Effects of Different Vitrification Solutions and Protocol on Follicular Ultrastructure and Revascularization of Autografted Mouse Ovarian Tissue

Document Type : Original Article

Authors

1 . Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran

2 .Department of Veterinary, Shabestar Branch, Islamic Azad University, Shabestar, Iran

3 .Department of Anatomical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran

4 4.Department of Immunology, Tabriz University of Medical Sciences, Tabriz, Iran

5 5.Department of Reproductive Biology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran

Abstract

Objective
Many attempts have been made to preserve fertility by improving the cryopreservation of the ovarian tissue. This current studyaimed to improve of direct cover vitrification (DCV) protocol on follicular preservation and angiogenesis in autografted ovarian tissue.
Materials and Methods
In this experimental study, sixty five female Balb/c mice (5-6 week-old) were anesthetized and their ovaries were dissected. The left ovaries were vitrified by DCV solution, thawed by descending concentrations of sucrose, and then autografted subcutaneously. The right ovaries were autografted with no vitrification procedure prior to transplantation. The animals were sacrificed under anesthesia on the 7thday after transplantation to obtain ovarian tissue. Follicular quality was assessed by histological and ultrastructure observations, and angiogenesis was examined by immunohistochemical staining and real-time polymerase chain reaction (PCR) analysis.
Results
The histological and ultrastructure features of the follicles preserved well after vitrification of the ovarian tissue by 10% ethylene glycol (EG) and 10% dimethyl sulfoxide (DMSO). Revascularizationwas manifested prominently in the DCV1-vitrified/grafted ovaries by von Willebrand factor (vWF) and alpha smooth muscle actin (α-SMA) immunostaining. The ovarian tissue vitrified in DCV1 protocol had higher expression levels of angiopoietin-2 (Ang-2) and vascular endothelial growth factor (VEGF) 7 days after autotransplantation (P < 0.01).
Conclusion
These findings suggest that DCV with 10% of both EG and DMSO, is an effective cryopreservation solution for preservation of good quality follicles as well an upregulation of angiogenic factors after ovarian tissue transplantation.

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