Supraphysiologic Temperature Enhances Cytotoxic Effects of Bupivacaine on Bovine Articular Chondrocytes in an In Vitro Study

PurposeThe purpose of this study was to determine the effects of temperature or 0.25% bupivacaine treatment in combination with supraphysiologic temperatures on chondrocyte viability.MethodsBovine articular chondrocytes in suspension culture were treated with phosphate-buffered saline solution at 20°C, 37°C, 40°C, 42°C, 45°C, 47°C, and 50°C for 15, 30, and 60 minutes or with phosphate-buffered saline solution at 37°C, 45°C, and 50°C for 30 and 60 minutes followed by 0.25% bupivacaine at 20°C for 60 minutes. Chondrocyte viability was analyzed by flow cytometry with the LIVE/DEAD Viability/Cytotoxicity Kit (Molecular Probes, Eugene, OR). Annexin V and ethidium double staining determined whether apoptosis or necrosis occurred.ResultsTemperatures from 20°C to 42°C did not cause chondrocyte death. Temperatures at or above 45°C caused significant chondrocyte death, particularly at 50°C for 60 minutes, compared with 37°C at 60 minutes (P < .01). When the chondrocytes were incubated at 50°C, subsequent exposure to bupivacaine significantly increased chondrocyte death compared with the saline solution–treated control group (P < .001). There were additive cytotoxic effects when bupivacaine was combined with supraphysiologic temperatures. It was also found that bupivacaine at supraphysiologic temperatures caused necrosis of articular chondrocytes.ConclusionsTemperatures at or above 45°C caused significant chondrocyte death. Bupivacaine treatment in the presence of 45°C and 50°C temperatures significantly increased necrosis of bovine articular chondrocytes in this in vitro study.Clinical RelevanceImmediate intra-articular injection of bupivacaine after heat-generating procedures may cause damage to the cartilage because of the additive cytotoxic effects of bupivacaine and elevated temperature.