Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-Xenotransplantation.

Abstract Background: Ovarian tissue cryopreservation has a wide range of cancerous indications. Avoiding relapse becomes a specific area of concern that clinicians frequently encounter. The data about the comparative viability of cancer cells after cryopreservation are limited. This study aimed to evaluate the effect of cryopreservation on breast cancer cells.Methods: Samples were prepared using ZR-75-1 and MDA-MB-231 cell lines and divided into cryopreserved and non-intervened groups, respectively. Biological properties and the related protein markers were investigated. Cell morphology was monitored under the microscope. Cell proliferation, migration, and invasion were characterized by CCK-8, wound-healing, and transmembrane assay, respectively. The expression of Ki-67, P53, GATA-3, E-cadherin, Vimentin, and F-Actin was measured by immunofluorescent staining and western blotting. Xenotransplantation was established on the chorioallantoic membrane (CAM) culture system to explore angiogenesis close to the grafts.Results: Lamellipodia and filopodia were observed in cryopreserved ZR-75-1 cells. Both cell lines demonstrated increased cell motility and invasive ability after cryopreservation. However, cell proliferation was invariable in accordance with a regular expression of Ki-67 and P53. In ZR-75-1 cells, data exhibited a downregulation of E-cadherin after the decreased expression of GATA3, indicating the loss of intercellular adhesion after cryopreservation. Vimentin and F-actin were both upregulated in cryopreserved sample cells. Angiogenesis in CAM was significantly activated by cryopreserved MDA-MB-231 cells.Conclusions: Cryopreservation causes an increasing malignancy of ZR-75-1 and MDA-MB-231 cells and thus raises the risk of metastasis.