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Innate Immune System
IMMUNE SYSTEM FUNCTION
To Protect against “entry” from microorganisms / pathogens
- Bacteria
- Viruses
- Fungi
- Parasites
- Toxins
- Venoms
Recognize Pathogens
Eliminate Pathogens
Enhance cellular synthesis and repair processes in the face of damage (or metabolic stress)
Two Basic “Systems”
- Innate Immune System: non-specific response directed against “anything that does not belong”
that also responds to any kind of cellular stress
- Adaptive Immune System: highly specific response directed towards a single pathogen or a tiny
component of a single pathogen
IMMUNE SYSTEM RESPONSE
Innate
- First “Line of Defense”
- Non- specifically blocks or attacks pathogens
Skin
- provides a waterproof barrier to prevent organisms from entering our “system”
Mucosal Epithelia
- can trap pathogenic particles and prevent them from getting through epithelial cells to
tissue
Leukocytes
Neutrophils, Eosinophils, Basophils & Mast Cells
Macrophages
NK-Cells
Vascular endothelial cells
Blood vessels have very tight junctions between
the cells that make up the vessel walls. Only when
inflammatory signals are produced locally will
pores open up in the vessel wall to allow
neutrophils, monocytes, and other immune cells
to squeeze out and enter the tissue.
Lymph vessels have relatively large gaps in the vessel walls so
the various immune cells can easily move from the lymph into
the tissues and back again into the lymph.
The cardiovascular system and lymph system provide a very
convenient transportation system for Neutrophils, monocytes,
NK cells, and other immune cells
INNATE IMMUNE SYSTEM (RESPONSE)
The term innate refers to the fact that these innate immune processes are not directed specifically toward any
particular pathogen but rather comprise part of a very general defense which defends against any type of foreign
pathogen. The innate system includes the skin and mucosal epithelial tissues. These tissues act as a “passive”
physical barrier to prevent the invasion of bacteria and viruses. The outer layer of skin is comprised of several layers
of dead skin cells which is simply a physical barrier. The epithelial cells lining the oral-GI tract and respiratory tracts
secrete mucus which traps bacteria & viruses and thereby prevents their entry into the “body”. The mucus is either
removed by coughing & spitting or by swallowing where the bacteria & viruses are destroyed by stomach acid or by
digestive enzymes in the intestine. The role played by the inflammatory processes of the innate system is the active
killing of those pathogens which make it through the physical barriers.
Inflammation
The processes of inflammation are actually considered to be a part of the innate immune system. A more modern
interpretation of inflammatory responses though is that they comprise a graded stress-response system designed to
enhance cellular functions to enable proper responses to cellular stresses and damage. The active killing process is
performed predominantly by neutrophils, macrophages, and NK-cells. These cells serve to protect us from invading
bacteria, viruses, fungi, and parasites (as well as some toxins and venoms).
Neutrophils
Neutrophils are a type of granulocyte (mast cells which live in our tissues, especially noticeable in the respiratory
epithelium, also are granulocytes) that exists predominantly in the blood although many also are found in the lymph.
Neutrophils respond to non-human cells by producing large amounts of oxygen radicals (among other toxic
molecules) which kill the invading pathogens essentially by oxidizing the lipids and proteins in their cell membranes
(rupturing and destroying them). Neutrophils are stimulated to do this by anything that is not smooth or which does
not have a protein coat (which describes most non-human cells) as well as to any entity which has antibody proteins or
compliment proteins attached to it. The antibodies and/or compliment stimulate the neutrophils to attack much
more aggressively than if there were no antibodies (a very important function relating to the adaptive immune system
response). In addition to killing cells quite efficiently, neutrophils also can phagocytize dead or dying cells.
INNATE IMMUNE SYSTEM (RESPONSE)
Inflammation
Macrophages
Macrophages are cells (derived from monocytes) that “live” within our tissues as well as in the lymph.
Macrophages recognize nonhuman pathogens in the same manner as neutrophils do and they are highly
stimulated by antibodies and compliment as well. Monocytes usually circulate in the blood and when they are
stimulated by various leukotrienes to enter the tissue they will differentiate to become macrophages.
Macrophages are specialized for phagocytizing and digesting dead cells, dying cells, and foreign (non-human)
cells. They also can produce oxygen radicals to kill things. As discussed below, they are one of the important
types of antigen-presenting cells necessary for the adaptive immune response.
Natural Killer Cells
NK cells are a type of lymphocyte that is specialized at killing non-human cells (as well as many cancer cells) by
lysing them. They recognize any cells which do not express the MHC as well as virus-infected cells and cells
coated with antibodies. They kill cells by “injecting” proteins (porins) into their outer membranes that then
cause the membranes to rupture.
All of these cells (neutrophils, macrophages, NK cells) are more or less inactive (although the NK cells are more
active all the time) and need to be activated by either antibodies, physical contact with pathogens, or eicosanoids
and cytokines produced by proinflammatory responses. The inflammatory process can be initiated by physical
damage to cells due to the effects of infectious agents, cellular damage due to mechanical or chemical
disruption, or simply the overproduction of reactive oxygen species (ROS) from normal metabolic stress.
Inflammation:
The Good
Overall Purpose of Inflammation
1. Increased blood flow to increase “in-flow” of inflammatory and immune cells,
2. Infiltration of the tissue with inflammatory cells to kill off infectious agents,
3. Blood clots to stop the bleeding,
4. Scar tissue formation to close up wounds,
5. Increased protein synthesis to help repair the damage,
6. Increased cell division (signals) to replace any dead cells,
7. Initiation of an immune response resulting in the production of antibodies (and other things) . . .
Overall Purpose of Inflammation – More Details
- First Line of Protection From Bad Things (microorganisms / pathogens)
- Bacteria
- Fungi
- Toxins
- Viruses
- Parasites
- Venoms
- A variety of cells can respond to pathogens to initiate proinflammatory signaling by producing proinflammatory
eicosanoids and cytokines (interleukins and chemokines) in order to initiate an inflammatory response, including
dendritic cells, NK cells, neutrophils, tissue macrophages, mast cells, endothelial cells, and fibroblasts. Essentially all
cells can produce proinflammatory eicosanoids in response to cellular stress and cellular damage.
- Inflammatory Cells Recognize and Eliminate Pathogens:
Neutrophils (circulate in the blood – chemoattraction into tissues)
Macrophages (live in tissue/circulate as monocytes – chemoattraction into tissues)
Kill things by eating and digesting them
Kill things by releasing oxygen radicals: O2- H2O2 ·OH
NK Cells (circulate in blood – peripheral tissues)
Kills things using perforins; activated on by:
- anything with no MHC
- antibodies attached
How Does This Inflammation Thing Work?
Specific receptors for pathogen associated molecular patterns (PAMPs) bind to components of pathogens and
initiate signal transduction pathways that lead to the synthesis of the various proinflammatory signaling
molecules. Specific receptors for damage associated molecular patterns (DAMPs) bind to components of dead (or
dying) cells to activate essentially the same signal transduction pathways to initiate the synthesis of
proinflammatory signaling molecules; signaling molecules that then activate the various inflammation processes:
1. P38-MAPK, ERK-MAPK, calcium, and ROS activate Phospholipase A which cuts off the fatty acid
arachidonic acid from membrane phospholipids
2. Arachidonic acid is made into prostaglandins, leukotrienes, and thromboxanes:
Prostaglandins: PGE-2 - causes vasodilation (among many other things)
Leukotrienes: LTB-4 - attraction/activation of neutrophils
Thromboxanes: TXA-2 – platelet aggregation
3. Activated macrophages, neutrophils, glial cells, dendritic cells, B-cells, fibroblasts, and endothelial
cells produce a variety of interleukins and other cytokines:
IL-1 → MCP-1: monocyte attraction, COX-2:PG/LT/TX
IL-6 → fibrinogen production
IL-8 → chemoattractant
TNF → HSP-47/NFkB / AP-1 (fibrosis/protein synthesis/cell division)
NFkB → ICAM
One of the major signaling process involved in initiating inflammation is the production of the eicosanoids:
prostaglandins, thromboxanes, and leukotrienes from arachidonic acid. The enzyme phospholipase A2 (PLA2)
is activated indirectly by either ROS or calcium; or directly by p38 MAPK and ERK-MAPK. Damage to cellular
membranes by bacterial toxins, bacterial digestive enzymes, viral damage, physical trauma, and chemical attack
by various chemical and oxygen radicals allows calcium to leak through the membrane and activate PLA-2. Any
metabolic stress (think exercise, high rates of tissue growth/repair) also leads to activation of PLA-2 through
calcium and ROS-mediated activation of the P38- and ERK-MAPK signal transduction pathways.
Activation of PLA-2 leads to removal of Arachidonic acid (or Eicosapentaenoic acid / Docosahexaenoic acid)
from the phospholipid: phosphatidyl choline (predominantly on ER and nuclear membranes). These fatty acids
are picked up by COX and LOX enzymes and synthesized into several prostaglandins, leukotrienes, and
thromboxanes, while the Lyso-PC is synthesized into Platelet Activating Factor. On binding to their appropriate
receptors these lipid-derived signaling molecules initiate various responses: Leukotriene B4 is a potent
chemotactic molecule which greatly enhances the accumulation of leukocytes within the inflammatory area
while thromboxane A2 activates platelets. Prostaglandin E2 (can cause vasodilation to enhance blood flow to
the infected (or damaged) area and for enhancing vasopermeability to allow neutrophils easy entry to the tissue
from the blood. Thus, one end result of the production of all of these signaling molecules is the efficient
recruitment and activation of neutrophils and macrophages to kill and digest any pathogens as well as dying (or
dead) cells.
In addition to the activation of neutrophils and macrophages, eicosanoid molecules also activate other
inflammatory cells, such as glial cells, dendritic cells, B-cells, fibroblasts, as well as other endothelial cells that
may be in the region. Although most of these other cells will be “ignored” here, they are still important in the
inflammatory process because when they are activated to produce inflammatory signals, the result is an
amplification of inflammatory signals being produced within the damaged region. Thus, they participate in a
cascade of effects: damaged cells produce inflammatory signals which stimulate other cells in the local region
to produce inflammatory signals, amplifying the production of inflammatory signals; resulting in the efficient
recruitment of inflammatory cells into the region.
(These signaling molecules have many other effects than the ones listed above... These examples simply
illustrate the concept that each change in cellular function demands a signaling molecule of some sort to
initiate the change in function)
Activation of PLA-2 by P38- and ERK-MAPKs, calcium, or ROS leads to removal of Arachidonic acid (or Eicosapentaenoic acid /
Docosahexaenoic acid) from the phospholipid: phosphatidyl choline. These fatty acids are picked up by COX and LOX enzymes
and synthesized into several prostaglandins, leukotrienes, and thromboxanes by the specific cell-types indicated, while the Lyso-PC
is synthesized into Platelet Activating Factor. On binding to their appropriate receptors these lipid-derived signaling molecules
initiate the responses indicated in the red boxes.
As a result of pathogen- or chemical-induced membrane damage (DAMPS; including: ssRNA, dsRNA, and CpG
DNA, and endogenous nucleotides), or contact with a pathogen (PAMPS; including: LPS, bacterial peptidoglycans,
bacterial flagellin, envelope proteins from viruses and hemagglutinin protein) a variety of signaling pathways are
activated which then stimulate the synthesis, or expression, (and extracellular release) of IL-1, IL-6, and TNF by the
pathogen-activated macrophages or the damaged endothelial cells (or pathogen-activated neutrophils). These
cytokines can then initiate the activation of transcription activators in many of the cells in the damaged or infected
region. They can stimulate the expression (transcription and translation) of the acute-phase inflammatory response
elements: cyclooxygenase-2 (COX2), IL-8, NFκβ and c-reactive protein. IL-6, TNF, and NFκβ in turn , activate other
transcription activator proteins which then induce the synthesis of P-selectin, E-selectin, VCAM (vascular cell
adhesion molecule), and ICAM (inter cellular adhesion molecule) by endothelial cells, especially those of the
capillaries (other effects of these cytokines are discussed below). These cellular adhesion molecules are necessary
for the recruitment of circulating neutrophils and monocytes into the infected or damaged area while the other
interleukins activate these (and other) inflammatory cells.
Once activated by the various PGs, PAMPs and/or DAMPs, neutrophils and macrophages can produce large
quantities of reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (other highly reactive
molecules are produced but they are being ignored here) which then react with each other to produce the highly
reactive (and damaging) hydroxyl radical. These ROS are responsible for killing the (pathogens). Unfortunately,
they also cause oxidative damage to membranes of surrounding cells and therefore create an enhanced risk for
inflammation-mediated cytotoxicity. ROS also can cause DNA-damage in the normal cells in the inflamed region
leading to enhanced risk for cancer. The activated monocytes recruited into the tissue (via adhesion molecules)
mature to become macrophages (to add to the resident macrophages) to add to this inflammatory response.
Another consequence resulting from the production of these signaling molecules is the formation of blood-clots
and/or scar tissue to close the wound. (This could, however, be a problem if there is no wound to close.) Blood-clot
formation is initiated by IL-6 which activates platelets to become sticky (clotting factors will conveniently be ignored
in this course). Scar formation is initiated by several of the cytokines produced by the stimulated neutrophils and
macrophages. Both IL-1 and TNF can stimulate the activation of the transcription activator AP-1 which, among
other things, induces expression of TGF-β; TGF-β then stimulates tissue fibroblasts to produce HSP-47. HSP-47
mediates enhanced collagen synthesis by these fibroblasts. IL-6 stimulates hepatocytes and epithelial cells to
produce fibrinogen which, along with collagen, provides the major proteins necessary fibrosis (scar formation).
Various components of dead cells
(Damage Associated Molecular
Patterns) and of pathogens (Pathogen
Associated Molecular Patterns) bind to
Toll-like Receptors, Nod-like receptors,
and/or Rig-like receptors.
Binding of ligands to RLRs leads to the
formation of IPS-1 complexes with
mitochondria and the subsequent
activation of the transcription factors
IRF3, IRF7, and NFκβ via Iκκζ/TBK1
and caspases 8&9, respectively.
Binding of ligands to TLRs leads to
activation of NFκβ (via TAK1) and
INF3/7 (via TBK1) via MyD88 and
MyD88-independent pathways,
respectively.
Binding of ligands to NLRs leads to the
formation of inflammasome complexes
and the activation of MEKK3 and
TAK1. MEK3 activates p38 MAPK
pathway while TAK1 activates both
JNK MAPK and NFκβ pathways.
Activation of the MAPK pathways
leads to the activation of various
transcription factors which, along with
the activated NFκβ and IRF3&7 lead to
the synthesis of the cytokines IL-1, IL6, TNFα, INF and the chemokines
RANTES, IL-8, MCP-1, and MIP-2 and
the adhesion molecules VCAM, ICAM,
E-Selectin, and V-selectin.
Binding of IL-1, IL-6, and TNFα to
receptors of the PAMP/DAMP activated
cells (autocrine stimulation) leads to
additional activation of the p38 and JNK
MAPK pathways (IL-1 TNFα: via ASK1
and TAK1) as well as additional
activation of NFκβ via MEK3/6 and
MEK1/2/3/4.
Activation of the ERK-MAPK pathway
and mTORC1 by IL-6 leads to additional
activation of various transcription
factors as well as accelerated protein
synthesis via mTORC1.
These autocrine-mediated signaling
events lead to accelerated synthesis of
the various pro-inflammatory molecules:
cytokines IL-1, IL-6, TNFα, INF, the
chemokines RANTES, IL-8, MCP-1,
and MIP-2, and the adhesion molecules
VCAM, ICAM, E-Selectin, and Vselectin.
Solid arrows indicate original activation
pathways; open arrows indicate
autocrine-mediated activation of signal
transduction pathways and resulting
synthesis.
An intact cell membrane acts as a barrier for
(among other things) charged particles such
as calcium.
A damaged cell membrane will allow (among
other things) charged particles to enter the
cell or organelle.
Entry of calcium into the cytosol
via normal signaling events
mediated by DAG or through
ROS-damaged membranes
activates PKC, which then may
activate PLA-2, and the ERK- and
p38-MAPK pathways. At the same
time calcium-activated increases
in cellular metabolism
subsequently increases
production of ROS, which also
may activate PLA-2 as well as p38MAPK and JNK-MAPK via TrxAsk1. ROS-caused single and
double-strand breaks in DNA
activates repair by ATM which
also activates NFκβ and ROSmediated PI3K activity also
enhances mTORC1 via Akt to
enhance overall rates of protein
synthesis. The result of all these
effects is the synthesis of a variety
of proinflammatory molecules in
response to cellular stress.
Another cellular response initiated by the inflammatory signals is stem cell division. When there is sufficient
damage to kill tissue cells, they then need to be replaced. Stem cells in the tissue respond to the various
inflammatory cytokines by increasing their rates of protein synthesis in order to grow and then they divide,
producing daughter cells which replace the dead tissue cells. Signaling molecules called growth factors are
important in producing this cell growth/cell division effect. The chemokine CXCL8 (formerly known as IL-8)
stimulates platelets (which are activated by IL-6) to synthesize and release platelet-derived growth factor
(PDGF) which stimulates all cells in the region to enhance rates of protein synthesis. Prolonged activation of
the transcription activator AP-1 also leads to greatly enhanced rates of protein synthesis and ultimately (in
stem cells) to cell division. Thus the inflammatory cytokines produced by stressed or damaged cells and
then by stimulated macrophages and neutrophils are responsible for initiating all of the steps necessary for
repair of wounds and protection from infection: enhanced rates of protein synthesis for cell-repair, enhanced
rates of fibrosis for wound healing, enhanced rates of stem cell division to replace dead cells, and activation
of neutrophils, macrophages, and dendritic cells to kill any invading pathogens. This last result of the
primary inflammatory response is very important in initiating the development of an adaptive immune
response.
The adaptive immune response ultimately results in the production of antibodies which recognize specific
antigens. Antibodies are proteins which are produced by antibody-producing B-lymphocytes. These
antibody proteins specifically bind to structures that are found only on the pathogen which initiated the
adaptive response in the first place. One antibody produced by one B-lymphocyte binds to one antigen while
a different antibody produced by a different B-lymphocyte binds to a different antigen. One pathogen (let’s
say a particular strain of bacteria such as Mycobacterium tuberculosis) may be capable of having hundreds
of different antigenic sites as part of the bacterium. One antigenic site may be composed of several amino
acids from a particular protein that corresponds to a shape that is definitely “not human” in origin. With
several hundred different proteins making up the bacterium it is possible that there could be hundreds of
short sequences of amino acids among those bacterial proteins which possibly represent different antigenic
sites. For many other strains of bacteria, some of the lipids contained in the bacterial cell wall are so different
from those in humans that they also might be capable of being antigens. How these antigens actually
stimulate the adaptive immune response is a real cool process; a process which starts with antigen-presenting
cells such as the macrophage or dendritic cell.
Regulation of Tissue Regeneration
PGE2 and other growth factors activate
various MAPK signal-transduction
pathways in Stem Cells to initiate the
activation and synthesis of a variety of
proteins that are necessary for activating
growth of the stem cell and the synthesis
of DNA to duplicate the chromosomes in
order to proceed to the
mitosis/cytokinesis phase.
A decline in the levels of PGE2 and of
growth factors induces G0 arrest via
reducing MAPK activities.
Progenitor cells divide more-or-less
continually in response to normal levels
of stress-response signaling with the
displaced daughter cells differentiating
into tissue cells.
In skin and intestine extensive tissue
damage greatly enhances rates of cell
division in stem cells and their daughter
cells can differentiate directly into tissue
cells.
Some of the recruited immune cells produce reactive oxygen species which may then cause damage to
other cells in the region…
With an appropriate inflammatory response bacteria are killed before they can cause extensive damage and the
wound is closed and tissue repaired before additional infections can occur and cause more damage.
A Brief Review:
As a result of all of the inflammatory signaling molecules being produced, a variety of cellular responses occur in
the affected tissues:
1. Increased blood flow to increase “in-flow” of inflammatory and immune cells,
2. Infiltration of the tissue with inflammatory cells to kill off infectious agents,
3. Blood clots to stop the bleeding,
4. Scar tissue formation to close up wounds,
5. Increased protein synthesis to help repair the damage,
6. Increased cell division (signals) to replace any dead cells,
7. Initiation of an immune response resulting in the production of antibodies (and other things) . . .
So . . . Exactly why do I want to know all this?
… why is all this inflammation and cell-function stuff important?
Well, we are made up of billions of cells and the proper function of our organs (each made up of a different
array of specialized cells) depends on the proper functioning of the individual cells.
Normal processes of metabolism in cells are the processes that ensure proper cell function. While these
processes are highly regulated, they can be increased by any additional calcium entering the cell. Normal
metabolic processes produce reactive oxygen specie (ROS) and therefor any additional metabolism will
result in the production of small amounts of additional reactive oxygen species ROS.
Normal functions of the protein synthesis enzymes and the lipid synthesis enzymes and the repair enzymes
are regulated by a variety of signaling molecules as well as by ROS and calcium through their combined
effects on signal transduction pathways; ensuring that any increase in damage from ROS due to any cell
stress is repaired and cell function continues.
When there is damage from, say, a cut to the skin, liver damage from drinking a little turpentine, or lung
damage from breathing in smoke from burning plastic, or damage to respiratory-tract tissues from a
bacterial or viral infection, then an inflammatory response will occur to accelerate the repair/replace
processes to ensure that the damage is repaired, dead cells are replaced, and tissue (or organ) function is
returned to normal.
Because normal signal transduction processes are regulated in part by both calcium and ROS;
this means that chronic elevation in both because of chronic stress to cells will result on
chronic over-activation of these S-T pathways…
… resulting in chronic proinflammatory signaling
The chronic proinflammatory signaling can then lead to the invasion of inflammatory cells into the tissue that
then cause lots of ROS-mediated damage and much more proinflammatory signaling. The damage can of
course, lead to cell death.
If only a few cells die then nobody cares… but if a whole bunch of cells in a single organ die at once, we just
might be in trouble: if a bunch of heart or brain cells die, we may go along with them. If a bunch of liver cells
die, we may suffer liver failure… if it is “mild” the inflammatory response may save our liver. The rest of our
cells in our other organs however, suffer until the liver is completely repaired because they depend on the liver
to produce many different molecules that are necessary for their proper function and to handle the wastes
produced by all of our cells. If the liver damage is too severe to repair quickly, then other organs can fail
because they don’t get what they need and again, we will die along with them. In essence, cells in each of the
different organs are all inter-dependent on the function of all the other cells in other organs.
Obviously without the inflammation response to start the repair & replace process you would die at the first
scratch… on the other hand, anything that inappropriately enhances this response, or sustains it for long
periods of time such as:
overexposure to toxic chemicals (cigarette smoke, toluene, carbon tetrachloride, ethanol, chlorine…) or to a
lack of specific nutrients and beneficial phytochemicals…
inevitably leads to continual local production of growth signals and ROS from the inflammatory cells and
eventually a slow & painful death from organ failure, cardiovascular disease, cancer, or some other
degenerative disease…
Ultimately, our health depends on optimal function of our cells! And this function can be impaired by
aberrant inflammation.
Normally resolved
inflammatory response
& healing
Non-resolved
inflammatory
responses lead to
enhanced risk for
chronic diseases
The only way to stop proinflammatory
signaling is to produce the
proresolution signaling molecules that
then stop the production of the
proinflammatory signaling molecules.
Proinflammatory signaling leads to an
induction of COX-2 and mPGES;
increasing PGE-2 synthesis and
subsequently an enhanced expression
of 12-LOX and 15-LOX. Increased
LOX in conjunction with COX-2
greatly enhances the synthesis of
lipoxins, resolvins, and PGJ-2.
Lipoxins and resolvins suppress
proinflammatory signaling via
inhibiting the activation of MAPKs
and NFκβ through enhanced
expression of SOCS in all
inflammatory cells while PGJ-2
activates apoptosis in neutrophils as
well as suppresses MAPKs and NFκβ.
At the same time, macrophages
phagocytize apoptotic neutrophils
(without production of ROS) and are
stimulated to egress from the tissue,
effectively ending an inflammatory
response.