Traditional ex vivo culture setups fail to imitate the native tissue niche, leading to cellular senescence, phenotypic drift, growth arrest and loss of stem cell multipotency. Growing evidence suggests that surface topography, substrate stiffness, mechanical stimulation, oxygen tension and localised density influence cellular functions and longevity, enhance tissue-specific extracellular matrix deposition and direct stem cell differentiation. In this review, we discuss how these cues will facilitate engineering of physiological in vitro microenvironments to enable clinical translation of cell based therapies and development of in vitro models for drug discovery applications.