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Objective: To evaluate the ability of human adiposederived stem cells (h-ASCs) to survive and differentiate in corneal stroma. Methods: Our experiment consisted of 2 phases. First, we cultured h-ASCs in different types of hyaluronic acid (HA)-derived synthetic extracellular matrixes (sECMs) to determine the capability of proliferation and survival of the cells in hydrogels. Second, h-ASCs were grown in plastic flasks, labeled with an intracytoplasmic membrane fluorescent molecule, transferred onto different types of sECMs or the native HA product, and then inserted into the corneal stroma of the rabbits. After 10 weeks, we assessed the viability of the stem cells and the expression of cornea-specific proteins. Results: The in vitro study showed that the HyStem-HP hydrogel had the highest yield of cells (1.1×106/mL) compared with other types of HA-derived sECMs culture media, and the cells grown in the HyStem-HP hydrogel appeared more elongated and fibroblastlike. The in vivo study demonstrated that the labeled h-ASCs could be identified in the stroma with any type of sECM. The HAderived sECMs, particularly the HyStem-HP hydrogel, showed better survival and cell morphologic features compared with pure HA. Immunostaining of keratocan, aldehyde dehydrogenase, and type I collagen revealed that the stem cells had expressed human cornea-specific proteins. Conclusion: Human adipose-derived stem cells can be successfully grown on HA-derived sECMs in vivo and can express human cornea-specific proteins. Clinical Relevance: Human ASCs on an HA-derived scaffold may be used as a source of keratocytes to regenerate extracellular matrix-like material in situations where the cornea stroma has been compromised. [ABSTRACT FROM AUTHOR]