NEUTROPHIL

By: Ali Mohamed

Supervisor: Prof.dr. Laila hamdy

Neutrophil
Granulopoiesis
One of the branches of haematopoietic tree, which prouduce 120 billion of
granulocytes daily (Crea et al., 2009). Because granulocytes have a short life time:
3-12 h at blood and 1-3 days in tissues, the rate of granulopoiesis is high (Crea et al.,
2009). The rate of neutrophil as example is 1-2 x 1011 cells / day (Malengier-Devlies
et al., 2021)

Figure 1show haematopoietic tree, HSC (haemtopoitic stem cell) develope into CLP
( common lymphoid progenitor) or CMP (common myloid progenitor, early
cytokines which control there pathways, CMP in presence of specific cytokines
develop to GMP which could be basophiles or eosinophils or neutophil
Myelocytes are characterized by the production of secondary or “specific” granules,
which allow the phenotypic differentiation between eosinophils, basophils and
neutrophils (Crea et al., 2009) the most important cytokine in neutrophil
development is G-SCF ( granulocyte colony-stimulating factor) (Bonecchi et al.,
2022). The human G-CSF gene is located on chromosome 17q21–22 and contains
five exons and four introns (Crea et al., 2009) while G-SCF control early steps in
hematopoietic it also with GM-CSF and IL-3 drive the earliest steps of
granulopoiesis, however G-SCF control last stages in Granulopoiesis (Nakajima &
Ihle, 2001) G-SCF also, influence on mature neutrophil acticity (Platzer et al., 1985)
Types of granulocytes
Neutronphil Basophil Eosinophi
Nucleus Poly nuclear lobulated nucleus Two loped nucleus
morpholgy
PH 7.6 (Hayes et al., 9.6 (Shah et al., 6-7.5 (Kottyan et
2020) 2021) al., 2009)
Abundance in 50-70% 0.5-1% 1-3%
blood
Phagocytosis yes no Yes (Cline et al.,
1968)
Major role Inflammation Allergic disease: Anti-parasites
Asthma
Skin allergy
Anaphylaxis
(Voehringer, 2011)
Life span 4-5 days 1-2 days (Min et 2-5 days (Park &
(Koenderman et al., 2012) Bochner, 2010)
al., 2022)
Neutrophil:
It was discovered by Elie Metchnikoff in starfish larvae (Segal, 2005)
The function and strategies that have to kill microbes was dicoverd after 1970 (Segal,
2005), it have three main mechanisms to kill microbe: 1- phagocytosis, granules,
NETs
Develop in bone marrow then run into blood 10-12 h then arrive target tissue which
know by migration, this process have multi step, neutrophil is the one of cells which
caused inflammation due inflammation factors like chemokines or NETs, after 4-7
days in tissue it die by apoptosis.
Neutrophil show great defense against virus, bacteria, fungi and tumor cells.
Basophil
Basophils are the least abundant leucocytes primarily found in the circulation. The
lifespan of basophils is short; recently estimated to be in the range of 1–2 days.
express the high-affinity receptor for IgE and can release a similar spectrum of
mediators upon IgE cross-linking (Min et al., 2012)
basophils are a primary source of IL-4 in vivo (Min et al., 2004)
IL-3, presumably derived from T cells, plays a crucial role of regulating basophil
responses (Min et al., 2004) due to secretion of IL-4 basophil may be have a role in
TH-2 differentiation which response to allergens (Sokol et al., 2007)
Eosinophil
Have a short life 8-18 h in blood and 2-5 days in tissue
How ever, cutokines increase eosinophil survival in vitro to 14 days (Park &
Bochner, 2010) it reside in GIT within lamina propria
It have role against helminth. bacteria: Eosinophil’s proteins have ribonuclease
activity (Rosenberg, 1995)
Eosinophil show promotion of b-cell proliferation, survival and antibody production
(Travers & Rothenberg, 2015)
Neutrophil recruitment
Five major steps: 1-tethering 2- rolling 3-adhesion 4- crawling 5- transmigration
(Kolaczkowska & Kubes, 2013)
It start by inflammatory mediators which attract neutrophils.
Tethering: at inflammation state P-selectin and E-selctin on endothelial cell is
increase which bond to PSGL-1 so neutrophil get slower to 5 um/s this case called
slow rolling (Futosi et al., 2013) (Kolaczkowska & Kubes, 2013)
Rolling: is made by bonds between surface molcules and neutrophil molecule, the
major molecule is selevtin and selectin family (Cugno et al., 2021) L-selectin on
neutrophil while rolling facilitate their secondry tethering
3-adhesion: he dissociation of a P-selectin–PSGL1 bond at the rear of the cell should
be balanced with the formation of another bond at the leading edge (Kolaczkowska
& Kubes, 2013)
CXCL8 signal via CXCR2 to activate neutrophils (Kolaczkowska & Kubes, 2013)
Neutrophil have heparan sulphate with negative charge which bond to chemokines
with positive charge, then activate G- coupeled protein which expressed integrins
that bind to ICAM receptors
Crawling: neutrophils cross several barriers consisting of: endothelium, basement
membrane and pericytes.
1- To cross endothelium via interaction between MAC1 and IMAC 1 then
partially digest basement membrane so it prefers to cross via low extracellular
matrix.
2- To migrate through pericytes, by interaction between ICAM-1 and MAC1
Transmigration: Transmigration requires integrins and CAMs (ICAM1, ICAM2 and
vascular cell adhesion protein 1 (VCAM1)) and other adhesion molecules,
ectoenzymes and others.
Transmigration have two methods: 1- paracellularly 2- transcellularly.
Paracellularly means: between endothelial and transcellularly means through
endothelial
NOTE: not all neutrophil recruitment requires selectins and/or integrins, as like
Neutrophils lacking selectin ligands and integrins still adhere to blood vessels in the
lungs (Kolaczkowska & Kubes, 2013)
Phagocytosis:
Many receptors activate phagocytosis, receptors send signal which enhance
phagosome vacule.
Fc receptors: The main Fc receptors of human resting neutrophils are FcγRIIA
(CD32) and FcγRIIIb (CD16) (Lee et al., 2003)
Receptor should distinguish between IgG ubiquitius and IgG associated with
particles.
“Phosphorylation of ITAM generates docking sites for proteins bearing SH2
domains, including the tyrosine kinase, Syk. Subsequent phosphorylation of Syk
leads to the recruitment of various signaling proteins to the activated FcyR complex.
Highlighting the importance of Syk in FcyR‐mediated phagocytosis, Jaumouillé et
al found that in macrophages, Syk regulated FcyR responsiveness by increasing
lateral receptor mobility and clustering through a reduction in actin polymerization”
(Naish et al., 2022)
Phagsome formation: NADPH oxidase generates ROS, these process is metabolic
process so neutrophil produce ROS from glycolysis. (Naish et al., 2022)
Granules play important role: delivery of antimicrobial granule proteins
Azurophil which fuse with phagosome.
Granules are heterogenous and can also be characterized by size, morphology,
electron density, or protein content.
Granules:
They are marker to transition from myeloblast to promyelocyte (Borregaard &
Cowland, 1997)
Are produced from golgi complex (Naish et al., 2022) they are three types: primary
which know by azurophilic which staining by azure A and azure B which know by
Romanowsky-Giesma stain. (Chistiakov et al., 2015) (Häger et al., 2010)
become blue- and purple-colored and it is peroxidase positive, Specific granules are
rich in antimicrobial substances, for example, lactoferrin and lipocalin‐2, and also
contain membrane proteins which are peroxidase negative. (Naish et al., 2022)
Granules type 3 which have rich in Gelatinases A and B (Chistiakov et al., 2015)
NETs
“leads to the death of neutrophils and is characterized by subsequent morphological
changes: disintegration of nuclear membrane, chromatin decondensation,
disappearance of plasma membrane, and finally the spill of DNA-based NETs into
the extracellular space” (Rada, 2019) how ever in some cases neutrophil release only
parts of it nucleus or DNA (Rada, 2019)
24 proteins were identified in NETs formed by stimulation of neutrophils with
phorbol 12-myristate 13-acetate (PMA) (Papayannopoulos, 2017) some researches
discuss that number of proteins depends on stimuli (Dwyer et al., 2014) NET
formation was most robust at 4 and 8 h and persisted beyond 24 h post infection
(Kolaczkowska et al., 2015) ROS, MPO and NADPH is dependent in NET (Rada,
2019) (Metzler et al., 2011)
Some examples of chemokines
IL-4: inhibitory effect on neutrophils. (Egholm et al., 2019)
IL-6 and IL-8: migration of neutrophil (Hashizume et al., 2011)
IL-12: promotes gamma interferon-dependent neutrophil recruitment in the lung
(Sun et al., 2007)
IL-15: promotes neutrophil cytoskeletal rearrangement by increasing actin
expression (Perera et al., 2012)
INFy: due to: (Gomez et al., 2015) Neutrophil does not produce INF-y how ever
there is a relation between INF-y and NETs
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