Alterations of hematological variations in rats exposed to extremely low frequency magnetic fields (50 Hz).

Abstract

The aim of this study was to evaluate the possible effects of in vivo exposure to extremely low frequency electromagnetic fields (ELF-EMF) on whole blood parameters (hematological parameters) in rats.
Forty eight female Wistar rats, obtained from the Medical Science Application and Research Center, Dicle University, Turkey in 2004 were divided into four separate groups: two exposed groups (0.97 mT, 50 and 100 days, 3h/day) and two controls (sham).
Eosinophil, hemoglobin and MPV levels significantly decreased in rats that were exposed to EMF for 50 days. When the data for rats exposed for 50 days and 100 days were compared, it was found that MPV levels in rats exposed for 100 days were significantly lower. There was no significant difference in total leukocyte, neutrofil, lymphocyte, monocyte, eosinophil and basophil counts, or in erythrocyte, Hct, MCH, MCHC, RDW, PLT and PDW levels between the exposed and sham-exposed groups. ELF-EMF exposure had no effect on body weight. Also, liver weight did not show any significant difference between groups.
Our results indicate that the applied ELF-EMF exposure may induce slight but statistically significant alterations in some hematological parameters of rats, within the physiological range.

Full text

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ORIGINAL ARTICLE
Alterations of Hematological Variations in Rats Exposed to Extremely
Low Frequency Magnetic Fields (50Hz)
Dilek Ulker Cakir,aBeran Yokus,bMehmet Zulkuf Akdag,cCemil Sert,dand Nuriye Metee
aDepartment of Clinical Biochemistry, Faculty of Medicine, University of OnsekizMart, Canakkale, Turkey
bDepartment of Biochemistry, Faculty of Veterinary Medicine,cDepartment of Biophysics,eDepartment of Biochemistry,
Faculty of Medicine, Dicle University, Diyarbakir, Turkey
dDepartment of Biophysics, Faculty of Medicine, University of Harran, Sanliurfa, Turkey
Received for publication August 23, 2008; accepted June 19, 2009 (ARCMED-D-08-00377).
Background and Aims. The aim of this study was to evaluate the possible effects of in
vivo exposure to extremely low frequency electromagnetic fields (ELF-EMF) on whole
blood parameters (hematological parameters) in rats.
Methods. Forty eight female Wistar rats, obtained from the Medical Science Application
and Research Center, Dicle University, Turkey in 2004 were divided into four separate
groups: two exposed groups (0.97 mT, 50 and 100 days, 3 h/day) and two controls (sham).
Results. Eosinophil, hemoglobin and MPV levels significantly decreased in rats that were
exposed to EMF for 50 days. When the data for rats exposed for 50 days and 100 days
were compared, it was found that MPV levels in rats exposed for 100 days were
significantly lower.
There was no significant difference in total leukocyte, neutrofil, lymphocyte, monocyte,
eosinophil and basophil counts, or in erythrocyte, Hct, MCH, MCHC, RDW, PLT and
PDW levels between the exposed and sham-exposed groups. ELF-EMF exposure had
no effect on body weight. Also, liver weight did not show any significant difference
between groups.
Conclusions. Our results indicate that the applied ELF-EMF exposure may induce slight
but statistically significant alterations in some hematological parameters of rats, within
the physiological range.
? 2009 IMSS. Published by Elsevier Inc.
Key Words: Blood cell, Electromagnetic field, Hemoglobin hematology, Platelet, Leukocyte.
Introduction
In modern society, the use of electricity is so widespread
that it is impossible to avoid exposure to magnetic fields
(MF) including extremely low frequency electromagnetic
fields (ELF-EMF) produced by power lines and many kinds
of electrical appliances (1e4).
Over the past two decades, there has been growing public
and scientific interest inpossible health risks associated with
exposure to ELF-EMF (!200e300 Hz), which may affect
biological systems and a number of cell functions (1e5).
The results of many different types of animal and human
studies dealing with the biological effects of exposure to low
frequency electromagnetic fields (EMFs) have consistently
been both positive and negative (6). Some of these studies
seem to show a relationship between exposure to power-
frequency (50e60 Hz) magnetic fields and subtle bioeffects
(7,8). However, other studies have not shown such a link (9).
ELF-EMF do not have enough photon energy to break
macromolecular bonds (8). One of the interaction mecha-
nisms of magnetic fields with biological systems is that
magnetic fields alter the spin states of reactive oxygen
species (ROS) (10) which, in turn, change the relative prob-
abilities of recombination and other interactions, with
possible biological consequences (10,11). If this is the case,
the proportion of radicals reacting with macromolecules
will increase, leading to possible adverse effects on cell
Address reprint requests to: Dr. Dilek Ulker Cakir, Departments of
Clinical Biochemistry, Faculty of Medicine, University of OnsekizMart,
Canakkale, Phone: þ900286 2180018; FAX: þ900286 2189303; GSM
þ90533 4817490;
yahoo.com
E-mail addresses: ducakir@comu.edu.tr or ducakir@
0188-4409/09 $esee front matter. Copyright ? 2009 IMSS. Published by Elsevier Inc.
doi: 10.1016/j.arcmed.2009.07.001
Archives of Medical Research 40 (2009) 352e356

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function (11). At the cellular level, lipids, proteins, carbo-
hydrates, and nucleic acids may be damaged by reactions
with ROS. Moreover, the increase in the concentration of
ROS may give rise to functional and morphological distur-
bances in the cell through oxidative stress, leading to
reversible or irreversible tissue injury, e.g., lipid peroxida-
tion and cell membrane and DNA damage (12e14,31).
Reports from in vitro research indicate that low-level
EMF may alter membrane structural and functional proper-
ties and trigger cellular responses. EMF can penetrate the
cell membrane and possibly influence cytoplasmic structure
and function. In the presence of a resonant field, migrating
cell length grows from 10 to |40 mm with greatly exagger-
ated neutrophil extension. It should be noted that this
change occurred only under very specific conditions and
may not be applicable to in vivo situations (15).
Exposure to extremely frequency electromagnetic field
(ELF-EMF) (50 Hz, 0.97 mT) may have subtle bioeffects
that could affect health, and ELF-EMF may also affect
the immune and hematological systems through these
mechanisms. Results obtained in studies on the effects of
ELF-EMF on hematological parameters are conflicting,
and this led us to carry out the present study. The aim of
the study was to evaluate the possible effects of in vivo
exposure to ELF-EMF on the hematological parameters
of whole blood in rats.
The magnetic field strength used in the present study is
within the limits contained in occupational and public envi-
ronment MF exposure guideline standards and it exists in
both public and occupational environments (16). MF inten-
sity in this study is within the range of MF emitted from
some electrical appliances (17,18).
Materials and Methods
The experiments were performed on 48 female Wistar rats
obtained from the Medical Science Application and
Research Center of Dicle University in Turkey. They were
aged 2 months at the beginning of the study, weighed
183e203 g, and were fed with standard pelleted food
(TAVASInc.Adana,Turkey).Theratsweredividedintofour
groups of 12: two control groups (sham) and two experi-
mental groups. The experimental groups were exposed to
0.97 mT ELF in methacrylate boxes (17 ? 17 ? 25 cm).
The first group (n 5 12) was exposed to EMF for 50 days
and the second group (n 5 12) for 100 days, for 3 h/day.
The third (n 5 12) and fourth (n 5 12) groups were sham
groups that were treated like the experimental group except
for ELF-EMF exposure (corresponding to the first and
second groups, respectively). ELF magnetic field exposures
were done during the light period. The experiments for
each group were performed at the same time (for 50 days
exposuregroupfrom09:00e12:00(AM)and100daysexpo-
suregroup 13:00e16:00 (PM) insimilar locations within the
laboratory. The animals were kept in a 14/10 h light/dark
environment at a constant temperature of 22 ?3?C,
with 45 ? 10% humidity. This protocol was approved by
the local ethics committee.
The MF was generated in a device designed by our
group that had two pairs of Helmholtz coils of 25 cm in
diameter. This magnet was constructed by winding 225
turns of insulated soft copper wire with a diameter of
1.0 mm. One pair of Helmholtz coils was placed vertically
and another pair of coils was placed horizontally. The
distance between coils was 25 cm. An AC current produced
by an AC power supply (DAYM, Turkey) was passed
through the device. The current in the wires of the ener-
gized exposure solenoid was 0.65 A, which resulted in
a 50-Hz MF. MF intensity was measured from 15 different
points in a methacrylate cage by a person who was not
involved in the animal experiment using a digital teslameter
(209101074, Phywe, Goettingen, Germany); the mean was
0.97 ? 0.136 mT. Notemperature differences were observed
between exposure and sham coils during the exposure.
Ambient fields varied from 0.1 to 0.2 mT (1e2 mG). We
measured ambient fields with a cell sensor (Cell Sensor
EMF detection meter, Tec HealthCorp), with a sensitivity
0.1e5 mT (1e50 mG). Animals were euthanized with keta-
lar (50 mg/kg) after the last exposure and blood was drawn
into sterile syringes containing EDTA. Blood was obtained
during the light period. After slaughtering, the liver was
removed and weighed.
The rats were slaughtered after the exposure periods, and
leukocyte formulae (neutrophil, lymphocyte, monocyte,
eosinophil, basophil), erythrocyte indexes (hemoglobin,
hematocrit, MCV, MCH, MCHC, RDW), and platelet
indexes (MPV, PCT, PDW) were determined in whole blood
samples. These analyses were carried out with an Abbott
Cell-DYN 3700 M.A.P.S.S. Laser Differential machine by
impedance (coulter) and ‘‘optic laser scatter’’ methods.
Statistical analysis was performed with SPSS 8.0
package (SPSS Inc., Chicago, IL). The analysis of differ-
ences between exposed groups and sham groups for each
variable on given days was performed using independent
sample t test.
Results
Dataconcerningbloodcountparameterlevelsinshamgroups
andgroupsexposedfor50and100daysareshowninTable1.
Results of the analysis are given as mean ? standard devia-
tion. Also shown in the same tables are statistical differences
determined according to exposure periods and groups.
Eosinophil, hemoglobin and MPV levels in rats exposed
to MF for 50 days decreased compared to the sham group
(p!0.001, p !0.01 and p !0.01 respectively); however,
we did not find a difference in the group exposed for 100
days. Although it did not reach statistical significance, there
353
Electromagnetic Field and Blood Parameters

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was a tendency to decrease total leukocyte count and
a decrease in relative neutrophil, lymphocyte, monocyte
and basophil counts in both exposed groups compared with
the sham groups. Mean corpuscular volume (MCV) levels in
rats exposed to EMF for 100 days decreased compared to
the group exposed for 50 days (p!0.05). ELF-EMF expo-
sure had no effect on body weight. Also, liver weights did
not show any significant difference between groups
(Table 2). No significant differences in the other investigated
variables were found between controls and exposed rats.
Discussion
The major findings were that ELF-EMF exposure induced
a decrease in eosinophil, Hb and MPV levels. Similarly,
Seto et al. (8) demonstrated that a high intensity (80 kV/m)
60-Hz electric field decreased eosinophil counts. They also
recorded decreased white cell counts, although red cell
parameters did not differ significantly.
Hemoglobin concentrations clearly decreased in rats that
were exposed to EMF for 50 days, and this was found to be
statistically significant. Lino (19) stated that Hb is sensitive
to EMF. Furthermore, other studies have shown that eryth-
rocytes orient with the applied MF (20). It is hypothesized
that these results are probably associated with the change in
the conformation of Hb under MF action (21).
Cabrales et al. (22) observed that hematocrit, Hb and
neutrophil levels decreased
compared to the control group after ELF-EMF exposure
(60 Hz, 0.11 mT, 6 months). The reason for the observed
hematological variability in response to ELF-EMF expo-
sure in rats was explained as spleen hyperfunction (23).
Spleen hyperfunction increases the rate of destruction of
red blood cells, leukocytes and platelets (24). By contrast,
Chater et al. (25) suggested that Hb levels increased after
subacute exposure to SMF.
Fiorani et al. indicated that 0.5 mT magnetic field had no
effect on intact RBC (10). Similar findings were observed
in this study: peripheral blood erythrocytes count was not
changed in exposed rats. In a study by Selmaoui et al., no
significant differences were observed between sham-
exposed (control) and exposed men for hemoglobin
concentration and total leukocytes and eosinophils (26).
Our investigation showed that long-term exposure to
ELF-EMF decreased the mean volume of thrombocytes
(MPV) in the group exposed for 50 days. Decreased
MPV can cause microthrombocyte states in the peripheral
blood. MPV decreases in thrombocytopenia of bone
marrow insufficiency. The observed tendency of a decrease
significantly(p !0.05)
Table 1. Blood count parameters of the exposed and sham-exposed groups at 50 and 100 days
Exposed groupsControl (sham) groups
(50 days)
2.24?1.14
0.4 ?0.35
1.56?0.89
0.12?0.09
0.06?0.03a
0.09 ?0.13
6.76 ?1.9
13.45?2.031
59.08?16.15
87.68?2.05a
22.3?9.74
25.371?10.67
15.5?0.99
703.2?303.6
8.84?0.812
0.66?0.25
18.53?1.53
(100 days)
2.49?1.38
0.43?0.11
1.55?1.08
0.2 ?0.11
0.27?0.35
0.05 ?0.04
7.59 ?0.54
14.11?0.99
64.69?4.6
85.33?2.19a
18.62?0.47
21.82?0.37
16.33?1.39
525.36?227.59
9.02?1.219
0.52?0.09
20 ?2.48
(50 days)
3.78?2.98
0.41?0.23
2.83?2.98
0.24?0.23
0.13?0.081
0.09?0.07
7.77?0.27
14.9?0.961
67.69?2.42
87.05?2.02
19.2?1.37
21.73?1.91
16.65?1.83
672.75?327.48
10?0.882
0.78?0.25
19.72?1.73
(100 days)
3.02?2.02
0.58?0.45
1.56?1.33
0.44?0.49
0.35?0.35
0.14?0.13
7.34?1.27
14.04?1.16
63.08?10.96
85.97?2.02
19.76?3.85
22.99?4.39
16.79?1.68
503.3?247.14
8.54?1.29
0.5?0.17
19.63?1.86
WBC
Neu
Lym
Mono
Eos
Baso
RBC
HGB
HCT
MCV
MCH
MCHC
RDW
PLT
MPV
PCT
PDW
Note: All values are given as mean ?SD. Statistical comparisons were made with the independent sample t-test.
1,2Indicate significant differences with the corresponding periods of the other group (p!0.05, p!0.01, respectively).
aDifferences between mean values having the same superscript in each group is significant (p!0.05).
Table 2. Total body weight and liver weight of experimental and control groups
Weight (g)Exposed groupsControl (sham) groups
(50 days)
8.84?0.78
184.71?16.24
(100 days)
9.12?0.89
198.5?15.27
(50 days)
8.75?0.81
183.5?15.24
(100 days)
9.24?0.93
199.4?17.82
Liver
Total weight
354
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in leukocyte count and leukocyte index supports the idea of
bone marrow insufficiency. However, MPV did not
decrease in rats exposed to EMF for 100 days.
Furthermore, there was a tendency for a decrease in
white blood cells in the exposed groups, but this did not
reach statistical significance. Svedenstal et al. investigated
the effect of 50 Hz EMFs in CBA mice exposed to 8 mT
(20 days) generated by a 220 kV transmission line and
observed a decreasing number of mononuclear leukocytes
after EMF exposure (9). Differently, Oroza et al. (27)
exposed rats to a magnetic field for 1, 2 and 4 weeks; only
the 4-week exposure induced an increase in leukocyte
count. Also, Osbakken et al. exposed mice to a 1.89 T
DC magnetic field but observed no consistent consequences
on their hematocrit or white blood cells counts (24).
In the present study, the decrease in MCV at 100 days
indicated the presence of hypochromic RBC in the form
of microcytes. This result is in disagreement with the data
reported by Elferchichi et al. (28) showing that SMF expo-
sure (128 mT, 1 h/day, 5 days) induced an increase in MCV
levels. Also, Mukewar and Baile (29) did not observe any
modification in MCV. Dasdag et al. suggest that ELF
electromagnetic fields do not affect the hematological and
immunological parameters of welders (30).
Lookingat the above-mentionedstudies asawhole, itcan
beseenthatsomestudiesreportedthatEMFcausedachange
in hematological parameters, whereas others reported no
change. The reason for these conflicting results probably
stems from differences in exposure setups, experimental
conditions such as an alternative or static MF, the frequency,
different intensities and duration time of MF, time of
recovery, investigation targets and assay methods (31).
The reason for the nonlinear time progression in the
results obtained (eosinophil, Hb and MCV) may be associ-
ated with physiological defense and/or hemopoetic system
adaptation.
Our data demonstrated that ELF-EMF has no influence
on body weight. Previous studies have shown similar
results. In a study where rats were exposed to EMF at an
intensity of 6.25 T for 8 h/day, body weights of experi-
mental and control groups showed no difference after 3
months (32). Similarly, Margonato et al. found no differ-
ence in the body weight of rats exposed to EMF (50 Hz,
5 T, 32 weeks) (33). In another study, rats were exposed
to EMF (9.4 T, 6 h/day for 10 weeks) without any change
in body weight in the experimental animals (34). Also,
Robertson et al. detected no significant differences in body
weight between exposed and unexposed (control) mice at
any time in the study, and the hematological parameters
were essentially unchanged (35). They reported that
although there were real changes, their magnitude was very
small and did not indicate serious biological effects.
The liver is a hematopoietic organ, and any anatomic
changes within this organ, including weight, cause alter-
ations in hematological parameters. In the present study,
no statistically significant difference in liver weight was
found between experimental and control groups. In the
same way, no difference in liver weight was observed in rats
exposed to EMF for a longer period (10 weeks, 3 months,
12 months) or at a greater intensity (32,34). However,
Boorman et al. (36) found that of rats exposed to EMF
for 8 weeks, only females showed an increase in liver
weight compared to the control group.
Although past investigations have suggested that ELF-
MF induces tissue injury and macromolecule damage such
as strand lipid peroxidation and oxidative DNA damages,
the identity of specific lesions responsible for these biolog-
ical effects of ELF-MF remains elusive. Some investigators
have reported that ROS may be active in the mechanisms of
ELF-EMF effects (12,14,31). If cellular or plasma ROS
levels are influenced by exposure to ELF electromagnetic
fields, the levels of oxidative DNA damage and lipid
peroxide in some tissues would be altered as well (37). It
has been found that electric and magnetic fields can influ-
ence lipid peroxidation under certain experimental condi-
tions (12e14). In a study by Aksen et al. it was reported
that malondialdehyde (MDA) concentration of the ovaries
and uterus increased in rats exposed to ELF magnetic field
(same exposure regime in the present study) (14). Higher
plasma TBARS levels were also detected in rats exposed
to ELF magnetic field for 50 and 100 days in another study
(31). These experiments indicate that exposure to magnetic
fields of a few mT results in an enhancement of lipid perox-
idation, although the relevance of this phenomenon and the
thresholds for its occurrence remain unknown (37). These
findings could explain the present results, which hypothe-
sized that MF affects cell structure by means of ROS.
In conclusion, a relationship was found between expo-
sure to an ELF-electromagnetic field and a decrease in
some whole blood count parameters (0.97 mT 3 h/day for
50 consecutive days). Our results indicate that the EMF
exposure applied may induce slight but statistically signif-
icant alterations in certain hematological parameters in rats,
within the physiological range.
Nevertheless, our results should be confirmed by further
studies. Because of the differences in body size, geometry
and physiological responses, extrapolation of these results
to humans is not straightforward and any such comparison
should be made with great caution. According to these
findings, we believe that ELF-electromagnetic fields have
subtle bioeffects on hematological parameters.
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