A completely new strategy for cell culture focusing on the design of three-dimensional (3D) smart surfaces by supercritical fluid technology has been developed. This approach might overcome the limitations on cell expansion and proliferation of currently existing techniques. An alternative technology, based on supercritical carbon dioxide, was used to polymerize poly(N-isopropylacrylamide) (PNIPAAm) and to foam poly(D,L-lactic acid) (P(D,L)LA), creating a thermosensitive 3D structure which has proven to have potential as a substrate for cell growth and expansion. We demonstrated that the thermosensitive matrices promoted cell detachment, thus P(D,L)LA scaffolds have the potential to be used as substrates for cell growth and expansion avoiding enzymatic and mechanical methods of cell harvesting. The harvested cells were replated to evaluate their viability, which was not compromised. A major advantage of this technology is the fact that the prepared materials can be recovered and reused. Therefore, the same substrate can be recycled and reused for different batches. An indirect impact of the technology developed is related to the field of biotechnology, as this novel technology for cell expansion can be applied to any adherent cell cultures.