Changes in Exosome Release in Thyroid Cancer Cells after Prolonged Exposure to Real Microgravity in Space
Authors: Stefan Riwaldt
Marcus Krüger
Manfred Infanger
Daniela Grimm
Petra M Wise
Paolo Neviani
Thomas Juhl Corydon
Markus Braun
Markus Wehland
Journal: International Journal of Molecular Sciences (2021)
Discipline: CHEMISTRY
Keywords: cell culture; exosomes; microgravity; spaceflight; tetraspanins
Affiliations: The Saban Research Institute, Children's Hospital Los Angeles, University of Southern California, 4650 Sunset Blvd, Los Angeles, CA 90027, USA., Department of Microgravity and Translational Regenerative Medicine, Clinic for Plastic, Aesthetic and Hand Surgery, Otto von Guericke University, Universitätsplatz 2, 39106 Magdeburg, Germany., Department of Biomedicine, Aarhus University, Ole Worms Allé 4, 8000 Aarhus C, Denmark., Department of Ophthalmology, Aarhus University Hospital, Palle Juul-Jensens Blvd. 99, 8200 Aarhus N, Denmark., Research Group "Magdeburger Arbeitsgemeinschaft für Forschung unter Raumfahrt- und Schwerelosigkeitsbedingungen" (MARS), Otto von Guericke University, 39106 Magdeburg, Germany., Deutsches Zentrum für Luft- und Raumfahrt (DLR), Raumfahrtmanagement Bonn-Oberkassel, 53227 Bonn, Germany.

Abstract:  Space travel has always been the man's ultimate destination. With the ability of spaceflight though, came the realization that exposure to microgravity has lasting effects on the human body. To counteract these, many studies were and are undertaken, on multiple levels. Changes in cell growth, gene, and protein expression have been described in different models on Earth and in space. Extracellular vesicles, and in particular exosomes, are important cell-cell communicators, being secreted from almost all the cells and therefore, are a perfect target to further investigate the underlying reasons of the organism's adaptations to microgravity. Here, we studied supernatants harvested from the CellBox-1 experiment, which featured human thyroid cancer cells flown to the International Space Station during the SpaceX CRS-3 cargo mission. The initial results show differences in the number of secreted exosomes, as well as in the distribution of subpopulations in regards to their surface protein expression. Notably, alteration of their population regarding the tetraspanin surface expression was observed. This is a promising step into a new area of microgravity research and will potentially lead to the discovery of new biomarkers and pathways of cellular cross-talk. 

Retracted: No

PubPeer comments:
 What could possibly be cooler than an article that combines hot topics like  exosomes , microgravity and sprinkles in phrases like  Since the dawn of manned spaceflight ? I congratulate the authors on a very interesting topic! At the risk of upending this very beautiful applecart, though, I have to make a few gentle corrections and offer some alternative interpretations. This paper is probably not about exosomes and may not be about microgravity either. The first issue I see is that the cells were grown in 10% fetal bovine serum (FBS or FCS). FBS is very rich in extracellular vesicles (EVs). In many cell cultures grown with FBS, EVs released from the cultured cells are only a minority of the total EV population. Second, related to this, no medium-only controls were flown on the rocket ISS, so there is no way of knowing how much of the reported differences were from cellular responses and from responses of  stuff including but not limited to EVs in the culture medium. The third point is that exposure to microgravity was just one of the things that distinguishes the ground and flight conditions. I haven t been to space, but I ve heard that there are some g forces associated with liftoff. Were the ground cells exposed to the same? And is it possible that oxygen tension or other air qualities were different? What about exposure to radiation out where the atmosphere is rather thin? I certainly feel for the authors in trying to develop a well controlled experiment, but there are multiple variables that distinguish ground from flight. Fourth, and I might have missed this, but was cell viability measured or controlled for? This is a major confounder of EV experiments even on the ground. If this or that inhibitor or agonist or any other condition causes a tiny bit more cell death, the ratio of internal and external membrane markers can change remarkably and drown out any EV-related changes. If just a few cells died and spewed their contents on liftoff, it could explain the results. Fifth, exosomes? Exosomes are EVs that come from the multivesicular body (inside the cell, endosomal). There are several other ways that EVs or similar particles can be birthed, such as  ectosomes from the plasma membrane (outer border of the cell). I grant that there are hundreds of reviews in the literature and also a lot of older studies that claim that all exosomes are smaller than other EVs (like when the authors say that the size range of 50-100 nm is exclusively exosomes). But ectosomes can be just as small as exosomes. Tetraspanins like the ones measured here are also claimed to be exclusive to true exosomes, but this is not supported by recent findings. Even the ESCRT machinery, once thought to be exclusive to internal compartments, can be found at the PM, not just in endosomes. There is no single marker that can distinguish exosomes from ectosomes or other EVs for all cell types. Even if you find one for your cell type, it might change with conditions (like taking off in a rocket or getting zapped with a little extra radiation?). Without very detailed and well controlled experiments with the cells under all tested variables, there is not even a way to know what might be differentially parceled out to EV subtypes. There s more, but I m being a bore already. I just want to address the characterisation still. The guidelines of the International Society of EVs are quoted in the paper, but they are not followed really, and maybe that would be impossible. Still, the complete reliance on one really kinda new platform is less than encouraging, and what look suspiciously like marketing claims (no processing needed, amounts needed for input,  exosomes ) are repeated in the paper without citation or data. The single platform used for the analysis of tetraspanins (no, not just exosomes) is a very interesting and I think promising one. But what do we know for sure? Still not much. At the lower end of the size range, beyond what the system and software can size, what are the fluorescent particles? How much do these  phantom bodies contribute? These could be membrane fragments of dead cells. Or bits of junk released from stressed or ecstatic cells. No one knows yet. Like above, more controls would be useful. In their absence, and goddess only knows how impossible it is to get an extra few grams on a spaceflight, claims might be amended slightly. Sorry to the authors for being so critical. I do value the study and also the fact that it was conceived in 2013 if not earlier. Onwards and upwards! ,