Effects of Me2SO and Trehalose on the Cell Viability, Proliferation, and Bcl-2 Family Gene (BCL-2, BAX, and BAD) Expression in Cryopreserved Human Breast Cancer Cells

摘要

Long-term cryopreservation of the viability and metabolic state of cells in cancer cell/tissue specimens has significant implications for diagnostic verification of disease progression in cancer patients and selection of effective treatment options via development of the patient-derived xenograft (PDX) models for drug screening. The purpose of this study is to investigate the effects of cryoprotectant agents (CPAs) on the expression of BCL-2 family genes (BCL-2, BAX, and BAD) that are involved in the growth and development of breast cancers. MCF-7 cells were cryopreserved in Dulbecco's modified Eagle's medium (DMEM) with 20% (v/v) fetal bovine serum, using 10% (v/v) Me2SO (dimethyl sulfoxide, DMSO) or 7.5% (v/v) Me2SO with 100is-300 mM trehalose as cryoprotectant solutions. After storage at -80 degrees C for 7 days, the cells were thawed for evaluation. The use of Me2SO and trehalose has affected cell survival, proliferation, apoptotic state, as well as BCL-2 family gene expression. The conventional 10% (v/v) Me2SO method yields similar to 80% post-thaw cell survival and good cell proliferation, but it drastically alters the pattern of the BCL-2 family gene expression. The antiapoptotic gene BCL-2 is downregulated, whereas two proapoptotic genes BAX and BAD are upregulated. The partial substitution of Me2SO with 200 or 300 mM trehalose enhances cell proliferation of survived cells after cryopreservation. The presence of trehalose upregulates the expression of both the antiapoptotic gene BCL-2 and proapoptotic genes BAX and BAD. Cryopreservation could tip off the checkpoint of the apoptotic pathway regulated by the BCL-2 family members, and the effect may be protectant dependent. The findings of this study demonstrate the importance of paying attention to the potential change of gene expression and metabolic state of cancer cells after cryopreservation in an attempt to development of the PDX models from cryopreserved cancer cells or tissue specimens.