptorMacrophage Scavenging Receptor-LPS and Gram neg. Scavenging Receptor-LPS and Gram neg. bacteriabacteriaToll-like ReceptorsToll-like Receptors

TLR

2

TLR

2

TLR

1

TLR

6

TLR

3

TLR

4

TLR

5

TLR

9

Pe p

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ept id

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dsR

NA

LP

S (

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TOLL-LIKE RECEPTORS

Ba c

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TLR

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TLR

8

Receptors and Responses of Phagocytes in Innate Immunity

Opsonins and Opsonization

Defect in killing activity in MQ resulted in

Chronic granulomatous disease,CGD

Defect in Phagocyte oxidase enzyme.

Gram Pos. bacteria can not be killed with

MQs.

Adherence and Opsonization

• Adherence to Surface Receptors– Mannose Receptor– Scavenger Receptor

• Opsonins: Greek- “Prepare food for”– Enhance Phagocytosis– Antibodies-Fc Receptors– Complement Proteins-Complement Receptors– MBL and CRP via C1q-R on MQs

• All multi-cellular organisms are able to recognize and eliminate pathogens

• Despite their extreme heterogeneity, pathogens share highly conserved molecules, called “pathogen-associated molecular patterns” (PAMPs)

• Host cells do not share PAMPs with pathogens• PAMPs are recognized by innate immune recognition receptors

called pattern-recognition molecules/receptors (PRMs/PRRs)

Innate Immune Recognition via Patterns

Pathogen-associated molecular patterns

CHALLENGE: How can the host discriminate large numbers of diverse pathogens from each other &/or from self using a restricted number of receptors?

SOLUTION: Evolve variety of receptors that recognize conserved motifs on pathogens that are not found on higher eukaryotes

Pathogen-associated molecular patterns (PAMPs) recognized by pattern recognition receptors (PRRs) or molecules (PRMs)

Two categories of PRRs: Those that mediate capture, uptake & presentation of antigen

(scavenger receptors) Those that lead to the activation of pro-inflammatory

pathways (Toll-like receptors)

TLR1 triacyllipopeptides

TLR2 Bact. Lipoproteins, LTA, Peptidoglycan

TLR3 dsRNA (viral)

TLR4 LPS

TLR5 Flagellin

TLR6 diacyllipopeptides

TLR7 Natural ligand unknown

TLR8 Natural ligand unknown

TLR9 Unmethylated CpG DNA

TLR10 ?

Activation of signal transduction pathways by TLRs induces genes that function in host defense

Pro-inflammatory cytokines

Chemokines

MHC & costimulatory molecules

iNOS & antimicrobial peptides that directly destroy pathogens

TLRs have “shared” & “specific” signal transduction pathways

Shared – all TLRs & IL1R

4 essential components – adaptor proteins MyD88, TOLLIP & TRAF6, & protein kinase IRAK

Specific – some, but not all TLRs…

TLR signaling pathways

Involvement of TLR in Linking Innate Immunity to Adaptive Immunity

Phagocyte response to infection• The SOS signals

– N-formyl methionine-containing peptides

– Clotting system peptides– Complement products– Cytokines released by tissue

macrophages• Phagocyte response

– Vascular adherence– Diapedesis– Chemotaxis– Activation– Phagocytosis and killing

Source: SOM PathMicro online textbook

Phagocytosis

A. Attachment via receptors– FcR, complement R,

scavenger R, Toll-like R

B. Pseudopod extensionC. Phagosome formationD. Granule fusion and

Phagolysosome formation

A

B

C

D

Respiratory burstO2-dependent MPO-independent reactions

Toxic compounds: superoxide anion O2- , hydrogen peroxide H2O2 ,

singlet oxygen 1O2 , hydroxyl radical OH*

G-6-P-dehydrogenase Pentose-P + NADPHGlucose + NADP+

NADPH oxidase

Cytochrome BNADP+ + O2 NADPH + O2

Superoxide dismutase H2O2 + 1O22O2- + 2H+

OH* + OH- + 1O22O2- + H2O2

Respiratory burstO2-dependent MPO-dependent reactions

Toxic compounds: hypochlorous acid OCl-, singlet oxygen 1O2

myeloperoxidase OCl- + H2O H2O2 + Cl-

1O2 + Cl- + H2O 2OCl- + H2O

Respiratory burstDetoxification reactions

Superoxide dismutase H2O2 + O2O2- + 2H+

Catalase H2O + O22H2O2

O2-independent killing

Effector molecule Function

Cationic proteins (cathepsin) Damage to microbial membranes

Lysozyme Hydrolyses mucopeptides in the cell wall

lactoferrin Deprives pathogens of iron

Hydrolytic enzymes (proteases) Digests killed organisms

Summary of intracellular killing pathways

Nitric oxide-dependent killing• Bacteria binds to macrophage• Production of TNF-alpha• Upregulates iNOS• Release of NO

• NO is toxic to infected cells in vicinity of macrophage

• NO sythetase is in the cytosol of activated MQ but noy naïve.

Macrophage

IFN-gammaTNF

O2 + L-arginine NO + citrullineNO synthetase

Phagocytosis“It is the toll-like receptors on macrophages that enable them to sense that the material is microbial in origin, and must therefore be eliminated quickly.”

Chemotaxis = movement toward infections.

FcR-mediated PhagocytosisThree types of FcR

• FcRI(CD16)– monocytes, M. IFN- stim neutrophils– high affinity binds monomeric IgG

• FcRII (CD32) A&B forms– low affinity binds to multimeric IgG– A - phagocytic and NK cells– B - B/T cells

• FcRIII (CD64)– monocytes, M, neutrophils

Innate immunity after combat with microbes, stimulates acquired immunity. So, it’s affect on the nature of acquired immunity and after then acquired immunity improves the function of innate immunity.

Y

Endosome

YNaive T Cell

MHC B7

Inflammatory and effectorcytokines

PRR

PAMP

Activated T Cell

CD40L, FasL, CD30L, CD27L

B Cell

Phagocytosis

APC

Direct Bactericidal ActivityPhagocytosisOxygen burstAnti-microbial peptides

Pathogen-specific Antibody

Complement

Determinants recognized by theinnate immune system

Immune response to damage• Dependent on what, where and how bad• Phased response with critical timing

– Requires chemokine signalling, receptor binding, etc

Days: 0 4 7 Weeks: 2 4 6

Non-specific killer cellsNK cells

EosinophilsMast cells

Innate response to virus infection and altered self

• Infected or altered self (transformed) cell downregulated MHC

• NK does not receive inhibitory signal

• Signals kill infected cell

NK cell

Infected/transformed cell

No MHC

Inhibitory RNK R

Innate response to extracellular microorganisms (parasites)

• Activated eosinophils release granule components– Major basic protein

• Major component of granules

– Eosinophil peroxidase• Cationic hemoprotein

– Eosinophil cationic protein• ribonuclease

Eosinophil

Natural Killer (NK) Cells- First identified by having the ability to lytically kill certain tumor cell lines

without prior sensitization

- Kill target cell by release of cytotoxic granules containing granzymes and perforin which penetrate target cell membrane and induce programmed cell death

- Can mediate Antibody-Dependent Cellular Cytotoxicity (ADCC) with CD16 that bind to IgG1,3

- Kill virally-infected cells with missing MHC class I

- Activated by IFN-/ or IL-12,18 (produced rapidly by activated macrophages) and proliferate with IL-15.

- Activated NK cells secrete IFN, acts on macrophages to increase microbial phagocytosis and killing

Antibody-Dependent Cellular Cytotoxicity

NK cell receptorsInhibitory Receptors:- germ-line encoded, no combinatorial diversity as seen with T/B cell receptors

- inhibit cytotoxicity to prevent killing of normal host cells

- specific for MHC class I alleles

- binding to class I sends inhibitory signal to NK cells

Activating Receptors:- germ-line encoded

- recognize carbohydrate structures on self proteins

NK Cell Cytotoxicity• Schematic representation

of NK cell receptors and killing. Normal cells are not killed because inhibitory signals from MHC class I molecules override activating signals. In tumor cells or virus-infected cells, reduced expression or alteration of MHC molecules interrupts the inhibitory signals, allowing activation of NK cells and lysis of target cells.

Virus-infected or

NK produces IFN-gamma for MQs and it produce IL-12 for NK.

Natural Killer-T Cells (NK-T Cells)- Minor subset of -T cells originally described by expression of NK-cell associated

markers

- Majority express invariant TCR (V14-J18/V8.2), remaining express diverse TCR

- Rapidly release large amounts of IL-4 and IFN, can interact with/influence other “innate immune” lymphocytes (NK cells, -T cells)

- Recognize self and foreign glycolipids presented by CD1

- Crystal structure analysis of CD1d indicates the presence of an MHC-like fold with a large, hydrophobic binding groove

- Due to the unique glycolipid antigen binding ability of CD1 molecules, it has been speculated that CD1 acts as an alternative mechanism for surveillance of foreign and altered-self glycolipids that would otherwise escape conventional class I and II pathways

“Innate” Lymphocytes- Unique, minor subsets of T and B lymphocytes that undergo receptor gene

rearrangements to generate receptor diversity (unlike NK cells)

- These subsets express limited receptor diversity, utilizing only a small number of receptor gene segments

- Tend to found in specific locations in the body, usually sites that encounter exogenous antigens or pathogens

Cytokines

-The most important cells in innate immunity

that produce cytokine are MQs, Neu, NK.

-IFN-a & B (control of viral infection)

-TNF-a & IL-1 (pro-inflammatory cytokines)

- IL-12 & IL-15 (NK activator)

-IFN-gamma (MQ activator)

- IL-6 (Neu production and from BM and CRP)

InterferonsProteins usually produced by virally infected cells

* Types of interferons:

1- Alpha interferon Secreted by Macrophages Induced by Viruses or Polynucleotide

2- Beta interferon Secreted by Fibroblasts, Viruses

3- Gamma interferon T- lymphocytes, Specific antigens

dsRNA

Interferon: An AntiviraldsRNA normally is not present in cells.

Thanks