Cryopreservation of artificial cartilage: Viability and functional examination after thawing

In biomedical research and in reconstructive surgery, preservation of intac t tissue has been an unsolved problem. In this study, we investigated the v iability of cryopreserved artificial cartilage and its synthetic activity o f cartilage-specific matrix proteins after thawing for in vitro use. A poly mer fleece cylinder (diameter = 3 mm; height = 3 mm) was loaded with a susp ension of bovine chondrocytes (25 x 10(6)/ml) and encapsulated with fibrin glue. After a culture period of 1 week, the artificial cartilage units were frozen in a cryoprotection solution containing 10% basal medium (RPM[ 1640 ), 10% DMSO and 80% FCS. The freezing procedure consisted of three steps: a 30-min period at +4 degreesC followed by a 24-hour storage at -80 degreesC . After that, the tissue units were transferred into liquid nitrogen (-196 degreesC) for final storage. Using histochemical staining techniques of cry ogenic slices, we investigated the ability of cryopreserved artificial cart ilage to produce its specific matrix after thawing. A modified MTT assay wa s used to determine the viability of frozen tissue units in comparison with unpreserved samples at different moments after thawing. Depending on the c hondrocytes used for the formation of artificial cartilage, the viability o f cryopreserved tissue varied between 65 and 85%. Both the intensity of alc ian blue staining for proteoglycans and the azan staining for collagens inc reased proportionally with incubation time after thawing. These findings in dicate that cryopreservation of small artificial cartilage units is possibl e with a minor loss of cell viability. Secondly, its synthetic activity of cartilage-specific matrix did not decline after the freezing process. Copyr ight (C) 2001 S. Karger AG, Basel.