Ovary and ovulation: Low temperature storage and grafting of human ovarian tissue

Abstract

Ovarian tissue storage at low temperatures is a promising new method for protecting young cancer patients from the sterilizing effects of chemotherapy and/or radiotherapy. Tissue can be stored and returned to the body in due course as a thin cortical graft since the primordial follicles are distributed peripherally and are relatively resistant to ischaemia. Slices of tissue donated by healthy patients were trimmed to a uniform size and preserved by slow freezing to liquid nitrogen temperatures for up to 2 months in one of the following cryoprotectants: dimethylsulphoxide, ethylene glycol, glycerol, propylene glycol. Their viability was assessed by counting follicles in histological sections 18 days after grafting under the kidney capsules of severe combined immunodeficiency (SCID) mice, and the results were expressed as percentages of the numbers in comparable pieces of ungrafted tissue. While only 10% of the total number of follicles was found in the glycerol group compared to controls, significantly higher percentages (44-84%) survived cryopreservation in the other media. The tissues were sterile when frozen and thawed without a cryoprotectant. These results suggest that if comparable results could be achieved by autografting, a patient's fertility should be safeguarded from cytotoxic treatment.

Cryopreservation of caprine ovarian tissue using glycerol and ethylene glycol

“…Based on the literature [12] and [16], EG was more effective at protecting cells during cryopreservation than GLY, perhaps due to its low toxicity and efficient permeability [51]. Although some authors [12], [16] and [18] have shown that GLY is inferior to the other cryoprotectants, in our study the percentage of normal preantral follicles after cryopreservation in GLY was similar to EG. It is noteworthy that we used a different slow freezing curve and a removal procedure for GLY similar to the employed by Candy et al [12].…”

“…However, results obtained by Candy et al [12] using GLY (4.3-28.3 and 16.3-32%: survival of primordial and primary follicles, respectively) were comparable to those for caprine preantral follicles in our experimental conditions. Another report in human ovarian tissue [18] using a freezing protocol similar to Candy et al [12], i.e. slow freezing curve and fast removal of GLY without sucrose, obtained only 10% viable preantral follicles after freezing and thawing.…”

“…Previous work with mature caprine oocytes indicated that the permeability of GLY was highly dependent on temperature, and at 20 °C [4] and 4 °C [16], this cryoprotectant has lower permeability than EG. Newton et al [18] compared the efficiency of four cryoprotectants (DMSO, propanediol, EG and GLY) in the cryopreservation of human ovarian tissue; they showed that EG had better results because it penetrated more quickly in the tissue, due probably to its lower molecular weight [50]. Based on the literature [12] and [16], EG was more effective at protecting cells during cryopreservation than GLY, perhaps due to its low toxicity and efficient permeability [51].…”

“…The vials were plunged directly into liquid nitrogen (−196 °C) and stored for up to 5 days before thawing. The freezing curve used in this study was chosen because most procedures currently used to cryopreserve oocytes [25], [26], [27], [28], [29] and [30] and ovarian tissue [10], [11], [12], [14], [15], [18], [24], [31], [32] and [33] stipulate a cooling rate of 0.3-0.5 °C/min from the seeding temperature (usually −5 to −9 °C) to a lower temperature, usually between −30 and −40 °C.…”

“…Some authors [12] and [18] affirm that GLY is less permeable than other cryoprotectan such as EG, and consequently, not suitable for the cryopreservation of ovarian tissue. However, none of the cryoprotectants noted have been tested on caprine ovarian tissue.…”