Orthotopic and heterotopic ovarian tissue transplantation




Although still experimental, cryopreservation and transplantation techniques for ovarian tissue have been well described, and a number of successful human pregnancies have occurred. Ovarian cryopreservation is the only fertility preservation procedure that can be offered to prepubertal children, and when cytotoxic treatment is urgent. There are two main approaches for autotransplantation of human ovarian tissue. In the heterotopic transplantation, cortical fragments can be grafted subcutaneously at various sites whereas in orthotopic transplantation cortical pieces are transplanted into its original location. Both approaches have their own advantages and disadvantages. While natural pregnancy can occur in orthotopic transplantation, heterotopic transplantation may be indicated if the pelvis is not suitable for transplantation due to previous radiation or severe scar formation. Furthermore, tissue monitoring may be easier in the heterotopic site. In this article, we reviewed the indications, limitations, risks and transplantation techniques for ovarian tissue.


Cancer is a major health problem in both developed and developing countries. Recent data estimate that 692,000 women and 745,000 men have been afflicted with cancer in 2007 in the United States. In women, cancer incidence rates for all sites increased by 0.3% per year from 1987 to 2003. However, there have been remarkable improvements in survival rates for various cancers, as a result of the developments in current treatment modalities, along with the ability to detect tumours in the early stages by well-established screening programmes. Furthermore, a cure is now possible for many childhood and adult cancers, especially for leukaemias and lymphomas. By the year 2010, one in every 250 person is estimated to have survived childhood malignancies. In addition to malignant diseases, increasing number of non-malignant systemic conditions has also been successfully treated with chemotherapy or haematopoietic stem cell transplantation.


As increasing number of women survive cancer each year, a growing segment of patients inevitably face the long-term sequel of cancer treatment that dramatically compromise their quality of life. Increased awareness of the impact of various cytotoxic treatments on gonadal function has now resulted in a surge in the number of patients seeking help to preserve their fertility. Cryopreservation of embryos is a standard technique for fertility preservation when there is adequate time for ovarian stimulation. If the patient has no partner or unwilling to use donor sperm, oocytes can be frozen instead. Although cryopreservation and transplantation techniques have been well described for ovarian tissue, currently the experience with this procedure is limited. Nevertheless, at present, it is the only fertility preservation procedure that can be offered to prepubertal children and can be implemented without any delay in treatment. In this article, the reader will find a comprehensive review of ovarian cryopreservation and transplantation techniques.


Ovarian tissue cryopreservation: indications


Studies with fresh ovarian transplants date back to the beginning of the 20th century. Cryopreservation of ovarian tissue and transplantation began in animal models in 1950s after the discovery of cryoprotective agents. Early studies showed limited success with only 10% of the primordial follicles surviving the freeze–thaw procedure, because of the availability of a poor cryoprotectant agent ‘glycerol’, and the absence of automated cryopreservation systems. Additional cryoprotectants became available during 1990s, including propandiol, dimethylsulphoxide and ethylene glycol, leading to successful ovarian tissue cryopreservation (OTC), transplantation and resumption of fertility in animals. In the second half of 1980s, heterotopic transpositon of human ovary with microsurgical anastomosis has been reported. In the first report of orthotopic ovarian transplantation with frozen banked ovarian tissue, menopause was temporarily reversed in a 29-year-old woman suffering from surgical menopause. After declaration of the first embryo from frozen–thawed ovarian fragments transplanted in a heterotopic site in 2004 , subsequent live births were reported using this technique.


There is a growing list of indications for OTC for fertility preservation ( Table 1 ). The indications now include not only neoplastic diseases but also non-neoplastic conditions requiring chemotherapy, radiotherapy or haematopoietic stem cell transplantation. Oophorectomy for benign ovarian tumours and for BRCA germline mutations can also be indications for OTC.



Table 1

Indications for ovarian tissue cryopreservation.





Cancer in children
Hodgkin and non-Hodgkin lymphoma
Leukemias
Ewing sarcoma
Wilms tumor
Neuroblastoma
Genital rhabdomyosarcoma
Pelvic osteosarcoma
Breast cancer
Infiltrative ductal histological subtype
Stage I–III
Cancer of the cervix
Autoimmune and hematological diseases treated with chemotherapy or HSCT a
Surgery for benign ovarian disease
Endometriosis
Benign ovarian lesions
Patients receiving pelvic radiation
Solid organ tumors presenting in the pelvis
Ewing sarcoma
Osteosarcoma
Tumors of the spinal cord
Retroperitoneal sarcoma
Rectal cancer
Benign bone tumors
Prophylactic oophorectomy
BRCA-I/II positive patients
Patients undergoing surgery for gynecological cancers

a Includes genetic, hematological, and autoimmune disorders.



Although the clinical indications are similar for ovarian tissue and oocyte cryopreservation, there are fewer logistical restrictions with the latter. OTC has broader applications and, in theory, a greater source of oocytes compared with oocyte cryopreservation as ovarian cortex may contain tens of thousands of oocytes. Several livebirths have been reported with oocyte freezing; however, the number of successful pregnancy reports has been growing from frozen–thawed ovarian tissue transplantation ( Table 2 ).



Table 2

Embryo development and pregnancies reported after transplantation of frozen thawed ovarian tissue.
















































































































Author Year Transplantation site Cryo indication IVF/spontaneous Age at ovarian cryo. Age at tx. Outcome
Oktay 2004 Heterotopic Breast cancer IVF 30 36 Embryo development
Donnez 2004 Orthotopic Hodgkin’s disease Spontaneous 25 31 Healthy live birth
Meirow 2004 Orthotopic Hodgkin’s disease IVF 26 28 Healthy live birth
Demeestere 2006 Orthotopic/heterotopic Hodgkin’s disease Spontaneous 24 29 One miscarriage at 7 weeks, one healthy live birth
Oktay 2006 Heterotopic Hodgkin’s disease Spontaneous 28 32 Healthy live birth
Rosendahl 2006 Orthotopic/heterotopic Hodgkin’s disease IVF from heterotopic site 28 30 Biochemical pregnancy
Anderson 2008 Orthotophic Non Hodgkin’s lympohoma IVF 32 34 Ebryo development
Anderson 2008 Orthotopic/heterotopic Hodgkin’s disease IVF 25 27 Clinical pregnancy
Anderson 2008 Orthotopic Hodgkin’s diseas IVF 26 28 Healthy live birth
Anderson 2008 Orthotopic Ewings sarkomu IVF 27 30 Healthy live birth
Silber a 2008 Orthotopic Idiopathic premature ovarian failure Spontaneous 14 28 Ongoing pregnancy

Tx: transplantation cryo: cryopreservation.

a Fresh and frozen thawed ovarian cortical strips are used.



Childhood cancers


Cancer ranks as the second leading cause of death in children between the ages of 1 and 14 years. Cure rates and life expectancy have dramatically improved for many childhood cancers. Despite increased survival rates, they are certainly not immune to the gonadotoxic effects of various cancer treatments.


Children have the highest number of primordial follicle reserve and the greatest benefit from OTC is expected in this group. As an advantage, no ovarian stimulation is required in OTC and therefore time restrictions are fewer, and no concerns exist as to the risk of stimulating hormone-sensitive cancers following ovarian stimulation. Since it avoids ethical concerns of ovarian stimulation and oocyte retrieval in children, cryopreservation of the ovarian tissue is the only acceptable option for prepubertal children undergoing any type of gonadatoxic therapy. In addition, this is the only fertility preservation technique that can reverse hormonal functions.


Cancers in adults


Breast cancer is the most common female cancer seen during reproductive ages. In 2007, as many as 182,460 new cases were estimated to be diagnosed with breast cancer, accounting for more than a quarter of all female cancers. Approximately, 25% of breast cancer cases occur before menopause, and 15% occur under the age of 45. More than 90% of all breast cancers are diagnosed at a local or regional disease stage, with corresponding 5-year survival rates of 97% and 79%, respectively. With improved cure rates from breast cancer, a greater attention has been focussed on the long-term adverse effects of breast cancer treatment, which can compromise the quality of life of breast cancer survivor. Along with the increase in the number of women who delay first childbirth beyond the age of 35, more liberal use of adjuvant chemotherapy has resulted in a large proportion of young women with breast cancer facing infertility and premature ovarian failure. The incidence of chemotherapy-induced amenorrhoea has been reported as 68% with classic oral CMF, while it ranges between 0% and 96% with antracyline-based regimens. When considering OTC, it should be underlined that the risk of ovarian involvement seems low in early-stage and locally advanced breast cancer.


Each year, nearly 500,000 women are afflicted by cervical cancer worldwide. Almost half of them are under the age of 35. Patients with advanced-stage disease and those with early-stage disease with high-risk factors receiving pelvic or pelvic/para-aortic radiation therapy are at high risk for developing ovarian failure. In the case of ovarian stimulation in patients with cervical cancer, there is risk of bleeding from the cancerous cervix during oocyte retrieval. Ovarian tissue can be harvested in selected patients for cryopreservation during primary cancer surgery; however, the risk of ovarian involvement has to be considered. Squamous cell cancer of the cervix, which is the most encountered subtype, rarely metastasises to the ovaries, whereas adenocarcarcinoma of the cervix involves ovaries at a rate as high as 12%. Patients receiving only pelvic radiotherapy can benefit from ovarian transposition; however, success rates vary greatly because of the possibility of vascular damage during the procedure. As such, patients receiving pelvic radiation therapy for any pelvic tumour such as rectal cancer, solid organ tumours presenting in the pelvis, osteosarcoma and tumours of the spinal cord can also resort to fertility preservation technologies including OTC.


Autoimmune diseases


A number of autoimmune diseases can affect women of reproductive age. Cytotoxic treatment has been used effectively to treat various autoimmune diseases, including systemic lupus erythematosus, steroid-resistant glomerulonephritis, Behcet’s disease, inflammatory bowel diseases and pemphigus vulgaris. Cortical fragments of ovary can be harvested to cryopreserve for possible future use in order to preserve fertility.


Patients undergoing haematopietic stem cell transplantation


Autologous or allogeneic haematopietic stem cell transplantation (HSCT) has become an important therapeutic tool in the management of some malignant and non-malignant systemic diseases. Diseases associated with genetically abnormal stem cells, some autoimmune diseases unresponsive to immunosuppressive therapy and diseases associated with the deficiency of bone marrow stem cell products are among the non-malignant diseases treated with HSCT. HSCT has also been used to treat breast cancer, multiple myeloma and lymphoma. If there are time restrictions for ovulation induction, or if the patient is single or a child, ovarian tissue can be frozen in these patients to preserve fertility.


Others


The cumulative lifetime risk of developing ovarian cancer is ∼ 60% in the presence of BRCA-1 mutation, and 10–20% in women with BRCA-2 mutation. Furthermore, lifetime risk of breast cancer in female carriers of BRCA-1 mutation is 80–90%. Despite the fact that the risk of peritoneal cancer cannot be totally eliminated in BRCA-positive patients, prophylactic oophorectomy is suggested as soon as childbearing is completed or by the age 35–40 years to decrease the risk of both breast and ovarian cancer. Cortical pieces of ovarian tissue can be harvested to freeze for future use to preserve fertility.


There is a risk that ovarian reserve can be compromised by any type of ovarian surgery for benign ovarian diseases, mostly including endometriosis and benign neoplastic ovarian lesions. Despite that follicular growth appears lower with frozen/thawed tissue, functional ovarian cortex surrounding benign ovarian cysts transplanted into the subcutaneous space of severe combined immunodeficient disease (SCID) mice can sustain ovarian tissue function. Healthy pieces of cortical ovarian tissue can be isolated and frozen in these patients for possible future use. A combination of OTC and in vitro maturation can be implemented to maximise the success rate of the procedure.




Xenografting studies of human ovarian tissue


Cortical fragments of human ovarian tissue can be xenotransplanted into T- and B-cell-deficient SCID mice. This animal model was first used by Gosden to observe follicle development in xenografted sheep and cat ovarian tissue. Subsequently, follicle development, ovulation, corpus luteum formation and metaphase II oocytes were demonstrated by several groups after gonadotropin stimulation of xenografted human ovarian tissue in SCID mice.


In a recent study, it was demonstrated that cryopreservation and xenotransplantation did not appear to greatly affect the human primordial/primary follicle ultrastructure. Interestingly, in frozen–thawed xenografts, secondary human ovarian follicles presented a well-preserved ultrastructure, but asynchrony between oocytes and granulosa cell development was detected. On the contrary, a previous study demonstrated that immature oocytes in human ovarian tissue xenotransplanted into SCID mice grew to maturity, whereas many oocytes, grown in host animals and further matured in vitro , showed aberrant microtubule organisation and chromatin patterns.


In a prospective study, the effect of a gonadotropin-releasing hormone (GnRH) agonist on the number of follicles in different developmental stages in cryopreserved human ovarian grafts transplanted into gonadotropin-stimulated or unstimulated SCID mice were investigated. GnRH agonist treatment did not prevent primordial follicle depletion after the xenografting of ovarian tissue in SCID mice with or without gonadotropin stimulation. Furthermore, GnRHa caused an additional loss of follicles if administered during the critical neo-vascularisation period after the transplantation. The same group also found that prolonged gonadotropin stimulation significantly reduces primordial follicles in xenografts of cryopreserved human ovarian tissue. Even though ovariectomy may improve the development of follicles after xenotransplantation of cryopreserved human ovarian grafts, exogenous use of gonadotropins seems essential for improved follicle survival in recipient SCID mice.


In summary, even though human ovarian xenografts provided a model to study human ovarian tissue autotransplantation, their use as a means to use banked ovarian tissue is in question. Concerns regarding cross-species retroviral infections should be addressed. Moreover, this technique will require large numbers of animals to be killed since only very small fragments of ovarian tissue can be xenografted. This may not only make the technique impractical, but may also raise further ethical concerns.




Xenografting studies of human ovarian tissue


Cortical fragments of human ovarian tissue can be xenotransplanted into T- and B-cell-deficient SCID mice. This animal model was first used by Gosden to observe follicle development in xenografted sheep and cat ovarian tissue. Subsequently, follicle development, ovulation, corpus luteum formation and metaphase II oocytes were demonstrated by several groups after gonadotropin stimulation of xenografted human ovarian tissue in SCID mice.


In a recent study, it was demonstrated that cryopreservation and xenotransplantation did not appear to greatly affect the human primordial/primary follicle ultrastructure. Interestingly, in frozen–thawed xenografts, secondary human ovarian follicles presented a well-preserved ultrastructure, but asynchrony between oocytes and granulosa cell development was detected. On the contrary, a previous study demonstrated that immature oocytes in human ovarian tissue xenotransplanted into SCID mice grew to maturity, whereas many oocytes, grown in host animals and further matured in vitro , showed aberrant microtubule organisation and chromatin patterns.


In a prospective study, the effect of a gonadotropin-releasing hormone (GnRH) agonist on the number of follicles in different developmental stages in cryopreserved human ovarian grafts transplanted into gonadotropin-stimulated or unstimulated SCID mice were investigated. GnRH agonist treatment did not prevent primordial follicle depletion after the xenografting of ovarian tissue in SCID mice with or without gonadotropin stimulation. Furthermore, GnRHa caused an additional loss of follicles if administered during the critical neo-vascularisation period after the transplantation. The same group also found that prolonged gonadotropin stimulation significantly reduces primordial follicles in xenografts of cryopreserved human ovarian tissue. Even though ovariectomy may improve the development of follicles after xenotransplantation of cryopreserved human ovarian grafts, exogenous use of gonadotropins seems essential for improved follicle survival in recipient SCID mice.


In summary, even though human ovarian xenografts provided a model to study human ovarian tissue autotransplantation, their use as a means to use banked ovarian tissue is in question. Concerns regarding cross-species retroviral infections should be addressed. Moreover, this technique will require large numbers of animals to be killed since only very small fragments of ovarian tissue can be xenografted. This may not only make the technique impractical, but may also raise further ethical concerns.

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Nov 9, 2017 | Posted by in OBSTETRICS | Comments Off on Orthotopic and heterotopic ovarian tissue transplantation
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