This article investigates the effects of simulated microgravity on immune function using single-cell analysis of human peripheral blood mononuclear cells (PBMCs). The study reveals that microgravity alters key immune pathways, including cytoskeleton, interferon signaling, pyroptosis, temperature-shock, innate inflammation, nuclear receptors, and sirtuin signaling. It also shows that microgravity affects monocyte inflammatory parameters and impairs T cell and NK cell functionality. Using machine learning, the study identifies compounds, such as the flavonol quercetin, that can reverse many of these abnormal pathways. The research validates findings using RNA sequencing, super-resolution microscopy, and data from space missions like Inspiration-4, JAXA, and the Twins study. The study highlights that microgravity induces immune dysfunction, including increased viral reactivation and shifts in immune responses toward Th2 immunity. It also shows that simulated microgravity alters gene expression in immune cells, leading to changes in pathways related to innate immunity, mitochondrial dysfunction, and cell stress. The study further demonstrates that simulated microgravity affects the cytoskeleton and mitochondrial morphology of immune cells, and that these changes may mimic those seen during TLR ligation. Overall, the research provides insights into the mechanisms of immune dysfunction in microgravity and suggests potential countermeasures to maintain normal immunity in space.This article investigates the effects of simulated microgravity on immune function using single-cell analysis of human peripheral blood mononuclear cells (PBMCs). The study reveals that microgravity alters key immune pathways, including cytoskeleton, interferon signaling, pyroptosis, temperature-shock, innate inflammation, nuclear receptors, and sirtuin signaling. It also shows that microgravity affects monocyte inflammatory parameters and impairs T cell and NK cell functionality. Using machine learning, the study identifies compounds, such as the flavonol quercetin, that can reverse many of these abnormal pathways. The research validates findings using RNA sequencing, super-resolution microscopy, and data from space missions like Inspiration-4, JAXA, and the Twins study. The study highlights that microgravity induces immune dysfunction, including increased viral reactivation and shifts in immune responses toward Th2 immunity. It also shows that simulated microgravity alters gene expression in immune cells, leading to changes in pathways related to innate immunity, mitochondrial dysfunction, and cell stress. The study further demonstrates that simulated microgravity affects the cytoskeleton and mitochondrial morphology of immune cells, and that these changes may mimic those seen during TLR ligation. Overall, the research provides insights into the mechanisms of immune dysfunction in microgravity and suggests potential countermeasures to maintain normal immunity in space.