Directing adipose-derived stem cells into keratinocyte-like cells: impact of medium composition and culture condition. 
Adipose-derived stem cells (ASC) are known to transdifferentiate into a wide range of different cell species in vitro including along the epidermal lineage.  This property makes them a promising tool for regenerative medicine to restore the epidermal barrier.  This study is dedicated to identify in vitro conditions enabling transdifferentiation to a keratinocyte-like phenotype.  In particular, the impact of different culture conditions (media compositions, 2D, 3D cultures) and extracellular matrix (ECM) molecules was evaluated.  Adipose-derived stem cells derived from subcutaneous abdominal fat were characterized by stemness-associated markers and subjected to different media.  Epithelial differentiation in 2D cultures was monitored by pan-cytokeratin expression using flow cytometry and immunocytochemistry.  To evaluate the impact of different ECM molecules on epidermal stratification, 3D cultures were produced, lifted to the air-liquid interface (ALI) and examined by histological analysis and quantitative real-time RT-PCR.  We identified a medium composition containing retinoic acid, hydrocortisone, ascorbic acid and BMP-4 enabling maximum pan-cytokeratin expression in 2D cultures.  Moreover, adhesion to type IV collagen further promotes the pan-cytokeratin expression.  When cultures were lifted to the ALI, significant stratification was observed, particularly in supports coated with type IV collagen or fibronectin.  Moreover, epidermal differentiation markers (involucrin, cytokeratin 1 and 14) become induced.  Conditions with hampered wound healing such as non-healing ulcers demand new treatment regimes.  The here introduced optimized protocols for transdifferentiation of ASC into keratinocyte-like cells may help to establish more effective treatment procedures.