Novel cell culture model using pure hydrostatic pressure and a semipermeable membrane pouch

Cell constructs and culture methods are essential tools in tissue engineering. The cell construct should be equivalent to the native cartilage it is intended to replace. Thus, three-dimensional cell constructs are usually composed of a high density of cells and dense extracellular matrix. However, dense constructs suffer from a lack of passive nutrient supply, gas exchange, and removal of degraded debris. We have developed a novel hydrostatic pressure/perfusion culture system that improves the quality of neo-tissues, providing an automated and affordable system for clinical applications. We have also developed a semipermeable membrane pouch that contains a fragile amorphous cell carrier. Although amorphous material is difficult to handle, it is a useful medium in which to deliver cells to the desired site via injection. We evaluated phenotypes of bovine articular chondrocytes embedded in a collagen type I gel enclosed within membrane pouches permeable to molecules of various sizes. Constant or cyclic hydrostatic pressure was externally applied to the medium phase with a new culture system. Accumulation of cartilage specific matrix was promoted with a 500-kDa cutoff membrane pouch and cyclic hydrostatic pressure at 0.5 MPa, 0.5 Hz. This new method will be useful in the delivery of engineered cells to a desired tissue in regenerative medicine.