The behavior of human blood vessel lining cells under a simulated space

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Why did we decide to investigate this?

Gravity has been a constant throughout the history of the earth. This simple fact masks the complexity of gravity as an evolutionary force. Gravity is a vector, i.e. a force (a.k.a. weight) that has a magnitude and a direction at each point in the space. Any change in the gravity field in the space or simulated space alters the behavior of cells. Since the beginning of space exploration, most of the studies were diverted towards combustion science, fluid physics, fundamental physics, and materials science. However, for the last few decades, researchers around the globe are actively working to address the problems related to space expeditions with the simulated hyper- or hypo (micro)-gravity (weightlessness). Declined gravity force offers a favorable environment for cells and tissue culture mimicking natural growth and easier self-association of cells, unlike traditional cultures. In the present era of space exploration for human habitation, it is important to understand how the change in the gravity field affects human physiology. Microgravity affects the heart’s ability to pump blood and therefore, blood pressure and its distribution in the organs. Endothelium, the inner lining of blood vessels, senses the changes in the blood pressure and helps the human body to adapt to the altered environment. Therefore, studying initial vascular signaling under low gravity is very critical to understand the microgravity-driven perturbations in vascular health and physiology of the subject. For more than a decade, our group has been interested to delineate the underlying effects of microgravity on cells of the inner lining of blood vessels, the endothelial cells. However, the effects of simulated microgravity on endothelial cell activation and its functional aspects like angiogenesis have not been explored. The quest for the ideal vascular substitute is still in its infancy and search is ON for an alternative system for manipulating cells or tissues to form blood vessels. We employed a customized 3D clinostat, which is an effective, ground-based tool to stimulate microgravity (0.003g) conditions. The equipment consists of an inner and outer frame with dimensions of 30x30x30 (cm), controlled by two different motors constructed based on the design of clinostat used by the Fokker Space, Netherlands.

In the recent study, we investigated the effects of short-term (2 hours) microgravity on cellular functions of endothelial cells, their mRNA and small RNA expressions using deep sequencing techniques to identify the mechano-sensor miRNA/genes. Deep high-throughput sequencing approach allowed us to analyze the miRNA and mRNA expression profiles in Human Umbilical Cord Vein Endothelial Cells (HUVEC), cultured under gravity, or simulated microgravity conditions.

Challenges and limitations

One of the concerns of the study is that micro RNAs have a half-life of ~12 hours, the cells were treated with microgravity for only 2 hours and then were incubated overnight, while in other literature reports cells are usually treated with microgravity for 24h or longer. Here, the stability of microRNA is a concern.  We mostly researched the effects of short-term simulated microgravity on vascular signaling and functions for the last 15 years. Our research aimed to explore biotechnological scopes of the sensitization of vascular cells with simulated microgravity for developing angiogenesis scaffolds in the laboratory for treating chronic wounds. This work also reveals the immediate effects of microgravity specifically on blood vessels during a space travel scenario. We discovered that short-term exposure to simulated microgravity ensures higher endothelial functions, like tube formation in the scaffold, tip cell formation, and pro-angiogenic activities even after 18 to 48 hours of incubation after the 2 hours exposure to simulated microgravity. Therefore, we envisage that 2 hours-simulated microgravity perturbs the endothelial microRNA profile and modulates endothelial transcriptome and signaling, which transpires functional manifestations even after 24 hours of incubation. 

Salient findings

We identified 1,870 differentially expressed miRNAs in HUVEC under microgravity when compared to the cells subjected to unitary gravity. The functional association of identified miRNAs targeting specific mRNAs revealed that miRNAs, differentially regulated the genes involved in cell adhesion, angiogenesis, cell cycle, JAK-STAT signaling, MAPK signaling, nitric oxide signaling, VEGF signaling and wound healing pathways. Additional experiments showed that the microgravity influenced the cell proliferation and vascular functions of the HUVEC effectively. Consensus on the interactome results indicates restricted fluctuations in the transcriptome of the HUVEC exposed to short-term MG that could lead to higher levels of endothelial functions like angiogenesis and vascular patterning.

Why is this work important?

The study shows the first interactome of transcriptome and micro RNA of vascular cells in modulated gravity and offers a comprehensive understanding of genome interactome that helps to envisage low gravity-induced vascular remodeling, which has a significant impact on various aspects of life like development, vascular homeostasis, and angiogenesis.

This post was jointly written with Prof. Suvro Chatterjee. The complete article can be found here: https://doi.org/10.1038/s41526-020-00108-6

Pavitra Kumar

Postdoc, University of Bern

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