Osteoimmunolgy describes the interactions between blood immune cells and human multipotent mesenchymal stromal cells (hMMSCs) as a basis of successful wound and fracture healing. The aim was to investigate in vitro interaction of adipose-derived hMMSCs and human blood mononuclear cells (hBMNCs) modulated by calcium phosphate (CaP) coating in 3D culture. Titanium plates (10 u 10 u 1 mm3) with micro-arc bilateral multilevel CaP coatings were used. Cell-IQ phase-contrast microscopy and the real-time cell analyzer (RTCA) showed the CaP coating and hBMNCs synergistic negative effects on hMMSC motility that could be conditioned by enhanced osteogenic differentiation of stromal cells. Indeed, a 10-fold increase in the bone mineralization around the CaP-coated samples was detected in the mixed (hBMNCs + hMMSCs) 3D culture. Thus, cellular and molecular crosstalk between hBMNCs and hMMSCs modulated by multilevel micro-arc CaP coating with bone-like topography is an effective 3D model to study in vitro the novel pathways of osteoimmunology.