Simulated Microgravity Promotes Angiogenesis through RhoA-Dependent Rearrangement of the Actin Cytoskeleton

Abstract

Background/aims: Microgravity leads to hydrodynamic alterations in the cardiovascular system and is associated with increased angiogenesis, an important aspect of endothelial cell behavior to initiate new vessel growth. Given the critical role of Rho GTPase-dependent cytoskeleton rearrangement in cell migration, small GTPase RhoA might play a potential role in microgravity-induced angiogenesis. Methods: We examined the organization of actin filaments by FITC-conjugated phalloidin staining, as well as the expression and activity of RhoA by quantitative PCR and Western blot, in human umbilical vein endothelial cells (HUVECs) under normal gravity and simulated microgravity. Effect of simulated microgravity on the wound closure and tube formation in HUVECs, and their dependence on RhoA, were also analyzed by cell migration and tube formation assays. Results: We show that in HUVECs actin filaments are disorganized and RhoA activity is reduced by simulated microgravity. Blocking RhoA activity either by C3 transferase Rho inhibitor or siRNA knockdown mimicked the effect of simulated microgravity on inducing actin filament disassembly, followed by enhanced wound closure and tube formation in HUVECs, which closely resembled effects seen on microgravity-treated cells. In contrast, overexpressing RhoA in microgravity-treated HUVECs restored the actin filaments, and decreased wound closure and tube formation abilities. Conclusion: These results suggest that RhoA inactivation is involved in the actin rearrangement-associated angiogenic responses in HUVECs during simulated microgravity.

Key Proteins Involved in Spheroid Formation and Angiogenesis in Endothelial Cells After Long-Term Exposure to Simulated Microgravity

“…Endothelial cells (ECs) play a key role in the pathogenesis of various diseases and are highly sensitive to microgravity. ECs were investigated in space on the International Space Station (ISS) [2, 3] and on Earth using techniques to simulate microgravity [4-7]. With the help of so-called ground-based facilities like clinostats or the RPM, which are devices constructed to simulate microgravity, some aspects of annulling gravity can be studied.…”

“…The mechanisms for this behavior are still not completely clear. A recent study showed that ras homolog gene family member A (RhoA) inactivation supports the actin rearrangement-associated angiogenic responses in human umbilical vein endothelial cells (HUVECs) during simulated microgravity [7]. Early cytoskeletal changes of microgravity-exposed ECs have previously been described in real and simulated microgravity [10, 11].…”

Enzyme-Crosslinked Electrospun Fibrous Gelatin Hydrogel for Potential Soft Tissue Engineering

“…In that study, the enhanced cell spreading on enzyme-crosslinked materials was proven after excluding the other biophysical and biological factors by comparing the cell morphology on the substrates with a similar stiffness and arginine–glycine–aspartate (RGD) density. The strong actin filaments and high cell spreading area usually represent phenotypes for angiogenesis and vascularization [ 51 , 52 ]. Overall, though both Gel–HPA fibrous hydrogel and gelatin fibrous scaffolds showed excellent biocompatibility, Gel–HPA fibrous hydrogel crosslinked by HRP had the ability to enhance cell adhesion, cell spreading and cell proliferation, which are necessary for cell activities and tissue regeneration.…”

Low Intensity Vibration Restores Nuclear YAP Levels and Acute YAP Nuclear Shuttling in Mesenchymal Stem Cells Subjected to Simulated Microgravity

“…Research aimed at studying the effects of microgravity at the cellular level often relies on simulated microgravity (SMG) devices designed to alter the gravitational conditions that cells experience by rotating on one or multiple axes at low speed 29-31 . SMG decreases MSC proliferation 32 and cytoskeletal contractility 29,33,34 . In this way, application of physical or soluble factors that induce cytoskeletal contractility are commonly used as countermeasures for SMG 35,36 .…”