A cryoinjury model using engineered tissue equivalents for cryosurgical applications

Cryosurgery is emerging as a promising treatment modality for various cancers, but there are still challenges to be addressed to improve its efficacy. Two primary challenges are determining thermal injury thresholds for various types of cell/tissue, and understanding of the mechanisms of freezing induced cell/tissue injury within a cryolesion. To address these challenges, various model systems ranging from cell suspensions to three-dimensional in vivo tissues have been developed and used. However, these models are either oversimplifications of in vivo tissues or difficult to control and extract precise experimental conditions from. Therefore, a more readily controllable model system with tissue-like characteristics is needed. In this study, a cryoinjury model was developed using tissue engineering technology, and the capabilities of the model were demonstrated. Engineered tissue equivalents (TEs) were constructed by seeding and culturing cells in a type I collagen matrix. Two different cell lines were used in this study, AT-1 rat prostate tumor cells and LNCaP human prostate cancer cells. The constructed TEs underwent a freeze/thaw cycle imitating in vivo cryosurgery. Thermal conditions within TEs during freeze/thaw cycles were characterized, and the responses of TEs to these thermal conditions including freezing induced cellular injury and extracellular matrix damage were investigated at three different time points. The results illustrate the feasibility to establish thermal thresholds of cryoinjury for different cell/tissue types using the presently developed model, and its potential capabilities to study cell death mechanisms, cell proliferation or migration, and extracellular matrix structural damage after a freeze/thaw cycle.