Manganese ferrite nanoparticles were synthesized by a facile, low-cost, environmentally friendly and high yield methodology based on the aqueous co-precipitation of proper salts. Firstly, structural, morphological and magnetic characterization schemes were performed to determine crucial factors for optimizing their heating potential, such as size, polydispersity, saturation magnetization and coercivity. In an effort to simulate the in vivo environment of animal tissue phantoms and study the thermal heating effects resulting from Brownian motion and hysteresis losses, nanoparticles at various concentrations were embedded in aqueous media of varying agar concentration. During the in vitro application healthy cells (primary bone marrow-derived osteoblasts and 3T3-L1 fibroblast-like preadipocytes) and human osteosarcoma Saos-2 cells were incubated with manganese ferrite nanoparticles. The heating profile of the particles was studied at different concentrations and in correlation with their potential cytotoxic effect. Our results revealed concentration dependent cytotoxicity profile and uptake efficiency together with variable specific loss power values yet with fast thermal response, opening novel pathways in material selection as hyperthermia agents.