Orbital Spaceflight Remodels miRNA Expression and Suppresses miRNA Biogenesis Machinery in Astronaut PBMCs

MicroRNAs (miRNAs) regulate cellular stress responses and immune homeostasis, but their temporal behavior and biogenesis machinery during human spaceflight remain poorly characterized. We performed a miRNA-focused transcriptomic analysis of peripheral blood mononuclear cells collected during the MESSAGE Science Mission, integrating terrestrial reference samples, longitudinal samples from three Axiom-3 astronauts before launch and on International Space Station Days 4, 7, and 10, and a descriptive post-flight sample from the Galactic-07 suborbital mission. Of 2,239 profiled miRNA-associated transcripts, 118 met the expression and analytical filtering criteria, and 36 were significantly altered across the replicated mission phases. Of these, 34 showed decreased expression, and 2 showed increased expression, indicating a predominantly suppressive response. Hierarchical clustering and trajectory analysis identified distinct temporal programs characterized by sustained repression, transient post-launch or early-orbital induction, and delayed changes during orbital residence. Functional enrichment associated the responsive genes with miRNA-mediated gene silencing, vascular development, angiogenesis, cell migration, extracellular organization, and metabolic regulation. Genes encoding core components of miRNA maturation and activity—including DROSHA, DGCR8, DICER1, AGO1–4, XPO1, XPO4–7, and XPOT—showed coordinated reductions from ISS Day 4 onward, with limited recovery at Days 7 and 10. Co-expression and Gene Ontology analyses further linked this network to pre-miRNA processing, nuclear export, nucleocytoplasmic transport, and RNA-induced silencing complex assembly. These findings demonstrate that orbital spaceflight is associated with coordinated remodeling of miRNA expression and suppression of transcripts required for miRNA processing, transport, and effector function in astronaut PBMCs. This regulatory signature provides a foundation for mechanistic validation and development of miRNA-based biomarkers for astronaut health monitoring during longer missions.