Determination of viability of human cartilage allografts by a rapid and quantitative method not requiring cartilage digestion.
Fresh osteochondral allograft transplantation is increasingly used for the treatment of cartilage pathologies of the knee. It is believed that transplantation success depends on the presence of viable chondrocytes in the graft, but methods to evaluate graft viability require the isolation of chondrocytes by enzymatic digestion of the cartilage and/or the use of radioactive precursors. We have adapted the well-known cell viability assay based on the reduction of tetrazolium derivatives to evaluate cartilage viability. We took advantage from the histological properties of cartilage tissue and the fact that some tetrazolium derivatives (e.g., WST-1, XTT) give soluble reduction products that can permeate the hyaline cartilage matrix. We have validated this assay in human cartilage explants from arthrotomy interventions and deceased donors, measuring the reduced product in the explant supernatant. Using this method we have compared the performance of several culture media in cartilage viability. From those tested, DMEM supplemented with fetal bovine serum results in higher viability of the cartilage and the explants remain viable at least 15 days in culture at 37 degrees C. Cartilage cells continued expressing chondrocyte-specific genes, suggesting the maintenance of chondrogenic phenotype. The described method offers a quantitative and convenient method to measure the viability of human cartilage grafts.
López
Carmen
C
Departmento de Biología Molecular, Instituto de Biomedicina y Biotecnologia, CSIC, Universidad de Cantabria, Santander, Spain.
Ajenjo
Nuria
N
Muñoz-Alonso
Maria J
MJ
Farde
Pilar
P
León
J
J
Gómez-Cimiano
J
J
D000818:Animals; D002356:Cartilage / Q000166:cytology* / Q000473:pathology; D002417:Cattle; D002470:Cell Survival; D019902:Chondrocytes / Q000637:transplantation*; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry / Q000378:metabolism; D015203:Reproducibility of Results; D013778:Tetrazolium Salts / Q000737:chemistry / Q000378:metabolism; D014184:Transplantation, Homologous / Q000379:methods*
D007202:Indicators and Reagents; D013778:Tetrazolium Salts