Objective To evaluate the cytotoxicity of celastrol on 3T3 cells, predict acute toxicity (LD50) based on the cytotoxicity data, and compare acute toxicity results by using mouse up-and-down method and Bliss method.
Methods In this study, we employed an in-house validated in vitro cytotoxicity model based on the 3T3 cells neutral red uptake (NRU) assay. Using eight concentrations of celastrol (0.01-2.05 mg/L) for 48 h incubation, we calculated IC50 values for celastrol. The LD50 value of celastrol for systemic toxicity was computed by using RC model to set the starting dose for up-and-down procedure. Seventy ICR mice were divided into seven groups, half male and half female, and administered with 3, 4, 5, 6, 7, 8, and 0 mg/kg40% dimethyl sulfoxide (DMSO) and sodium chloride of celastrol intravenously. After observation for 14 days, we obtained LD50 values for celastrol using Bliss method based on the mortality rate.
Results The IC50 values for celastrol using 3T3 cells NRU assay was (0.119 7& #177;0.018 7) mg/L (n=2), with coefficient of determination R2 > 0.96. The computed LD50 value for celastrol systemic toxicity was 5.375 mg/kg. The starting dose of up-and-down procedure was 3.3 mg/kg, and the derived LD50 value was 3.157 mg/kg with 95% confidence interval of 3.1-5.5 mg/kg. The LD50 value derived from Bliss method was 4.899 mg/kg with 95% confidence interval of 4.483-5.354 mg/kg.
Conclusion The acute toxicity of celastrol predicted by 3T3 cytotoxicity test is consistent with the results by in vivo up-and-down procedure and Bliss method.
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