ArticlePDF Available

Microgravity simulation by diamagnetic levitation: Effects of a strong gradient magnetic field on the transcriptional profile of Drosophila melanogaster

Authors:

Abstract and Figures

Many biological systems respond to the presence or absence of gravity. Since experiments performed in space are expensive and can only be undertaken infrequently, Earth-based simulation techniques are used to investigate the biological response to weightlessness. A high gradient magnetic field can be used to levitate a biological organism so that its net weight is zero. We have used a superconducting magnet to assess the effect of diamagnetic levitation on the fruit fly D. melanogaster in levitation experiments that proceeded for up to 22 consecutive days. We have compared the results with those of similar experiments performed in another paradigm for microgravity simulation, the Random Positioning Machine (RPM). We observed a delay in the development of the fruit flies from embryo to adult. Microarray analysis indicated changes in overall gene expression of imagoes that developed from larvae under diamagnetic levitation, and also under simulated hypergravity conditions. Significant changes were observed in the expression of immune-, stress-, and temperature-response genes. For example, several heat shock proteins were affected. We also found that a strong magnetic field, of 16.5 Tesla, had a significant effect on the expression of these genes, independent of the effects associated with magnetically-induced levitation and hypergravity. Diamagnetic levitation can be used to simulate an altered effective gravity environment in which gene expression is tuned differentially in diverse Drosophila melanogaster populations including those of different age and gender. Exposure to the magnetic field per se induced similar, but weaker, changes in gene expression.
Content may be subject to copyright.
ms-2
BA
z (mm)
C
Γ (ms-2)
Microgravity simulation by diamagnetic levitation:
effects of a strong gradient magnetic field on the
transcriptional profile of Drosophila melanogaster
Herranz et al.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52 (1 February 2012)
RESEARCH ARTICLE Open Access
Microgravity simulation by diamagnetic levitation:
effects of a strong gradient magnetic field on the
transcriptional profile of Drosophila melanogaster
Raul Herranz
1,2*
, Oliver J Larkin
3
, Camelia E Dijkstra
3
, Richard JA Hill
4
, Paul Anthony
3
, Michael R Davey
3
,
Laurence Eaves
4
, Jack JWA van Loon
5
, F Javier Medina
1
and Roberto Marco
2
Abstract
Background: Many biological systems respond to the presence or absence of gravity. Since experiments
performed in space are expensive and can only be undertaken infrequently, Earth-based simulation techniques are
used to investigate the biological response to weightlessness. A high gradient magnetic field can be used to
levitate a biological organism so that its net weight is zero.
Results: We have used a superconducting magnet to assess the effect of diamagnetic levitation on the fruit fly D.
melanogaster in levitation experiments that proceeded for up to 22 consecutive days. We have compared the
results with those of similar experiments performed in another paradigm for microgravity simulation, the Random
Positioning Machine (RPM). We observed a delay in the development of the fruit flies from embryo to adult.
Microarray analysis indicated changes in overall gene expression of imagoes that developed from larvae under
diamagnetic levitation, and also under simulated hypergravity conditions. Significant changes were observed in the
expression of immune-, stress-, and temperature-response genes. For example, several heat shock proteins were
affected. We also found that a strong magnetic field, of 16.5 Tesla, had a significant effect on the expression of
these genes, independent of the effects associated with magnetically-induced levitation and hypergravity.
Conclusions: Diamagnetic levitation can be used to simulate an altered effective gravity environment in which
gene expression is tuned differentially in diverse Drosophila melanogaster populations including those of different
age and gender. Exposure to the magnetic field per se induced similar, but weaker, changes in gene expression.
Background
SincethebeginningoflifeonEarth,organismshave
developed under the influence of Earthsgravity.Evolu-
tion has provided a number of different solutions to the
mechanical challenge of supporting the weight of a liv-
ing organism [1-4]. In general, the mechanical stresses
induced by gravity on an organism increase with its
mass, although for organisms living in water, the effect
of gravity is to some extent mitigated by buoyancy.
Gravity has an important effect on the development of
seedlings and studies show that the gravitational sense
mechanism acts at the cellular level (geotropism) [5].
Another well-known effect of altered gravity on living
organismsisthereductionofthestrengthofthebones
of astronauts after they have undertaken long missions
in orbiting spacecraft. The gravitational acceleration,
which is g=9.8ms
-2
on the Earthssurface,exertsa
force of 9.8 N on a mass of 1 kg. The reduced gravity
on the surfaces of Mars (0.37 g), the Moon (0.18 g), and
the microgravity conditions in orbiting space stations
may have important effects on astronauts manning the
first space colonies, and on the development of animals
and plants. It is also possible that zero- and reduced-
gravity influences the behaviour of micro-organisms,
either directly or through the effect of reduced gravity
on the environment, e.g. modified convection in fluids
and gases could affect bacterial physiology [6].
One of the current challenges is to advance our
understanding of the way genomic information is
* Correspondence: r.herranz@csic.es
Contributed equally
1
Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, E-28040,
Madrid, Spain
Full list of author information is available at the end of the article
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
© 2012 Herranz et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creative commons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, pro vided the original work is properly cited.
modulated by different physical and environmental
forces to produce the diverse phenotypes that are
encountered in biology. The fruit fly Drosophila is an
ideal organism with which to explore environmental
effects on the genome; the genome of more than twelve
species of Drosophila have had their genomes sequenced
and are available in Flybase [7] in different gene
annotations.
Several methods exist to simulate weightlessnesson
Earth, such as the Random Positioning Machine (RPM)
[8,9]. Another approach to study the response of organ-
isms to changes in gravity is the use of diamagnetic levi-
tation (See, for example, [10-14]). Diamagnetic material,
which includes water and biological tissue, is repelled
from magnetic fields. Within the bore of a powerful Bit-
terelectromagnet or superconducting solenoid, the
repulsive diamagnetic force on water can be enough to
balance the weight of the water so that it levitates
[15-18]. Most soft biological tissues can be levitated
under the same conditions, since water is the main con-
stituent of the tissue by mass, and most of the remain-
ing material has a magnetic susceptibility and a density
similar to that of water [19]. This technique differs from
floatation, in that the diamagnetic force acts throughout
the body of the levitating object, at the molecular level,
not just at its surface, as is the case in buoyancy. In this
respect, the diamagnetic force can be compared to the
centrifugal force that balances the force of gravity on an
object orbiting the Earth or another planet.
Several of the authors have experience in testing the
effects of gravity using space laboratories [20-22] and
also in ground simulation facilities, such as the RPM
[23-25]. In the fruit fly, Drosophila,extensivegene
expression changes, as well as changes in the motility
behaviour of imagoes, occur in both real and simulated
microgravity. Conversely, hypergravity (up to 10 g) has a
relatively weak effect on motility and gene expression.
These changes can affect additional traits, such as the
life-span of the flies [26]. In addition, differences have
been observed in the motility and behaviour of the flies
exposed to real or simulated microgravity [20,26]
including the behaviour of levitating flies in a supercon-
ducting magnet [27].
In this paper, we investigated the effect of levitation (0
g*) and simulated hypergravity (twice Earth-gravity, 2 g*)
on the fruit fly Drosophila melanogaster using a spe-
cially-designed superconducting magnet with a closed-
cycle cryogenic system at the University of Nottingham
(Additional file 1, Figure S1). We levitated the flies in
continuous experiments of up to 22-days duration.
Effects were observed on the overall gene expression
patterns, and also a significant delay in the development
of the fruit flies from embryo to adult, compared with
control conditions in Earths gravity. The results were
compared with those of similar experiments in the
RPM. We also studied the effect on the flies of a spa-
tially uniform strong magnetic field (16.5 Tesla), in
which the diamagnetic forces were negligible so that
flies experienced normal gravity.
Methods
Levitation magnet and experimental arrangement
The superconducting magnet used to levitate the flies
has a 5-cm diameter vertical bore, which is tempera-
ture-regulated by forced air flow through the bore
(Additional file 1, Figure S1). The magnetic field is
strongest at the centre of the bore. The force on dia-
magnetic material in the magnetic field is proportional
to the product of the field strength, measured in Tesla
(T), and the field gradient, measured in Tm
-1
.The
expression for the effective gravity (g*) as a function of
magnetic field is given in Additional file 1[28]. The field
gradient is zero at the centre of the bore, and rises to a
maximum 85 mm above and below the centre.
When the field at the centre of the solenoid is 16.5 T,
the diamagnetic force on a sample of liquid water placed
approximately 80 mm above the centre is sufficient to
balance the force of gravity, so that the water levitates
as a freely-suspended droplet (Figure 1 and Additional
file 1, Figure S2). At the levitation point, the effective
gravity acting on the liquid is zero. The weight of the
same sample of water placed approximately 80 mm
below the centre of the solenoid is twice that outside
the magnet; in this sense, the effective gravity acting on
the water is twice Earths gravity (2 g).
Stable, containerless levitation of water and biological
material is possible using this magnet, but for a biologi-
cal specimen such as Drosophila, performing container-
less experiments is not practical, since it is impossible to
perform equivalent controls. Instead, the flies were con-
tained in three 25 mm-diameter, 10 mm-tall arenas
constructed within 25 ml clear plastic tubes (Figure 1
and Additional file 1, Figure S3). The tubes were placed
to enclose the stable levitation point, the centre of the
solenoid where the magnetic field was strongest and the
2gpoint of water respectively. For convenience we label
the three arenas, from top to bottom, 0 g*, 1 g*and2
g*. The asterisk (*) indicates that the g-value(0 g,1g,2
g) refers to the effective gravity on water at the centre
of each arena. It also serves as a reminder that there
was a strong magnetic field present in these arenas. The
magnetic field was 11.5 T at the centre of the 0 g* and 2
g* arenas and 16.5 T at the centre of the 1 g*arena.
Note that, in using the label 0g*to refer to the upper
arena in the magnet bore, we do not mean that the
effective gravity acting on liquid water within the 0 g*
arena was precisely zero everywhere. In fact, the effec-
tive gravity acting on water varied by a few percent
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 2 of 13
within all three arenas in the magnet bore, owing to the
spatial variation of the magnetic field. The effective
gravity acting on water inside the 0 g* tube was zero at
the levitation point, rising to approximately 5 × 10
-2
gat
the walls of the arena (Figure 1C and Additional file 1,
Figure S3). We use the term 0 g* only as a convenient
label for the arena and the tube in which it was
contained.
A fourth arena, labelled 1 g(i.e. without *), was placed
in an incubator well away from the solenoid, where the
magnetic field of the solenoid is small compared to the
Earths magnetic field. Video images of the flies in each
container were recorded using CCD cameras; white LED
lighting on a 12 hour photoperiod cycle provided illumi-
nation for the videos.
There have been reports that static high magnetic
fields (several Tesla fields with no levitation) can pro-
duce effects on bacteria [29], plants [30,31], mammals
[32], flies [33] and the development of frog eggs [34]. By
performing experiments simultaneously at 0 g*, 1 g* and
2g* and in the control outside the magnet (1 g), we
were able to distinguish unambiguously between the
effects on the flies of altering their net effective weight,
and any other effects of the high magnetic field.
Biological experiments
To maximise the output of our magnet experimental
time we performed three experiments under different
environmental conditions inside the magnet, which, for
convenience, we refer to by the duration of the experi-
ment (short-term,medium-term,long-term).
1) Short-term(1 day) experiment
Virgin females were mated overnight with an excess of
young males. Male and female 1-2 day old Oregon R
Drosophila melanogaster flies were both exposed to dia-
magnetic levitation for the following 26 hours at 14°C.
During this time, their behaviour was monitored using
CCD cameras. After exposure to the magnetic field, the
eggs laid inside the magnet were counted and were
allowed to hatch in order to monitor the normality of
the developmental process of eggs formed during levita-
tion conditions. Both males and females were homoge-
nised separately to study their gene expression profiles
by microarray analysis. Our choice of 14°C for the bore
ms-2
A B
z (mm)
C
ī (ms-2)
Figure 1 Effective gravity along the magnet bore and arena container properties.A) Side view of an arena contained within a transparent
plastic tube. The flies were constrained to the volume indicated by the red rectangle, between a cellophane disc (retained by two black o-rings
visible in the picture) and a semi-solid culture medium (off-white material at bottom of tube). B) Effective gravity acting on water within the 0
g* tube. The colour indicates the magnitude of the effective gravity (ms
-2
). Arrows show the magnitude and direction of the effective gravity.
The residual gravity on the flies (within the red rectangle) is less than 5 × 10
-2
g. C) Effective gravity acting on water on the solenoid axis, Γ,asa
function of vertical position, z. The centre of the solenoid is at z= 0 mm. The arenas were placed between the pairs of horizontal red lines
shown on the plot.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 3 of 13
temperature was based on preliminary observations indi-
cating that any effects may be amplified by suboptimal
growth conditions [21]. For comparison, we performed
an additional control well away from the magnet (i.e. far
enough away that it was not influenced by the field of
the superconducting magnet), but otherwise under simi-
lar conditions.
2) Medium-term(5 day) experiment
Pupae (OregonR Drosophila early pupae) collected in
Madrid and transported to Nottingham at 14°C were
incubated in the magnet for 5 days at 24°C, to analyse
the effect on the metamorphosis of the flies replicating
the design of a real space experiment performed on-
board the ISS [21]. Pupae were allowed to develop into
adults inside the magnet to study phenotypic alterations.
Recently-hatched flies were separated into males and
females and then homogenised in bulk. RNA was
extracted and assayed using microarrays to determine
the effect of the magnetic field on the expression
profile.
3) Long-term(22 day) experiment
The samples (recently-laid OregonR Drosophila eggs)
were exposed to the magnetic field for 22 days at 19°C
to analyse the effect of the magnetic field and altered
effective net weight of the flies on their entire life cycle,
from embryo to embryo. The larvae were allowed to
pupate in situ and a second generation of flies hatched
under these conditions. The second generation imagoes
were removed from the magnet and separated into
males and females. The males were homogenised to
extract the RNA and assayed for gene expression using
microarrays. The females were allowed to lay eggs in
order to monitor embryo development. In addition to
the 19°C experiments, we also performed an additional
external control (i.e. outside the magnet) at 24°C, and a
replicate long-term experiment (i.e.22daysat19°C)in
the RPM (0 g
RPM
,1g
RPM
) at the Dutch Experiment Sup-
port Center, DESC at OCB-ACTA, VU-University of
Amsterdam, the Netherlands.
Table 1 summarises the experimental design of the
short-term (1 day), medium-term (5 days) and long-
term (22 days) experiments. The 54 hybridised RNA
samples were identified with a CEL file name and classi-
fied by experimental duration, gender and environmen-
tal conditions (g* force, magnetic field, temperature).
This microarray dataset has been published in the
Table 1 Description of the 54 microarray samples used in this paper
Experimental design Gender Ground
Based Facility
Condition (effective force, g*) Name of CEL file replicates
Short-term Magnet 0 g* S0A S0B S0C
(26 h/14°C) Magnet 1 g* S1A S1B S1C
Exposure: Females Magnet 2 g* S2A S2B S2C
From 1-2 days to 1g(14°C) SCA SCB SCC
2-3 days imagoes 1g(24°C) SDA SDB SDC
Magnet 0 g* X0A X0B X0C
Magnet 1 g* X1A X1B
Males Magnet 2 g* X2A X2B X2C
1g(14°C) XCA XCB
1g(24°C) XDA XDB
Medium-term Magnet 0 g* M0A
#
——
(5d/24°C) Females Magnet 1 g* M1A
#
——
Exposure: 1 g MCA
#
——
From early pupae Magnet 0 g* N0A N0B
to recently Males Magnet 1 g* N1A N1B N1C
hatched imagoes 1gNCA NCB NCC
Long-term Magnet 0 g* L0A L0B L0C L0D
(22d/19°C) Magnet 1 g* L1A L1B L1C
Exposure: Males Magnet 2 g* L2A
#
——
From embryo to 1gLCA LCB LCC
mature imagoes RPM 0 g
RPM
R0A R0B
1g
RPM
RCA RCB RCC
From the group of 54 samples, 50 have been used in the main analysis, while four (not replicated, orphan
#
, microarrays shown dashed in the table) have been
used only for comparisons or as internal controls. Submission to the MIAME compliant Array Express Archive at EMBL_EBI has been done [EMBL_EBI:E-MEXP-
2082].
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 4 of 13
MIAME compliant Array Express Archive [EMBL_EBI:
E-MEXP-2082].
Gene expression analysis using the Drosophila microarray
Total RNA extracted from fruit flies was hybridised
using Affymetrix
®
Drosophila 2.0 whole genome Gene-
Chip arrays. We analysed two to four independent bio-
logical replicates of the experiments including RNA
from ten flies per array; the number of arrays in each
condition is indicated in Table 1. Only one array was
hybridised for females from the medium-term experi-
ment and for male flies from the 2 g* tubes in the long-
term experiment, owing to reduced quality of these
samplesreplicates. Differentially-expressed genes were
detected using a Volcano-plot comparison with Gene-
Spring GX 10 software (version 2.1), with a p-value cut-
off of 0.05 (except in the four single-array conditions
highlighted in italics on Table 1) and a fold difference >
1.7 including a parametric test that assumed equal
variances.
A global and integrative analysis using gene expres-
sion dynamics inspector(GEDI) self-organising maps,
was performed using the above software [35]. Firstly, we
applied the Robust Multichip Average (RMA) algorithm
for background correction, normalisation and expres-
sion-level summarisation of the arrays (see above). Sec-
ond, we identified 18921 probe sets that showed an
expression above the 20
th
percentile, relative to the 1 g
control, in half of the experimental conditions. For these
sets, we calculated the average signal ratio, and used
this value for GEDI analysis automatic clustering.
Mosaics of 20 × 16 grid size (average of 59 genes/tile)
were obtained for each condition using the standard set-
tings of the software (more methodological information
can be found in Additional file 1).
Results
A) Diamagnetic levitation of Drosophila melanogaster
In the 0 g* tube, we observed flies levitating freely (i.e.
not in contact with any surface, or flying) within 1-2
mm of the levitation point of liquid water (Additional
file 2, and [27]). This is not unexpected because the
flies have a high water content. The net effective
weight of a freely levitating fly is zero, in the sense
that there is no net (gravitational plus magnetic) force
on the fly. Although we observed a few flies levitating
freely, the majority remained in contact with the
walls, floor and ceiling of the arena enclosed within
the tube.
The net effective weight of an individual fly on the
walls, floor or ceiling depends on its position within
the arena (Figure 1C). The effective weight is less than
5% of its weight outside the magnet throughout the
arena.
We observed that the levitation position of the freely
levitating flies varied with their hydration. Dehydrated
(dead) flies levitated a few millimetres lower in the mag-
net than living flies. There was a 1-2 mm difference in
levitation position between each of the Drosophila
stages. This is consistent with the greater water content
of the embryos and early larvae (more than 80% of total
mass), reaching the lower level at late pupae (less than
70% of total mass) in comparison with an average 75%
of water content in adults [36,37].
B) Delay of development due to the magnetic field
The results of the long-term(22-day) experiments
demonstrated that 1-12 hour old embryos can develop
fully, progressing from larvae to pupae to imagoes, both
in the RPM and in a strong magnetic field up to 16.5 T.
Development in the magnet (0 g*, 1 g*and2g*) was
slightly but reproducibly delayed by one day, compared
to the 1 gcontrol outside the magnet, suggesting that
metamorphosis can be delayed in one or more develop-
mental checkpoints. A less evident delay in development
was observed in the RPM.
Table 2 shows the number of flies that developed
from eggs laid in the magnetic field in a short term
(1-day) experiment. After the females were mated, 25
males and 25 females were selected randomly and
placed together in the same container in the magnet for
26 hours at 14°C. The eggs laid during those 26 hours
were incubated outside the magnet and the flies that
developed from the eggs were counted. The results
show that exposure to the strong magnetic field during
oogenesis and laying caused a large reduction in the
number of adult flies that developed from the eggs. The
number of flies that developed from eggs laid in the
16.5 T magnetic field (1 g* tube) was just 31% of the
number that developed from eggs laid in the 1 g control
tube outside the magnet. In the 0 g*and2g*tubes,
where there was a significant field gradient, the
Table 2 Imagoes developed from eggs laid during the
medium term experiment
Control at
24°
Control at
14°C
1g*
14°C
0g*
14°C
2g*
14°C
Total number of flies 152 86 27 5 4
% of flies in relation to
14°C
177% 100% 31% 6% 5%
% of flies in relation to
24°C
100% 57% 18% 3% 3%
25 females and 25 males were exposed to the magnetic field for 26 hours
continuously. Prior to this, the males and females had been kept together for
24-48 hours. The table shows the number of eggs, deposited by the females
whilst inside the magnet (0 g*, 1 g*, 2 g*), that went on to develop into
imagoes outside the magnet. Two controls, one at 24°C and one at 14°C
outside the magnet were performed.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 5 of 13
reduction in the number of flies was even greater, being
only 5-6% of the number resulting from the 1 g control.
C) Magnetic field affects gene expression
Using Affymetrix whole genome microarrays (Droso-
phila version 2.0 with 18952 probesets and GeneSpring
GX), we analysed the gene expression profile of Droso-
phila exposed to the magnetic field, and compared the
results with Drosophila in a temperature-controlled
incubator placed well away from the magnet (1 g). We
also compared the results of short-term,medium-
termand long-termexperiments. In most experi-
ments we performed three replicates, except in a small
number of cases (identified by dashes in Table 1) where
we were unable to obtain one or more replicates owing
to random contamination of the extractions, or time
constraints on the use of the magnet. We also compared
microarray results from the long-termmagnet experi-
ment with 5 microarrays from a RPM microgravity
simulation experiment. The transcriptional profiles of
each experiment are shown in Figure 2. Here, the pro-
files have been presented as a condition tree,inwhich
similarities between transcriptional profiles are reflected
in the grouping of the experiments within the tree. For
example, the transcriptional profiles from the 0 g*, 1 g*
and 2 g*tubesinthelong-termexperiment display
readily identifiable similarities and so are grouped
together in the diagram. Likewise, common features can
also be identified in the profiles from the medium-
termand short termexperiments. There were signifi-
cant differences in the profiles from experiments of dif-
ferent duration. For example, there were clear
differences between the profile of flies in a 0 g*tubein
a long-term experiment and the profile of flies in a 0 g*
tube in a short-term experiment. The variation between
experiments of different durations was smaller for
females than for males. These results showed that the
precise biological state of the organism (i.e. age, gender,
temperature) was important in the magnetic field effect.
The Venn diagram in Additional file 1, Figure S4
shows the number of genes that changed their expres-
sion levels in the magnet compared with the 1 gexternal
controls located well away from the magnet. The num-
ber of genes affected was nearly independent of the
duration of the experiment, or the location (0 g*, 1 g*or
2g* arena) inside the magnet. Interestingly, the group of
genes affected in the long-term experiment was different
from the group affected by the medium-term experi-
ment. Similarly, the group of genes affected by the
short-term experiment was different from both the
long-term and medium term experiments, although
there was some overlap in the three groups of genes
affected. We identified 496 genes that were sensitive to
the strong magnetic field in males, i.e. genes that are
up- or down-regulated in one or more of the long-,
medium- and short-term experiments in the magnetic
field. Of this group of 496 genes, 105 were common to
two different experiments. We found only one gene
common to all three experiments (long, medium and
short-term), namely CG33070-RB, Sex Lethal, encoding
an RNA binding protein.
In females, 474 genes were sensitive to the magnetic
field, of which 115 were common to flies in two differ-
ent experiments. Fourteen genes were common to flies
in all three experiments; three of these genes have been
annotated as heat shock proteins.
Less than 10% of the magnetic field-sensitive genes
were common to both males and females; 5 were
observed in two or more experiments, with 47 observed
in one or more experiments.
We also analysed the short-term, medium-term and
long-term experiments separately, in order to identify
the differentially-expressed genes induced by the mag-
netic field. The number of genes sensitive to the mag-
netic field in the individual experiments is shown in a
series of Venn diagrams in Additional file 1, Figure S4.
D) Isolating the effect of magnetically-altered effective
weight from other effects of the strong magnetic field
The above results indicate that the strong magnetic field
present in all three tubes (0 g*, 1 g*and2g*) had a
Ƃ
LONG TERM
Ƃ
ƃ
ƃ
ƃ
19ºC
24ºC
14ºC
MEDIUM TERM
SHORT TERM
SHORT TERM
MEDIUM TERM
0g*
1g*
2g*
1g
1g*
0g*
1g
0g*
2g*
1g*
1g14ºC
1g
1g24ºC
0gRPM
1g*
0g*
1g
0g*
2g*
1g*
1g14ºC
1g24ºC
LONG TERM ƃ
19ºC
24ºC
14ºC
Figure 2 Clustering of the 22 analysed transcriptional profiles.
Clustering of the transcriptional profiles is revealed by a condition
tree calculated with a hierarchical cluster algorithm, using Pearson
absolute distance metric and the average linkage rule. The length of
the branch is an indicator of the number of gene expression
variations found between each condition and experiment (shorter
distances indicate a greater resemblance).
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 6 of 13
significant effect on gene expression of the flies. In order
to locate genes that could play a role in gravisensing or
adaption, it was necessary to account for the effects of
the strong magnetic field. We attempted to isolate the
effect of the vertical diamagnetic force on flies (which
alters the effective net weight of the flies), from any
other effects of the magnetic field by comparing the
gene response of the flies in the 0 g*and2g*tubesto
those in the 1 g* tube
Two different approaches were used:
Approach 1: A list of genes that were up- or
down-regulated in the 0 g* tube in the magnet was
compiled and compared with the external control
outside the magnet (1 g). We repeated this for the 1
g*and2g* tubes, and then removed from the 0 g*
vs 1 gand 2 g*vs1glists those genes that appeared
in the 1 g*vs1glist.
This approach was based on the results in Addi-
tional file 1, Figure S4, first and second rows, in
which it is evident that few genes were common to
flies in all tubes in the magnet (0 g*, 1 g*, 2 g*) and
the 1 gcontrol tube. We found that between 20%
and 50% of the genes affected by a change in effec-
tive gravity (0 g*or2g*) were present in flies in
both 0 g* and 2 g* tubes.
Approach 2: We compiled two lists of genes: one
of genes that were up- or down-regulated in 0 g*
compared with 1 g*,andoneofgenesthatwere
altered in 2 g* compared with 1 g*.
In both approaches, we made the prior assumption
that the magnetic field effects observed in the 1 g*tube
(at 16.5 T) had nearly the same influence on the genes
in the 0 g*and2g* tubes where the field was smaller
(11.5 T).
The numbers of genes in the lists resulting from the
two procedures described are shown in the highlighted
third row of Additional file 1, Figure S5. The results
from the RPM experiment (described in the Methods
section) are also shown for comparison. The five gene
ontology groups with the highest statistical significance
in each gene list are in the table in Additional file 1, Fig-
ure S5. Most of the individual genes affected were not
the same in the different tubes (0 g*, 1 g*, 2 g*). How-
ever, we could identify a common theme in the affected
gene groups, which consisted of defence/immune/stress
response and cell signalling gene ontology (GO) groups.
Figure 3A lists those genes with an increase in expres-
sion of 2.5 fold or more, or decrease of 0.4 fold or
more, at 1 g*, compared to the 1 gcontrol (at 0 T).
Those genes with the largest change in expression are
listed towards the top of the table. In Figure 3B, we list
genes that have a similar significant change in expres-
sion at 0 g*, compared to 1 g*. Figure 3C shows the
samelistforthechangeingeneexpressionat2g*
A
) B)
Probe set Name Description
1g* vs 1g control
outside magnet
Fold
Change Probe set Name Description 0g* vs 1g*
Fold
Change
1634409_at CG31775-RA
A
DH region gene
L
ong-term males + 2.677 1633471_at CG11765-RA
P
eroxiredoxin 2540
M
ed-term males + 5.279
1623489_at CG11091-RA Sphinx Long-term males 2.492 1629530_at CG15066-RA Immune induced molecule 23 Long-term males 3.804
1630568_at Dm P. unknown func. DUF753 Med-term males 0.394 1634395_at CG13804-RA Royal jelly protein Short-term females 3.731
1626319_a_at CG18279-RB Immune induced mol. 10 Long-term males 0.394 1625061_at CG17298-RA Circadian rithm related
M
ed-term males + 3.660
1637059_s_at CG10578-RA DnaJ-like-1 Med-term males 0.391
1630291_at CG4312-RA Metallothionein B Short-term males 3.566
1626429_at CG6821-RA
arval serum prot.1gamma
M
ed-term males + 0.381 1627659_at CG32510-RA Unknown Short-term females + 3.232
1640035_at CG3763-RA
F
at body protein 2 (fbp2)
M
ed-term males + 0.375 1630523_at CG12106-RA Unknown Short-term females 2.835
1625985_at CG31446-RA Calycin -ike Binding funct. Long-term males 0.369 1639912_x_at CG40323-RB Defense response Short-term males 2.713
1634143_at CG8345-RA Monooxigenase Med-term males 0.364 1634591_at CG11878-RA P. Kinase-like Long-term males 2.660
1639019_s_at CG18279-PA Immune ind. mol. 10 Long-term males 0.355 1634265_at CG17924-RA Accessory gland-spec.pept.95EF Short-term females 2.576
1640755_at CG10248-RA Cytochrome P450-6a8 Med-term males 0.348 1637261_at CG15351-RA Chorion protein c at 7F Short-ter m females 2.459
1630291_at CG4312-RA Metallothionein B Short-term males 0.334 1632271_a_at CG7971-RA Spliceosome complex Med-term males 0.392
1633607_at CG2444-RA SVC secreted protein
M
ed-term males + 0.323 1630723_a_at CG8708-RB ȕ-1.3-galactosyltransferase a. Short-term males 0.384
1630800_s_at AM01323_revcomp Med-term males 0.315 1629356_at CG10122-RA RNA polymerase I subunit Med-term males 0.382
1626351_at CG4471-RA Tetraspanin 42Ep
M
ed-term males + 0.314 1639036_at CG8857-RC Ribosomal protein S11 Med-term males 0.375
1625063_a_at CG31606-RB Unknown Long-term males 0.313 1641207_s_at CG4991-RB Aa-polyamine transporter Med-term males 0.361
1630561_at CG13641-RA Unknown (Nuclease b.) Med-term males 0.312 1637499_s_at CG5953-RA MADF-like (cell cycle) Long-term males 0.359
1629061_s_at CG32041-RB Unknown
M
ed-term males + 0.308 1638000_at CG4922-RA Spalt-adjacent Short-term females 0.356
1624101_at CG10242-RA Monooxigenase Long-term males 0.304 1623601_at CG8221-RA Amyrel (alfa-amylase) Short-term females 0.334
1623950_s_at CG2198-RB
A
malgam
M
ed-term males + 0.303 1629845_at CG13835-RA Phospholipase A2 activity Long-term males 0.286
1626439_at CG15353-RA Mating/Starvation related Long-term males 0.298 1622946_at CG6908-RA DUF227 (P. Kinase-like) Long-term males 0.286
1630590_at CG18417-RA Proteolyses related Long-term males 0.298 1637357_at CG9463-RA
N
-glycosyl Hydrolase Short-term females 0.223
1631701_a_at CG8502-RC
L
arval cuticle c.
M
ed-term males + 0.295 1625373_at CG6966-RA Cell proliferation Long-term males 0.209
1623815_at CG8100-RA
L
arval serum c.
M
ed-term males + 0.286 1633834_at CG9701-RA
N
-glycosyl Hydrolase Long-term males 0.206
1631803_at
1631962_at
CG2559-RA
CG33192-RA
L
arva
l
serum prot. 1alpha
Metallothionein D
M
ed-term males +
Short-term females
0.272
0.245 C) Probe set Name Description 2g* vs 1g*
Fold
Change
1638872_at CG5436-RA
H
eat shock protein 68
M
ed-term males + 0.209 1626821_s_at CG6489-RA Heat-shock-p70Ba,b,c Short-term females 7.312
1629530_at CG15066-RA Immune induced mol. 23 Long-term males 0.182 1632841_x_at CG6489-RA Heat-shock-p70Bc Short-term females 5.637
1627271_at CG13422-RA Glucosidase activity Long-term males 0.175 1638872_at CG5436-RA Heat shock protein 68 Short-term females 4.685
1633471_at CG11765-RA Peroxiredoxin 2540 Med-term males 0.160 1634187_x_at Transposon ī-element:CR32865 Short-term females 4.250
1636293_at CG2217-RA Defense to bacteria Long-term males 0.107 1637516_at CG15449-RA Unknown Long-term males 3.999
1627551_s_at CG18372-RA
A
ttacin-A & B
L
ong-term males + 0.069 1630258_at CG4181-RA Glutathione S transferase D2 Short-term females 3.852
1626821_s_at CG6489-RA
H
eat-shock-p70B,b,c
M
ed-term males + 0.044 1636524_at Dm Hsp70 like Long-term males 3.717
1639571_s_at CG18743-RA
H
eat-shock-p70Aa,b
M
ed-term males + 0.041 1641055_at CG4463-RA Heat shock protein 23 Short-term females 3.449
1632841_x_at CG6489-RA
H
eat-shock-p70Bc
M
ed-term males + 0.034 1627271_at CG13422-RA Glucosidase activity Short-term females 3.184
1625124_at CG10146-RA
A
ttacin-A
L
ong-term males + 0.028 1639571_s_at CG18743-RA Heat-shock-p70Aa,b Short-term females 2.782
A) Magnetic effect: 1g* versus control outside the magnet 1g. B) and C) Gravity related effects, 0g* and 2g*
versus 1g* respectively. Genes in which the expression is also altered in females medium-term experiment
are marked with a ‘+’ (the fold change in med-term females is not shown due to a lack of statistical support).
Arrows identify genes common to both lists. Dotted lines are used where the gene expression responds in the
same way to the magnetic field in both lists. Solid lines are used where the genes behave in opposite ways.
1625673_at CG12983 Unknown Long-term males 2.779
1623068_at CG4105-RA Cytochrome P450-4e3 Long-term males 0.342
1632683_s_at Transposon Gag-int-pol (copiaGIP) Long-term males 0.321
1640755_at CG10248-RA Cytochrome P450-6a8 Long-term males 0.286
1622946_at CG6908-RA DUF227 (P. Kinase-like) Long-term males 0.187
1633378_at CG9976-RA Galactose-specific C-type lectin Long-term males 0.145
Figure 3 List of genes that respond to the magnetic field with a fold increase of more than 2.5 or fold decrease of more than 0.4.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 7 of 13
compared to 1 g*. The genes listed in Figure 3B and 3C
are those which were affected by the vertical diamag-
netic force alone, and not by any other effects of the
magnetic field. Since the vertical diamagnetic force
altered the effective net weight of the flies, these genes
may have a role in gravisensing or altered gravity
adaption.
There was only one gene with a similarly significant
change in expression in the RPM experiment (an
unknown function gene, CG15065 with a 0.365 fold
expression change in the RPM). Some of the groups of
genes identified in section C appear again in the lists
showninFigure3.Ofparticularinterestaregenesthat
appear in more than one experiment (for instance, those
in italics that are also present in the medium-term
experiment on females, orphan arrays) owing to their
additional statistical significance.
As shown in Figure 3A, there were only two genes (of
unknown function) that were significantly up-regulated
at 1 g*comparedtothe1gcontrol. Of those that were
significantly down-regulated, the types of genes that
appeared most frequently were those related with heat
shock, immune/defence response, oxidation and lipid
processes. It is interesting that the same heat shock
genes that appear in this list also appeared in the 2 g*vs
1g* list (Figure 3C), but in the latter they were over-
expressed; the Heat-shock-p70gene was severely
down-regulated in 1 g* compared to 1 gin males in a
medium-termexperiment, and up-regulated in 2 g*
compared to 1 g* in females in a short-termexperi-
ment. None of the heat-shock genes that appeared in
Figure 3A appeared in the 0 g*vs1g* list (Figure 3B),
but two immune response genes (Peroxiredoxin 2540
and Immune Induced molecule 23) were common to
both of them, being down-regulated in 1 g*andup
regulated in 0 g* experiments.
The sensitivity of these genes to the magnetic field
was made more evident when we integrated these genes
into a virtual cellular pathway. We used Pathway Studio
6.01 for a graphical output of the relations among the
genes in each group. Additional file 1, Figure S6 shows
the magnet affected genes (Figure 3A), including some
connector genes in order to fill the gaps amongst them.
From the pathway, most of the genes were linked in less
than two steps with other affected genes, suggesting that
their functions were connected in the cell.
Many of the genes listed in Figure 3 have unknown
function, and it would be revealing to identify the func-
tion of each of these genes. One of these unknown-
function genes deserves special attention. CG6908
appeared in both 0 g* vs. 1 g* and 2 g* vs. 1 g* lists (Fig-
ure 3B &3C), and was strongly repressed (4 to 5 fold) in
both conditions. Gene CG6908 encodes a relatively large
protein with a PKC-like domain in the middle and was
not changed due to magnetic field (Figure 3A). There-
fore, this gene seems specifically sensitive to the mag-
netic field gradient (net gravity change).
We assessed how many of these magnetically-affected
geneswerealsopresentinthegroupof36genesthat
were up- or down-regulated in the RPM long-term
experiments. This result is shown in the grey box in
Additional file 1, Figure S4. We found that only one
gene was up- or down-regulated in both the RPM and
the long-termmagnet experiments, compared to the
relevant control (CG32641-RA; GenBank accession
number). This gene encodes a protein with Heat shock
protein/Chaperone DNAJ domains. Curiously, this gene
was altered only at 1 g* in the magnet, but not at 0 g*
tube, as one might expect. When we compared the
group of 36 RPM-altered genes with those altered in
any of the short-term experiments in the magnet, we
found that three genes were commonly affected. One of
these was Yuri Gagarin,a gene selected previously as
one of the gravity-response genes [38]. The other two,
automatically annotated unnamed genes, remain to be
analysed further.
E) Global transcriptional states in the magnet: GEDI
analysis
Taking into account that there are very few individual
genes that were affected consistently between short-,
medium- and long-term experiments, we analysed the
transcriptome status as a whole. We analysed the micro-
array data with the Gene Expression Dynamics Inspec-
tor(GEDI) program [35]. The GEDI software organises
the gene expression patterns into mosaics of nxmtiles.
Each tile corresponds to a cluster of genes that behave
similarly across conditions, designated a centroid. Differ-
ent colours reflect the expression intensity of a centroid
in each condition (in our case the average ratio of inten-
sities compared to 1 gcontrols). Additionally, GEDI
places similar centroids close to each other in the
mosaic, creating an image of the transcriptome and
allowing its analysis as an entity by simple visual com-
parison of the mosaics corresponding to different condi-
tions. For this analysis, we normalised the expression
data as indicated in the supplementary methods (Addi-
tional file 1). We used 18921 probe-sets for the GEDI
analysis. They were placed in 20 × 16 mosaics with an
average of 59 genes per centroid. The results obtained
are available as GEDI original files in Additional file 3
and summarised in Figure 4.
As expected from Results sections C and D, the tran-
scriptome obtained from the 0 g*tubesrespondeddif-
ferently in each of the experiments (short-, medium-
and long-term), and the response in 0 g* (compared to 1
gand 1 g* controls) depended on the sex of the flies. In
all cases, the response to the magnetic field was weak;
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 8 of 13
most of the genes showed a log to the base 2 ratio
change in expression of < 1.35 versus the control
expression, as indicated by the colour scale (Figure 4).
We observed a similarly weak response in the other two
tubes (1 g*, 2 g*) in the magnet. We note that the 0 g*
and 2 g* images seem to be more closely related to each
other than to the 1 g* image. A RPM produced an effect
superficially similar to that of the magnetic field (in the
1g* arena), in that the number of genes affected and
the expression change of these genes were similar to
that observed in the magnet, but the RPM affected dif-
ferent gene clusters (Additional file 1, Figure S4 and S5).
A change in temperature to 24°C produced a similar
effect. However, the sensitivity of the transcriptome to
the external physical parameter change (magnetic field,
field gradient, temperature, RPM movements) depended
on the biological state of the sample. For example,
female flies seem to be less sensitive to short-term
exposure to strong magnetic fields than males (the med-
ium-term experiment had the opposite trend, but this
could be due to the fact that the medium-term experi-
ment with females was carried out with insufficient
replicates due to time constraints). If we compare male
and female patterns in the 1 g* tube of the medium-
term experiment (in Additional file 1, Figure S4), it is
possible to localise a group of genes that are up-regu-
lated by the magnetic field in males, but down-regulated
in females, suggesting differential transcriptome adap-
tion responses to stress in selected populations of the
same species.
Interestingly, the last two columns on the right of Fig-
ure 4, showing the 0 g*vs.1g*responseand2g*vs.1
g* response do not show opposite trends in gene expres-
sion, as one might expect initially. This is especially
clear in experiments with male flies that produced very
similar transcriptome images in the GEDI analyses. An
GEDI (20x16 training)
18921 probesets
(Average tile size= 59 probesets)
Ƃ
LONG TERM
0g*1g*2g*
MEDIUM TERMSHORT TERM
ƃ
Ƃ
ƃ
ƃ
versus 1
g
control placed outside the GBF versus 1
g
* control
Mi
crograv
i
ty
(0g* vs 1g*)
H
ypergrav
i
ty
(2g* vs 1g*)
0gRPM
24ºC
24ºC
1.35
0.67
1.00
19ºC
24ºC
14ºC
Average tile signal log2 ratio
Figure 4 GEDI 20 × 16 clustering analysis based on the three magnet experiments and one experiment at RPM. One experiment per
row is shown for separate male and female data, in different Ground Based Facilities (GBF). The colour scale on the right indicates the average
log to the base 2 ratios of each cluster compared to the parallel 1 gcontrol for conditions 0 g*, 1 g*, 2 g* and others (first four columns), and
versus the 1 g* control in the fifth (0 g*vs1g*) and sixth (2 g*vs1g*) columns. The centre panel indicates the number of probesets (18921
probesets) included in each cluster (20 × 16 clusters with an average size of 59 probesets per cluster). Panels obtained from an orphan array are
indicated by a black dot. Panels not linked to the magnetic field effect (i.e. those in the fourth column) are enclosed by a dashed line.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 9 of 13
opposite trend has been observed in mechanical simula-
tors, RPM vs. centrifuge [21].
Discussion
A) Development and behaviour is modified by high
gradient magnetic fields
We have observed both embryonic development and
pupal metamorphosis under diamagnetic levitation con-
ditions (short- and medium-term experiments). Addi-
tionally, we observed a complete Drosophila life cycle,
from embryo to embryo, in a magnetic field of up to
16.5 T, over a 22 day long term experiment. In this
experiment, the number and rate of eggs laid could not
be evaluated due to the lack of in situ imaging. In a
later, short-term experiment, we observed that the early
stages of embryogenesis were affected by the strong
magnetic fields present in the magnet bore (a one third
decrease in fertility in the 1 g* tube, Table 2).
The observation of a decrease in the number of flies
resulting from eggs laid in the 1 g*tube,butagreater
decrease in flies resulting from eggs laid in the 0 g*and
2g* tubes, indicates that both magnetic field and the
field gradient played a role in this effect. We emphasise
that the magnetic field in the 0 g* and 2 g*tubesisless
than in the 1 g* tube; the field in these two tubes is 11.5
T, compared to 16.5 T in the 1 g* tube. The large field
gradient (~100 Tm
-1
)inthe0g* and 2 g* tubes (absent
in the 1 g* tube) altered the effective net weight of the
flies, which affected their motility, as reported in [27]. A
magnetic field in the absence of a field gradient (i.e.the
conditions in the 1 g* tube) can also affect the organism,
for example through magnetic alignment of biostruc-
tures in the magnetic field (see, for example,[34,39-42]),
or through induction of an electromotive force (by Fara-
days law) as the organism moves through the strong
magnetic field [39].
It is important to point out here that behavioural
changes, delays in development, and altered rate of ovi-
position have been described in several experiments in
orbiting spacecraft with Drosophila [43-46]. The final
conclusion of those experiments was that normal devel-
opment is possible in space[43-46], despite the micro-
gravity effects. Consistently, in our long term
experiments some flies were able to develop in each
magnet condition although their development was even
slower than in the RPM or in real space mission experi-
ments, so the effects observed in the short term experi-
ment were not completely deleterious. Changes in the
motility of diamagnetically levitated flies [27] could be
an explanation for a change in the oviposition rate;
embryo retention is a typical behavioural response to
environmental stress. This could explain the reduced
number of imagoes that developed from eggs laid in the
magnetic field, and in particular, at 0 g* and 2 g*.
B) Global transcriptional effects are observed indicating
that magnetic field affects some external stress response
elements
Microarray analysis of the RNAs indicated that there
were changes in the gene expression of imagoes that
were exposed transiently to, or developed during meta-
morphosis in the presence of a diamagnetic force (0 g*
and 2 g* tubes), and/or a high magnetic field (Figure 4).
The main conclusions from the gene expression analysis
are that the exposure of the samples to the strong mag-
netic field, with or without a field-gradient, caused sig-
nificant changes in the expression of immune, stress,
and temperature response genes (several heat shock pro-
teins, for instance, appear affected). It is notable that
experiments of different durations activated or repressed
different genes, although in similar GO groups (Figure
3), and also that these GO groups are similar to the
ones found in real microgravity conditions in space
[21,23]. Our results indicate that the vertical diamag-
netic force (that levitated the flies in the 0 g*tubeand
enhanced the net effective weight of the flies in the 2 g*
tube) had less of an influence on the transcription pro-
file than other effects of the strong magnetic field,
which was present in all three tubes (11.5 T in 0 g*and
2g*, and 16.5 T in 1 g*). Since the differences between
the samples in the magnet and samples placed well
away from the magnet increased with the duration of
the experiment, this suggests that the effects of the mag-
netic field accumulated with time. This could be
explained if the population was less synchronised in
long-termexperiments, and could also be due to dif-
ferent sensitivities of males and females to the magnetic
field, i.e. affecting to sexual or reproductive parameters.
C) Magnetic levitation is an alternative to microgravity
simulation on Earth allowing isolation of magnetic and
gravitational effect
By comparing the samples from the 0 g* and 2 g* arenas
(in which there was a strong magnetic field with large
field gradient) with samples from the 1 g* arena (strong
magnetic field, insignificant gradient), we have identified
a number of genes that were affected only by the pre-
sence of a strong magnetic field gradient (Figure 3). One
such gene, CG6908, was altered only by the presence of
the large field gradient, i.e. it was unaffected by a mag-
netic field with no field gradient. Since the field gradient
alters the flieseffective weight, we speculate that
CG6908 could play a role in a putative signalling cas-
cade involved in the gravity response of Drosophila.
Some interesting genes were altered by the magnetic
field,withorwithoutthefieldgradientinoursamples,
one of which was the Yuri Gagarin gene, a gravity-
response gene described previously in Drosophila [38].
We also located a number of genes in which ion/metal-
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 10 of 13
related enzymes were encoded (e.g. CG4312-RA). These
alterations could be related directly to the influence of
the magnetic field on the ions [47], or only by the dele-
tion/delocalisation of the ions/metals in the cytosolic or
extracellular medium. The gene expression experiments
showed that the effect of the magnetic field was com-
parable to the effect of altered gravity. With careful
design of the control experiments, we have demon-
strated how the effect of magnetically-altered gravity on
gene expression can be distinguished from other effects
of the strong magnetic field.
D) Effect of strong magnetic field on oogenesis
One of the most striking results of this study was an
inhibition of oogenesis, or at least its attenuation, by
magnetic fields, a fact already observed with weaker
and short-lasting magnetic fields [33] and consistent
with the gender-related differences in microarray ana-
lysis. Several decades ago it was suggested that during
the first stages of embryogenesis some electrical poten-
tial differences around the embryo may be involved in
the origin/maintenance of axis formation, which allows
the developmental program to continue [48]. These
axes are believed to be created or oriented by the grav-
ity vector (at least originally). Our results suggest that
this process is still active in the embryos, and that dia-
magnetic levitation can partially suppress the genera-
tion of the charge distribution at the embryo surface
[34]. Additionally, the effect on the phenotype is much
more severe when gravity plus charges are simulta-
neously lost, causing more than 90% of the eggs to fail
to complete embryogenesis and metamorphosis to ima-
goes (Table 2). A similar synergistic effect has been
observed already in spaceflight and simulated micro-
gravity experiments under suboptimal environmental
conditions [21].
Conclusions
In summary, we have performed a series of experiments
to examine the effects on fruit flies of magnetically-
induced weightlessness (0 g*) and simulated hypergravity
(2 g*), using exposure times up to 22 days. The data for
0g* (diamagnetically levitated) conditions show signifi-
cant similarities with those obtained in related experi-
ments in which a Random Positioning Machine was
used to simulate microgravity.
For both 0 g*and2g* conditions, a significant delay
was observed in the development of the flies from
embryo to adult, compared to normal gravity condi-
tions and also significant changes in the expression of
immune, stress and temperature response genes. We
detected similar but weaker effects on the flies
exposed to the strong magnetic field only at the 1 g*
condition.
Experiments on gene transcription are sensitive to
small variations in environmental conditions. The ability
to perform experiments simultaneously under the same
conditions of lighting, air temperature, pressure and
humidity, enables us to attribute unambiguously the
effects we observe in the magnet to the altered effective
gravity environment and magnetic field.
The effects of the magnetic field and magnetic field
gradient reported here suggest that the transcriptome is
finely tuned to the environmental conditions and that
relatively small differences in the design of an experi-
ment or the population chosen could lead to different
gene expression profiles.
Additional material
Additional file 1: Supplementary online material including
additional methods and figures S1 to S6.
Additional file 2: Ten minute-long clip of levitating flies in the 0 g*
arena (QuickTime format) also available with a Drosophila walking
discussion at [27].
Additional file 3: GEDI analysis files, including each cluster list of
probesets and their expression ratio for each condition (zip file).
Acknowledgements
Professor Roberto Marco died on June 27th, 2008 from cancer. He made a
major contribution to planning and carrying out this project, and this paper
is dedicated to his memory. This work was supported by grants from the
Spanish Space Program in the Plan Nacional de Investigacion Cientifica y
Desarrollo TecnologicoESP2006-13600-C02-01 and -02, AYA2009-07792-E
awarded to RM, FJM and RH. RPM access was possible due to the Dutch
Space Research Organization NWO-ALW-SRON grant MG-057 to JJWAvL.
Research using the superconducting magnet was supported by grants from
the UK Engineering and Physical Sciences Research Council (EPSRC), Nos.
GR/S83005/01 and EP/G037647/1. RJAH acknowledges EPSRC for support of
a Research Fellowship (EP/I004599/1). RH acknowledges CSIC for support of
a Research Fellowship (JAE-Doc 2008 program).
Author details
1
Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, E-28040,
Madrid, Spain.
2
Departamento de Bioquímica, Universidad Autónoma de
Madrid (UAM), Arzobispo Morcillo s/n, E-28029, Madrid, Spain.
3
School of
Biosciences, University of Nottingham, Sutton Bonington Campus,
Loughborough, LE12 5RD, UK.
4
School of Physics & Astronomy, University of
Nottingham, Nottingham NG7 2RD, UK.
5
Dutch Experiment Support Center,
DESC at OCB-ACTA, VU-University and University of Amsterdam, Amsterdam,
the Netherlands.
Authorscontributions
RH carried out RPM experiments, both RPM and magnet sample processing,
molecular genetic studies and data analyses and drafted the manuscript.
OJL, CED and RM conceived the study, designed the experimental apparatus
for the magnet system and carried out the experiments in the magnet. OJL
constructed the apparatus. RJAH developed the experimental apparatus with
OJL, performed the calculations of the magnetic field and effective gravity,
operated and advised on the use and effects of the magnet system, and
edited the manuscript. PA, MRD, LE (magnet experiments), FJM and JJWAL
(RPM experiment) participated in the study design and coordination and
helped to draft the manuscript. All authors, except the late Professor RM,
read and approved the final manuscript.
Received: 15 July 2011 Accepted: 1 February 2012
Published: 1 February 2012
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 11 of 13
References
1. Dubinin N, Vaulina E: The evolutionary role of gravity. Life Sci Space Res
1976, 14:47-55.
2. Ross M: The influence of gravity on structure and function of animals.
Adv Space Res 1984, 4:305-314.
3. Tairbekov MG, Klimovitskii V, Oganov VS: [The role of gravitational force in
the evolution of living systems (the biomechanical and energy aspects)].
Izv Akad Nauk Ser Biol 1997, 517-530.
4. Volkmann D, Baluska F: Gravity: one of the driving forces for evolution.
Protoplasma 2006, 229:143-148.
5. Rosen E, Chen R, Masson PH: Root gravitropism: a complex response to a
simple stimulus? Trends in Plant Science 1999, 4:407-412.
6. Leys NM, Hendrickx L, De Boever P, Baatout S, Mergeay M: Space flight
effects on bacterial physiology. J Biol Regul Homeost Agents 2004,
18:193-199.
7. Drysdale R: FlyBase: a database for the Drosophila research community.
Methods Mol Biol 2008, 420:45-59.
8. van Loon JJWA: Some history and use of the Random Positioning
Machine, RPM, in gravity related research. Adv Space Res 2007,
39:1161-1165.
9. van Loon JJWA, Veldhuijzen JP, Kiss J, Wood C, vd Ende H, Guntemann A,
Jones D, de Jong H, R W: Microgravity Research starts on the ground.
Apparatuses for long-term ground based hypo-and hypergravity studies.
Proceedings of the 2nd European Symposium on the Utilisation of the
International Space Station ESTEC, Noordwijk, The Netherlands,16-18 November
1998 1999, ESA SP-433:415-420.
10. Coleman CB, Gonzalez-Villalobos RA, Allen PL, Johanson K, Guevorkian K,
Valles JM, Hammond TG: Diamagnetic levitation changes growth, cell
cycle, and gene expression of Saccharomyces cerevisiae. Biotechnol
Bioeng 2007, 98:854-863.
11. Dijkstra CE, Larkin OJ, Anthony P, Davey MR, Eaves L, Rees CE, Hill RJ:
Diamagnetic levitation enhances growth of liquid bacterial cultures by
increasing oxygen availability. J R Soc Interface 2011, 8:334-344.
12. Guevorkian K, Valles JM Jr: Swimming Paramecium in magnetically
simulated enhanced, reduced, and inverted gravity environments. Proc
Natl Acad Sci USA 2006, 103:13051-13056.
13. Valles JM Jr, Lin K, Denegre JM, Mowry KL: Stable magnetic field gradient
levitation of Xenopus laevis: toward low-gravity simulation. Biophys J
1997, 73:1130-1133.
14. Valles JM Jr, Maris HJ, Seidel GM, Tang J, Yao W: Magnetic levitation-based
Martian and Lunar gravity simulator. Adv Space Res 2005, 36:114-118.
15. Beaugnon E, Tournier R: Levitation of organic materials. Nature 1991,
349:470.
16. Beaugnon E, Tournier R: Levitation of water and organic substances in
high static magnetic fields. J Phys III France 1991, 1:1423-1428.
17. Berry MV, Geim AK: Of flying frogs and levitrons. Eur J Phys 1997,
18:307-313.
18. Hill RJ, Eaves L: Nonaxisymmetric shapes of a magnetically levitated and
spinning water droplet. Phys Rev Lett 2008, 101:234501.
19. Schenck JF: Health and physiological effects of human exposure to
whole-body four-tesla magnetic fields during MRI. Ann N Y Acad Sci 1992,
649:285-301.
20. de Juan E, Benguría A, Villa A, Leandro LJ, Herranz R, Duque P, Horn E,
Medina FJ, Loon Jv, Marco R: The AGEINGExperiment in the Spanish
Soyuz Mission to the International Space Station. Microgravity Sci Technol
2007, 19:170-174.
21. Herranz R, Benguria A, Lavan DA, Lopez-Vidriero I, Gasset G, Medina FJ, van
Loon JJ, Marco R: Spaceflight-related suboptimal conditions can
accentuate the altered gravity response of Drosophila transcriptome.
Mol Ecol 2010, 19:4255-4264.
22. Matía I, González-Camacho F, Herranz R, Kiss JZ, Gasset G, Loon JJWAv,
Marco R, Medina FJ: Plant cell proliferation and growth are altered by
microgravity conditions in spaceflight. Journal of Plant Physiology 2010,
167:184-193.
23. Herranz R, Benguria A, Fernández-Pineda E, Medina FJ, Gasset G, van
Loon JJ, Zaballos A, Marco R: Gene Expression Variations During
Drosophila Metamorphosis in Space. The GENE Experiment in the
Spanish Cervantes Mission to the ISS. J Gravit Physiol 2005, 12:253-254.
24. Herranz R, Laván D, Benguría A, Duque P, Leandro L, Gasset G, Zaballos A,
Medina FJ, Loon JJv, Marco R: The GeneExperiment in the Spanish
Soyuz Mission to the International Space Station. Effects of cold
transportation. Microgravity Sci Technol 2007, 19:196-200.
25. Herranz R, Laván DA, Dijkstra C, Larkin O, Davey M, Medina FJ, van
Loon JJWA, Marco R, Schiller P: Drosophila Behaviour & Gene expression
in altered gravity conditions: Comparison between Space and ground
facilities. Proc of the Life in Space for Life on Earth Symposium, Angers,
France, 22-27 June 2008, (ESA SP-663, December 2008) 2008.
26. Benguria A, Grande E, de Juan E, Ugalde C, Miquel J, Garesse R, Marco R:
Microgravity effects on Drosophila melanogaster behavior and aging.
Implications of the IML-2 experiment. J Biotechnol 1996, 47:191-201.
27. Hill RJA, Larkin O, Dijkstra C, Manzano AI, Juan Ed, Davey M, Anthony P,
Eaves L, Medina FJ, Marco R, Herranz R: Effect of magnetically simulated
zero-gravity and enhanced gravity on the walk of the common fruitfly.
Journal of the Royal Society Interface .
28. Hill RJ, Eaves L: Vibrations of a diamagnetically levitated water droplet.
Phys Rev E Stat Nonlin Soft Matter Phys 2010, 81:056312.
29. Tsuchiya K, Okuno K, Ano T, Tanaka K, Takahashi H, Shoda M: High
magnetic field enhances stationary phase-specific transcription activity
of Escherichia coli. Bioelectrochem Bioenerg 1999, 48:383-387.
30. Adair RK: Comment: Influence of stationary magnetic fields on water
relations in lettuce seeds. Bioelectromagnetics 2002, 23:550, discussion 551-
552.
31. Reina FG, Pascual LA, Fundora IA: Influence of a stationary magnetic field
on water relations in lettuce seeds. Part II: experimental results.
Bioelectromagnetics 2001, 22:596-602.
32. Valiron O, Peris L, Rikken G, Schweitzer A, Saoudi Y, Remy C, Job D: Cellular
disorders induced by high magnetic fields. J Magn Reson Imaging 2005,
22:334-340.
33. Ramirez E, Monteagudo JL, Garcia-Gracia M, Delgado JM: Oviposition and
development of Drosophila modified by magnetic fields.
Bioelectromagnetics 1983, 4:315-326.
34. Denegre J, Valles JJ, Lin K, Jordan W, Mowry K: Cleavage plans in frogs
eggs are altered by strong magnetic fields. Proc Natl Acad Sci 1998,
95:14729-14732.
35. Eichler GS, Huang S, Ingber DE: Gene Expression Dynamics Inspector
(GEDI): for integrative analysis of expression profiles. Bioinformatics 2003,
19:2321-2322.
36. Church RB, Robertson FW: A biochemical study of the growth of
Drosophila melanogaster. J Exp Zool 1966, 162:337-352.
37. Gefen E, Marlon AJ, Gibbs AG: Selection for desiccation resistance in adult
Drosophila melanogaster affects larval development and metabolite
accumulation. J Exp Biol 2006, 209:3293-3300.
38. Armstrong JD, Texada MJ, Munjaal R, Baker DA, Beckingham KM: Gravitaxis
in Drosophila melanogaster: a forward genetic screen. Genes Brain Behav
2006, 5:222-239.
39. Glover PM, Cavin I, Qian W, Bowtell R, Gowland PA: Magnetic-field-induced
vertigo: a theoretical and experimental investigation. Bioelectromagnetics
2007, s28:349-361.
40. Kaur H, Kumar S, Kaur I, Singh K, Bharadwaj LM: Low-intensity magnetic
fields assisted alignment of actin filaments. Int J Biol Macromol 2010,
47:371-374.
41. Maret G, Dransfield K: Biomolecules and polymers in high steady magnetic
fields NY: Springer; 1985.
42. Maret G: Recent biophysical studies in high magnetic fields. Physica B
1990, 164:205-212.
43. Marco R, Benguria A, Sanchez J, de Juan E: Effects of the space
environment on Drosophila melanogaster development. Implications of
the IML-2 experiment. J Biotechnol 1996, 47:179-189.
44. Marco R, Gonzalez-Jurado J, Calleja M, Garesse R, Maroto M, Ramirez E,
Holgado MC, de Juan E, Miquel J: Microgravity effects on Drosophila
melanogaster development and aging: comparative analysis of the
results of the Fly experiment in the Biokosmos 9 biosatellite flight. Adv
Space Res 1992, 12:157-166.
45. Marco R, Vernos I, Gonzalez J, Calleja M: Embryogenesis and aging of
Drosophila melanogaster flown in the space shuttle. Preliminary analysis
of experiment fly 15E. Naturwissenschaften 1986, 73:431-432.
46. Vernos I, Gonzalez-Jurado J, Calleja M, Marco R: Microgravity effects on the
oogenesis and development of embryos of Drosophila melanogaster
laid in the Spaceshuttle during the Biorack experiment (ESA). Int J Dev
Biol 1989, 33:213-226.
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 12 of 13
47. Gegear RJ, Casselman A, Waddell S, Reppert SM: Cryptochrome mediates
light-dependent magnetosensitivity in Drosophila. Nature 2008,
454:1014-1018.
48. Palmer J, Slack C: Some bio-electric parameters of early Xenopus
embryos. J Embryol Exp Morphol 1970, 24:535-553.
doi:10.1186/1471-2164-13-52
Cite this article as: Herranz et al.: Microgravity simulation by
diamagnetic levitation: effects of a strong gradient magnetic field on
the transcriptional profile of Drosophila melanogaster.BMC Genomics
2012 13:52.
Submit your next manuscript to BioMed Central
and take full advantage of:
Convenient online submission
Thorough peer review
No space constraints or color figure charges
Immediate publication on acceptance
Inclusion in PubMed, CAS, Scopus and Google Scholar
Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit
Herranz et al.BMC Genomics 2012, 13:52
http://www.biomedcentral.com/1471-2164/13/52
Page 13 of 13
... There have been several studies using D. melanogaster to profile transcriptional changes during spaceflight and in spaceflight analogs. These studies have shown that spaceflight and simulated spaceflight conditions cause significant changes to immunity (Hateley et al., 2016;Herranz et al., 2010Herranz et al., , 2012Hosamani et al., 2016;Marcu et al., 2011;Taylor et al., 2014), metabolism (Ma et al., 2015;Ogneva et al., 2016), and development and proteolysis (Ogneva et al., 2016;Walls et al., 2020). China's space agency launched male flies for 13 days in Shenzhou-9 spaceship (2012), and post-flight studies (48 hours after return to Earth) were conducted on space-flown flies and two controls (ideal lab incubator control and the other mimicking spaceflight conditions including simulated launch process) (Ma et al., 2015). ...
... In a study by the same research team, microarray analysis on control flies and flies subjected to diamagnetic levitation to induce weightlessness (0 g) or HG (2 g) using short term, medium-term, and long-term exposures, revealed significant upregulation of genes related to heat shock and immunity (Herranz et al., 2012). In male flies exposed to levitation, there is overexpression of genes related to immune response (immune induced molecule 23), oxidative stress (peroxiredoxin 2540), and cellular damage clearance (metallothionein B) compared to the control male flies. ...
... Interestingly, the genes altered in diamagnetic levitation (0 g) are not the same as those affected in the RPM experiments. Comparing a group 36 genes that are differentially altered in RPM long-term experiment (22 day exposure to RPM) to the genes affected by diamagnetic levitation (0 g), the authors found one common gene CG32641 (Heat shock protein/Chaperone DNAJ gene group) in the long-term experiment group (22 days exposure to diamagnetic levitation), and three genes in the short-term experiment group (1 day exposure to diamagnetic levitation) including yuri gagarin, a gravity response gene (Herranz et al., 2012). ...
Article
NASA is planning to resume human-crewed lunar missions and lay the foundation for human exploration to Mars. However, our knowledge of the overall effects of long-duration spaceflight on human physiology is limited. During spaceflight, astronauts are exposed to multiple risk factors, including gravitational changes, ionizing radiation, physiological stress, and altered circadian lighting. These factors contribute to pathophysiological responses that target different organ systems in the body. This review discusses the advancements in gravitational biology using Drosophila melanogaster, one of the first organisms to be launched into space. As a well-established spaceflight model organism, fruit flies have yielded significant information, including neurobehavioral, aging, immune, cardiovascular, developmental, and multi-omics changes across tissues and developmental stages, as detailed in this review.
... This grants access for several hours and longer for microgravity simulation on the ground for preliminary spaceflight microgravity screening studies (Wuest et al., 2015). The limitation to average gravity effects by horizontal rotation resulted to the evolution of random positioning machines (RPMs) (Herranz et al., 2012). ...
... Although, most RPM systems are based on two axes so that the two axes are mounted perpendicular to each other. This arrangement therefore sufficiently positions the sample on the experimental platform in any desirable direction (i.e., can point in any direction) (Herranz et al., 2012). ...
... Several biological parameters such as microbial responses, plant cells and mammalian cell cultures can be experimented on the RPM (Herranz et al., 2012). Cell growth and cell proliferation during early plant development has been experimented on RPM (Kiss et al., 2019), while RPM has also been used to develop plant seedlings (Herranz et al., 2012) (Table 2). ...
Article
Gravity is always present on Earth. However, the influence of gravity can be modified or compensated. Generally, the products created under microgravity have important properties that usually surpasses the best terrestrial counterparts. Commercially, these products have striking features that aids marketing. Fast responding microgravity platforms can be used for short-term experiments, where the reactions or observations takes less time. This can be provided in drop towers, parabolic flights and sounding rockets. To study long-term effects of microgravity, human tended space laboratories have been used, such as the International Space Station. Scientists have developed various kinds of ground-based facilities and equipment for some of them to achieve the condition of functional weightlessness as a result of limited access to space laboratories. Slow responding microgravity platforms can be used for long-term experiments, where the reactions or observations takes more time. Microgravity analog, model or simulations are slow responding platforms and are able to provide long-term effects of microgravity. This includes long-duration bedrests, water immersions and in magnetic levitation, clinostats, random positioning machines, rotating wall vessels and even centrifuges. This study compares and gives an overview of currently known real, analog, model or simulated microgravity environments and platforms, and demonstrates their individual capacities, benefits and limitations.
... Since Geim and coworkers [2] demonstrated levitation of a live frog and Valles et al. [4] studied a levitating frog's egg in 1997, diamagnetic levitation has been used in experiments on a variety of biological organisms, including, for example, Paramecia [21], yeast [22,23], Arabidopsis plants [23][24][25] and cell cultures [26][27][28], bacteria [29][30][31], bone cells [32][33][34][35][36], a live mouse [37], and fruit flies [38,39]. ...
... In other experiments, effects of the strong field (∼10 T) are observed, besides that of levitation; see, for example, Refs. [31,38,47,48]. ...
... For the more detailed quantitative estimations of the damage, the integrals of normal forces (pressures), pressure oscillations, and deformations of surface may be included into DAF with corresponding weighting coefficients. As the threshold value the critical wall shear stress τ * ~120 mPa for mouse embryos [73,74] is used here. ...
... Exposition of the fruit fly D. melanogaster in the 0 g physical conditions generated by B = 16.5 T magnetic field during 22 days produced a delay in the development of the fruit flies from embryo to adult [74]. Microarray analysis indicated changes in overall gene expression after the exposition. ...
Book
This book contains fourteen chapters dealing with various aspects of the biomechanics of today. The topics covered are glimpses of what modern biomechanics can offer scientists, students, and the general public. We hope this book can be inspiring, helpful, and interesting for many readers who are not necessarily concerned with biomechanics daily.
... The effects of microgravity on organisms mainly include changes in gene expression, chromosome aberration, apoptosis, immunosuppression, cardiovascular disease, skeletal muscle atrophy and bone loss (Girardi et al., 2012;Lin et al., 2020;Neelam et al., 2020). The commonly used equipment and models for ground simulation of microgravity include rotating wall vessels (RWV), 2D and 3D clinostat, random positioning machine (RPM), hindlimb unloading (HLU) model, antimagnetic levitation model and parabolic flying aircraft (Herranz et al., 2012;Herranz et al., 2013;Ghosh et al., 2016;Stervbo et al., 2018;Paul et al., 2021). For humans, the head-down tilt bed rest model is usually used to study the effect of microgravity on bone, muscle, cardiovascular system (Liang et al., 2014;Moreno-Villanueva et al., 2017). ...
Article
Full-text available
Distinct from Earth’s environment, space environmental factors mainly include space radiation, microgravity, hypomagnetic field, and disrupted light/dark cycles that cause physiological changes in astronauts. Numerous studies have demonstrated that space environmental factors can lead to muscle atrophy, bone loss, carcinogenesis, immune disorders, vascular function and cognitive impairment. Most current ground-based studies focused on single environmental factor biological effects. To promote manned space exploration, a better understanding of the biological effects of the spaceflight environment is necessary. This paper summarizes the latest research progress of the combined biological effects of double or multiple space environmental factors on mammalian cells, and discusses their possible molecular mechanisms, with the hope of providing a scientific theoretical basis to develop appropriate countermeasures for astronauts.
... Magnetic levitation is widely used in engineering and technology. The magnetic levitation approach was applied to animals, cells, and microorganisms, including bacteria [13,[15][16][17][18]. Recently, Parfenov et al. described a magnetic levitation system that uses relatively low magnetic fields due to the supplementation of paramagnetic nutritive medium with the salt of the paramagnetic rare-earth element gadolinium [15]. ...
Article
Full-text available
Changes in bacterial physiology caused by the combined action of the magnetic force and microgravity were studied in Escherichia coli grown using a specially developed device aboard the International Space Station. The morphology and metabolism of E. coli grown under spaceflight (SF) or combined spaceflight and magnetic force (SF + MF) conditions were compared with ground cultivated bacteria grown under standard (control) or magnetic force (MF) conditions. SF, SF + MF, and MF conditions provided the up-regulation of Ag43 auto-transporter and cell auto-aggregation. The magnetic force caused visible clustering of non-sedimenting bacteria that formed matrix-containing aggregates under SF + MF and MF conditions. Cell auto-aggregation was accompanied by up-regulation of glyoxylate shunt enzymes and Vitamin B12 transporter BtuB. Under SF and SF + MF but not MF conditions nutrition and oxygen limitations were manifested by the down-regulation of glycolysis and TCA enzymes and the up-regulation of methylglyoxal bypass. Bacteria grown under combined SF + MF conditions demonstrated superior up-regulation of enzymes of the methylglyoxal bypass and down-regulation of glycolysis and TCA enzymes compared to SF conditions, suggesting that the magnetic force strengthened the effects of microgravity on the bacterial metabolism. This strengthening appeared to be due to magnetic force-dependent bacterial clustering within a small volume that reinforced the effects of the microgravity-driven absence of convectional flows.
... 1141 10. Changes in MFs alter gene expression, especially in immune, stress and temperature response genes, 1142 and affect melanoma at 1-5 nT. 1143 Non-thermal DNA breaks, discovered in 1994, include ELF MF at 50 Hz, 1 mT. ...
Book
Electromagnetic Hypersensitivity is categorised as a multisymptomatic 'el-allergy' in the Nordic classification of 2000 (R.68.8). Its symptoms are 'certainly real' and it can be a 'disabling condition' (W.H.O., 2005). It was first recorded in the mid 20th century as an occupational illness, but it has now spread into the general population through environmental exposure from increasing levels of electromagnetic fields and radiation. This Summary covers current research on this syndrome, covering EM Sensitivity and EM Hypersensitivity. It includes tables of symptoms, EMF sources and exposure guidelines, along with references to scientific studies. This New Edition adds updates, international doctors' protocols, aspects of quantum biology, evidence for sensitivity in animals and plants, case studies, disability issues and human rights.
... ii. La lévitation magnétique La lévitation magnétique ( Figure 15A) est un modèle permettant de simuler la microgravité en créant une force allant à l'encontre du vecteur gravitaire et per ettant de co penser l'attraction terrestre. Il per et de réaliser la lévitation de cellules [349] mais également d'organis es [350,351]. De plus, ce modèle peut être utilisé non seulement pour simuler la microgravité [352] ou une gravité partielle [353], mais également pour réaliser de la culture de cellules en 3D [354,355]. ...
Thesis
Au cours des vols spatiaux, les astronautes sont sujets à de nombreux stress perturbant leur organisme et notamment leur système immunitaire. Afin d’étudier ces altérations et du fait du nombre restreint de missions spatiales, il est nécessaire d’utiliser des modèles permettant de simuler, sur Terre, les stress rencontrés en vol. Au cours de cette thèse, nous avons étudié les effets de stress associés aux vols spatiaux sur le système du complément et les cellules dendritiques (DC). Dans un premier temps, nous avons étudié les effets d’une combinaison de stress ou de stress individuels, sur l’expression de la molécule C3 du complément chez l’amphibien et la souris. Nous avons montré que certains de ces stress associés aux vols spatiaux, dont la microgravité simulée, provoquent une augmentation de C3 dans des larves de P. waltl. Toutefois, ces variations ne sont pas retrouvées chez des souris placées en microgravité simulée par suspension anti-orthostatique. Dans un deuxième temps, nous avons étudié in vitro les effets d’une microgravité simulée (RPM) sur le phénotype et la fonction de DC murines. Nous avons montré que la structure du cytosquelette d’actine et la survie des DC étaient altérées par la microgravité simulée. De plus, les DC exposées à la RPM présentent un phénotype plus immature caractérisé à la fois par une diminution de l’expression membranaire des molécules de co-stimulation mais également de leur capacité à sécréter des cytokines pro-inflammatoires. Bien que ces caractéristiques soient indispensables aux fonctions des DC, les modifications mises en évidence ne semblent toutefois pas altérer leur capacité à présenter l’antigène. Pris ensemble, ces résultats montrent l’importance d’étudier les effets de stress associés aux vols spatiaux, comme la microgravité, sur le système immunitaire. Une meilleure compréhension des mécanismes mis en jeu permettra de comprendre les effets du stress sur la santé et de développer des contremesures adaptées aux vols spatiaux.
... In many cases, clinostat and real microgravity experiments lead to identical impact on cell or plant growth [18], while the rotation speed of the clinostat always has to be taken into account [19]. Nevertheless, it must be mentioned that "microgravity" and related terms are not always applied consistently [20], which is especially complicated in the case of magnetic levitation which naturally changes not only gravity, but also the magnetic field in which experiments are performed, and influences different parts of the plants in different ways, depending on their magnetic properties [21][22][23]. ...
Conference Paper
Investigating seedling germination, plant growth and flowering in simulated microgravity conditions in a clinostat is not only of academic interest, but also a highly important issue for long-term space missions. Another factor influencing plant growth and flowering, from that point of view, is the earth magnetic field which cannot be switched off simply, either, but is thus less often investigated. Here we give an overview of recent technologies and findings related to the influence of gravity - or its real or simulated absence - and magnetic fields on germination, growth and flowering of typical model plants like Arabidopsis thaliana and more space-travel related plants useful for feeding astronauts. In addition, we suggest a novel combined system to investigate plants in combinations of microgravity and freely definable magnetic fields.
Article
Full-text available
The European research community, via European Space Agency (ESA) spaceflight opportunities, has significantly contributed towards our current understanding of spaceflight biology. Recent molecular biology experiments include “omic” analysis, which provides a holistic and systems level understanding of the mechanisms underlying phenotypic adaptation. Despite vast interest in, and the immense quantity of biological information gained from space omics research, the knowledge of ESA-related space omics works as a collective remains poorly defined due to the recent exponential application of omics approaches in space and the limited search capabilities of pre-existing records. Thus, a review of such contributions is necessary to clarify and promote the development of space omics among ESA and ESA state members. To address this gap, in this review we: i) identified and summarised omics works led by European researchers, ii) geographically described these omics works, and iii) highlighted potential caveats in complex funding scenarios among ESA member states.
Article
Full-text available
Previous experiments in space (unmanned satellites, space shuttle and the International Space Station, ISS), have shown that adult Drosophila flies change their motile behaviour in microgravity. A consistent increase in motility in space was found in these experiments, but mature flies (two weeks old) showed less increase than recently hatched flies. In the case of relatively long exposure to microgravity, the aging of male flies measured upon return to Earth was increased, with flies dying earlier than the corresponding in-flight 1g centrifuge or ground controls. The older flies, which experienced a smaller increase in motility, did not show this acceleration in the aging process. More recently we have performed comparative experiments using ground simulation facilities. Preliminary experiments using a random positioning machine (RPM) indicate that the effects of this simulation approach on the behavior of Drosophila are of smaller magnitude than the corresponding exposure to real microgravity. Further experiments are in progress to confirm this effect. However, when exposed to magnetic levitation, flies exposed to simulated weightlessness increased markedly their motile behavior compared with 1g controls both inside and outside the magnet. This altered gravity-related increase in motility was also less pronounced in more mature flies. This motility effect at the levitation position reproduces the results in real microgravity indicating the interest for space science of this simulation approach. Similar experiments are being performed in the Larger Diameter Centrifuge (LDC) located in ESTEC (the Netherlands) and indicate that 6g, 12g and 20g are key points in the hypergravity response in flies. Our experiments have shown that developmental processes from embryo to adult proceeded normally in the magnet, the RPM and the LDC. In terms of gene expression, preliminary results indicate that the affected set of genes under hypergravity responds in general in an opposite direction than that induced by the real or simulated microgravity exposure. The interest in conducting comparative parallel experiments in the complete spectrum of ground simulation methods is shown in the above studies and will be achieved in the near future.
Article
Full-text available
The ISS expedition 8, a 10 days "taxi" flight Soyuz Mission to the International Space Station (ISS) to replace the two-member ISS crew, took place during October 2003. Within the Spanish Cervantes Scientific Mission, in this crew exchanging flight, some biological experiments were performed. The third member of the expedition, the Spanish born ESA astronaut Pedro Duque, returned with the Soyuz 7 capsule and the experiment containing transport box after 11 days on microgravity. In the GENE experiment, we intended to determine how microgravity affects the organism rebuilding processes that occurs during Drosophila metamorphosis. In addition to the ISS samples, some control experiments were performed including a 1g Ground control parallel to the ISS flight samples, a Random Position Machine microgravity simulated control and a parallel Hypergravity (10g) exposed samples experiment. We have used extracted RNA from these samples to test the differences among gene expression during Drosophila development with one of the current more powerful technology, a Drosophila complete genome microarray (version 1.0, Affymetrix TM). A preliminary analysis of the results indicates that around five hundred genes change their expression profiles being especially affected the mitochondrial ribosomal ones.
Conference Paper
Full-text available
With the nearing utilization phase of the International Space Station (ISS) scientists will have the possibility to perform more sophisticated microgravity experiments and experiments of longer duration than could be performed on board pervious orbiting spacecraft. To thoroughly prepare for these ISS studies, ground based experiment tools for simulated (or real) microgravity and hypergravity will be essential. Recently a Random Positioning Machine, which generates simulated microgravity, became available. The Free Fall Machine, which generates short but constantly repeated period of real microgravity, is in use for some time already. The latest version of this FFM has its own dedicated 1xg control. To broaden the scope for acceleration studies a tissue culture centrifuge (Midi-CAR) can generate static and dynamic accelerations up to 100xg and allows culture in standard tissue culture plates as well as standard flight hardware. Finally, a large radius 'animal centrifuge' is used to study the effects of hypergravity on e.g. small animals like rodents or fish. In general it might be stated that ground based research is essential in gravitational biology. Facilities for ground based research should be used to identify possible effects of gravity before performing costly and time consuming real microgravity experiments for space station.
Article
Drosophila flies placed in a habitat with two lateral boxes demonstrated sensitivity to magnetic fields: Oviposition decreased by exposure to pulsated extremely low frequency (ELF) (100)Hz, 1.76 miliTesla (mT) and sinusosidal fields (50 Hz, 1 mT), while there was no initial effect of exposure to a static magnetic field (4.5 mT). Drosophila eggs treated for 48 h with the above described fields showed that (1) mortality of eggs was lower in controls than in eggs exposed to all tested magnetic fields; (2) mortality of larvae increased when a permanent magnet was used; (3) mortality of pupae was highest when a permanent magnet was used; and (4) general adult viability was highest in controls (67%) and diminished progressively when eggs were exposed to pulsated (55%), sinusoidal (45%), and static (35%) magnetic fields.
Article
Nature is the international weekly journal of science: a magazine style journal that publishes full-length research papers in all disciplines of science, as well as News and Views, reviews, news, features, commentaries, web focuses and more, covering all branches of science and how science impacts upon all aspects of society and life.
Article
Diamagnetic objects are repelled by magnetic fields. If the fields are strong enough, this repulsion can balance gravity, and objects levitated in this way can be held in stable equilibrium, apparently violating Earnshaw's theorem. In fact Earnshaw's theorem does not apply to induced magnetism, and it is possible for the total energy (gravitational + magnetic) to possess a minimum. General stability conditions are derived, and it is shown that stable zones always exist on the axis of a field with rotational symmetry, and include the inflection point of the magnitude of the field. For the field inside a solenoid, the zone is calculated in detail; if the solenoid is long, the zone is centred on the top end, and its vertical extent is about half the radius of the solenoid. The theory explains recent experiments by Geim et al, in which a variety of objects (one of which was a living frog) was levitated in a field of about 16 T. Similar ideas explain the stability of a spinning magnet above a magnetized base plate. Stable levitation of paramagnets is impossible. Samenvatting. Magnetische velden stoten diamagnetische voorwerpen af. Zulke velden kunnen zo sterk zijn dat zij de zwaartekracht opheffen. Het is op deze wijze mogelijk zulke voorwerpen te laten zweven. Dit vormt een stabiel evenwicht, wat in tegenspraak schijnt te zijn met Earnshaw's Theorema. Echter Earnshaw's Theorema is niet langer geldig als het magnetisme veld geinduceerd is. De totale energie (bevattende bijdragen van het magnetisme en de zwaartekracht) kan toch een lokaal minimum vertonen. Algemene criteria voor zo'n minimum zullen worden opgesteld. Verder zal worden aangetoond dat voor een cilindrisch symmetrisch veld, langs zijn symmetrie as altijd een zone gevonden kan worden waarin een stabiel evenwicht bestaat. Voor het veld binnen een soleno?de zal deze zone in detail bepaald worden. Als deze spoel voldoende land is bevindt deze zone zich aan het uiteinde van de spoel. De lengte van deze zone langs de symmetrie as van het veld is ongeveer de helft van de straal van de spoel. Deze theorie geeft een goede verklaring voor de experimenten van Geim et al. In deze experimenten werden een grote verscheidenheid aan verschillende voorwerpen (waaronder een levende kikker) tot zweven gebracht in velden van ongeveer 16 T. Analoge theori?n verklaren de stabiliteit van een roterend permanent magneetje boven een magneetische grondplaat. Het is onmogelijk om paramagnetische voorwerpen stabiel te doen zweven.
Article
1. The growth of a wild strain of Drosophila melanogaster has been studied from the time of hatching from the egg to the adult. Wet and dry weights, protein and protein fractions, lipids, RNA and DNA have been determined at successive intervals. 2. All estimates have been carried out on animals grown axenically on defined medium. The medium has been shown to permit growth which is as fast as on live yeast medium when the larvae can burrow freely. 3. Insoluble protein and chitin nitrogen reach a maximum of about 70% total nitrogen in the late pupa, at which time the soluble nitrogen reaches its lowest level. The amino acid fraction reaches a maximum in the early third instar. 4. The water content declines from 70–75% of the live weight at the end of the first instar, rises to a maximum during the second instar and declines sharply to about 66% at the prepupal stage. 5. Total lipids represent an increasing proportion of body weight during larval growth, increasing from about 6% in young larvae to about 15% at pupation. 6. The curve of RNA increase follows the dry weight and protein curves during larval life. The ratio of RNA to protein declines during the first two instars. 7. The increase in DNA content follows a path similar to that of RNA. The RNA/DNA ratio reaches a maximum 24 hours after hatching from the egg, declines to about half this value by 84 hours, and remains more or less constant thereafter. 8. The rates of increase of protein, RNA and DNA during successive periods of larval life show striking differences. For protein the rate is high and increasing until about 36 hours after which it declines rapidly during the rest of larval life. For both nucleic acids the rate of increase falls to a minimum in the early third instar, about the time of the critical size, then increases during proliferation of the imaginal discs and finally declines at the end of larval life. 8. The data are compared with evidence from other strains of Drosophila and from other species of insect. In the latter case, it appears that all the Diptera, so far examined, follow a similar trend in the increase of the nucleic acids during growth. 9. RNA synthesis has been followed during development from egg to adult with the aid of pulse-labeling and sucrose gradient analysis. Developmental stages differ in the amount of rapidly labeled high molecular weight material which is most evident in newly hatched larvae and in mid-third instar larvae, at times when the rate of synthesis of total RNA is high.
Article
An experimental study on water absorption by lettuce seeds previously treated in a stationary magnetic field of 0–10 mT is presented. A significant increase in the rate with which the seeds absorb water is observed in the interval 0–10 mT of magnetic treatment. An increment in the total mass of absorbed water in this interval is also observed. These results are consistent with the reports on the increase of germination rate of the seeds, and the theoretical calculation of the variations induced by magnetic fields in the ionic currents across the cellular membrane. The fields originate in changes in the ionic concentration and thus in the osmotic pressure which regulates the entrance of water to the seeds. The good correlation between the theoretical approach and experimental results provides strong evidence that the magnetic field alters the water relations in seeds, and this effect may be the explanation of the reported alterations in germination rate of seeds by the magnetic field. Bioelectromagnetics 22:596–602, 2001. © 2001 Wiley-Liss, Inc.
Article
PurposeTo evaluate whether static high magnetic fields (HMFs), in the range of 10–17 T, affect the cytoskeleton and cell organization in different types of mammalian cells, including fibroblasts, epithelial cells, and differentiating neurons.Materials and Methods Cells were exposed to HMF for 30 or 60 minutes and subsequently assessed for viability. Cytoskeleton arrays and focal adhesions were visualized using immunofluorescence microscopy.ResultsCell exposure to HMF over 10 T in the case of cycling cells, and over 15 T in the case of neurons, affected cell viability, apparently because of cell detachment from culture dishes. In the remaining adherent cells, the organization of actin assemblies was perturbed, and both cell adhesion and spreading were impaired. Moreover, in the case of neurons, exposure to HMF induced growth cone retraction and delayed cell differentiation.Conclusion Cell exposure to HMF (over 10T and 15 T in the case of cycling cells and neurons, respectively) affects the cell cytoskeleton, with deleterious effects on cell viability, organization, and differentiation. Further studies are needed to determine whether such perturbations, as observed here in cultured cells, have consequences in whole animals. J. Magn. Reson. Imaging 2005. © 2005 Wiley-Liss, Inc.