Cell cultivation under different gravitational loads using a novel random positioning incubator

Abstract

Important in biotechnology is the establishment of cell culture methods that reflect the in vivo situation accurately. One approach for reaching this goal is through 3D cell cultivation that mimics tissue or organ structures and functions. We present here a newly designed and constructed random positioning incubator (RPI) that enables 3D cell culture in simulated microgravity (0 g). In addition to growing cells in a weightlessness-like environment, our RPI enables long-duration cell cultivation under various gravitational loads, ranging from close to 0 g to almost 1 g. This allows the study of the mechanotransductional process of cells involved in the conversion of physical forces to an appropriate biochemical response. Gravity is a type of physical force with profound developmental implications in cellular systems as it modulates the resulting signaling cascades as a consequence of mechanical loading. The experiments presented here were conducted on mouse skeletal myoblasts and human lymphocytes, two types of cells that have been shown in the past to be particularly sensitive to changes in gravity. Our novel RPI will expand the horizon at which mechanobiological experiments are conducted. The scientific data gathered may not only improve the sustainment of human life in space, but also lead to the design of alternative countermeasures against diseases related to impaired mechanosensation and downstream signaling processes on earth.

Simulated Microgravity: Critical Review on the Use of Random Positioning Machines for Mammalian Cell Culture

“…We have recently reported another approach to upgrading the RPM by installing a commercial CO 2 incubator onto the rotating frames. This RPM, called “random positioning incubator” (RPI) [ 9 , 10 ], has the advantage of being independent of large laboratory incubators ( Figure 1 ). Furthermore, the closed chamber of the incubator isolates the environment of the culture flasks and thus prevents exposure of biological samples to vapor and wear from the machinery, for example, [ 10 ].…”

“…We have also been working on an upgrade of the software responsible for controlling the motion of the RPM. Three different algorithms were introduced recently that simulated partial Earth gravity (0 to 0.6 g), allowing simulation of moon- or Mars-like gravity conditions [ 9 ]. All algorithms are adaptations of the random walk algorithm originally designed to simulate microgravity [ 10 ].…”

“…In one case the timing is flexible and adjusted online as the experiment runs. In the other case the timing is strictly periodic and predefined before the experiment starts [ 9 ].…”

“…All three algorithms were tested on suspended human T cells and adherent mice myoblasts. Chemically activated T cells showed a decreased activation rate that correlates strongly to the decreasing simulated mean gravity values [ 9 ]. The results were similar for all algorithms.…”

Calcium's Role in Mechanotransduction during Muscle Development

“…In this study, we have also shown that hypergravity at 4 g does not affect myoblast proliferation or cell cycle progression. Furthermore, by using a novel RPM that simulates partial gravity, we observed a linear correlation between the slowed cell growth and the accumulation in the G2/M phase corresponding to the partial gravity level of exposure [88]. We attribute these effects to an impaired regulation of calcium through mechanosensitive channels disturbed by the removal of the force of gravity.…”