Mechanical loading directs the differentiation of mesenchymal stem cells (MSCs) in vitro and it has been hypothesized that the mechanical environment plays a role in directing the cellular fate of MSCs in vivo. However, the complex multicellular composition of trabecular bone marrow means that the precise nature of mechanical stimulation that MSCs experience in their native environment is not fully understood. In this study, we developed a multiscale model that discretely represents the cellular constituents of trabecular bone marrow and applied this model to characterize mechanical stimulation of MCSs in vivo.We predicted that cell-level strains in certain locations of the trabecular marrow microenvironment were greater in magnitude (maximum e12¼24,000 le) than levels that have been found to result in osteogenic differentiation of MSCs in vitro (>8000 le),which may indicate that the native mechanical environment of MSCs could direct cellular fate in vivo. The results also showed that cell–cell adhesions could play an important role in mediating mechanical stimulation within the MSC population in vivo. The model was applied to investigate how changes that occur during osteoporosis affected mechanical stimulation in the cellular microenvironment of trabecular bone marrow. Specifically,a reduced bone volume (BV) resulted in an overall increase in bone deformation, leading to greater cell-level mechanical stimulation in trabecular bone marrow (maximume12¼48,000 le). An increased marrow adipocyte content resulted in slightly lower levels of stimulation within the adjacent cell population due to a shielding effect caused by the more compliant behavior of adipocytes (maximum e12¼41,000 le). Despite this reduction, stimulation levels in trabecular bone marrow during osteoporosis remained much higher than those predicted to occur under healthy conditions. It was found that compensatory mechanobiological responses that occur during osteoporosis, such as increased trabecular stiffness and axial alignment of trabeculae, would be effective in returning MSC stimulation in trabecular marrow to normal levels. These results have provided novel insight into the mechanical stimulation of the trabecular marrow MSC population in both healthy and osteoporotic bone, and could inform the design three dimensional(3D) in vitro bioreactor strategies techniques, which seek to emulate physiological conditions.