Corneal disease remains to be one of the leading causes of blindness in the world and limbal stem cell (LSC) therapy is a promising therapy for LSC deficiency, which is associated with the diseased corneal epithelium repair. Soft substrate could effectively promote the stemness maintenance of LSC and thus modification of cell culture substrate would help in the potential LSC deficiency therapy. Both Hippo-Yes-associated protein (YAP) and Notch pathway have been reported to affect the LSC function, however, the detailed mechanisms remain unclear. Instead of some soft but biologically toxic substrates, we present a hypothesis on the application of soft substrate generated by HA/PTX3, an FDA approved nontoxic drug, on the LSC culture in this current study. Soft substrate could help in the stemness maintenance and thus promote the LSC deficiency treatment. In more detailed mechanism detection, we hypothesize that soft substrate would block the activation of Hippo-YAP pathway and thus decrease the activity of Notch pathway. This proposed hypothesis should be evaluated by both a series of in-vitro experiments based on soft and stiff substrates and in-vivo treatment with LSC cultured in different conditions. Advanced experiments on related cellular behaviors and detailed molecular mechanisms would provide us more knowledge on the molecular mechanism detection as well as cell transplantation therapy.Copyright © 2021 Elsevier Ltd. All rights reserved.