p-nonylphenol(NP)stimulated proliferativeresponses and cy- tokine productionsof murine spleen cells and thymus cells invitro . lnthispaper we report the effect.
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of ofHealth Health Sciences Sciences
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(l):1-10 C2oo5)
UOEH27
(Original]
Endocrine Disrupters
Effeet
on
Macrophage
Functions In
V)itroof
Uki YAMASHITA, Tsutomu SuGIuRA, Yasuhiro YosHDA and EtsushiKuRoDA Dqpartment qf immunology,Schoot ofMedicine, UniversityofOccupationaland Environmentat Health, Jdpan.
}lahatanishi-ku, Kitakyushu 807-8555, Jttpan
(EDs)such as diethylstilAbstract: In previouspaperswe reported that endocrine disrupters A : BPA) bis(2-ethylhexyl) besterol(DES), 4,4'-isopropylidene diphenol(Bisphenol ,
responses proliferative
stimulated and p-nonylphenol(NP) phthalate(EHP) and thymus of murine spleen cells tokine productions we
report
the effect
of
EDs
on murine
macrophage
and cy-
in vitro . ln thispaper functionsin vitro, DES,
cells
exudate cells NP stimulated thioglycolate-induced peritoneal IL-6,IL-12, tumor (IL)-1, (macrophages)to produce cytokines such as interleukin and macrophage chemotactic necrosis factor protein-1. The effect of theseEDs was response of 1O'fi-1O'8 M, and the maxirnum was observed at the concentrations M. The accessory functionforT cell activation and CD86 exobtained at 10'7 in,EPsfor accessory function,was enhan.ced pression,an importantmolecule acuvmes These results suggest that EDs havemodulatmg treated macrophages. functions. of macrophage mouse. cytokine, accessory function, macrophage, Keywords : endocrine disrupter, 23 August 2004, accepted 30 September2004 ) (Received
BPA, EHP
and
Introduction used fbr compounds to show that many kindsof chemical Evidence has been accumulated estrogen or for making plasticarticles have an estrogen-1ike activity which mimics pesticide are called and may haveadverse effects on human beings. Thus, these compounds functions estrogens environmental [1-6].Their effects on mammals endocrine disrupters(EDs)or systems in rats and mice [7r and reproductive have been mainly studied using endocrine
have been suggested to also work on many other organs. 11]. However, these compounds The effect of EDs on the immune system has not been extensively studied, although immuno[12-17]. in preactivities of EDs havebeen suggested by some investigators modulating 4, 4'-isopropylidene stilbesterol(DES), vious papers we reported thatEDs such as diethyl diphenol(BisphenolA : BPA), bis(2-ethylhexyl)phthalate(EHP)and p-nonylphenol(NP) stimulated
responses proliferative
and
cytokine
productionsof
murine
spleen
cells
and thy-
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Disruptersen Macrophage of Endocrine Effect
'
/j',`ih'.C,ZM,,9.ke Mg.C6's,.l\1.i`hM./r･g.r,Ya CBeckman /r. :.k ]\illl: :?s : a {l#lr,i i, li,laY g:.2illfn,I gga g: "B'
e3
l
by a flowcytorneter cells were detected FITC-positive S
P
'
"a,r, Ogi:ib
S
C t'
a 0i;,Z,/h
M
"gn;9.:.aA,{e
5X
e
FL, Hialea, Coulter, h U b at'
$
c,
,Ze
,"ge]r ,r ,,Afl ,,hlouW
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,lh,
,:,i.2
l i i# $ s k $ g x $ iil,i i i 3 i i 1 /:.i. eii[E I' lk I, kii.i i,lli,ii lil lilr /liS li i/d :i ll. il,, lifi ik/Wts1 i/{ts S. pa. :f g:lhg
as
% positivecells.
of
l, ffe
'
%m
p
m'
?m
)
a "k
.
,.
fo
.Illlil
.:3
,
ef radioactivity amounts harvester(AbeKagaku Co.,Chiba, Japan). The spectrophotome!er intoDNA inthe cells were measured with a liquidscintillation corporated " da h P Th C ' k P ' 2` ' ' '' ' hs m ' y ' ' 'w c/k ',/ ' 'o 'il o
automated
cell
z･
-
" ,/J ,IS
'k' ,g
･
,i,
ei
m-
'
r
,/i ,il[a ,ll ,i,ilii
,l,
,..l･i
,./
by ELISA.
analysis Statistical
.il)kLe,eerigl,:･ks,,w.irEhle,ee,,a･ge.d,ztJ.ell'lltrmthr,zes,`yilfi,s･,ftI.l < O.05 was considered significant [25]. Results Dose response ofEDson IL-1 productionby macrophages were cultured with Peritonealexudate macroPhages was measured hours,and IL-1 in the culture supematant
.
several concentrations
by ELISA.
As
shown
of
EDs fbr in Table241,
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EDs
U YAMASHrrA
et at
DES, BPA, EHP and NP stimulated macrophages 10"-10'S M. The maximum response of EDs, of 10'7 M. Inthe fo11owingexperiments we
to produceIL-1at the concen-
such as
trations of concentration
EOiectofEDs on
EHP,
except
was
observed
at the
10"M EDs.
used
productionsby macrophages We then studied the effect of EDs on other cytokine productionsby macrophages. As shown in Table 2,these EDs stimulated macrophages to produce not only IL-1,butalso IL6,IL-12,TNF and MCP-1. cytokine
Table 1.Effeet
of
endocrine
disrupters on IL-1production by macrophages
IL-1production by
macrophages(ng/ml)
cultured
Reagents
with
10-6M
o
10-7M
10-SM
PBSDESBPAEHPmo 16 ± 3
Peritoneal exudate
45
± 4*
230± 14*
35 ± 2*
95
± 2*
180± 4*
31 ± 2*
750 ± 2*
340± 41*
80 ± 18*
25 ± 8
240 ± 4*
30 ± 4*
macrophages(5xl05
t'ml)
were
cultured
with
each
reagent(104-
10'8M) for 24 hours, and the amounts of IL-1orin the culture supernatants were as" indicates sayedby ELISA. significantly enhanced from PBS group
