Figure 1.

Experimental outline for tri-layer culture. Skin and adipose tissues were cultured independently prior to being combined at ‘day 0’. At day -21, endothelial cells are seeded onto silk sponges, while adipose derived stem cells undergo adipogenic differentiation in 2D. At day 14, the adipocyte-like cells are added to the endothelial seeded silk sponges and the co-cultures were maintained in a 1:1 adipogenic:endothelial cell media. Also at day 14, fibroblasts are cast within a collagen gel. After the cell contracts, keratinocytes are seeded on top of the collagen gel. After epithelialization has occurred the construct was raised to the air-liquid interface for 3 days. On day 0, the 2 constructs were combined with a thin layer of collagen gel as a binder. At this point the tri-layer construct was cultured in a 1:1 mix of the co-culture media to skin media.