CASE 1 - Partners in crime: Neutrophils and macrophages in infection,
inflammation and disease
Problem statement:
How do neutrophils and macrophages collaborate?
Brainstorm:
• Neutrophils are first line of defense → innate
• Macrophages (M1 and M2) present antigen and activate adaptive immune system
• M1: in overflow in inflammation, scares away M2 (type switching)
• interleukins
• NETs: Neutrophils release DNA
• Macrophages have MHC1 and MHC2 → complex presents antigen
• Neutrophils have MHC1
• inflammation: cell accumulation, increased blood flow, cytokines, innate cell recruitment
• neutrophils have granules
• few ways to get rid of pathogens
• production of neutrophils in bone marrow (production in embryos?)
• PCAM ICAM anchor proteins diapedesis
• respiratory burst to kill bacteria (radicals)
• PRR (DAMPs and PAMPs)
• B cells + T cells define adaptive immune system because they mutate the receptors
• Toll-like receptor, nod-like receptor, intra- and extracellular receptors
• inflammasome
Learning goals:
Neutrophils
1. What are neutrophils (features, functions, structure)
Neutrophils are the most abundant type of granulocytes and make up 40% to 70% of all white blood
cells in humans. They form an essential part of the innate immune system, with their functions
varying in different animals.
They are formed from stem cells in the bone marrow and differentiated into subpopulations of
neutrophil-killers and neutrophil-cagers. They are short-lived and highly mobile, as they can enter
parts of tissue where other cells/molecules cannot. Neutrophils may be subdivided into segmented
neutrophils and banded neutrophils (or bands). They form part of the polymorphonuclear cells
family (PMNs) together with basophils and eosinophils
Neutrophils are a type of phagocyte and are normally found in the bloodstream. During the
beginning (acute) phase of inflammation, particularly as a result of bacterial infection, environmental
exposure, and some cancers, neutrophils are one of the first responders of inflammatory cells to
migrate toward the site of inflammation. They migrate through the blood vessels and then
through interstitial tissue, following chemical signals such as interleukin-8 and H2O2 in a process
called chemotaxis. They are the predominant cells in pus, accounting for its whitish/yellowish
appearance.
1
,Neutrophils are recruited to the site of injury within minutes following trauma and are the hallmark
of acute inflammation; however, due to some pathogens being indigestible, they might not be able
to resolve certain infections without the assistance of other types of immune cells.
When circulating in the bloodstream and inactivated, neutrophils are spherical (bolvormig). Once
activated, they change shape and become more amorphous (vormloos) or amoeba-like and can
extend pseudopods as they hunt for antigens
Neutrophils undergo a process called chemotaxis via amoeboid movement, which allows them to
migrate toward sites of infection or inflammation. Cell surface receptors allow neutrophils to detect
chemical gradients of molecules such as interleukin-8 (IL-8), interferon gamma (IFN-γ), C3a, C5a,
and Leukotriene B4, which these cells use to direct the path of their migration.
Neutrophils have a variety of specific receptors, including ones for complement, cytokines
like interleukins and IFN-γ, chemokines, lectins, and other proteins. They also express receptors to
detect and adhere to endothelium and Fc receptors for opsonin
Being highly motile, neutrophils quickly congregate at a focus of infection, attracted
by cytokines expressed by activated endothelium, mast cells, and macrophages. Neutrophils
express and release cytokines, which in turn amplify inflammatory reactions by several other cell
types.
In addition to recruiting and activating other cells of the immune system, neutrophils play a key role
in the front-line defense against invading pathogens. Neutrophils have three methods for directly
attacking micro-organisms: phagocytosis (ingestion), degranulation (release of soluble anti-
microbials), and generation of neutrophil extracellular traps (NETs)
Neutrophils help prevent infections by blocking, disabling, digesting, or warding off invading particles
and microorganisms. They’re constantly searching for signs of infection, and quickly respond to trap
and kill pathogens.
They also communicate with other cells to help them repair damaged cells and mount an immune
response. Neutrophils play an important role in regulating the immune system and inflammation in
your body.
2. How are neutrophils developed? (Place where they reside)
Granulopoiesis. Neutrophils are produced in the bone marrow. From a self-renewing hematopoietic
stem cell (HSC), a multipotent progenitor (MPP) cell is formed. MPPs give rise to lymphoid-primed
multipotent progenitors (LPMP), which differentiate into granulocyte-monocyte progenitors (GMP).
These GMPs, under control of the granulocyte colony-stimulating factor (G-CSF) commit to
neutrophil generation by turning into myeloblasts, which then follow a maturation process that
includes the stages of promyelocyte, myelocyte, metamyelocyte, band cell, and finally a mature
neutrophil.
2
, 3. How do neutrophils get activated?
Neutrophils from the blood can be mobilized to sites of infection or inflammation through the
process known as the leukocyte adhesion cascade. Endothelial cells of blood vessels close to the
affected site get activated and express adhesion receptors such as E-, and P-selectins. These
receptors bind glycoprotein ligands on neutrophils, causing them to roll on the endothelium. Next,
the neutrophil is activated by chemokines, which induce a high affinity state in β2 integrins. Binding
of integrins to their ligands such as intercellular adhesion molecule-1 (ICAM-1) and ICAM-2 on
endothelial cells causes firm adhesion of the neutrophil. Next the neutrophil transmigrates into
peripheral tissues
4. What is the role of neutrophils in an infection? (NETs, oxidative burst)
Neutrophils have a well-established role during fungal and extracellular bacterial infections where
they promote bacterial clearance through phagocytosis, production of reactive oxygen and nitrogen
species (ROS/RNS), neutrophil extracellular trap (NET) formation, and production of pro-
inflammatory cytokines
Neutrophils are phagocytes, capable of ingesting microorganisms or particles. For targets to be
recognized, they must be coated in opsonins – a process known as antibody opsonization. They can
internalize and kill many microbes, each phagocytic event resulting in the formation of
a phagosome into which reactive oxygen species and hydrolytic enzymes are secreted. The
consumption of oxygen during the generation of reactive oxygen species has been termed the
"respiratory burst", although unrelated to respiration or energy production.
The respiratory burst involves the activation of the enzyme NADPH oxidase, which produces large
quantities of superoxide, a reactive oxygen species. Superoxide decays spontaneously or is broken
down via enzymes known as superoxide dismutases (Cu/ZnSOD and MnSOD), to hydrogen peroxide,
which is then converted to hypochlorous acid (HClO), by the green heme enzyme myeloperoxidase. It
is thought that the bactericidal properties of HClO are enough to kill bacteria phagocytosed by the
neutrophil, but this may instead be a step necessary for the activation of proteases
activation of neutrophils causes the release of web-like structures of DNA; this represents a third
mechanism for killing bacteria. These neutrophil extracellular traps (NETs) comprise a web of fibers
composed of chromatin and serine proteases that trap and kill extracellular microbes. It is suggested
that NETs provide a high local concentration of antimicrobial components and bind, disarm, and kill
microbes independent of phagocytic uptake. In addition to their possible antimicrobial properties,
NETs may serve as a physical barrier that prevents further spread of pathogens. Trapping of bacteria
may be a particularly important role for NETs in sepsis, where NETs are formed within blood
vessels. Finally, NET formation has been demonstrated to augment bactericidal activity during
infection macrophage
5. How do neutrophils recognize healthy cells? (PRRs)
Neutrophils recognize pathogens via classes of cell surface and intracellular receptors that bind to
microbe-specific molecules. Neutrophils also use numerous receptors that recognize host proteins
(such as IgG and complement) opsonizing the microbe. These receptors induce intracellular signals
that lead to full pathogen-killing capacity. The magnitude, quality, and duration of the elicited
response are dictated by the repertoire of receptors engaged at any one time, which in turn defines
the set point (i.e., activation status) of the neutrophil.
3
, Pathogen-associated molecular patterns (PAMPs) are recognized by neutrophil pattern-recognition
receptors (PRRs). Many of these receptors are also engaged by damage-associated molecular
patterns (DAMPs), which are released by necrotic during sterile inflammation such as in burns or
hypoxia. Engagement of PRRs by either PAMPs or DAMPs likely has similar outcomes
So, neutrophils basically recognize host cells since they don’t have any PAMPs or DAMPs on them.
6. Cytokines
4
inflammation and disease
Problem statement:
How do neutrophils and macrophages collaborate?
Brainstorm:
• Neutrophils are first line of defense → innate
• Macrophages (M1 and M2) present antigen and activate adaptive immune system
• M1: in overflow in inflammation, scares away M2 (type switching)
• interleukins
• NETs: Neutrophils release DNA
• Macrophages have MHC1 and MHC2 → complex presents antigen
• Neutrophils have MHC1
• inflammation: cell accumulation, increased blood flow, cytokines, innate cell recruitment
• neutrophils have granules
• few ways to get rid of pathogens
• production of neutrophils in bone marrow (production in embryos?)
• PCAM ICAM anchor proteins diapedesis
• respiratory burst to kill bacteria (radicals)
• PRR (DAMPs and PAMPs)
• B cells + T cells define adaptive immune system because they mutate the receptors
• Toll-like receptor, nod-like receptor, intra- and extracellular receptors
• inflammasome
Learning goals:
Neutrophils
1. What are neutrophils (features, functions, structure)
Neutrophils are the most abundant type of granulocytes and make up 40% to 70% of all white blood
cells in humans. They form an essential part of the innate immune system, with their functions
varying in different animals.
They are formed from stem cells in the bone marrow and differentiated into subpopulations of
neutrophil-killers and neutrophil-cagers. They are short-lived and highly mobile, as they can enter
parts of tissue where other cells/molecules cannot. Neutrophils may be subdivided into segmented
neutrophils and banded neutrophils (or bands). They form part of the polymorphonuclear cells
family (PMNs) together with basophils and eosinophils
Neutrophils are a type of phagocyte and are normally found in the bloodstream. During the
beginning (acute) phase of inflammation, particularly as a result of bacterial infection, environmental
exposure, and some cancers, neutrophils are one of the first responders of inflammatory cells to
migrate toward the site of inflammation. They migrate through the blood vessels and then
through interstitial tissue, following chemical signals such as interleukin-8 and H2O2 in a process
called chemotaxis. They are the predominant cells in pus, accounting for its whitish/yellowish
appearance.
1
,Neutrophils are recruited to the site of injury within minutes following trauma and are the hallmark
of acute inflammation; however, due to some pathogens being indigestible, they might not be able
to resolve certain infections without the assistance of other types of immune cells.
When circulating in the bloodstream and inactivated, neutrophils are spherical (bolvormig). Once
activated, they change shape and become more amorphous (vormloos) or amoeba-like and can
extend pseudopods as they hunt for antigens
Neutrophils undergo a process called chemotaxis via amoeboid movement, which allows them to
migrate toward sites of infection or inflammation. Cell surface receptors allow neutrophils to detect
chemical gradients of molecules such as interleukin-8 (IL-8), interferon gamma (IFN-γ), C3a, C5a,
and Leukotriene B4, which these cells use to direct the path of their migration.
Neutrophils have a variety of specific receptors, including ones for complement, cytokines
like interleukins and IFN-γ, chemokines, lectins, and other proteins. They also express receptors to
detect and adhere to endothelium and Fc receptors for opsonin
Being highly motile, neutrophils quickly congregate at a focus of infection, attracted
by cytokines expressed by activated endothelium, mast cells, and macrophages. Neutrophils
express and release cytokines, which in turn amplify inflammatory reactions by several other cell
types.
In addition to recruiting and activating other cells of the immune system, neutrophils play a key role
in the front-line defense against invading pathogens. Neutrophils have three methods for directly
attacking micro-organisms: phagocytosis (ingestion), degranulation (release of soluble anti-
microbials), and generation of neutrophil extracellular traps (NETs)
Neutrophils help prevent infections by blocking, disabling, digesting, or warding off invading particles
and microorganisms. They’re constantly searching for signs of infection, and quickly respond to trap
and kill pathogens.
They also communicate with other cells to help them repair damaged cells and mount an immune
response. Neutrophils play an important role in regulating the immune system and inflammation in
your body.
2. How are neutrophils developed? (Place where they reside)
Granulopoiesis. Neutrophils are produced in the bone marrow. From a self-renewing hematopoietic
stem cell (HSC), a multipotent progenitor (MPP) cell is formed. MPPs give rise to lymphoid-primed
multipotent progenitors (LPMP), which differentiate into granulocyte-monocyte progenitors (GMP).
These GMPs, under control of the granulocyte colony-stimulating factor (G-CSF) commit to
neutrophil generation by turning into myeloblasts, which then follow a maturation process that
includes the stages of promyelocyte, myelocyte, metamyelocyte, band cell, and finally a mature
neutrophil.
2
, 3. How do neutrophils get activated?
Neutrophils from the blood can be mobilized to sites of infection or inflammation through the
process known as the leukocyte adhesion cascade. Endothelial cells of blood vessels close to the
affected site get activated and express adhesion receptors such as E-, and P-selectins. These
receptors bind glycoprotein ligands on neutrophils, causing them to roll on the endothelium. Next,
the neutrophil is activated by chemokines, which induce a high affinity state in β2 integrins. Binding
of integrins to their ligands such as intercellular adhesion molecule-1 (ICAM-1) and ICAM-2 on
endothelial cells causes firm adhesion of the neutrophil. Next the neutrophil transmigrates into
peripheral tissues
4. What is the role of neutrophils in an infection? (NETs, oxidative burst)
Neutrophils have a well-established role during fungal and extracellular bacterial infections where
they promote bacterial clearance through phagocytosis, production of reactive oxygen and nitrogen
species (ROS/RNS), neutrophil extracellular trap (NET) formation, and production of pro-
inflammatory cytokines
Neutrophils are phagocytes, capable of ingesting microorganisms or particles. For targets to be
recognized, they must be coated in opsonins – a process known as antibody opsonization. They can
internalize and kill many microbes, each phagocytic event resulting in the formation of
a phagosome into which reactive oxygen species and hydrolytic enzymes are secreted. The
consumption of oxygen during the generation of reactive oxygen species has been termed the
"respiratory burst", although unrelated to respiration or energy production.
The respiratory burst involves the activation of the enzyme NADPH oxidase, which produces large
quantities of superoxide, a reactive oxygen species. Superoxide decays spontaneously or is broken
down via enzymes known as superoxide dismutases (Cu/ZnSOD and MnSOD), to hydrogen peroxide,
which is then converted to hypochlorous acid (HClO), by the green heme enzyme myeloperoxidase. It
is thought that the bactericidal properties of HClO are enough to kill bacteria phagocytosed by the
neutrophil, but this may instead be a step necessary for the activation of proteases
activation of neutrophils causes the release of web-like structures of DNA; this represents a third
mechanism for killing bacteria. These neutrophil extracellular traps (NETs) comprise a web of fibers
composed of chromatin and serine proteases that trap and kill extracellular microbes. It is suggested
that NETs provide a high local concentration of antimicrobial components and bind, disarm, and kill
microbes independent of phagocytic uptake. In addition to their possible antimicrobial properties,
NETs may serve as a physical barrier that prevents further spread of pathogens. Trapping of bacteria
may be a particularly important role for NETs in sepsis, where NETs are formed within blood
vessels. Finally, NET formation has been demonstrated to augment bactericidal activity during
infection macrophage
5. How do neutrophils recognize healthy cells? (PRRs)
Neutrophils recognize pathogens via classes of cell surface and intracellular receptors that bind to
microbe-specific molecules. Neutrophils also use numerous receptors that recognize host proteins
(such as IgG and complement) opsonizing the microbe. These receptors induce intracellular signals
that lead to full pathogen-killing capacity. The magnitude, quality, and duration of the elicited
response are dictated by the repertoire of receptors engaged at any one time, which in turn defines
the set point (i.e., activation status) of the neutrophil.
3
, Pathogen-associated molecular patterns (PAMPs) are recognized by neutrophil pattern-recognition
receptors (PRRs). Many of these receptors are also engaged by damage-associated molecular
patterns (DAMPs), which are released by necrotic during sterile inflammation such as in burns or
hypoxia. Engagement of PRRs by either PAMPs or DAMPs likely has similar outcomes
So, neutrophils basically recognize host cells since they don’t have any PAMPs or DAMPs on them.
6. Cytokines
4
