The dynamic adaptation of primary human endothelial cells to simulated microgravity

Abstract

Culture of human endothelial cells for 10 d in real microgravity onboard the International Space Station modulated more than 1000 genes, some of which are involved in stress response. On Earth, 24 h after exposure to simulated microgravity, endothelial cells up‐regulate heat shock protein (HSP) 70. To capture a broad view of endothelial stress response to gravitational unloading, we cultured primary human endothelial cells for 4 and 10 d in the rotating wall vessel, a U.S. National Aeronautics and Space Administration–developed surrogate system for benchtop microgravity research on Earth. We highlight the crucial role of the early increase of HSP70 because its silencing markedly impairs cell survival. Once HSP70 up‐regulation fades away after 4 d of simulated microgravity, a complex and articulated increase of various stress proteins (sirtuin 2, paraoxonase 2, superoxide dismutase 2, p21, HSP27, and phosphorylated HSP27, all endowed with cytoprotective properties) occurs and counterbalances the up‐regulation of the pro‐oxidant thioredoxin interacting protein (TXNIP). Interestingly, TXNIP was the most overexpressed transcript in endothelial cells after spaceflight. We conclude that HSP70 up‐regulation sustains the initial adaptive response of endothelial cells to mechanical unloading and drives them toward the acquisition of a novel phenotype that maintains cell viability and function through the sequential involvement of different stress proteins.—Cazzaniga, A., Locatelli, L., Castiglioni, S., Maier, J. A. M. The dynamic adaptation of primary human endothelial cells to simulated microgravity. FASEB J. 33, 5957–5966 (2019). http://www.fasebj.

Mitophagy contributes to endothelial adaptation to simulated microgravity

“…15 Also autophagy, a process which allows the cell to raze redundant and/or dysfunctional components, is considered a survival adaptive response, thus contributing to stress resistance. It is now clear that cells and tissues activate adaptive responses to cope with the absence of gravity, the only environmental factor which has remained constant since when life began on Earth.…”

“…All culture reagents were from Gibco (Thermo Fisher Scientific, Waltham, MA, USA). 15 Therefore, some samples were removed from the RWV 2 and 4 hours after the beginning of the experiment to measure the levels of ROS using 2′-7′-dichlorofluorescein diacetate (DCFH) as described, 15 while the others were maintained in the RWV for 10 d to evaluate mitochondrial mass. 7,10,24 As 1g condition control (CTR), HUVEC grown on beads were cultured in the vessels not undergoing rotation.…”

“…Since we have detected a mild and transient increase of ROS in simulated microgravity, 15 we performed experiments in the presence of NAC (1 mM). Since we have detected a mild and transient increase of ROS in simulated microgravity, 15 we performed experiments in the presence of NAC (1 mM).…”

“…21 Because research in space is limited by the high costs and low flight opportunities, several ground-based tools have been developed for simulating microgravity on Earth, 22 all based on eliminating a preferential direction of the gravity vector by continuous rotation. 15 Here, we utilize the same time points and demonstrate that simulated microgravity activates mitophagy, thus reducing mitochondrial content and oxygen consumption in HUVEC. 10,15,22,23 We have utilized the NASA-developed rotating wall vessel (RWV), which generates a vector-averaged gravity similar to that of near-Earth free-fall orbit.…”

“…Mitochondrial content is determined by the balance between mitochondrial biogenesis and degradation through the process called mitophagy, ie, the autophagic removal of damaged or superfluous mitochondria, while their organization, fundamental in determining their function, is the result of the balance between fusion and fission. 10,15,22,23 We have utilized the NASA-developed rotating wall vessel (RWV), which generates a vector-averaged gravity similar to that of near-Earth free-fall orbit. 21 Because research in space is limited by the high costs and low flight opportunities, several ground-based tools have been developed for simulating microgravity on Earth, 22 all based on eliminating a preferential direction of the gravity vector by continuous rotation.…”