Updated project metadata. Bone loss and immune dysregulation are among the main adverse outcomes of spaceflight challenging astronaut’s health and safety. However, consequences on B cell development and responses are still under-investigated. Up to now, most studies addressing these questions were performed using an amphibian species. Consequently, we used advanced proteomics analysis of femur bone and marrow of mice flown for one month on board the BION-M1 biosatellite, to determine whether extreme conditions encountered during a real spaceflight affect B cell development in mice and to examine reversibility of the effects upon return to Earth. Our data revealed that adverse effects on B lymphopoiesis were more marked one week after landing and that this phenomenon was associated with a 41% reduction of B cells in the spleen. Thus, the effects of spaceflight persisted during at least one week after landing. These reductions may contribute to explain increased susceptibility to infection even if we confirmed that animals were able to mount a humoral immune response.