Intervertebral disc cell protection of triamcinolone against bupivacaine toxicity

Abstract

Background context: Local anesthetics combined with corticosteroids are commonly used for management of back pain in interventional spinal procedures. Several recent studies suggest cytotoxicity of bupivacaine, whereas others report protective and cytotoxic effects of corticosteroids on chondrocytes and intervertebral disc cells. Considering the frequent use of these agents in spinal interventions, it is meaningful to know how they affect intervertebral disc cells.Purpose: This study was conducted to assess the effects of bupivacaine and triamcinolone, both alone and in combination, on viability of intervertebral disc cells in vitro.Methods: Nucleus pulposus cells were isolated from human disc specimens from patients undergoing surgery due to disc herniation or degenerative disc disease. They were grown in three-dimensional alginate beads for one week to maintain their differentiated phenotypes and to allow for matrix formation before analysis. After one week of culture, the cells were exposed to bupivacaine (0.1%, 0.25%, 0.5%, and 1%) or bupivacaine (0.1%, 0.25%, 0.5%, and 1%) with 1mg triamcinolone for 1, 3, or 6 hours. Cell viability was measured using trypan blue exclusion assay and flow cytometry. Live-cell/dead-cell fluorescent imaging was assessed using confocal microscopy.Results: Trypan blue exclusion assays demonstrated dose- and time-dependent cytotoxic effects of bupivacaine on human nucleus pulposus cells. Similar but reduced cytotoxicity was observed after exposure to the combination of bupivacaine and 1mg of triamcinolone. Flow cytometry showed a dose-dependent cytotoxic effect of bupivacaine on nucleus pulposus cells after three hours of exposure. The reduced cytotoxicity of bupivacaine combined with 1mg triamcinolone was also confirmed in flow cytometry. Confocal images showed that the increase in dead cells correlated with the concentration of bupivacaine. Nevertheless, fewer cells died after exposure to several different concentrations of bupivacaine combined with 1 mg triamcinolone than did after exposure to bupivacaine alone.Conclusions: The combination of bupivacaine and triamcinolone induced dose- and time-dependent cytotoxicity on human intervertebral disc cells in vitro, but the cytotoxicity was much weaker than that of bupivacaine alone. This study shows a potential protective influence of triamcinolone on intervertebral disc cells.