A pre-clinical model combining cryopreservation technique with precision-cut slice culture method to assess the in vitro drug response of hepatocellular carcinoma

Models considering hepatocellular carcinoma (HCC) complexity cannot be accurately replicated in routine cell lines or animal models. We aimed to evaluate the practicality of tissue slice culture by combining it with a cryopreservation technique. We prepared 0.3-mm-thick tissue slices by a microtome and maintained their cell viability using a cryopreservation technique. Slices were cultured individually in the presence or absence of regorafenib (REG) for 72 h. Alterations in morphology and gene expression were assessed by histological and genetic analysis. Overall viability was also analyzed in tissue slices by CCK-8 quantification assay and fluorescent staining. Tissue morphology and cell viability were evaluated to quantify drug effects. Histological and genetic analyses showed that no significant alterations in morphology and gene expression were induced by the vitrification-based cryopreservation method. The viability of warmed HCC tissues was up to 90% of the fresh tissues. The viability and proliferation could be retained for at least four days in the filter culture system. The positive drug responses in precision-cut slice culture in vitro were evaluated by tissue morphology and cell viability. In summary, the successful application of precision-cut HCC slice culture combined with a cryopreservation technique in a systematic drug screening demonstrates the feasibility and utility of slice culture method for assessing drug response.