<p>An ideal implant surface should promote cell attachment and tissue integration while preventing bacterial colonization. Engineering the surface topography of a biomaterial implant to elicit a differential response is a promising approach to dealing with both issues. To achieve this, we integrated Moth-Eye (ME) nanocones with micrometric features such as gratings and pillars in various hierarchical configurations, leading to different responses in bacteria and cells. These hierarchical topographies were tested on different polymers to compare their biological responses. Mesenchymal stem cells (MSCs) were employed as a model system to study cell behavior, given their ability to proliferate, self-renew, and differentiate. The bactericidal effect was tested using gram-negative <i>Escherichia coli</i> and gram-positive <i>Staphylococcus aureus</i>. Results show that these topographies-maintained cell viability and influenced cell morphology, orientation, migration and differentiation, which are crucial processes for regeneration and osseointegration. Finite Element Method (FEM) simulations estimated the traction forces generated by cells on these surfaces. Overall, the hierarchical topographies enhanced the expression of osteogenic markers in the MSCs while retaining the bactericidal properties of the ME nanostructures.</p>