The in vitro growth of a three-dimensional human dermal replacement using a single-pass perfusion system

A human dermal replacement has been developed by seeding human neonatal dermal fibroblasts onto a biosorbable polyglactin (polyglycolide/polylactide) mesh and culturing in a bioreactor. The mesh provides the proper environment for the cells to attach, grow in a three-dimensional array, and establish a tissue matrix over a 2- to 3-week culture period. The dermal replacement has been characterized and found to contain a variety of naturally occurring dermal matrix proteins, including fibronectin, glycosaminoglycans, and collagen types I and III. To efficiently and reproducibly produce this dermal tissue equivalent, a closed, single-pass perfusion system was developed and compared with a static process. In the single-pas perfusion system, growth medium (containing ascorbic acid) was perfused around the 4 x 6 in. pieces of mesh at specific flow rates determined by nutrient consumption and waste production rates. The flow rates used for this system indicate that a diffusion-limited regime exists with a mean residence time greater than 1 h for essential nutrients and factors. By controlling glucose concentrations in the system to a delta of 0.70 g/L from the inlet to the outlet of the bioreactor, it took 6 fewer days to grow a tissue similar to that produced by the static system.