Role of Pronase E in Isolation of Chondrocytes from Cartilage of Elderly Patients Receiving Total Joint Replacement

Abstract Purpose To examine the specific role of pronase E in isolation of human chondrocytes derived from total joint arthroplasty. Methods Cartilage were shaved from femoral head or tibial plateau of patients receiving total hip or knee joint replacement surgery (16 hips, 8 knees). Cartilage were subjected to 0.02% collagenase IA digestion for 16 hrs with (N = 19) or without (N = 5) the pre-treatment of 0.4% pronase E for 1.5 hrs. At the end of collagenase digestion, chondrocyte yield and viability were assessed. Chondrocyte phenotype was determined by the expression ratio of collagen type II to I. The morphology of cultured chondrocytes was monitored with a light microscope. Results Cell yield from pronase E pre-treatment group was significantly higher than that from the group without the pre-treatment (3,399 ± 1,637 cells/mg wet cartilage vs. 1,895 ± 688 cells/mg wet cartilage; P = 0.0067). Cell viability in the former group was also significantly higher than that in the latter (94% ± 2% vs. 86% ± 6%; P = 0.03). When cultured in monolayers, cells from cartilage with pronase E pre-treatment grew in a single plane showing rounded shape while cells from the other group grew in multi-planes and exhibited irregular shape. The mRNA expression ratio of collagen type II to I was 13.2 ± 7.5 in cells isolated from cartilage pre-treated with pronase E, indicating a typical chondrocyte phenotype. Conclusion Pronase E pre-treatment of human cartilage was crucial for chondrocyte isolation with collagenase IA and in vitro culturing.