Cryopreservation of vascularized ovary: An evaluation of histology and function in rats

Abstract

Cryopreservation of organs has been investigated to sustain the reproductive function of patients undergoing sterilizing chemotherapy and radiotherapy or reproductive surgery. A modified protocol for whole organ cryopreservation was described and the outcome of cryopreservative ovaries was evaluated, and apoptosis of cryopreservative cells stored for different time period and the viability of cryopreserved cells stored at different temperature was examined in rats. Lewis rat ovarian grafts were perfused for 30 min at 0.35 ml/min with M2 medium containing 0.1M fructose and increasing concentrations of 0-1.5M dimethylsulfoxide, cooled to -140 degrees C controlled by a computerized program, and stored in liquid nitrogen (-196 degrees C) for 24 hours. After being thawed, ovaries were transplanted to syngeneic recipients after bilateral oophorectomy. Graft functions were monitored postoperatively. The major findings were that: 1) A 100% survival rate of rat ovaries was achieved in this study. Ovarian hormone secretion recovered in 80% rats which had received cryopreservative ovarian grafts. Postoperative serum estradiol levels in the cryopreservative graft group were lower than in the sham surgery control, but much higher than in the bilateral oophorectomy group. 2) Histological examination of cryopreservative ovarian grafts showed preantral and antral follicles. Two gestations were obtained. 3) Estradiol levels remained low in ovariectomized rats while in the oophorectomized rats given cryopreservative ovarian grafts levels started to rise after 14 +/- 3 days. 4) The average viability in the cells from cryopreservative ovary organ (-196 degrees C) was about 71 +/- 18% compared to 90 +/- 9% of fresh cells. This success should encourage further improvement of cryopreservative techniques for large organs.

The human postmenopausal ovary as a tool for evaluation of cryopreservation protocols towards whole ovary cryopreservation

“…In our experimental approach we did not have to dissect the ovarian veins, which usually are 2-3 in numbers. It should be emphasized that careful cannulation, to avoid damage to the blood vessel walls, is important in organ cryopreservation since the blood vessels seem to be especially vulnerable to the freezing procedure [19].…”

Production of Donor-Derived Offspring from Cryopreserved Ovarian Tissue in Japanese Quail (Coturnix japonica)1

“…Apart from the cryopreservation strategy, failure to retain reproductive potential of grafts may also be associated with the transplantation procedure itself (e.g., ischemia-reperfusion injury and failure of the reestablishment of hormonal cycling) [38,39]. In the progeny test in this study, only two of nine recipients in the slow-freezing group produced donor-derived offspring, and the offspring from both of them were only donor derived, indicating complete removal of host ovaries but a low rate of recovery of the grafts.…”

Cryopreservation of Human Ovarian Tissue

“…Combinations of vascular cryoprotectant perfusion and immersion in liquid nitrogen, also by vitrification have been tested, but the histology of the ovaries has not yet been optimal. The technique is demanding and still at early experimental stage in animal models as rat (Qi et al, 2008) and sheep (Onions et al, 2009;Wallin et al, 2009;Demeestere et al, 2009). Promising six-year follow-up results have been published (Arav et al, 2010).…”