Microgravity-Induced Immune Plasticity and Temporal Modulation of Gene Networks and T Cell Subsets During the MESSAGE Science Mission in Axiom-3 on the ISS

Microgravity presents significant challenges to human physiology, particularly the immune system, during spaceflight. T lymphocytes, central regulators of adaptive immunity, are known to be sensitive to gravitational changes; however, the molecular mechanisms underlying immune modulation in space remain incompletely understood. This study, conducted as part of the MESSAGE (Microgravity Associated Genetics) Science Mission during the Axiom-3 mission, investigated how short-term microgravity influences immune regulation (https://www.nasa.gov/mission/station/research-explorer/investigation/?#id=9100). Blood samples were collected from three astronauts at Day 4, Day 7, and Day 10 of the mission, as well as from one suborbital astronaut and three healthy controls. Hemogram profiling, flow cytometry-based T cell subpopulation analysis, and RNA sequencing (RNA-Seq) were performed to characterize immune dynamics at cellular and molecular levels. Hemogram analysis showed no statistically significant changes in leukocytes, erythrocytes, haemoglobin, or hematocrit levels, suggesting stable hematological profiles during short-duration spaceflight. Flow cytometry revealed non-significant shifts in effector memory T cell subsets (TemEARLY and TemLATE), with a trend toward increased late effector memory differentiation by Day 10. Transcriptomic profiling identified time-dependent modulation of immune-related genes, including downregulation of CD69, IL2RA, LAT, and IFNG, indicating early suppression of T cell activation pathways. Heatmap analysis revealed coordinated transcriptional changes across 48 immune-associated genes, demonstrating that molecular adaptations occur before detectable alterations in immune cell counts or hematological profiles. These findings provide evidence of early, coordinated transcriptional reprogramming of immune-regulatory pathways under microgravity, highlighting the potential vulnerability of immune function during spaceflight. While cellular and haematological changes remained limited, transcriptomic signatures indicate that molecular responses precede phenotypic shifts. Understanding these mechanisms is critical for developing targeted countermeasures to protect astronaut immune health during long-duration space missions.