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USE OF THE ROTATING BIOREACTOR TO STUDY SKELETAL MUTATIONS: HEREDITARY MULTIPLE EXOSTOSIS.    P.J. Duke¹, D. Montufar-Solis¹, J.T. Hecht². Dept. of Orthodontics¹, Dental Branch, Dept. of Pediatrics² Medical School, University of Texas Health Science Center, Houston TX.

Osteochondromas are the most common bone tumors. These cartilage capped, bony projections are benign, and in individuals with Hereditary Multiple Exostoses (HME), are related to mutations in the putative tumor suppressor genes EXT1 and EXT2. Ultrastructural and confocal microscopy studies of cultured chondrocytes from HME excised exostoses identified abnormal amounts of α-actinin microfilaments throughout the cell. These fibrils were also found in freshly excised exostoses. HME chondrocytes are difficult to study because they will not differentiate in the usual systems developed for chondrocyte culture, i.e., agar, agarose, and alginate beads. 

The rotating bioreactor provides an environment for 3-D development of tissues.  In the present study, human costochondral chondrocytes and chondrocytes from excised HME exostoses were expanded in monolayer culture, trypsinized, aggregated on a rotating table, and then cultured for 3 weeks in a bioreactor. The aggregation of the two cell types differed: HME cells formed smaller, tighter aggregates, and continued to aggregate when placed in the bioreactor. Control chondrocytes formed cartilage as shown by positive staining with type II collagen antibody, and Toluidine blue metachromasia. HME chondrocytes did not stain metachromatically, nor did cells have the histological appearance of cartilage, but the fibrous components previously observed were seen intracellularly. These studies suggest that the formation of exostosis projections from the growth plates of HME individuals is related to the cell’s inability to form normal cell-cell or cell-matrix contacts, and to receive and respond correctly to cartilage differentiation signals.

(Supported by Shriners Hospital for Children grant #15955 to JTH and by UT Dental Branch Pilot Research Grant to JD.)