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Tendon injuries continuously rise, and regeneration is not only slow, but also limited due to the poor endogenous healing ability of the tendon tissue. Tissue grafts constitute the clinical gold standard treatment for severe injuries, but inherent limitations drive the field toward tissue engineering approaches to create suitable tissue constructs. Recapitulation of the native microenvironment represent a key challenge for the development of tendon tissue equivalents in vitro that can be further utilized as implantable devices. Methods to maintain cellular phenotype and to enhance extracellular matrix deposition for accelerated development of tissue-like modulus should be developed. Herein, we assessed the combining effect of surface topography and macromolecular crowding in human tenocyte culture. Our data demonstrated that bidirectionally aligned electrospun fibers induce physiological cell growth, while macromolecular crowding enhanced and accelerated tissue-specific extracellular matrix deposition. Collectively, these data advocate the use of multifactorial approaches for the accelerated development of functional tissue-like surrogates in vitro. The authors received financial support for this work from Science Foundation Ireland, Career Development Award (Grant Agreement Number: 15/CDA/3629); Science Foundation Ireland / European Regional Development Fund (Grant Agreement Number: 13/RC/2073); H2020, Marie Skłodowska-Curie Actions, Innovative Training Networks 2015 Tendon Therapy Train project (Grant Agreement Number: 676338).