The objective of this study was to optimize titanium surfaces by means of Ti:Sapphire femtosecond laser to improve the attachment of human cartilage cells on titanium prosthesis in middle ear surgery. The application of microstructures on titanium samples was evaluated and the influence of these microstructures on human auricular chondrocytes was studied in-vitro. After establishing the ear chondrocyte cell culture, cells were seeded on titanium platelets with selected microstructure patterns. Whereas the phenotype of cells seeded on unstructured titanium was similar to cells grown on standard tissue culture surfaces, the morphology of chondrocytes grown on structured titanium samples was influenced by the pattern. For future titanium middle ear prosthesis structural optimizations will be developed to promote chondrocyte growth and adhesion while impeding fibrocyte proliferation to avoid scarring on implant interfaces.