The study evaluated Ciona robusta extract (CRE) derived from an invasive marine tunicate, for its potential to improve skin hydration and collagen production in human dermal fibroblasts (HDFn) and keratinocytes (HaCaT). CRE increased procollagen levels in fibroblasts and hyaluronic acid production in keratinocytes, while also increasing the expression of skin-related genes including COL1A1, COL1A2, COL3A1, HAS2, HAS3, FLG, and AQP3. These changes suggest enhanced collagen synthesis, extracellular matrix formation, and maintenance of skin moisture. CRE also influenced TGF-β and signaling pathways such as MAPK, PI3K/AKT, and STAT3, which are involved in collagen production and skin hydration. Ciona robusta extract may help improve skin moisture and structural integrity by supporting collagen and hyaluronic acid production.