Tendon injuries are a common problem in humans and horses. There is a high re-injury rate in both species due to the poor regeneration of adult tendon and the resulting formation of scar tissue. In contrast, fetal tendon injuries undergo scarless regeneration, but the mechanisms which underpin this are poorly defined. It is also unclear if tendon cells derived from embryonic stem cells (ESCs) would aid tendon regeneration. In this study we determined the responses of adult, fetal and ESC-derived equine tenocytes to a range of cytokines, chemokines and growth factors that are upregulated following a tendon injury using both 2-dimensional (2D) and 3-dimensional (3D) in vitro wound models. We demonstrated that in 2D proliferation assays, the responses of fetal and adult tenocytes to the factors tested are more similar to each other than to ESC-tenocytes. However, in 2D migration assays, fetal tenocytes have similarities to both adult and ESC-tenocytes. In 3D wound closure assays the response of fetal tenocytes also appears to be intermediary between adult and ESC-tenocytes. We further demonstrated that while TGFβ3 increases 3D gel contraction and wound healing by adult and fetal tenocytes, FGF2 results in a significant inhibition by adult cells. In conclusion, our findings suggest that differential cellular responses to the factors upregulated following a tendon injury may be involved in determining if tendon repair or regeneration subsequently occurs. Understanding the mechanisms behind these responses is required to inform the development of cell-based therapies to improve tendon regeneration.