Formation of human fibroblast aggregates (spheroids) by rotational culture

In the current study, we attempted to form aggregates of fibroblasts by rot ationally shaking, declining fibroblast-material interactions, and augmenti ng cell-cell interactions. In addition, to promote cell-cell interactions, the medium was supplemented with insulin, dexamethasone, and basic fibrobla st growth. Under such improved culture conditions, normal neonatal human de rmal fibroblasts formed spheroidal aggregates within 1 day of rotation on a rotational shaker. The aggregates that formed had irregular shapes and wer e composed from only several cells after 12 h. However, they became nearly spheroidal after 24 h of shaking. The aggregates were approximately 240 mum in diameter. After 36 h of shaking, their shape became more rounded and th eir surfaces became smoother. No evidence of necrosis in the center of the aggregates was observed, although a small number of dead cells was scattere d throughout the aggregates. After 24-36 h, aggregates of normal human fibr oblasts were collected and reinoculated onto a scaffold composed of polygly colic acid, which is used commercially as a scaffold for artificial skin, c oated with collagen. The aggregates were successfully trapped to the mesh o f polyglycolic acid and became attached within 24 h. Therefore, the aggrega tes could provide an alternative method for seeding fibroblasts to scaffold for art artificial skin, such as a mesh of polyglycolic acid.