Cellular and molecular mechanisms of cell death following cryo-injury

Cryosurgery is a form of cryotherapy involving the application of freezing temperatures for the destruction of unwanted tissues. Since its initial emergence mainly as a topical treatment for dermal abnormalities, technological developments have significantly impacted the cryosurgical field. To further advances in cryotherapy, the work presented herein was conducted in an effort to characterize the underlying cell death processes likely to occur following the injuries caused by cryosurgery. We employed an in vitro cryosurgical model to investigate the contribution of apoptosis to cell death in cell types relevant to prostate cryoablation and endovascular cryotherapy.; To investigate the role that apoptotic cell death plays during prostate cryoablation, a human prostate cancer cell line (PC-3) was subjected to a wide range of temperatures (+37 to -75°C). Subsequent assessment revealed apoptotic DNA fragmentation following exposures at and above -15°C, which could be inhibited through the addition of an apoptotic pan-inhibitor. Inhibitor addition not only blocked visible apoptotic laddering, but also conferred detectable levels of PC-3 protection at all target temperatures ranging from -10 to -75°C. This inhibition occurred even at temperatures at which necrosis appeared to be the primary cause of death. The deduced apoptotic tendency revealed by the PC-3 studies presented herein has since resulted in the development of a promising new rationale for an adjunctive cryo-chemo approach to prostate cancer.; The role of apoptotic cell death was further investigated in an endovascular cryotherapy model. Two non-cancer cellular components of the human coronary artery vasculature, coronary artery endothelial cells (CAEC) and smooth muscle cells (CASMC), were briefly exposed to a range of temperatures (+37 to -60°C). CAEC exhibited superior viability and proliferative capacity relative to CASMC following cold exposure between 0 and -15°C. DNA fragmentation analysis and caspase activity assessment confirmed an apoptotic contribution to cell death in both CAEC and CASMC following a mild freeze exposure (-5°C). Protein expression levels of Bcl-2 family members (Bcl-2, Bcl-xL, and Bax) did not appear to be a factor in apoptotic cell death in these study. These molecular findings should help advance research in apoptotic cell death and aid in the optimization of endovascular cryotherapy.