BIOMEDICAL ENGINEERING VOLUME: 18 |NUMBER: 2 |2020 |JUNE
F equency Dependen Al e a ions o S. Ce e isiae
P oli e a ion Due o LF EMF Exposu e
Roman RADIL1, Jan BARABAS 1, Ladisla JANOUSEK 1, Ma in BERETA2,3
1Depa men o Elec omagne ic and Biomedical Enginee ing, Facul y o Elec ical Enginee ing and
In o ma ion Technology, Uni e si y o Zilina, Uni e zi na 1, 010 26 Zilina, Slo ak Republic
2Ins i u e o Measu emen Science, Slo ak Academy o Sciences,
Dub a ska ces a 9, 841 04 B a isla a, Slo ak Republic
3Depa men o Radiologic Technology, Facul y o Heal h, Ca holic Uni e si y in Ruzombe ok,
Names ie And eja Hlinku 1159, 034 01 Ruzombe ok, Slo ak Republic
[email p o ec ed], [email p o ec ed], ladisla[email p o ec ed], ume be e@sa ba.sk
DOI: 10.15598/aeee. 18i2.3461
Abs ac . The p esen ed pape deals wi h low e-
quency elec omagne ic ield applica ion on Saccha-
omyces ce e isiae cells. Expe imen s pe o med
h ough wide equency ange ha e shown selec i e e-
quency dependen biological esponse, which could be
success ully p edic ed by ion pa ame ic esonance he-
o y p oposed by V. V. Ledne . Al hough obse ed e-
sul s gi e sa is ying answe o ques ions whe he o no
elec omagne ic ields could a ec cell cul u es e en a
non- he mal le els, he esea ch p esen ed he ein opens
a mul i ude o ques ions abou he exac physical mech-
anisms unde lying he obse ed mic oo ganism beha -
io , as he heo y discussed wi hin he scope o his a -
icle is s ill no comple ely unambiguous.
Keywo ds
Elec omagne ism, low equency elec omag-
ne ic ields, p oli e a ion, cell memb ane, ion
pa ame ic esonance.
1. In oduc ion
I is well known ha he elec omagne ic condi ions
in which we li e nowadays a e di e en om hose in
which people used o li e in he pas . The majo i y
o li ing beings a e con on ed wi h a i icial Elec o-
Magne ic Fields (EMFs), which complemen he na u-
ally occu ing EMF o he Ea h, and ques ions pe -
aining o in luence he eo on biological sys ems a e
mo e equen han e e .
The numbe o expe imen al s udies in es iga ing
EMF e ec s on li ing cells, issues o on human heal h
in gene al, has g own apidly o e he las ew decades.
Much a en ion is cu en ly being paid o esea ch e-
ga ding po en ial connec ion be ween EMF exposu e
and cance o o he diseases, which has been dis-
cussed in a ious s udies [1], [2], [3], [4], [5], [6] and [7].
Nume ous wo ks ocusing on expe imen s wi h li ing
cells ha e shown di e ences in he p oli e a ion p o-
cess be ween cells exposed o EMF and con ol (non-
exposed) cells [8], [9], [10], [11], [12], [13] and [14].
O he wo ks in es iga e animal beha iou when ex-
posed o EMF [15] and [16] o possible genomic ins a-
bili ies due o EMF exposu e [17], [18], [19] and [20].
The esul s o hese wo ks a e o en inconsis en and,
as s a ed in Buchachenko [21], he biological elec o-
magne ic e ec s some imes seem o be i ep oducible
and con adic o y.
As poin ed ou in Ma ko [22], he p oblem is o en
discussed wi hin he scope o he mal e ec s wi h mi-
no in e es on possible non- he mal mechanisms, e en
when conside ing cu en ly accep ed s anda ds and
ecommenda ions (e.g. om In e na ional Commi ee
o Nonionizing Radia ion P o ec ion - ICNIRP, 1998).
Ac ually, he e alua ion o EMF e ec s on biological
objec s is much mo e complica ed due o p incipal di -
e ences in each app oach, and no pa o he p ob-
lem should be neglec ed, especially when Low F e-
quency EMF (LF EMF) and weak signals, ha a e a
below he ICNIRP ecommenda ions, a e conside ed.
The complexi y o his p oblem is documen ed by many
au ho s [23], [24], [25] and [26], who ied o p opose
physical mechanism o biological LF EMF impac , bu
he e is s ill lack o gene al accep ance o any p oposed
heo y ac oss he scien i ic communi y. This ac is a
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he same ime a s ong mo i a ional ac o o u he
in es iga ion in his ield o esea ch.
The Ion Pa ame ic Resonance (IPR) heo y, p o-
posed by Ledne [25], [26] and [27], is one o he mos
discussed models o LF EMF in e ac ion wi h bi-
ological objec s. The assump ions o his heo y
a e based on he Ion Cyclo on Resonance heo y
(ICR), i s desc ibed by Libo [24], specula ing ha
he physiological ac i i y o ce ain impo an ions
can be al e ed when he equency o applied ime-
a ying magne ic ield is equal o he equency o ion
mo ion in a s a ic magne ic ield. Ions in IPR
model a e ep esen ed by a ha monic oscilla o , bound
o a speci ic loca ion a he su ace o he cell mem-
b ane, and applica ion o combined magne ic ield
should al e i s oscilla ions. The men ioned physical
model included se e al impe ec ions, pa icula ly due
o he p oblem o he mal noise, and was c i icized
by Adai [28] and [29]. The heo y was de ended by
Engs om [30] and men ioned impe ec ions we e ad-
d essed by Ledne [26] and [31]. Despi e c i icism,
he e emains an imp essi e body o expe imen al e i-
dence ha can be aken as an empi ical basis o ICR
and IPR hypo heses [8], [9], [10], [11], [12] and [13].
In gene al, as Halgamuge e al. concluded [33]:
“. . . models based on elec ic in e ac ions ha e di icul-
ies o ob ain a high enough signal- o-noise a io a low
ield s eng hs. The ields ha a e needed o explain
he opening o a memb ane channel p o ein a e un e-
alis ically high. On he o he hand, models based on
magne ic in e ac ions can easily ob ain a la ge enough
signal- o-noise a io, because he le el o magne ic he -
mal noise in issue is low. The eason ha hese models
wo k, is he same eason ha makes he MRI echnique
possible.”
Ano he a emp o shed mo e ligh in o his e-
sea ch a ea is p esen ed wi hin his a icle. The au-
ho s op ed o expe imen ally in es iga e non- he mal
e ec s o LF EMF on Saccha omyces Ce e isiae cells,
a a ious uncommon equencies, o en neglec ed in
o he esea ch wo ks. To gain be e ep oducibili y
o conduc ed expe imen s, au ho s used hei inno a-
i e exposi ion sys em desc ibed in [33], p oposed wi h
he aim o achie ing he highes possible homogene-
i y o magne ic lux densi y wi hin he exposed ol-
ume, aking in o accoun sample dimensions and in-
cuba o p opo ions. This sys em enables expe imen s
wi h 3 pe i dishes (diame e o 9 cm, heigh o 2 cm
- each), each i adia ed by a homogeneous magne ic
ield. Ob ained esul s and obse ed biological eac-
ions a e hen discussed wi hin he scope o he IPR
heo y.
2. Ma e ials and Me hods
Saccha omyces ce e isiae cells (b and VIVO – com-
me cially a ailable om Lesa e Slo akia, Inc.) we e
selec ed as he a ge biological objec o i adia ion
by LF EMF. The eason o his choice is he g ow h
analogy wi h cance cells [34], as well as p e iously
published wo ks ega ding LF EMF e ec s on yeas
cells [35] and [36].
2.1. Expe imen al P o ocol
Chemical labo a o y es ing p o ocol o he men ioned
VIVO s and o S. Ce e isiae is publicly a ailable om
company Mik olab, LLC. This means ha he mic obi-
ological, physical and chemical p ope ies and pa am-
e e s o in es iga ed mic oo ganisms we e known p io
o he expe imen s.
Cells we e cul i a ed on GKCH aga , in acco -
dance wi h STN EN ISO 7218 (560104) and STN ISO
7954 (560087) s anda ds and speci ica ions. Cul i-
a ion was pe o med unde iden ical ambien condi-
ions o 72 hou s, wi h empe a u e o bo h exposed
and con ol samples se sligh ly abo e oom empe -
a u e, speci ically o 27 ◦C. Con inuous empe a u e
moni o ing was pe o med using h ee Nega i e Tem-
pe a u e Coe icien (NTC) he mis o s and p ocessed
and logged in eal- ime. Two o he NTC he mis o s
we e placed wi hin he incuba o chambe s o moni o
he empe a u e o bo h exposed and con ol samples,
and he hi d he mis o was placed ou side he in-
cuba o as a e e ence senso . To a oid any e ec s
om ai low, humidi y, o changes o ligh condi ions,
he exposed samples we e held in a solid plas ic holde ,
ma ching he diame e o he coil ca i y.
2.2. Expe imen al Se up and
E alua ion Me hods
The expe imen al se up, used o all o he p esen ed
expe imen s and de ailed in [37], is schema ically
shown in Fig. 1. The se up was speci ically designed
o pai ed sample expe imen s, conduc ed on wo se s
o h ee Pe i dishes inocula ed by S. Ce e isiae.
One se was placed wi hin one chambe o incu-
ba o , housing he mu-me al magne ic shielding box.
This se was he con ol, o e e ence sample - being
shielded om he gene a ed LF EMF. The second se
o samples, ep esen ing he exposed cells, was placed
a speci ic posi ion wi hin he al eady men ioned de-
eloped coil sys em, depic ed wi hin he eal scheme
o expe imen al se up on Fig. 2. The coil sys em was
d i en by ha monic cu en (gene a ed by he Agilen
33220A signal gene a o , manu ac u e Agilen Tech-
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nologies, Inc.), and ampli ied using a linea ampli ie
(Hube A1110-05, manu ac u e D . Hube GmbH)
- he esul ing ampli ude Imbeing in he 0.96–1.09 A
ange. The exac alue o exci a ion cu en depended
on he impedance ma ching and equency used, so as
o achie e he equi ed magni ude o magne ic lux den-
si y. The expe imen was epea ed o 5 imes a said
equency.
PC NI El is
AmG
MS
CS ES
Incuba o
A
Fig. 1: Expe imen al se up: G - signal gene a o Agilen
33220A; Am - ampli ie ; A - ampe e me e ; Incuba o
wi h wo chambe s: ES - exposed samples placed wi hin
he exci a ion coil, CS - con ol samples placed wi hin
he MS - magne ic shielding box; NI El is II - de el-
opmen boa d o empe a u e da a acquisi ion;
PC - empe a u e moni o ing and isualiza ion.
Fig. 2: P opopsed and cons uc ed expe imen al se up show-
ing he applica o used in p elimina y expe imen s and
he newly designed applica o .
F om he a o e men ioned i is e iden ha samples
we e i adia ed by LF EMF wi h magne ic lux densi y
~
B, which a ies spa ially wi hin he exposed olume.
The exac alue o ~
B- ield p oduced by he coil sys em
was alida ed by measu emen s, using sensi i e lux-
ga e senso s. Since his me hod is desc ibed in mo e
de ail in [33] and [38], o he pu pose o his a icle,
i is su icien o men ion he conclusion ha eal-li e
measu emen s e-con i med alues ob ained om nu-
me ical simula ions, documen ing minimum magne ic
lux a ia ions (max. 5 %) wi hin he i adia ed ol-
ume, when using he p oposed exci a ion coil sys em.
The alue o magne ic lux densi y a ied be ween
~
B=2.33–2.45 mT, and was equency independen .
A e con inuous i adia ion, he g ow h dynam-
ics pa ame e o bo h con ol and exposed samples
was quan i ied. Speci ically, g ow h dynamics we e
compu ed ia semi-au oma ed so wa e called Pe i-
Coun e , which was de eloped as a eplacemen o
comme cially a ailable coun e s. Algo i hm he eo is
de ailed in [39]. P incipally, i allows de ec ion and
coun ing o yeas g ow h a eas, using me hods o im-
age p ocessing, analysis, and ea u e ex ac ion, o en-
able expe imen al esul s quan i ica ion and compa i-
son o obse ed e ec s. S a is ical signi icance o ob-
se ed esul s was inally e alua ed using S uden ’s
T- es o pai ed samples, wi h signi icance le el
α= 0.05.
3. Resul s and Discussion
As was men ioned in p e ious sec ion o his a icle,
expe imen s we e ocused on he beha iou o Sac-
cha omyces ce e isiae cells in luenced by exogenous
LF EMF o di e en equencies p oduced by he de-
signed coil sys em. Ou wo k ollows p e iously con-
duc ed expe imen s published in [40], [41] and [42], bu
he equency ange di e s, and is wide in compa -
ison wi h p elimina y expe imen s. A i s , we e-
e alua ed he base equency o 1.6 kHz, which was ex-
pe imen ally p o en as s a is ically ele an o he in-
hibi o y e ec on S. Ce e isiae p oli e a ion. The i s
se o expe imen s was hen conduc ed wi h equen-
cies o 1.6, 0.8, and 0.4 kHz o e i y hypo hesis ega d-
ing e ec s o subha monic equencies and co ela ion
he eo wi h IPR heo y, p esen ed in [30]. Resul s
o hese expe imen s a e shown in Tab. 1.
Tab. 1: Compa ison be ween expe imen al esul s obse ed a
i adia ion signal equencies and IPR p edic ed e-
quencies a speci ied magne ic lux densi y a ge ing
Ca2+ ion.
Magne ic lux 2.39 2.39 2.39 2.39
densi y (mT)
F equency 1.600 0.800 0.400 0.200
gene a ed (kHz)
F equency 1.596 0.798 0.398 0.204
p edic ed (kHz)
G ow h a ea a io 0.59 0.54 0.64 0.80
(exposed/con ol)
F om he expe imen al esul s i is clea ha all sub-
ha monic equencies ha e shown inhibi o y e ec s and
s a is ically signi ican biological esponses a e p esen
a equencies 0.8 and 0.4 kHz, as well as he base e-
quency. These equencies co espond o he ion pa a-
me ic esonance equency o Ca2+ ions, and ha monic
componen s he eo in acco dance wi h Ledne ’s IPR
heo y om [25], [26] and [31] and he Eq. (1):
= c
n=1
n
·
q
2πm
·Bgen,(1)
whe e c ep esen s he cyclo on esonance equency
o he ion, qis he elemen a y cha ge o he ion, mis
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0.850
0.800
0.890
0.640
0.899
0.690
0.540
0.750
0.864
0.590
0.720
0.650
0.100 0.200 0.300 0.400 0.700 0.750 0.800 0.850 0.900 1.600 1.800 2.000
F equency (kHz)
0
0.2
0.4
0.6
0.8
1
1.2
G ow h a ea a io (exposed/con ol)
Fig. 3: G aphical in e p e a ion o p oli e a ion esponse o i adia ed samples wi hin he in es iga ed equency ange o applied
ime- a ying LF EMF.
mass o he ion and ndeno es he co esponding ha -
monic componen . Bgen is he mean alue o gene a ed
magne ic lux densi y, which in ou case was 2.39 mT.
P edic ed equencies o he biological esponse
based on his model a e also shown in Tab. 1. I is
e iden ha expe imen al equencies a which s a is-
ically signi ican biological e ec s we e obse ed a e
closely ela ed o hei heo e ically p edic ed coun e -
pa s.
Also appa en om he p esen ed se o expe i-
men s is ha he biological e ec ends o disappea a
lowe equencies, as was he case in expe imen s using
0.4 kHz. To e i y his obse a ion, we op ed o in es-
iga e a wide equency ange, wi h ocus on low e-
quencies and also on equencies a ound he expe imen-
ally p o en biologically ac i e equency o 0.8 kHz.
This se o expe imen s is p esen ed g aphically in
Fig. 3, o be e illus a ion and in e p e a ion he eo .
Obse ed esul s once again show inhibi o y e ec s.
Mo e in e es ingly, he e a e signs o pe iodici y wi hin
he expe imen al da a, wi h s ong peaks a p e iously
desc ibed esonance equencies o Ca2+ ha could be
p edic ed by IPR model. Fu he mo e, he p e ious as-
sump ion, ega ding he biological e ec ex inc ion a
lowe equencies, seemed o occu a equencies below
0.4 kHz.
Ano he in e es ing obse a ion conce ns he neigh-
bou hood o he equency 0.8 kHz, whe e he p oli e a-
i e esponse is weak in compa ison wi h he esonance
(base) equency, albei s ill p esen . This obse a ion
poin s o he window heo y, men ioned by Ma ko
in [22], bu could be also explained as a cumula i e e -
ec o o he ions, which play an impo an pa in cell
p oli e a ion p ocesses.
Since equency dependen biological esponse is e -
iden om he p esen ed esul s, we op ed o also
in es iga e he in luence o magne ic lux densi y ~
B
o he gene a ed LF EMF. Fo his pu pose, we chose
he equency o 0.8 kHz, whe e he biological esponse
seemed o be mos signi ican , and al e ed he alue
o ~
B o 50 %, 25 % and 10 % o i s o iginal alue.
These al e a ions we e echnically pe o med by am-
pli ude changes o e ec i e alue o d i ing cu en ,
which di ec ly in luences he ampli ude o ~
Bin he coil
ca i y and hus he exposed olume.
Resul s p esen ed in Fig. 4 show ha signi ican ly
s ong p oli e a i e esponse is p esen only wi h o igi-
nal magne ic lux densi y o 2.39 mT. The biological e-
sponse is weak o no longe obse ed a gi en equency
due o modi ied magne ic lux densi y o he applied
LF EMF.
F equency 800 Hz
2.39 1.195 0.5975 0.239
Bgen (mT)
0
0.2
0.4
0.6
0.8
1
1.2
G ow h a ea a io (exposed/con ol)
0.56
0.79
0.94 0.90
Fig. 4: G aph o p oli e a ion dependence o S. Ce e isiae on
Bgen, a he equency o 800 Hz.
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4. Conclusion
The p esen ed a icle discusses chosen heo ies ela ed
o LF EMF mechanisms o in e ac ion wi h biologi-
cal objec s, ep esen ed he ein by Saccha omyces ce e-
isiae cells. Repe i ion o p e iously conduc ed expe -
imen s published in [8], [38], [40], [41] and [42] con-
i med inhibi o y e ec on he p oli e a ion esponse
a he equency o 1.6 kHz. Fu he mo e, he expe -
imen al esul s clea ly demons a ed non- he mal e -
ec s o ex e nally applied ime- a ying LF EMF on
he p oli e a ion p ocess o cul i a ed cells. Ou ind-
ings ega ding he Ca2+ a e also in co ela ion wi h
esea ch p esen ed in [43]. The obse ed biological e-
sponses could be e e ed o as equency selec i e be-
cause s a is ically signi ican e ec s occu only a spe-
ci ic equencies, which poin s o he esonance cha ac-
e o he obse ed bio-e ec s.
The mechanism o ac ion in luencing he beha iou
o he i adia ed biological samples could be heo e i-
cally explained by he IPR heo y. This model success-
ully p edic s obse ed biologically ac i e equencies,
conside ed esonance equencies o Ca2+ ions and ha -
monic componen s he eo . F om his poin o iew, we
can d aw he conclusion ha Ca2+ ions play an im-
po an ole in cellula p oli e a ion p ocesses, and
could be exploi ed as a ge s o i adia ion by ex e -
nal LF EMF uned o pa ame ic esonance equency
he eo , which co esponds wi h indings p esen ed by
Belo a [27]. The ele ance o V. V. Ledne ’s heo y
can be also con i med by esul s o ou in es iga ion
o magne ic lux densi y changes wi h espec o p oli -
e a ion esponse a unchanged equency. This means
ha when expec ing biological esponse in acco dance
wi h he IPR heo y and Eq. (1), any change o ~
Bnec-
essa ily leads o change o he ion pa ame ic esonance
equency, bu no e e y change o equency mus lead
o change o magne ic lux densi y.
Despi e he indings p esen ed wi hin his a icle,
an unambiguous explana ion o EMF in luence on li -
ing s uc u es is s ill lacking. Thus, many ques ions a e
s ill open in his ield o esea ch and can only be an-
swe ed by g ea amoun o success ul and epe i i ely
ob ained expe imen al esul s om which conclusions
and heo ies can be d awn.
Acknowledgmen
This wo k was suppo ed by p ojec ITMS:
26210120021, co- unded om EU sou ces and Eu o-
pean Regional De elopmen Fund and by he Slo ak
Resea ch and De elopmen Agency unde he con ac
no. APVV-16-0190.
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Abou Au ho s
Roman RADIL was bo n in T encin, Slo ak
Republic, in 1984. G adua ed om he Facul y
o Elec ical Enginee ing, Uni e si y o Zilina, in
2008 om Biomedical Enginee ing and ecei ed
he Ph.D. deg ee in Theo e ical Elec omagne ics a
he same uni e si y, in 2012. A p esen he wo ks as
assis an p o esso a he Dep . o Elec omagne ic
and Biomedical Enginee ing, Facul y o Elec ical
Enginee ing and In o ma ion Technology, Uni e si y
o Zilina. His esea ch ac i i ies a e ocused on in es-
iga ion o LF EMF e ec s on biological samples and
biomedical image p ocessing. Resul s o his esea ch
ha e been published in se e al jou nals and p esen ed
in na ional and in e na ional con e ences.
Jan BARABAS was bo n in Ba dejo , Slo ak
Republic, in 1982. G adua ed om he Facul y
o Elec ical Enginee ing, Uni e si y o Zilina, in 2008
om Biomedical Enginee ing and ecei ed he Ph.D.
deg ee in Theo e ical Elec omagne ics a he same
uni e si y, in 2011. A p esen he wo ks as esea che
a he Depa men o Elec omagne ic and Biomedical
Enginee ing, Facul y o Elec ical Enginee ing and
In o ma ion Technology, Uni e si y o Zilina. His
cu en esea ch endea ou s include ongoing in es-
iga ion o elec omagne ic ields and in e ac ion
he eo wi h biological objec s, he indings o which
ha e been published in se e al jou nals and in e na-
ional con e ences. O he ac i i ies include biomedical
image p ocessing, signal p ocessing and p og amming.
Ladisla JANOUSEK was bo n in Michalo ce,
Slo ak Republic, in 1974. G adua ed om he Facul y
o Elec ical Enginee ing, Uni e si y o Zilina, in 1997
om Elec ic Powe Sys ems and ecei ed he Ph.D.
deg ee in Elec ic Machines and Appa a uses a
he same uni e si y, in 2002. A p esen he is head
o he Depa men o Elec omagne ic and Biomedical
Enginee ing, Facul y o Elec ical Enginee ing and In-
o ma ion Technology, Uni e si y o Zilina. The main
ields o his esea ch ac i i ies a e elec omagne ic
me hods o ma e ials’ non-des uc i e e alua ion and
non- he mal e ec s o EMF on li ing o ganisms. His
achie emen s ha e been acknowledged by se e al
p es igious awa ds.
Ma in BERETA was bo n in Ruzombe ok,
Slo ak Republic, in 1990. G adua ed om he Facul y
o Elec ical Enginee ing, Uni e si y o Zilina, in 2014
om Biomedical Enginee ing and ecei ed he Ph.D.
deg ee in Theo e ical Elec omagne ics a he same
uni e si y, in 2017. A p esen he wo ks as esea che
a he Ins i u e o Measu emen Science, Slo ak
Academy o Sciences in B a isla a. His esea ch
ac i i ies a e ocused on LF EMF biological e ec s
and moni o ing o ul a-weak pho on emission om
biological samples. Resul s o his esea ch ha e been
published in se e al jou nals and p esen ed in na ional
and in e na ional con e ences.
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