Respective role of polymorphonuclear leukocytes and their integrins (CD-huB) in the local or systemic toxicity of lipopolysaccharide. Hern#{225}nR. Chang,4.
Respective role of polymorphonuclear leukocytes and their integrins (CD-huB) in the local or systemic toxicity of lipopolysaccharide Hern#{225}nR. Chang,4 C. Vesin,t Malcolm Strath,t Jean-Claude *Department tDivision
of Genetics
and
of Parasitology,
Georges E. Grau,1 P. Pointaire,’ Pech#{232}re,*and Pierre-Francois
Microbiology
National
and
Institute
for
tDepartment Medical
of Pathology, Research,
Denis Piguett
of Geneva,
University
London,
Arsenijevic,5
United
Geneva,
and
Switzerland,
Kingdom
Abstract: The role of neutrophils (PMNs) and leukocyte integrins was investigated in two models of lipopolysaccharide (LPS)-induced toxicity: the systemic lethality assay in D-galactosamine-sensitized mice and the local reaction elicited by intradermal injection of LPS and tumor necrosis factor (TNF) at 24-h intervals. In the local reaction, depletion of PMNs with an anti-PMN mono-
earlier studies, using agents such as benzene, hydroxyurea, nitrogen mustards, or x-ray irradiation, have indicated that depletion of short-lived cells (principally PMNs and platelets) prevents expression of the local reaction [7].
clonal LFA1)
tion, the effect because there
antibody (mAb) and CD-lib (or
and CR3)
mAbs against completely
CD-ha prevented
hemorrhagic necrosis. Evaluation of histological and myeloperoxidase levels suggested different nism of protection because PMNs were abundant
(or the
sections mechain anti-
CD-liand absent in anti-PMN-treated mice. In the systemic assay, depletion of PMNs ensured 100% survival, whereas after administration of anti-CD-ha or b mAb, the percentages of survivors were 6 and 59%, respectively. One hour after LPS injection, the serum TNF-a level was higher in PMN-depleted mice than in controls. These studies provide evidence that neutrophils are essential for the expression of local or systemic LPS-induced injury, whereas the requirement for their leukocytic integrins is obvious 636-639;
only in 1993.
the
local
J.
reaction.
Leukoc.
Biol.
53:
In
the
using two
sion [8].
Words:
integrins
neutrophils
lipopolysacc/zaride
toxicity
INTRODUCTION Bacterial
lipopolysaccharides
(LPSs)
derived pressed
from gram-negative by a complex host
bacteria. response that
tion tumor [1-3].
of
cytokines necrosis A critical
are
toxic
compounds
Their toxicity involves the
that also have toxic factor a (TNF-a) and role of these cytokines
is exproduc-
properties, such as interleukin-1 (IL-I) in the expression of
LPS toxicity is indicated by the prevention of LPS-induced mortality by antagonists to TNF-a or IL-l. Thus, the toxicity of LPS is expressed first by the production of “toxic cytokines” (the so-called afferent limb) and subsequently by a complex response (the efferent limb), involving notably the complement and coagulation cascade, the production of metabolites
of
the
arachidonic
acid
pathway
and
various
The
Journal
of Leukocyte
Biology
Volume
53, June
investigated
or
systemic
cell
adhesion
results
MATERIALS
AND
injury.
molecules
induced
where
that
both cases, the requirement parent only for the local reaction.
PMNs
(mAb)
in
In
addi-
integrin mAbs was explored as a result of the overexpres-
endothelium,
indicate
of
role
antibody
LPS-induced
of antileukocytic is evidence that, to the
the
monoclonal
although
by
they
produce
PMNs
are
for the leukocytic and not the systemic
TNF-a,
damage essential
in
integrmns is apLPS-induced
METHODS
Mice Male BALB/c
CBA/J mice
mice (Bomholtgard, (Ifa Credo, France),
Denmark) 8 weeks old,
and were
female used in
experiments.
Monoclonal
antibodies
NIMP-R14, a rat immunoglobulin 02b (IgG2b) anti-mouse PMN mAb that produces selective in vivo PMN depletion, has been described [9]. This antibody recognizes a protein of 25-30 kd on the membrane of mouse PMNs (data not shown). 5C6, a rat anti-CDI1b (CR3, Mac-I) IgG2b mAb, and H35.89.9, a rat anti-CDlla (LFA-l) IgG2b mAb, have also
been
of
a mouse
described
[10,
Assay for mouse TNF-a
was
assay
using
In
as
this
work,
500
a control
was
tg
of
mAb
injected
or
i.p.
TNF-a
assayed a
11].
IgG
polyclonal
in
hamster
an
enzyme-linked
antimouse
immunosorbent
TNF
mAb
[12]
as
a first
cells
such as neutrophils (PMN5), platelets, and endothelial cells, as reviewed in ref. 4. Since Shwartzman, two models of LPS-induced injury have been explored: the systemic reaction, leading eventually to shock and death, and the local reaction, characterized by hemorrhagic necrosis (HN) [5]. In the local reaction, it is known that HN is preceded by sequestration of PMNs and platelets along the small blood vessels [6]. Furthermore,
636
local
the
stick
we
anti-PMN
of
of
PMNs
studies
specific
models
these
Key
present
a
1993
Abbreviations: IL-I,
HN,
interleukin-l;
MPO,
hemorrhagic LPS,
myeloperoxidase;
Reprint Medical HR. Medecine, Singapore
Received
PMN,
neutrophil;
IgG2b,
immunoglobulin
mAb, TNF-a,
G2b;
monoclonal tumor
antibody;
necrosis
factor
a.
F. Piguet, Department of Pathology, Centre University of Geneva, 1211 Geneva 4, Switzerland.
requests:
Pierre
Universitaire, Chang’s
necrosis;
lipopolysaccharide;
current
National
address: University
Department of
of
Singapore,
microbiology,
Kent
Lower
0511.
December
23,
1992;
accepted
February
11,
1993.
Faculty
Ridge
of
Road,
TABLE
I.
Effect
of Anti-PMN
and
Anti-CDI
la
or -CD!
lb
mAbs
Against
LPS-Induced
Mortality
Survivors/total Pretreatment
LPS”
Saline
Exp.
Anti-PMN
Anti-CDlla Anti-CD1Ib
“Mice
were
step, body
given
Exp.
10/10
-
IgG IgG IgG IgG
Normal
I
Exp.
2
-
-
5/5
-
1/5
0/6
1/17
2/6
4/5
10/17
6/6
-
-
+
-
-
4/6
followed by a rabbit and a biotinylated
antimouse TNF-a polyclonal goat antirabbit IgG.
anti-
CBA/J injection
mice of
0.5
of
ml
Local The
lethality received 0.5 mg of mAb 10 g of Escherichia coli
saline
containing
18 mg
IgG 055:B5
4 h before (Difco)
the i.p. LPS in
of D-galactosamine
phocytes control
was OP/7M/93L compared to 35P/9M/56L and the disappearance of circulating up to 5 days. Platelet counts were not After administration of anti-CDIIa the distribution of leukocytes was,
of BALB/c
mice
were
shaved
and
a priming
Hematological
Histological the
and
plexus
dose
of TNF-a
Mortality
of tissue
In PMN-depleted
survival
compared, Mann-Whitney
for
pg/mI
HN
lungs, or in paraffin
studies,
skins were or methyl
mice fixed with methacrylate.
were
of LPS, red was
mAb
(P
followed
24
discoloration completely
of the prevented
.01)
1). This
