2. ENVIRONMENTAL ChEMISTRy & TEChNOLOGy 2.1. Lectures

Chem. Listy, 102, s265–s1311 (2008) Environmental Chemistry & Technology
L04 COMPLEx ESSESSMENT OF
ORGANOPhOSPhATES LOw DOSE
ChRONIC ExPOSuRE ON ENDOThELIuM,
MACROPhAGES, PLATELETS AND
ESTERASES
e. ERMOLAEVA, n. GOnCHAROV, A. RADILOV,
L. GLASHKInA, I. MInDUKSHEVa , P. AVDOnInb ,
I. DOBRYLKO and V. REMBOVSKIY
Research Institute of Hygiene, Occupational Pathology and
Human Ecology, Saint-Petersburg, Russia,
aI. M.Sechenov Institute of Evolutionary Physiology and Biochemistry,
Saint-Petersburg, Russia,
bN. K. Koltzov Institute of Developmental Biology, Moscow,
Russia,
nvgoncharov@mail.ru
Introduction
Morphofunctional disturbances of circulation in rat
embryos intoxicated by derivatives of organophosphates
(OP), a role of endothelium directly affected by OP under
acute intoxications 1,2 , and various data on the role of endothelium
in development of peripheral neuropathies of various
genesis 3,4,5 were the basis for consideration of endothelium
as one of the main targets under chronic intoxication with OP.
In this research work, an attempt has been undertaken to clarify
an impact of circulatory disturbances, the cellular component
of haemostasis, and neuropathy target esterase (nTE)
activity upon reaction of rat organism under intoxication with
low doses of OP.
Materials and Methods
In these experiments diisopropylfluorophosphate (DFP),
which can induce the delayed polyneuropathy, and paraoxon
s297
which has no such effect, were applied with drinking water
at doses 10 –2 mg kg –1 (1/100 LD 50 ) and 10 –4 mg kg –1 (1/10,000
LD 50 ). The intoxication was conducted daily 5 times in a week
for 3 months. Esterases of choline and non-choline substrate specificity
6,7,8,9 were studied; for rat brain, the enzyme activity was
calculated per mg of protein 10 . To investigate the OP effects upon
nADPH-oxidase system, we used the functional state of peritoneal
macrophages with fluorescent microscope and fluorescent
probe dichlorfluorescein diacetate (DCF) 11 . Kinetic parameters
of platelet aggregation were studied by low angle light scattering
technique 12 . The functional activity of endothelium under OP
intoxication was studied with rat aorta by method of endotheliumdependent
relaxation 13,14 . The results obtained were processed by
variation statistics and MS Exсel software.
Results and Discussion
Comparing the level of inhibition of esterases in blood
plasma after 3 months’ intoxication with the OP, we have
found that the residual activity of nTE was much lower than
that of red blood cells’ acetylcholine esterase (AChE). Inhibition
of nTE activity in rat brain 2 months after stopping
intoxication with DFP gives an indirect evidence for development
of neurotoxic effects. Activity of esterases after chronic
intoxication demonstrates a switch of inhibition from one
enzyme to another, embracing both choline specific and nonspecific
esterases. It is noticeable that the most prolonged
neurotoxic effect (20–30 % reduction of nTE activity after
6 months’ period of the post-intoxication recovery) remained
in the platelet-rich plasma (PRP).
The next method demonstrating its efficacy for diagnostics
of the low dose intoxication with OP was estimation
of the level of generation of reactive oxygen species
(ROS) during “respiratory burst” by professional phagocytes
(macrophages). It is a widely known fact that macrophages
Fig. 1. Generation of ROS by mice macrophages after 3 months’ chronic intoxication and 2 and 4 months after cessation of the intoxication
with paraoxon and DFP at doses 10 –4 mg kg –1 (min) and 10 –2 mg kg –1 (max)