Since the beginning of life on Earth, organisms have lived under the influence of the Earth's physical parameters including its almost constant gravitational and magnetic fields. Therefore, evolution has had to provide a number of different solutions to meet the mechanical challenge of supporting the weight of a living organism . In general, the influence of gravity on the physiology of an organism increases with its mass, although for organisms living in water, the effect of gravity is to some extent mitigated by buoyancy. In plants, gravity has an important effect on the development of small seedlings via the sedimentation of heavy components (statoliths), but gravitational effects in non-specialized cells have also been reported [2, 3]. The reduced gravity on the surfaces of Mars (0.38 g) and the Moon (0.17 g) may significantly affect not only astronauts manning the first space colonies, but also the development of plants, which would be an essential part of life support systems. It is also possible that zero- and reduced-gravity might have unexpected effects on the behaviour of bacteria, viruses and other micro-organisms, either directly or through the effect of reduced gravity on the environment, e.g. through a modified convection in gases and fluids . The natural magnetic field strength at the surface of the Earth varies from 30 to 60 μT , but magnetic fields of up to 12 T are being used regularly in diagnostic non-invasive techniques, such as magnetic resonance imaging [6, 7], without any apparent long-lasting effects on cells. Nevertheless, there have been reports that high magnetic fields affect bacteria , plants [9–11], mammals , and flies . The exposure of Arabidopsis seedlings to magnetic fields in excess of about 15 T for 6.5 h has been linked with 2.5 fold changes in the expression levels of 114 genes .
To study the effect of an environment with altered gravitational forces, we have to place samples in orbit by means of space flights, sounding rockets, or use simulation facilities on the ground. Mechanical facilities such as 2D-clinostats or random positioning machines (RPMs), and for enhancement, centrifuges like the Large Diameter Centrifuge (LDC) [15–18], have been used for decades in well-equipped laboratories to average out the gravitational force. A critical evaluation and quantification of the exact physical effects involved in these simulators is, however, often missing. Nevertheless, such an evaluation would help to answer the question as to whether using such machines creates either a stimulus-free environment with respect to gravity (= simulated weightlessness) or an omnilateral gravistimulation with strong mechanical disturbances. An alternative and relatively new approach to study the response of organisms to changes in gravity is the use of diamagnetic levitation [19–24]. Since diamagnetic material is repelled by magnetic fields, when a diamagnetic object is positioned in a magnetic field gradient it experiences a magnetic force away from regions of high field. The size of the force is proportional to the product of the field strength (B) and the field gradient (the spatial derivative of the field B'). When B times B' is strong enough, this magnetic force can be used to counterbalance the gravitational force, leading to the stable levitation of a large variety of materials such as water and other fluids. Since the bulk of living organisms is composed of diamagnetic material, mostly water, organisms can be magnetically levitated, requiring BB' to be equal to about 1400 T2/m, which in commonly used magnets occurs at a field strength of about 16 T. Such conditions can now be readily produced in a dozen facilities around the world.
In this paper, we have studied the effect on the overall transcriptional state of Arabidopsis thaliana semi-solid cell cultures (callus) exposed to an environment of altered gravitational and magnetic forces for 200 min. This biological system, composed of undifferentiated proliferating cells, was chosen in the context of our previous investigations on the effects of gravity alteration on cell growth and proliferation [25, 26]. Our first goal in this analysis was to test what kind of facility would be most suitable and reliable as an altered gravity simulator. Previous investigations used different experimental settings for each facility, limited their analysis to a particular collection of genes or used a low number of seedlings per sample compromising statistical outcomes [3, 14, 27, 28]. The present study is the first systematic multi-facility, high-throughput, environmentally controlled collection of experiments that has been performed with the same set-up, almost simultaneously in two mechanical facilities (RPM for μg and LDC for 2 g) and a magnet-based facility using five different effective gravity (g*) conditions (from μg* to 2 g*). This was to allow sustainable inter-experiment comparisons of the results and a pooled analysis with multiple inner controls at similar magnetic fields. During our research, we detected a synergic effect when more than one environmental parameter was affecting the samples (gravitational and magnetic fields) that could be linked to the collapse of cellular strategies to support environmental stresses already observed in space samples .