Clinical Biochemistry of Domestic Animals (Sixth Edition) - UMK ...

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Chapter | 11 Neutrophil Function
are present within the lung lesions but not within intestinal
ulcers or lesions in other tissues. Therefore, neutrophils
can enter the lung by integrin-independent processes
(Ackermann et al. , 1996 ).
Canine leukocyte adhesion deficiency (CLAD) occurs
in Irish setter and Irish setter-cross-bred dogs ( Giger
et al. , 1987 ; Gu et al. , 2004 ; Kijas et al. , 1999 ; Renshaw
et al. , 1975 ). As with BLAD, the molecular basis of CLAD
is due to a single point mutation; however, in the dog, a
cysteine at amino acid 36 is substituted for a serine. This
cysteine is a highly conserved residue in the extracellular
domain of CD18, and the structural defect from the
mutation results in the failure to express the CD11/CD18
complex on the leukocyte surface. Affected dogs develop
severe bacterial infections shortly after birth and usually
die or are euthanized.
More recently, variant forms of LAD-1 have been
reported in humans in which CD18 integrins are expressed
on the surface of leukocytes at normal levels but fail to
support their adhesion ( Wehrle-Haller and Imhof, 2003 ).
In one case, LAD-1 was caused by a point mutation in the
β 2 -integrin, rendering it nonresponsive to inside-out activation
of leukocytes ( Hogg et al. , 1999 ). In other cases, the
leukocyte adhesion deficiency was extended to various
integrin families as well, including β 1 -, β 2 -, and β 3 -integrin
function. Interestingly, these latter cases result from a failure
by intracellular factor(s) or signaling pathway(s) essential
for integrin activation/clustering. Under physiological
shear stress, these LAD-affected leukocytes are unable to
generate firm adhesion and arrest.
Another human adhesion deficiency (LAD-2) was first
described in 1992 in two children of Arab origin ( Etzioni
et al. , 1992 ). LAD-2 is a rare congenital disease resulting
in mutations in a GDP-fucose transporter and a generalized
fucosylation defect that affects α1,2-, α1,3-, α1,4-, and
α 1,6-linkages of fucose in glycoproteins and glycolipids
( Wild et al. , 2002 ). Among other effects, LAD-2 neutrophils
demonstrate strongly reduced sLe X expression, which
diminishes their adhesivity with the selectin family members
and their accumulation on endothelium ( Fig. 11-2 ).
Thus, this disorder disrupts leukocyte extravasation at an
earlier stage than LAD-1.
B . Phagocytosis
The process of phagocytosis has been extensively studied in
macrophages but less well studied in neutrophils. Therefore,
we will present a general overview of phagocytosis based
on knowledge gained primarily from macrophages. Phagocytic
cells ingest bacteria and other particulate material by
receptor-mediated phagocytosis ( Peyron et al. , 2001 ; Stossel,
1974 ). This process forms a vesicle around the organism
and internalizes it ( Fig. 11-3 ). After internalization,
the vesicle containing the organism undergoes a stepwise
process of maturation in which the vesicle fuses with early
Receptor binding
Phagolysosome
Phagocytosis
Late endosome
Rab 7
Lysosome
Lamp-1
Late Phagosome
Early Phagosome
Early endosome
Fusion
Rab 5
FIGURE 11-3 Receptor-mediated phagocytosis of organisms by leukocytes.
After binding of organisms to cell membrane receptors, organisms
are endocytosed into a phagosome. After fusion with early endosomes, the
early phagosome is characterized by the presence of Rab5. Acidification
of the phagosome occurs at this stage. Thereafter, fusion with late endosomes
results in acquisition of Rab7 and shedding of Rab5. The late
phagosome then fuses with lysosomes, forming the phagolysosome that is
characterized by the presence of Rab7 and Lamp-1.
and late endosomes and lysosomes to create a phagolysosome.
Within the phagolysosome, lysosomal enzymes and
reactive oxygen radicals digest organisms. The process of
phagolysosome fusion and organism killing is complex,
and the details are still being investigated. Fusion of early
endosomes coincides with initiation of vesicle acidification.
This phase is marked by the presence of high levels of the
GTP-binding protein Rab5 on the phagosome membrane.
It is not clear whether Rab5 is brought to phagosome by
the early endosome or is acquired from the cytoplasm ( Via
et al. , 1997 ). Fusion of the phagosome with late endosomes
is marked by acquisition of Rab7 and loss of Rab5.
Subsequent fusion of lysosomes to the late phagosome is
characterized by acquisition of lysosome-associated membrane
protein (LAMP)-1 and LAMP-2.
Neutrophils contain primary, secondary, and, in some
species, tertiary granules in addition to lysosomes ( Stossel,
1974 ). Primary granules contain oxygen-dependent killing
mechanisms and a variety of cationic proteins and bactericidal/permeability-increasing
(BPI) protein ( Klebanoff and
Clark, 1978 ). Secondary granules contain most of the lysozyme
and lactoferrin. Secondary granules appear to fuse first
followed by primary granules ( Klebanoff and Clark, 1978 ).
Phagosome-to-endosome fusion is a highly controlled
process. The fusion system is typically composed of specific
receptor-mediated processes. An ATPase N-ethyl-maleimidesensitive
factor (NSF) and α soluble NSF attachment
protein ( α SNAP) interact with vesicle and target integral
membrane SNAP receptors (v- and t-SNAREs) that serve
as docking and fusion devices ( Vieira et al. , 2002 ).
v- and t-SNAREs form complexes spontaneously, and NSF
and SNAP provide the energy necessary to separate