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General
Proliferation / Differentiation / Apoptosis (Life / Growth / Death)
Orientation / Behavior
Calcium incorporation in cultured chondroblasts perturbed by an electromagnetic field.
Norton LA, Rovetti LA.
Department of Orthodontics, University of Connecticut Health Center, School of Dental Medicine, Farmington 06032.
We tested the hypothesis that electric perturbation influences 45Ca
incorporation in extracellular matrix (ECM) of cartilage in vitro.
Hypertrophic chondroblasts of tibial epiphyses (HC), sternum (SC), and
skin fibroblasts (F) were cultured from chick embryos. HC, SC, and F
cells were micromass seeded three times per week and maintained at 37.5
degrees C with 5% CO2 for two weeks. Cultures were randomly designated
control (C) or exposed (E) to a pulsed electromagnetic field (PEMF). A
time course experiment of calcium incorporation for all cultured groups
showed that 24 h of exposure produced the largest biological response in
chondroblasts. Calcium incorporation required supplemental phosphate.
Autoradiography data indicated that the calcium incorporation into
macromolecules largely occurred in the ECM. 45Ca steady-state
perturbation was enhanced by Streptomyces hyaluronidase (SH) but not by
testicular hyaluronidase (TH). 45Ca incorporation experiments tested the
effects of phosphate, SH, TH, and PEMF alone and in various
combinations on these cultures. Only PEMF or SH plus PEMF with phosphate
enhanced 45Ca incorporation. Other experiments examined the effect of
rotenone or freeze-thawing on cells exposed to PEMF. PEMF plus
freeze-thaw enhanced calcium incorporation in HC only. PEMF appeared to
cause disruption of the ECM, enhancing the probability of matrix
calcification.
J Orthop Res. 1988;6(4):559-66.
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Effect of pulsed electromagnetic fields (PEMF) on osteoblast-like cells. Alterations of intracellular Ca2+.
Satake T.
Department of Oral Biochemistry, Kanagawa Dental College.
Low-energy electromagnetic fields pulsed at frequencies of 60-90 Hz
significantly increase healing of chronic fracture nonunions in man.
These fields are effective at tissue current levels as low as several
orders of magnitude lower than those required for transmembrane
depolarization of normal cells. In this study, the effects of PEMF on
culture of rat osteoblast-like cells have been examined. The PEMF
promoted the growth of these cells, were also found to increase the
basal level of [Ca2+]i, and to decrease the responses towards epidermal
growth factor (EGF) and serum, when the degree of response was based on
the intracellular Ca2+ transient. These effects of PEMF were mimicked by
12-O-tetradecanoyl phorbol 13-acetate (TPA), a potent activator of
protein kinase C. Pretreatment of TPA enhanced the cell growth and
suppressed the intracellular Ca2+ transient induced with EGF and then
serum to about 170% of the control. Then, present study investigated how
the PEMF and TPA modulate EGF receptors of these cells. Both PEMF and
TPA decreased the level of EGF binding to these cells down to about 65%
and 75%, respectively. Scatchard analysis revealed the decrease of EGF
receptor without a significant change in the affinity for EGF by both.
In conclusion, it was indicated that PEMF acts at cell membrane and
modulates the receptors which is essential for cell growth and DNA
synthesis.
Kanagawa Shigaku. 1990 Mar;24(4):692-701.
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Membrane response to static magnetic fields: effect of exposure duration.
Rosen AD.
Department of Neurology, School of Medicine, State University of New York, Stony Brook 11794-8121.
The time-course for the reversible alteration in presynaptic membrane
function associated with exposure to a 123 mT static magnetic field was
examined in an attempt to help define the mechanism whereby these
fields influence biomembranes. Miniature endplate potentials (MEPPs)
were recorded in the isolated murine neuromuscular junction preparation,
maintained at a temperature of 35.5 degrees C. A minimum field duration
of 50 s was found to be necessary for MEPP inhibition, with the
efficacy of the field in inducing further inhibition being a function of
its duration, but only for periods up to 150 s. Longer durations were
not associated with additional inhibition. The time required for MEPP
frequency to return to baseline, following deactivation of the field,
was found to be linear for field durations up to 150 s. At and above
this limit, recovery time remained constant at 135 s. These findings are
consistent with the slow reorientation of diamagnetic molecular domains
within the membrane and suggest tight coupling to the mechanism
responsible for neurotransmitter release. The limits on this effect are
compatible with the mechanical constraints imposed by the membrane's
cytoskeleton.
Biochim Biophys Acta. 1993 Jun 5;1148(2):317-20.
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Resting potential of excitable neuroblastoma cells in weak magnetic fields.
Sonnier H, Kolomytkin OV, Marino AA.
Department of Orthopedic Surgery, LSU Medical Center in Shreveport, Louisiana 71130-3932, USA.
The mechanism by which static and low-frequency magnetic fields are
transduced into biological signals responsible for reported effects on
brain electrical activity is not yet ascertained. To test the hypothesis
that fields can cause a subthreshold change in the resting membrane
potential of excitable cells, we measured changes in transmembrane
current under voltage clamp produced in SH-SY5Y neuroblastoma cells,
using the patch-clamp method in the whole-cell configuration. In
separate experiments, cells were exposed to static fields of 1, 5, and
75 G, to time-varying fields of 1 and 5 G, and to combined static and
time-varying fields tuned for resonance of Na+, K+, Ca2+, or H+. To
increase sensitivity, measurements were made on cells connected by gap
junctions. For each cell, the effect of the field was evaluated on the
basis of 100 trials consisting of a 5-s exposure immediately followed by
a 5-s control period. In each experiment, the field had no discernible
effect on the transmembrane current in the vicinity of zero current (-
50 mV voltage clamp). The sensitivity of the measuring system was such
that we would have detected a current corresponding to a change in
membrane potential as small as 38 microV. Consequently, if sensitivity
of mammalian cells to magnetic fields is mediated by subthreshold
changes in membrane potential, as in sensory transduction of sound,
light, and other stimuli, then the ion channels responsible for the
putative changes are probably present only in specialized sensory
neurons or neuroepithelial cells. A change in transmembrane potential in
response to magnetic fields is not a general property of excitable
cells in culture.
Cell Mol Life Sci. 2000 Mar;57(3):514-20.
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Pulsed electromagnetic fields affect the intracellular calcium concentrations in human astrocytoma cells.
Pessina GP, Aldinucci C, Palmi M, Sgaragli G, Benocci A, Meini A, Pessina F.
Institute of General Physiology and Nutritional Science, University of Siena, Italy. pessina@unisi.it
Experiments assessed whether long term exposure to 50 Hz pulsed
electromagnetic fields with a peak magnetic field of 3 mT can alter the
dynamics of intracellular calcium in human astrocytoma U-373 MG cells.
Pretreatment of cells with 1.2 microM substance P significantly
increased the [Ca(2+)](i). The same effect was also observed when
[Ca(2+)](i) was evaluated in the presence of 20 mM caffeine. After
exposure to electromagnetic fields the basal [Ca(2+)](i) levels
increased significantly from 143 +/- 46 nM to 278 +/- 125 nM. The
increase was also evident after caffeine addition, but in cells treated
with substance P and substance P + caffeine we observed a [Ca(2+)](i)
decrease after exposure. When we substituted calcium-free medium for
normal medium immediately before the [Ca(2+)](i) measurements, the
[Ca(2+)](i) was similar to that measured in the presence of Ca(2+). In
this case, after EMFs exposure of cells treated with substance P, the
[Ca(2+)](i), measured without and with addition of caffeine, declined
from 824 +/- 425 to 38 +/- 13 nM and from 1369 +/- 700 to 11 +/- 4 nM,
respectively, indicating that electromagnetic fields act either on
intracellular Ca(2+) stores or on the plasma membrane. Moreover the
electromagnetic fields that affected [Ca(2+)](i) did not cause cell
proliferation or cell death and the proliferation indexes remained
unchanged after exposure. Copyright 2001 Wiley-Liss, Inc.
Bioelectromagnetics. 2001 Oct;22(7):503-10.
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The effect of strong static magnetic field on lymphocytes.
Aldinucci C, Garcia JB, Palmi M, Sgaragli G, Benocci A, Meini A, Pessina F, Rossi C, Bonechi C, Pessina GP.
Department of Physiology, University of Siena, Siena, Italy.
We investigated whether static electromagnetic fields (EMFs) at a
flux density of 4.75 T, generated by an NMR apparatus (NMRF), could
promote movements of Ca2+, cell proliferation, and the eventual
production of proinflammatory cytokines in human peripheral blood
mononuclear cells (PBMC) as well as in Jurkat cells, after exposure to
the field for 1 h. The same study was also performed after activation of
cells with 5 mg/ml phytohaemagglutinin (PHA). Our results clearly
demonstrate that static NMRF exposure has neither proliferative, nor
activating, nor proinflammatory effects on both normal and PHA activated
PBMC. Moreover, the concentration of interleukin-1beta, interleukin-2,
interleukin-6, interferon, and tumour necrosis factor alpha (TNFalpha)
remained unvaried in exposed cells. Exposure of Jurkat cells
statistically decreased the proliferation and the proliferation indexes,
which 24 and 48 h after exposure were 0.7 +/- 0.29 and 0.87 +/- 0.12,
respectively. Moreover, in Jurkat cells the [Ca2+]i was higher than in
PBMC and was reduced significantly to about one half after exposure.
This is consistent with the decrease of proliferation and with the low
levels of IL-2 measured. On the whole, our data suggest that NMRF
exposure failed to affect the physiologic behaviour of normal
lymphomonocytes. Instead in Jurkat cells, by changing the properties of
cell membranes, NMRF can influence Ca2+ transport processes, and hence
Ca2+ homeostasis with improvement of proliferation. Copyright 2003
Wiley-Liss, Inc.
Bioelectromagnetics. 2003 Feb;24(2):109-17.
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Interaction between weak low frequency magnetic fields and cell membranes.
Baureus Koch CL, Sommarin M, Persson BR, Salford LG, Eberhardt JL.
Department of Radiation Physics, Lund University Hospital, Lund, Sweden.
The question of whether very weak low frequency magnetic fields can
affect biological systems, has attracted attention by many research
groups for quite some time. Still, today, the theoretical possibility of
such an interaction is often questioned and the site of interaction in
the cell is unknown. In the present study, the influence of extremely
low frequency (ELF) magnetic fields on the transport of Ca(2+) was
studied in a biological system consisting of highly purified plasma
membrane vesicles. We tested two quantum mechanical theoretical models
that assume that biologically active ions can be bound to a channel
protein and influence the opening state of the channel. Vesicles were
exposed for 30 min at 32 degrees C and the calcium efflux was studied
using radioactive (45)Ca as a tracer. Static magnetic fields ranging
from 27 to 37 micro T and time varying magnetic fields with frequencies
between 7 and 72 Hz and amplitudes between 13 and 114 micro T (peak)
were used. We show that suitable combinations of static and time varying
magnetic fields directly interact with the Ca(2+) channel protein in
the cell membrane, and we could quantitatively confirm the model
proposed by Blanchard. Copyright 2003 Wiley-Liss, Inc.
Bioelectromagnetics. 2003 Sep;24(6):395-402.
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Effect of a 125 mT static magnetic field on the kinetics of voltage activated Na+ channels in GH3 cells.
Rosen AD.
Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907, USA. arosen@bilbo.bio.purdue.edu
Voltage activated Na(+) channels were examined in GH3 cells, using
the whole cell patch clamp method. Channel currents were recorded
before, during, and after a 150 s exposure to a 125 mT static magnetic
field. There was a slight shift in the current-voltage relationship and a
less than 5% reduction in peak current during magnetic field exposure.
More pronounced, however, was an increase in the activation time
constant, tau(m), during and for at least 100 s following exposure to
the field. This change in tau(m) was seen primarily at lower activation
voltages. No change was noted in the inactivation time constant, tau(h).
Changes were clearly temperature dependent, being evident only at and
above 35 degrees C. These findings are consistent with the hypothesis
that reorientation of diamagnetic anisotropic molecules in the cell
membrane are capable of distorting imbedded ion channels sufficiently to
alter their function. The temperature dependence of this phenomenon is
probably due to the greater ease with which a liquid crystal membrane
can be deformed. Copyright 2003 Wiley-Liss, Inc.
Bioelectromagnetics. 2003 Oct;24(7):517-23.
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Effect of a static magnetic field on ion transport in a cellulose membrane.
Ohata R, Tomita N, Ikada Y.
Institute for Frontier Medical Sciences, Kyoto University, 53
Kawahara-cho, Shogoin, Sakyo-ku, 606-8507 Kyoto, Japan.
rohata@mech.kyoto-u.ac.jp
A cellulose membrane was exposed to the static magnetic field (SMF)
in the presence of KCl solution and ion transport through the membrane
was measured before and after the SMF exposure. SMF at 0.24 T
significantly enhanced the rate of ion transport, especially after the
first exposure (p<0.05), while the increased ion transport rate did
not return to the initial basal level after exchange of the aqueous
medium. These results suggest that an irreversible, temporal
conformation change took place on the cellulose membrane or on the water
bound to the cellulose surface. The accelerating effect of SMF on the
ion transport seems to have occurred as a result of stabilized hydration
layer on the cellulose surface.
J Colloid Interface Sci. 2004 Feb 15;270(2):413-6.
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Effects of pulsed electromagnetic fields on human chondrocytes: an in vitro study.
Pezzetti F, De Mattei M, Caruso A, Cadossi R, Zucchini P, Carinci F, Traina GC, Sollazzo V.
Dipartimento di Morfologia ed Embriologia, Universita di Ferrara, via Fossato di Mortara 64, 44100 Ferrara, Italy.
(3)H-thymidine incorporation was studied in cultured human nasal and
articular chondrocytes exposed to low-energy, low-frequency pulsed
electromagnetic fields (PEMFs) (75 Hz, 2.3 mT). The reverse
transcriptase polymerase chain reaction (RT-PCR) analysis shows that
human secondary chondrocytes derived from both nasal and articular
cartilage express collagen type II mRNA, which is a specific marker of
the chondrocyte phenotype. In a preliminary series of experiments, cells
were exposed to PEMF for different time periods ranging from 6 to 30
hours (time-course), in medium supplemented with 10% or 0.5% fetal calf
serum (FCS) and in serum-free medium. The ratios between the
(3)H-thymidine incorporation in PEMFs and control cultures show an
increase of the cell proliferation in cultures exposed to PEMFs when
serum is present in the culture medium, whereas no effect was observed
in serum-free conditions. The increase in DNA synthesis, induced by
PEMFs, was then evaluated only at the times of maximum induction and the
results were analyzed by the three-factor analysis of variance (ANOVA).
The data presented in this study show that even if (3)H-thymidine
incorporation is higher in nasal than in articular chondrocytes, PEMF
induce an increase in the proliferation of both cell types. Moreover,
the concentration of FCS in the culture medium greatly influences the
proliferative response of human chondrocytes to the PEMF exposure.
Though normal human osteoblast cells increase their proliferation when
exposed to PEMFs if only 10% FCS is present in the medium, human
chondrocytes are able to increase their cell proliferation when exposed
to PEMFs in the presence of both 0.5% and 10% of FCS in the medium. The
results obtained may help to explain the basic mechanisms of PEMF
stimulation of fracture healing.
Calcif Tissue Int. 1999 Nov;65(5):396-401.
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Effects of low-frequency pulsed electromagnetic fields on the proliferation of chondrocytes.
Indouraine A, Petersen JP, Pforringer W.
Symbion Science Park (Symbion-Wissenschaftspark), Kopenhagen, Danemark.
Chondrocytes isolated from the human cartilage of 5 patients between
the ages 23 and 56 were exposed to low frequency pulsed electromagnetic
fields (9 mT; 3 Hz) for a daily period of 60 minutes on 5 consecutive
days and then every 48 hours for the next 6 days (11 days in total).
Cell viability was estimated using trypan blue exclusion and
proliferation was estimated by counting the cells in a haemacytometer.
Cell morphology was compared for control purposes by directly observing
the cells under a light microscope after staining cells in a
haematoxylin and eosin solution. The results were statistically analysed
and compared to a control sample. Data revealed that exposing cells
isolated from human cartilage to pulsed electromagnetic fields (9 mT; 3
Hz) led to a significantly higher number of cells in comparison to the
control sample. Among the cells from the 5 patients, growth varied
between 1.1 to 3.0 folds compared to the control sample. The difference
in cell viability between the exposed cells and the control sample was,
however, not significant. Some morphological variations were revealed
when the cells were observed under a light microscope. The exposed cells
were thinner and longer than the control cells which were large and
flat. The exposed cells tended to grow in a more uniform direction while
the control cells grew in all directions. These differences in
morphology and growth may be related to the higher density of the
exposed cells.
Sportverletz Sportschaden. 2001 Mar;15(1):22-7.
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Pulsed electromagnetic fields increase growth factor release by nonunion cells.
Guerkov HH, Lohmann CH, Liu Y, Dean DD, Simon BJ, Heckman JD, Schwartz Z, Boyan BD.
Department of Orthopaedics, University of Texas Health Science Center at San Antonio, 78229-3900, USA.
The mechanisms involved in pulsed electromagnetic field stimulation
of nonunions are not known. Animal and cell culture models suggest
endochondral ossification is stimulated by increasing cartilage mass and
production of transforming growth factor-beta 1. For the current study,
the effect of pulsed electromagnetic field stimulation on cells from
human hypertrophic (n = 3) and atrophic (n = 4) nonunion tissues was
examined. Cultures were placed between Helmholtz coils, and an
electromagnetic field (4.5-ms bursts of 20 pulses repeating at 15 Hz)
was applied to 1/2 of them 8 hours per day for 1, 2, or 4 days. There
was a time-dependent increase in transforming growth factor-beta 1 in
the conditioned media of treated hypertrophic nonunion cells by Day 2
and of atrophic nonunion cells by Day 4. There was no effect on cell
number, [3H]-thymidine incorporation, alkaline phosphatase activity,
collagen synthesis, or prostaglandin E2 and osteocalcin production. This
indicates that human nonunion cells respond to pulsed electromagnetic
fields in culture and that transforming growth factor-beta 1 production
is an early event. The delayed response of hypertrophic and atrophic
nonunion cells (> 24 hours) suggests that a cascade of regulatory
events is stimulated, culminating in growth factor synthesis and
release.
Clin Orthop. 2001 Mar;(384):265-79.
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Cell proliferation/cell death balance in renal cell cultures after exposure to a static magnetic field.
Buemi M, Marino D, Di Pasquale G, Floccari F, Senatore M, Aloisi C, Grasso F, Mondio G, Perillo P, Frisina N, Corica F.
Dipartimento di Medicina Interna, Facolta di Medicina e Chirurgia, Universita di Messina, Italy. Buemim@unime.it
The effect of a static magnetic field (MF) of 0.5 mT of intensity on
the cell proliferation/cell death balance was investigated in renal
cells (VERO) and cortical astrocyte cultures from rats. Magnetic
stimulation was delivered by magnetic disks at known intensities. The
percentage of apoptotic and necrotic cells was evaluated using flow
cytometry and morphological analysis following Hoechst chromatin
staining. An index of cell proliferation was determined using sulfonated
tetrazolium (WST-1). Control cultures were prepared without exposure to
MFs. After 2, 4 and 6 days of exposure to a MF, we observed a gradual
decrease in apoptosis and proliferation and a gradual increase in cells
with a necrotic morphology with respect to the control group. In
astrocyte cultures, over a 6-day exposure period, a gradual increase was
observed in apoptotic, proliferating, and necrotic cells. Our findings
suggest that the effect of exposure to MFs varies, depending on the cell
type; MFs may also have a nephropathogenic effect. Copyright 2001 S.
Karger AG, Basel
Nephron. 2001 Mar;87(3):269-73.
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Effects of pulsed electromagnetic fields on human articular chondrocyte proliferation.
De Mattei M, Caruso A, Pezzetti F, Pellati A, Stabellini G, Sollazzo V, Traina GC.
Dipartimento di Morfologia ed Embriologia, Universita di Ferrara, Italy.
Low-energy, low-frequency pulsed electromagnetic fields (PEMFs) can
induce cell proliferation in several cell culture models. In this work
we analysed the proliferative response of human articular chondrocytes,
cultured in medium containing 10% FBS, following prolonged exposure to
PEMFs (75 Hz, 2.3 mT), currently used in the treatment of some
orthopaedic pathologies. In particular, we investigated the dependence
of the proliferative effects on the cell density, the availability of
growth factors and the exposure lengths. We observed that PEMFs can
induce cell proliferation of low density chondrocyte cultures for a long
time (6 days), when fresh serum is added again in the culture medium.
In the same conditions, in high density cultures, the PEMF-induced
increase in cell proliferation was observed only in the first three days
of exposure. The data presented in this study show that the
availability of growth factors and the environmental constrictions
strongly condition the cellular proliferative response to PEMFs.
Connect Tissue Res. 2001;42(4):269-79.
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Low frequency EMF regulates chondrocyte differentiation and expression of matrix proteins.
Ciombor DM, Lester G, Aaron RK, Neame P, Caterson B.
Department of Orthopaedics, School of Medicine, Brown University, Providence, RI, USA. deborah_ciombor@brown.edu
This study describes the enhancement of chondrogenic differentiation
in endochondral ossification by extremely low frequency pulsed
electric/magnetic fields (EMFs). The demineralized bone matrix
(DBM)-induced endochondral ossification model was used to examine the
effects of EMF stimulation. [35S]-Sulfate and [3H]-thymidine
incorporation and glycosaminoglycan (GAG) content were determined by
standard methods. Proteoglycan (PG) and GAG molecular size and
composition were determined by gel chromatography and sequential enzyme
digestion. Immunohistochemical and Western blot analysis of PGs were
done with antibodies 2B6, 3B3, 2D3 and 5D4. Northern analysis of total
RNA extracts was performed for aggrecan, and type II collagen. All data
was compared for significance by Student's t- or analysis of variance
(ANOVA)-tests. The EMF field accelerated chondrogenesis as evidenced by
an increase in: (1) 35SO4 incorporation and GAG content, (2) the number
of chondrocytes at day 8 of development, (3) the volumetric density of
cartilage and (4) the extent of immunostaining for 3B3 and 5D4. No
differences in DNA content or [3H]-thymidine incorporation were observed
between control and stimulated ossicles, suggesting the absence of
enhanced cell proliferation or recruitment as a mechanism for the
acceleration. PG and GAG molecular sizes and GAG chemical composition
were similar in stimulated and control ossicles, indicating that
stimulation resulted in an accelerated synthesis of normal cartilage
molecules. The increased expression of PG and type II collagen mRNA as
well as a greater immunoreactivity of 3B3 and 5D4 suggest an increase in
the rate of differentiation of chondrocytes and enhanced phenotypic
maturation.
J Orthop Res. 2002 Jan;20(1):40-50.
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Effects of pulsed electromagnetic field (PEMF) stimulation on bone
tissue like formation are dependent on the maturation stages of the
osteoblasts.
Diniz P, Shomura K, Soejima K, Ito G.
Department of Orthodontics, Kagoshima University Dental School, Kagoshima, Japan.
The effects of pulsed electromagnetic field (PEMF, 15 Hz pulse burst,
7 mT peak) stimulation on bone tissue-like formation on osteoblasts
(MC3T3-E1 cell line) in different stages of maturation were assessed to
determine whether the PEMF stimulatory effect on bone tissue-like
formation was associated with the increase in the number of cells and/or
with the enhancement of the cellular differentiation. The cellular
proliferation (DNA content), differentiation (alkaline phosphatase
activity), and bone tissue-like formation (area of mineralized matrix)
were determined at different time points. PEMF treatment of osteoblasts
in the active proliferation stage accelerated cellular proliferation,
enhanced cellular differentiation, and increased bone tissue-like
formation. PEMF treatment of osteoblasts in the differentiation stage
enhanced cellular differentiation and increased bone tissue-like
formation. PEMF treatment of osteoblasts in the mineralization stage
decreased bone tissue-like formation. In conclusion, PEMF had a
stimulatory effect on the osteoblasts in the early stages of culture,
which increased bone tissue-like formation. This stimulatory effect was
most likely associated with enhancement of the cellular differentiation,
but not with the increase in the number of cells. Copyright 2002
Wiley-Liss, Inc.
Bioelectromagnetics. 2002 Jul;23(5):398-405.
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Nitric oxide mediates the effects of pulsed electromagnetic field
stimulation on the osteoblast proliferation and differentiation.
Diniz P, Soejima K, Ito G.
Department of Orthodontics, Kagoshima University Dental School, 8-35-1 Sakuragaoka, Kagoshima 890-8544, Japan.
The purpose of this research was to investigate whether the effects
of pulsed electromagnetic field (PEMF) stimulation on the osteoblast
proliferation and differentiation are mediated by the increase in the
nitric oxide (NO, nitrogen monoxide) synthesis. The osteoblasts
(MC3T3-E1 cell line) were cultured in the absence (-NMMA group) or in
the presence (+NMMA group) of the NO synthase inhibitor L-NMMA. First,
osteoblasts were subjected to PEMF stimulation (15 Hz and 0.6 mT) up to
15 days. The DNA content and the NO concentration in the conditioned
medium were determined on the 3rd, 7th, and 15th days of culture.
Following, osteoblasts were stimulated in the proliferation (P-NMMA and
P+NMMA groups) or in the differentiation (D-NMMA and D+NMMA groups)
stages of maturation, and the alkaline phosphatase (AlPase) activity was
determined on the 15th day of culture for all groups. PEMF stimulation
increased significantly the nitrite concentration in the -NMMA group on
the 3rd, 7th, and 15th days of culture. However, this effect was
partially blocked in the +NMMA group. The DNA content in the -NMMA
group, but not in the +NMMA group, increased significantly on the 3rd
and 7th days of culture. The AlPase activity in the P-NMMA and D-NMMA
groups, but not in the P+NMMA and D+NMMA groups, also increased
significantly. In conclusion, the PEMF stimulatory effects on the
osteoblasts proliferation and differentiation were mediated by the
increase in the NO synthesis. Copyright 2002 Elsevier Science (USA)
Nitric Oxide. 2002 Aug;7(1):18-23.
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Exposure of cells to static magnetic field accelerates loss of integrity of plasma membrane during apoptosis.
Teodori L, Grabarek J, Smolewski P, Ghibelli L, Bergamaschi A, De Nicola M, Darzynkiewicz Z.
Department of Biomedicine and Toxicology, UTS BIOTEC, ENEA-Casaccia, Rome, Italy.
BACKGROUND: Much attention is being paid to the biologic effects of
magnetic fields (MFs). Although MFs enhance tumorigenesis, they are
neither mutagenic nor tumorigenic. The mechanism of their tumorigenic
effect has not been elucidated.
METHODS: To investigate the effect of MFs on apoptosis in HL-60
cells, we exposed the cells to static MFs of 6 mT generated by a
magnetic disk of known intensity. Apoptosis was triggered by the DNA
topoisomerase I inhibitor, camptothecin (CPT). Activation of caspases in
situ using the fluorochrome-labeled inhibitor (FLICA) method and
determination of plasma membrane integrity by excluding propidium iodide
(PI) were measured by both laser scanning cytometry (LSC) and flow
cytometry (FC). LSC and FC identified cells at three sequential stages
of their demise: early apoptosis (cells with activated caspases and PI
negative); late apoptosis (cells with activated caspases but unable to
exclude PI); secondary necrosis (cells with apoptotic morphology no
longer stained with FLICA, not excluding PI).
RESULTS: MF alone did not induce any apoptogenic or necrogenic
effect. CPT exposure led to the sequential appearance of apoptotic
cells. In the presence of CPT and MF, the overall proportion of cells
undergoing apoptosis was not significantly changed. However, we
consistently observed a significant increase in the frequency of late
apoptotic/necrotic cells when compared with samples treated with CPT
alone (P < 0.001), as well as a decrease in the percentage of early
apoptotic cells (P = 0.013). The data obtained by FC and LSC were
consistent with each other, showing a similar phenomenon.
CONCLUSION: Whereas MF alone or with CPT did not affect overall cell
viability, it accelerated the rate of cell transition from apoptosis to
secondary necrosis after induction of apoptosis by the DNA-damaging
agent, CPT. Modulation of the kinetics of the transition from apoptosis
to secondary necrosis by MF in vivo may play a role in inflammation and
tumorigenesis. Copyright 2002 Wiley-Liss, Inc.
Cytometry. 2002 Nov 1;49(3):113-8.
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Evaluation of the morphology of the human placental cotyledon following dual in vitro perfusion in variable magnetic field.
Lopucki M, Lancut M, Rogowska W, Czerny K, Jedrych B, Pietruszewski S, Kornarzynski K, Kotarski J.
I Katedry i Kliniki Ginekologii AM w Lublinie.
Objectives and the aim of the study was electron-microscopy
morphological estimation of the human placental cotyledon after 180
minutes of dual closed perfusion in vitro.
MATERIALS AND METHODS: In the experimental group the cotyledons were
exposed to variable magnetic field of 2 mT magnetic induction and 50 Hz
frequency. The control group K (10 perfusions) was not subjected to
magnetic field while the experimental group B (10 perfusions) was
influenced by magnetic field.
RESULTS: It was found that homogeneous variable magnetic field
disturbs the ultrastructure of the nuclei and cytoplasma and it
increases the density of the vascular-epithelial membrane of villi cells
of human placenta in vitro.
CONCLUSION: Variable sinusoildal, magnetic field of 2 mT magnetic
induction and 50 Hz frequency disturbs the ultrastructure of the nuclei
and cytoplasma and it increases the density of the vascular-epithelial
membrane of villi cells of human placenta in vitro after 180 minutes of
dual closed perfusion in vitro.
Ginekol Pol. 2003 Oct;74(10):1187-93.
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50-Hz magnetic field exposure influences DNA repair and
mitochondrial DNA synthesis of distinct cell types in brain and kidney
of adult mice.
Schmitz C, Keller E, Freuding T, Silny J, Korr H.
Department of Anatomy and Cell Biology, RWTH Aachen University, Pauwelsstrasse/Wendlingweg 2, 52057, Aachen, Germany.
Despite several recent investigations, the impact of whole-body
magnetic field exposure on cell-type-specific alterations due to DNA
damage and DNA repair remains unclear. In this pilot study adult mice
were exposed to 50-Hz magnetic field (mean value 1.5 mT) for 8 weeks or
left unexposed. Five minutes after ending exposure, the mice received
[(3)H]thymidine and were killed 2 h later. Autoradiographs were prepared
from paraffin sections of brains and kidneys for measuring unscheduled
DNA synthesis and mitochondrial DNA synthesis, or in situ nick
translation with DNA polymerase-I and [(3)H]dTTP. A significant (
P<0.05) increase in both unscheduled DNA synthesis and in situ nick
translation was only found for epithelial cells of the choroid plexus.
Thus, these two independent methods indicate that nuclear DNA damage is
produced by long-lasting and strong magnetic field exposure. The fact
that only plexus epithelial cells were affected might point to possible
effects of magnetic fields on iron transport across the
blood-cerebrospinal fluid barrier, but the mechanisms are currently not
understood. Mitochondrial DNA synthesis was exclusively increased in
renal epithelial cells of distal convoluted tubules and collecting
ducts, i.e., cells with a very high content of mitochondria, possibly
indicating increased metabolic activity of these cells.
Acta Neuropathol (Berl). 2003 Dec 19. [Epub ahead of print]
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Effects of 0.2 T static magnetic field on human skin fibroblasts.
Pacini S, Gulisano M, Peruzzi B, Sgambati E, Gheri G, Gheri Bryk S, Vannucchi S, Polli G, Ruggiero M.
Department of Human Anatomy, Histology and Forensic Medicine, University of Firenze, viale Morgagni 85, 50134, Firenze, Italy.
Human skin fibroblasts were exposed to 0.2 T static magnetic field
generated by a magnetic resonance tomograph. After 1h exposure, cell
morphology was modified in association with a concomitant decrease in
the expression of some sugar residues of glycoconjugates. Study of cell
proliferation and mitogenic signal transduction showed a decrease of
thymidine incorporation and of second messenger formation. However, cell
viability, assessed by colony forming assay, was unaffected. These
results demonstrate that the static magnetic field generated by
routinely used magnetic resonance tomograph induces alterations on human
skin fibroblasts.
Cancer Detect Prev. 2003;27(5):327-32.
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Exposure to 900 MHz electromagnetic field induces an unbalance
between pro-apoptotic and pro-survival signals in T-lymphoblastoid
leukemia CCRF-CEM cells.
Marinelli F, La Sala D, Cicciotti G, Cattini L, Trimarchi C, Putti S, Zamparelli A, Giuliani L, Tomassetti G, Cinti C.
Institute for Organ Transplantation and Immunocytology, ITOI-CNR, Bologna unit, c/o IOR, Bologna, Italy.
It has been recently established that low-frequency electromagnetic
field (EMFs) exposure induces biological changes and could be associated
with increased incidence of cancer, while the issue remains unresolved
as to whether high-frequency EMFs can have hazardous effect on health.
Epidemiological studies on association between childhood cancers,
particularly leukemia and brain cancer, and exposure to low- and
high-frequency EMF suggested an etiological role of EMFs in inducing
adverse health effects. To investigate whether exposure to
high-frequency EMFs could affect in vitro cell survival, we cultured
acute T-lymphoblastoid leukemia cells (CCRF-CEM) in the presence of
unmodulated 900 MHz EMF, generated by a transverse electromagnetic (TEM)
cell, at various exposure times. We evaluated the effects of
high-frequency EMF on cell growth rate and apoptosis induction, by cell
viability (MTT) test, FACS analysis and DNA ladder, and we investigated
pro-apoptotic and pro-survival signaling pathways possibly involved as a
function of exposure time by Western blot analysis. At short exposure
times (2-12 h), unmodulated 900 MHz EMF induced DNA breaks and early
activation of both p53-dependent and -independent apoptotic pathways
while longer continuous exposure (24-48 h) determined silencing of
pro-apoptotic signals and activation of genes involved in both
intracellular (Bcl-2) and extracellular (Ras and Akt1) pro-survival
signaling. Overall our results indicate that exposure to 900 MHz
continuous wave, after inducing an early self-defense response triggered
by DNA damage, could confer to the survivor CCRF-CEM cells a further
advantage to survive and proliferate. Copyright 2003 Wiley-Liss, Inc.
J Cell Physiol. 2004 Feb;198(2):324-32.
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Low electromagnetic field (50 Hz) induces differentiation on primary human oral keratinocytes (HOK).
Manni V, Lisi A, Rieti S, Serafino A, Ledda M, Giuliani L, Sacco D, D'Emilia E, Grimaldi S.
Istituto di Neurobiologia e Medicina Molecolare (INeMM), CNR, Rome, Italy.
This work concerns the effect of low frequency electromagnetic fields
(ELF) on biochemical properties of human oral keratinocytes (HOK).
Cells exposed to a 2 mT, 50 Hz, magnetic field, showed by scanning
electron microscopy (SEM) modification in shape and morphology; these
modifications were also associated with different actin distribution,
revealed by phalloidin fluorescence analysis. Moreover, exposed cells
had a smaller clonogenic capacity, and decreased cellular growth.
Indirect immunofluorescence with fluorescent antibodies against
involucrin and beta-catenin, both differentiation and adhesion markers,
revealed an increase in involucrin and beta-catenin expression. The
advance in differentiation was confirmed by a decrease of expression of
epidermal growth factor (EGF) receptor in exposed cells, supporting the
idea that exposure to electromagnetic field carries keratinocytes to
higher differentiation level. These observations support the hypothesis
that 50 Hz electromagnetic fields may modify cell morphology and
interfere in differentiation and cellular adhesion of normal
keratinocytes. Copyright 2004 Wiley-Liss, Inc.
Bioelectromagnetics. 2004 Feb;25(2):118-26.
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Pulsing electromagnetic fields induce cellular transcription.
Goodman R, Bassett CA, Henderson AS.
Weak, pulsing electromagnetic fields can modify biological processes.
The hypothesis that responses to such induced currents depend on pulse
characteristics was evaluated by using transcription as the target
process. Two pulses in clinical use, the repetitive single pulse and the
repetitive pulse train, were tested. These pulses produced different
results from each other and from controls when transcription in dipteran
salivary gland cells was monitored with tritiated uridine in
transcription autoradiography, cytological nick translation, and
analysis of isolated RNA fractions. The single pulse increased the
specific activity of messenger RNA after 15 and 45 minutes of exposure.
The pulse train increased specific activity only after 45 minutes of
exposure.
Science. 1983 Jun 17;220(4603):1283-5.
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Pulsed electromagnetic fields alter phenotypic expression in chondroblasts in tissue culture.
Norton LA, Witt DW, Rovetti LA.
Department of Orthodontics, School of Dental Medicine, University of Connecticut Health Center, Farmington 06032.
We hypothesize that pulsed electromagnetic fields (PEMF) alter
phenotypic expression of chondroblasts by promoting the production of
alkaline phosphatase (AP) and altering the structure of proteoglycans.
Chondroblasts from the hypertrophic zone of tibial epiphyses (HC),
sternum (SC), and skin fibroblasts (F) were cultured from 16 day chick
embryos. Cultures were randomly designated control (C) or experimental
(E). E received PEMF for 24 h in a 6 h on, 6 h rest sequence. The
controls were in the same incubator shielded by Mu metal. Assays for AP
activity were performed and normalized to protein content. Proteoglycan
synthesis assay involved labeling with 35S fractionating in a 5% to 20%
surcrose gradient determining total protein and chondroitin sulfate
content. PEMF showed no change of AP activity on F. A high AP basal
activity was found in HC, but was not increased above the control. PEMF
increased AP in the SC samples (E/C ratio). The sucrose gradient data
showed a shift in peaks for SC only altering the ratio of carbohydrate
to protein for the SC. Analysis of carbohydrate and protein indicated
that the effect was decreased synthesis or degradation of protein. We
conclude that PEMF alters the phenotypic expression of sternal
chondroblasts in our in vitro system.
J Orthop Res. 1988;6(5):685-9.
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Magnetic behavior of human erythrocytes at different hemoglobin states.
Sakhnini L, Khuzaie R.
Physics Department, College of Science, University of Bahrain, State of Bahrain. l_sakhnini@yahoo.com
The effect of a static magnetic field on human erythrocytes at
different hemoglobin states (normal, oxidized and reduced hemoglobin)
was investigated. Three different blood samples, normal, iron deficiency
anemic and beta thalassemia minor, were studied. Measurements of the
magnetization curves of the erythrocytes for all blood samples in all
states showed diamagnetic behavior; however, oxidation was found to
enhance this behavior. These measurements have also shown that the
normal and iron deficiency samples in the reduced states exhibit a less
diamagnetic response in comparison with the normal state. This result
indicates that the reduction process gave rise to a paramagnetic
component of the magnetization. Analysis of the measured paramagnetic
behavior, using a Brillouin function, gave an effective magnetic moment
of 8 muB per reduced hemoglobin molecule for both normal and anemic
samples. This result shows that both anemic and normal blood have
similar magnetic behavior and the only difference is the number of
hemoglobin molecules per erythrocyte. For the beta thalassemia minor
blood sample, magnetic measurements showed that both the normal and
reduced states have almost the same diamagnetic behavior. However, this
diamagnetic response is less than that for the normal state of the iron
deficiency anemic sample. This result may indicate a low oxygen intake
for the blood in the normal state for the beta thalassemia minor blood.
All magnetic measurements were made using a vibrating sample
magnetometer using field steps of 0.001 T from 1 T to -1 T.
Eur Biophys J. 2001 Oct;30(6):467-70.
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Effects of static magnetic field on human leukemic cell line HL-60.
Sabo J, Mirossay L, Horovcak L, Sarissky M, Mirossay A, Mojzis J.
Department of Medical Biophysics, Medical Faculty, P.J. Safarik
University, Trieda SNP 1, 04011 Kosice, Slovak Republic.
grosiar@central.medic.upjs.sk
A number of structures with magnetic moments exists in living
organisms that may be oriented by magnetic field. While most
experimental efforts belong to the area of effects induced by weak and
extremely low-frequency electromagnetic fields, we attempt to give an
attention to the biological effects of strong static magnetic fields.
The influence of static magnetic field (SMF) on metabolic activity of
cells was examined. The metabolic activity retardation is observed in
human leukemic cell line HL-60 exposed to 1-T SMF for 72 h. The
retardation effect was observed as well as in the presence of the
mixture of the antineoplastic drugs 5 fluorouracil, cisplatin,
doxorubicin and vincristine.
Bioelectrochemistry. 2002 May 15;56(1-2):227-31.
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The effect of extremely low frequency magnetic fields on cytochrome oxidase subunit 1 mRNA transcription.
Zhong T, Chen Q, Wu R, Yao G, Lu D, Chiang H.
Bioelectromagnetic Lab., Zhejiang University School of Medicine, Hangzhou 310031, China.
OBJECTIVE: To clone and identify MF-1 gene which responded to
extremely low frequency magnetic fields(ELF MF) in Daudi cells, and
explore the response universality of MF-1 gene in several MF-sensitive
cell lines, so as to provide experimental basis for revealing the
mechanism of biological effects induced by magnetic field.
METHODS: The DNA fragment of MF-1 was cloned and sequenced; the mRNA
level of MF-1 gene were analysed in MF-sensitive cell lines(HL-60, L1210
and CHL) by Northern blot after these cells being treated with 0.1 mT
and 0.8 mT MF for 20 minutes and 24 hours, respectively.
RESULTS: The MF-1 cDNA sequence had 100% homology with cytochrome
oxidase subunit 1 gene(CO1) by searching Gene Bank database; the
transcription of CO1 in HL-60, L1210 and CHL cell lines which exposed to
0.1 mT and 0.8 mT MF for 20 minutes were significantly lower(0.38 +/-
0.12 and 0.37 +/- 0.04) than that of control(0.58 +/- 0.12) and so did
for 24 hours exposure(0.46 +/- 0.09 and 0.45 +/- 0.09 vs 0.65 +/- 0.06)
(P < 0.05).
CONCLUSION: CO1 is a MF-responsive gene. Cytochrome oxidase activity
may be affected through low level of CO1 transcription by magnetic
fields, thus induce bioeffects in organisms.
Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi. 2002 Aug; 20(4): 249-51.
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Orientation of human glioblastoma cells embedded in type I collagen, caused by exposure to a 10 T static magnetic field.
Hirose H, Nakahara T, Miyakoshi J.
Laboratory of Radiation Biology, Graduate School of Science, Kyoto
University, Kitashirakawa-Oiwake, Sakyo, Kyoto 606-8502, Japan.
We investigated the preferred orientation of human glioblastoma cells
(A172) following exposure to static magnetic fields (SMF) at 10 Tesla
in the presence or absence of collagen. A172 cells embedded in collagen
gel were oriented perpendicular to the direction of the SMF. A172 cells
cultured in the absence of collagen did not exhibit any specific
orientation pattern after 7 days of exposure to the SMF. Thus we
succeeded in evoking the magnetic orientation of human glioblastoma
cells by exposure to the SMF. Our results suggest that the orientation
of glioblastoma cell processes may be due to the arrangement of
microtubules under the influence of magnetically oriented collagen
fiber. Copyright 2002 Elsevier Science Ireland Ltd.
Neurosci Lett. 2003 Feb 20;338(1):88-90.
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Magnetic field effects on assembly pattern of smooth muscle cells.
Iwasaka M, Miyakoshi J, Ueno S.
Department of Biomedical Engineering, Graduate School of Medicine,
University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan (M.
I., S. U.) and Department of Radiological Technology, School of Health
Science, Faculty of Medicine, Hirosaki University, 66-1 Hon-cho,
Hirosaki 036-8564, Japan (J. M.).
Under a strong magnetic field, the diamagnetic properties of
biological cells modulate the behavior of the cells themselves, under
conditions of both floating and adherence. The morphological effects of
strong static magnetic fields on adherent cells are less well understood
than the effects of magnetic fields on red blood cells. In the present
study, a high-intensity magnetic field of 14 T affected the morphology
of smooth muscle cell assemblies, and the shapes of the cell colonies
extended along the direction of the magnetic flux. The phenomenon was
most notable under magnetic fields of more than 10 T, where an
ellipsoidal pattern of smooth muscle cell colonies was clearly observed.
The ellipticity of the cell colony pattern with a 14-T magnetic field
was 1.3, whereas that with a field of 0-8 T was close to a circle at
about 1.0. The evidence that smooth muscle cells detect high-density
magnetic flux and thus change their cell orientation was shown as a
visible pattern of cellular colonies. The speculated mechanism is a
diamagnetic torque force acting on cytoskeleton fibers, which are
dynamically polymerizing-depolymerizing during cell division and cell
migration.
In Vitro Cell Dev Biol Anim. 2003 Mar;39(3):120-123.
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Control of orientation of rat Schwann cells using an 8-T static magnetic field.
Eguchi Y, Ogiue-Ikeda M, Ueno S.
Department of Biomedical Engineering, Graduate School of Medicine,
University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Schwann cells aid in neuronal regeneration in the peripheral nervous
system via guiding the regrowth of axons. In this study, we investigated
the magnetic orientation of Schwann cells, and of a mixture of Schwann
cells and collagen, after an 8-tesla magnetic field exposure. We
obtained cultured Schwann cells from dissected sciatic nerves of
neonatal rats. After 60 h of magnetic field exposure, Schwann cells
oriented parallel to the magnetic fields. In contrast, the mixture of
Schwann cells and collagen, Schwann cells oriented in the direction
perpendicular to the magnetic field after 2 h of magnetic field
exposure. In this case, Schwann cells aligned along the collagen fiber
oriented by magnetic fields. The magnetic control of Schwann cell
alignment is useful in medical engineering applications such as nerve
regeneration.
Neurosci Lett. 2003 Nov 13;351(2):130-2.
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Detection of intracellular macromolecule behavior under strong magnetic fields by linearly polarized light.
Iwasaka M, Ueno S.
Department of Biomedical Engineering, Graduate School of Medicine,
University of Tokyo, Tokyo, Japan. iwasaka@medes.m.u-tokyo.ac.jp
Strong static magnetic fields on the order of 10 T have a diamagnetic
force on cell components and generate a clear alignment of a smooth
muscle cell assembly, parallel to the direction of the magnetic fields
within an exposure period of 3 days. This work shows the effects of
diamagnetic torque forces on cell component motion. Linearly polarized
light was utilized to detect the displacement of intracellular
macromolecules. The polarized light passed through a mass of cells in a
magnetic field, and transmission of the light increased and reached a
plateau 2 h after the beginning of magnetic field exposure at 14 T.
However, no distinct change was observed in transmission of the light
under zero magnetic field exposure. The change in polarized light
intensity through the lamellar cell assembly under magnetic fields
corresponds to behavioral changes in cell components. It was speculated
that intracellular macromolecules rotated and showed a displacement due
to diamagnetic torque forces during 2-3 h of magnetic field exposure at
14 T. Copyright 2003 Wiley-Liss, Inc.
Bioelectromagnetics. 2003 Dec;24(8):564-70.
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