Restoration of Reproductive Potential after Autotransplantation of Frozen Ovaries in Mice Pretreated with Cyclophosphamide

Dong Ho Lee; Jong Ok Kim; Young Kyoung Yoo; Kwan Ju Lee; Jung Su Kim; Chang Joon Ahn

doi:10.4048/jbc.2008.11.1.10

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Lee, Kim, Yoo, Lee, Kim, and Ahn: Restoration of Reproductive Potential after Autotransplantation of Frozen Ovaries in Mice Pretreated with Cyclophosphamide

Original Article

J Breast Cancer 2008;11(1):10-17.

Published online: 31 March 2008

DOI: https://doi.org/10.4048/jbc.2008.11.1.10

Restoration of Reproductive Potential after Autotransplantation of Frozen Ovaries in Mice Pretreated with Cyclophosphamide

Dong Ho Lee, Jong Ok Kim¹, Young Kyoung Yoo, Kwan Ju Lee, Jung Su Kim, Chang Joon Ahn

Department of Surgery, College of Medicine, The Catholic University of Korea, Seoul, Korea

¹Department of Surgical Pathology, College of Medicine, The Catholic University of Korea, Seoul, Korea

E-mail : cjahn1220@hanmir.com

^* This article was supported by the Clinical Research Institute of Daejeon St. Mary Hospital, Catholic University.

^* This article was released at 10th Korean conference on clinical oncology in March 2007.

Received 25 August 2007 Accepted 21 November 2007

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Purpose

Infertility due to ovarian failure that is caused by antineoplastic chemotherapeutic agents is one of the primary problems of female cancer patients who are in their reproductive years. It has become important to preserve the reproductive potential of female cancer patients. This study was conducted to determine whether autotransplantation of frozen ovaries can restore reproductive potential.

Methods

This study included 30 female mice that had normal reproductive potential. The mice were divided into 4 groups: the positive control, the negative control, the comparison group, and the experimental group. The positive control group received right total oophorectomy, and the negative control group received bilateral total oophorectomy. Greater than or equal to 90% of the left ovary was removed in the mice of the comparison group, and then cyclophosphamide was administered. In the experimental group, the right ovary taken out by right total oophorectomy, and this was crypreserved using the vitrification method. And then cyclophosphamide was administered. The cryopreserved ovary was autotransplanted to the left gonadal fat pad after greater than or equal to 90% of the left ovary was removed. The reproductive performance in each group was analyzed according to the pregnancy rate after mating.

Results

In the positive control group, all five mice became pregnant, and the number of fetuses was 4 to 5 (mean= 4.60±0.55). In the comparison group, the pregnancy rate was 50%, and the mean number of fetuses was 1.40±0.55. In the experimental group, 7 of 10 (70%) mice became pregnant, and the mean number of fetuses was 4.71±2.56. There was no significant difference in the number of fetuses between the positive control and the experimental group (p=0.093), but there was a significant difference in the number of fetuses between the comparison group and the experimental group (p=0.019).

Conclusion

The results of this study suggest that autotransplantation of frozen ovaries using the vitrification method may restore the impaired ovarian function induced by antineoplastic chemotherapeutic agents.

Keywords: Antineoplastic chemotherapeutic agent, Ovarian failure, Cryopreserved ovary, Vitrification method, Autotransplantation

REFERENCES

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Fig 1.

Gross findings in the auto-transplanted ovary in the experimental group. (A) There are several fetuses in the left uterus after successful autotransplantation of cryopreserved ovary, (B) Even after successful autotransplantation of the cryopreserved ovary, no fetus is in the left uterus, (C) After autotransplantation of the cryopreserved ovary, there is no feus in the left uterus due to atrophic change of the autotransplanted ovary.

Fig 2.

A primary follicle in the the positive control group (A) and the experimental group (B) after transplantation (H&E stain, original magnification ×200). A primary follicle of the experimental group is alive and there is no significant difference in the structural components between the two groups.

Fig 3.

H&E-stained ovaries. (A) An ovary in the positive control group has about 13 living oocytes, (B) an autografted ovary (right ovary) in the experimental group has about 5 living oocytes, (C) an atrophied ovary treated with cychlophosphamide (left ovary) in the experimental group has no oocytes (original magnification ×40).

Table 1.

Time-frame of the experiment

	D1	D3	D4	D5	D7	D15	D21	D28	D37
Positive	RT.OX				Sham			Mating
Negative	BO.OX						Mating
Comparison	NT.OX	CY 250	CY 250	CY 250				Mating
Experimental	RT.OX, CP	CY 250	CY 250	CY 250		LT.OX, AT			Mating

D=postoperative day; RT.OX=Right total ovariectomy; BO.OX=Both total ovariectomy; NT.OX=Right total and left near-total ovariectomy; CP=Cryopreservation of resected right ovary; CY250=Intraperitoneal injection of cychlophosphamide 250 mg/kg; Sham=sham operation; LT.OX=Left neartotal ovariectomy; AT=Autotransplantation of cryopreserved ovary.

Table 2.

The number of the offsprings of successfully pregnant mice

Number	Positive		Negative		Comparison		Experimental
Number	P	Pups	P	Pups	P	Pups	P	Pups	Atrophy
1	○	4	Ｘ	0	○	2	○	9	Ｘ
2	○	5	Ｘ	0	○	1	○	3	Ｘ
3	○	4	Ｘ	0	○	1	○	6	Ｘ
4	○	5	Ｘ	0	○	1	○	4	Ｘ
5	○	5	Ｘ	0	○	2	○	1	○
6					Ｘ	0	○	6	Ｘ
7					Ｘ	0	○	4	Ｘ
8					Ｘ	0	Ｘ	0	Ｘ
9					Ｘ	0	Ｘ	0	○
10					Ｘ	0	Ｘ	0	○
P rate (%)	5/5 (100%)		0/5 (0%)		5/10 (50%)		7/10 (70%)

P=Pregnancy (○) or non-pregnancy (Ｘ); Pups=the number of the offsprings of successfully pregnant mice; Atrophy=atrophy auto-grafted ovary (○) or not (Ｘ).

Table 3.

Number of pups of successfully pregnant mice

	Positive control (Unilateral ovariectomized group)	Comparison group (Partial ovariectomized, Cy [750 mg/kg] IP)	Experimental group (Cy [750 mg/kg] IP, cryopreserved ovary transplant)
Number of pups of successfully pregnant mice (Mean±SD)	4.60±0.55	1.40±0.55 (p<0.001)	4.71±2.56 (p=0.093)

There was a significant difference between the comparison and the experimental group (p=0.019).

Table 4.

The difference in the number of pups of successfully pregnant mice between the comparison group and the experimental group comparison group

	Comparison group (Partial ovariectomized, Cy [750 mg/kg] IP)	Experimental group (Cy [750 mg/kg] IP, cryopreserved ovary transplant)
Number of pups of successfully pregnant mice (Mean±SD)	1.40±0.55	4.71±2.56 (p=0.019)

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