Jelly Knees - An Astronaut's Malady

The Effects of Microgravity on Human Cartilage Tissue

By Maria Volosatov

Final z-projected image of chondrocyte differentiated human mesenchymal stem cell 3D spheroid

Final z-projected image of chondrocyte differentiated human mesenchymal stem cell 3D spheroid taken using Cytation 5 Cell Imaging Multi-Mode Reader. DAPI: Hoechst 33342 stained nuclei, GFP: Alexa Fluor 488 phalloidin stained actin filaments, CY5: Collagen II expression. Courtesy of Wikimedia Commons via CC BY-SA 3.0 DEED.

As our attempts to brave the cosmic frontier grow increasingly bolder, the impact of spaceflight on the human body becomes imperative to understand. Flight in outer space dramatically impacts nearly every aspect of human health, as humans adapt to conditions foreign to those on terra firma. With evidence-based research, however, we have devised methods to counteract and reverse some of these drastic impacts. But as we improve our understanding of how these enterprises affect us on the molecular level, could we perhaps also reveal a deeper truth on the genesis of certain diseases?

The University of Alberta Space Design Group, one of the winning groups of the 2021 Reduced Gravity Experiment Design Challenge (CAN-RGX)—run by SEDS-Canada—delved into these questions. With the aid of NRC’s Falcon 20 shuttle, the UASDG team, comprised of University of Alberta students Amira Aissiou, Shankar Jha, Kirtan Dhunnoo, Kingston Wong, Sherry Gao, Vasu Gupta, Vince Montero, Nasifa Hasan, Darsh Trivedi, and faculty advisor Dr. Adetola Adesida, were able to investigate the effects of short-term microgravity exposure on tissue-engineered cartilage constructs on the transcriptomic and proteomic levels [1].

Studies show that bone density and geometry develop to resist mechanical stress that is inherent to Earth gravity, as well as through mechanical stimulation or exercise [2]. As the pull of Earth’s gravity reduces with the square of the distance, the local strains on the musculoskeletal system ease, leading to deterioration and the development of cartilage-degrading diseases such as osteoarthritis (OA). These effects have been observed in astronauts on the cellular and tissue level [3]. To understand how the near absence of gravity, or microgravity, affects tissue on the molecular level is a step further, delving into the underlying factors that determine how living tissue reacts to these otherworldly stressors.

Using human bone marrow mesenchymal stem cells (hBMSC) of three male and three female donors, the team expected to observe a perturbation in the transcriptome—the sum of the cellular genetic readout—of the cartilage constructs that would differentiate based on sex. The expectation of observing differences between the biological sexes stems from earlier studies observing osteoarthritic alterations in the articular and meniscal cartilages of mice, as well as extensive literature evidence of the differences in OA development between male and female patients. Separating the cellular colonies into three groups, a lab control, ground control, and air group (microgravity-exposed), the team used a purpose-built three-way valve syringe to link the cartilage cells with RNAlater fluid to preserve it for further processing. After only 140 seconds of true microgravity, in the span of 11 parabolic flights, significant changes were observed in the gene expression between the ground and air samples.

A total of 94 differently-expressed genes (DEGS) for the male group and 74 for the female group were observed—however only 5 of said DEGS were communal. Clearly the response was not independent of the biological sex. Molecular agents involved in protein activity and regulation of protein modification processes appeared to be reacting to the microgravity-induced stress differently. 

In genomics, the expression of a certain gene can be characterized numerically by observing the fold change: a ratio quantifying the increase or decrease in gene expression on a logarithmic scale. For instance, a positive 2-fold change value for gene X between group A and group B would indicate that the expression of gene X is higher in group A than in group B by a factor of 22. Conversely, one can have a negative fold change, indicating a decrease in expression [4]. The alteration of cellular structure appeared in both sexes, with a noted upregulation of genes involved in the regulation of actin dynamics and cytoskeletal rearrangements. This confirms that both groups are susceptible to cytoskeletal structural changes induced by microgravity. The mechanism of the response, however, was the key difference.

Dr. Adetola Adesida, the group’s faculty advisor and professor in the Department of Surgery at the University of Alberta, elaborated on the differences in stress response between the sexes: “Heat shock proteins tend to be expressed when cells are stressed…[t]hat tells us that the molecular entities that are involved in cellular stress are different between the sexes. This may also suggest why males experience OA later in life [as opposed to females].” The differences in the stress response between the female and male cartilage cells were quantitatively derived from the upregulation of heat shock proteins, particularly the gene HSP70. According to Mayer and Bukau’s publication in the Journal of Cellular and Molecular Life Sciences, HSP70 is expressed when cells undergo extreme stress, such as heat shock. This gene acts as a protein chaperon, aiding in the folding of other proteins and essentially acting as a temporary shut-down procedure that protects the cell and aids in its recovery post-stress. The response time appears to be different, as the study highlights the sex-dependent response to parabolic flight microgravity lies with the transcription of members of the HSP70 family [5][6].

One of the most intriguing aspects of this study was the group’s ability to pinpoint the initiation of OA on the molecular level. Of the five topmost upregulated secretome genes, WNT7B and WNT9A were upregulated in female-derived hBMSC. Both genes are implicated in OA development and encode proteins which are involved in developmental, physiological and disease processes in cells. Dr. Adesida points out that we tend to look at the development of OA at later stages in affected individuals; “We tend to analyze cartilage from the affected joints [and] look at the progression of the disease. In this study, we actually made cartilage from healthy [donor] cells, and then exposed them to an environment predicted to induce OA in healthy cartilage. That induction…is causing a cascade of pathways that are going to degrade cartilage over a longer period of time.” The group’s results shine light on the potential beyond the scope of the experiment for further studies. Firstly, the molecular response of male and female human stem cells to microgravity, and secondly to bridge the experiment to clinical studies and investigate the regenerative potential of hBMSC. 

This is a unique method of looking at the pathogenesis of OA. Rather than interfering with the progression of the disease when it has already taken root, we may get a clear picture of its initiation before it progresses into clinical symptoms. In discussing the clinical implications of the study, Dr. Adesida explains: “Looking at the disease from a healthy individual’s context enables us to determine what stage we can intervene.” It is necessary to consider ethical methods of intervention. Whether that entails a therapeutic solution or a pharmaceutical one, Dr. Adesida believes that timing is vital. “I think it will be a combination of detecting early signs and translating that to Earth conditions and seeing what is happening—if there are clinical benefits [to a pharmaceutical agent].”

Investigating methods to target pathways with inhibitor agents in male and female cells may be the key not only to potential cures for OA patients, but also in developing preventive methods that will allow astronauts to adapt and better rehabilitate from spaceflight of longer durations.

About: This opportunity was facilitated by SEDS-ÉEDS Canada’s Reduced Gravity Experiment Design Challenge, an annual contest for students across Canada to design an experimental proposal, whereby the chosen teams can conduct their experiment in an induced microgravity environment. The Canadian Reduced Gravity Experiment Design Challenge, along with the Canadian Stratospheric Balloon Design Challenge, Canadian Analog Research Expedition, and Young Space Entrepreneurs Competition, are all part of SEDS’ mission to enable the next generation of Canadian space sector professionals. The CAN-RGX flight campaign will be conducted in August. We are excited to continue to work with student teams and allow young researchers to make direct contributions in technology, medicine, environmental studies, astrophysics, and more.

Sources & Additional Reading

[1] Aissiou, A. K., Jha, S., Dhunnoo, K., Ma, Z., Li, D. X., Ravin, R., Kunze, M., Wong, K., & Adesida, A. B. (2023, January 19). Transcriptomic response of bioengineered human cartilage to parabolic flight microgravity is sex-dependent. Nature News. https://www.nature.com/articles/s41526-023-00255-6

[2] Bergmann, P., Body, J.J., Boonen, S., Boutsen, Y., Devogelaer, J.P., Goemaere, S., Kaufman, J., Reginster, J. Y., Rozenberg, S., (2010, December 20). Loading and Skeletal Development and Maintenance. Journal of Osteoporosis. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3010667/

[3] Abadie, L. J., Cranford, N., Lloyd, C. W., Shelhamer, M. J., Turner, J. L., (2021, February 2). The Human Body in Space. NASA Human Research Program page. https://www.nasa.gov/hrp/bodyinspace

[4] Bonnin, S., Ponomarenko, J., Cozzuto, L., Hermoso, T. (2020, March 6). Comparing experimental conditions: differential expression analysis. Bioinformatics for Biologists course page.
https://biocorecrg.github.io/CRG_Bioinformatics_for_Biologists/differential_gene_expression.html#:~:text=The%20Fold%20change%20indicates%20whether,Type%20compared%20to%20Knock%20Out.

[5] Mayer, M. P., Bakau, B. (2005, March ). Hsp70 chaperones: Cellular functions and molecular mechanism. Journal of Cellular and Molecular Life Sciences. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2773841/#:~:text=Hsp70%20proteins%20are%20central%20components,segments%20within%20their%20substrate%20proteins.

[6] Orphazyme US. (2021, May 17). “The Science of Heat Shock Proteins in Proteostasis”, YouTube, uploaded by Orphazyme US. https://www.youtube.com/watch?v=-h6MhD_kUPo

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