Sex-specific cartilage biomarker response to 5 days of dry immersion

INTRODUCTION
Human spaceflight is associated with the deconditioning of multiple physiological systems due to the absence of
Earth gravity (Shelhamer M. et al., 2020). To unravel the underlying mechanisms of the adaptation to microgravity,
ground-based models have been developed, with the objective of simulating physiological effects of spaceflight
on the human body on Earth, primarily by immobilization and unloading. Commonly applied ground-based models
are dry immersion (DI) or -6° head-down-tilt bed rest (HDT). Cartilage remodeling can be evaluated by
quantifying metabolites, so called biomarkers, generated during cartilage synthesis and degradation (Lotz et al.,
2013) and microgravity can result in alterations in biomarker assessed cartilage metabolism, that have the potential
to initiate catabolic processes (Niehoff et al., 2016). Type II collagen is a major component of articular cartilage
and the serum biomarkers C2C and CPII indicate degradation and synthesis of this protein. Previous studies
indicate changes in cartilage biomarker concentrations in male study participants exposed to HDT (Liphardt et al.,
2018; Liphardt et al., 2020). However, it is unknown if the impact of immobilization by DI on serum biomarker
concentration is different in women compared to men. Thereto, the purpose of the study was to investigate sexdependent effects of five days of immobilization by DI on serum concentrations of C2C and CPII.
METHODS
The study was part of a research program of the European Space Agency (ESA) and was carried out at the Institute
de Médecine et de Physiologie Spatiales (MEDES) in Toulouse, France. Eighteen healthy female participants
(29±5 years, 165±6 cm, 59±6 kg) and nineteen male participants (28±4 years, 177±4 cm, 72±7 kg) were subjected
to four days of baseline data collection (BDC), five days of DI, and three days of recovery (R). During DI,
participants were fully immersed in a tub filled with water for 24 hours per day, while being protected against
direct water contact by a specialized elastic fabric. This immobilization model limits movement, in particular of
the lower extremities and reduces ground support. Fasting, venous blood samples were taken on four days at 7 am:
BDC-26h, DI46h, DI118h and R+46h (Figure 1). We analyzed serum concentrations of C2C and CPII with
commercially available ELISAs (IBEX Pharmaceuticals Inc., Montreal, Québec, Canada). A two-factor repeated
measures ANOVA, followed by a LSD post-hoc test, were carried out to detected significant differences over time
and between sexes.
RESULTS
The serum C2C concentration was significantly different between sexes at all time points (p=0.006), with higher
concentrations for male participants compared to females (e.g., BDC-26h: 184.92±43.85 ng/mL (female),
223.33±47.22 ng/mL (male), p=0.015). The serum C2C concentration of female participants increased
significantly (p=0.013) from BDC-26h to DI46h by +10% and by +7% from DI46h to DI118h (p=0.02), resulting
in an overall increase of +17% compared to baseline (DI118h vs. BDC-26h, p=0.002). During recovery the serum
C2C concentration remained elevated compared to baseline (R+46h vs. BDC-26h, p=0.006). In male participant,
progression of serum C2C concentration was similar, starting with a significant increase (p=0.012) from BDC26h to DI46h by +10% and an additional significant increase (p=0.029) of +9% from DI46h to DI118h, causing
an overall increase of +19% compared to baseline (DI118h vs. BDC-26h, p<0.001). Surprisingly, serum C2C
concentration decreased by -9% during recovery in male participants (R+46h vs. DI118h, p=0.025), but values
were still significantly higher compared to baseline (R+46h vs. BDC-26h, p<0.001).
The serum CPII concentration was not significantly different between sexes at all time points (p=0.853), thereto
female and male concentration were considered together (e.g., BDC-26h: 1434.50±473.74 ng/mL (female),
1421.51±347.44 ng/mL (male), p=0.924). The serum CPII concentration, mean of both sexes, increased
significantly (p=0.004) from BDC-26h to DI46h by +8%. Concentrations further increase (p=0.005) by +12%
from DI46h to DI118h, resulting in an overall increase of +20% compared to baseline (DI118h vs. BDC-26h,
p<0.001). During recovery the serum CPII concentration remained elevated and was still significantly increased
compared to baseline (R+46h vs. BDC-26h, p<0.001).
DISCUSSION
The C2C biomarker concentrations were significantly higher in male than in female participants, which might be
caused by higher overall cartilage volume in men compared to women (Otterness I.G. and Eckstein F., 2007).
Both, serum C2C and CPII concentrations increased significantly after only 46 hours of DI and increased further
until the end of DI (DI118h). These findings indicate that five days of DI elicits substantial changes in the assessed
cartilage biomarkers. However, the observation that both degradation and synthesis biomarkers of type II collagen
were increased may imply a rescue mechanism to balance catabolic and anabolic processes.
Nevertheless, both biomarkers stayed elevated even after 46 hours of recovery. These results suggest that even a
brief period of immobilization and unloading can have sustained consequences on articular cartilage metabolism
in both healthy females and males. This emphasizes the importance of implementing effective countermeasures to
preserve cartilage function for long-duration space missions.
REFERENCES
Shelhamer M., Bloomberg J., LeBlanc A., et al., 2020, NPJ Microgravity 6, 5.
Niehoff A., Brüggemann G.-P., Zaucke F., et al., 2016, Osteoarthritis Cartilage, 24(1), S144-S145.
Lotz, M., Martel-Pelletier J., Christiansen C., et al., 2013, Ann Rheum Dis, 72(11), 1756-1763.
Liphardt A.-M., Mündermann A., Andriacchi T.P., et al., 2018, J. Orthop. Res., 36(5), 1465-1471.
Liphardt A.-M., Mündermann A., Heer M., et al., 2020, J. Orthop. Res., 38(11), 2373-2382.
Otterness I.G. and Eckstein F., 2007, Osteoarthritis Cartilage, 15(6), 666-672.
Funding: Federal Ministry of Economic Affairs and Climate Action, Germany (DLR 50WB2021, DLR 50WB2022).
Figure 1: Study timeline to illustrate blood sampling time points. BDC: Baseline Data Collection (grey area), DI:
Dry Immersion (blue area), R: Recovery (green area)