Fertility preservation in cancer patients: in vivo and in vitro options

Oocyte, embryo and ovarian tissue cryopreservation are being increasingly proposed for fertility preservation among cancer patients undergoing therapy to enable them to have babies after the cancer is cured. Embryo cryopreservation is not appropriate for single girls without any spermpartner. It is impossible in cases requiring immediate cancer cure because oocyte retrieval is an extended procedure. Thus ovarian tissue cryopreservation has been suggested for fertility preservation especially in cancer patients. The main goal of ovarian cryopreservation is re-implanting the tissue into the body to restore fertility and the hormonal cycle. Different cryopreservation protocols have been examined and established for vitrification of biological samples. We have used Cryopin to plunge ovarian tissue into the liquid nitrogen and promising results have been observed. The possibility of recurrence of malignancy in the reimplanted tissue could be a problem. Xenografting-implantation of the preserved tissue in another species-also has its drawbacks such as molecular signaling from the recipient. In vitro follicle culturing is a safer method to obtain mature oocytes for fertilization and the various studies that have been carried out in this area are reviewed in this paper

Dietary vitamin E is more effective than omega-3 and omega-6 fatty acid for improving the kinematic characteristics of rat sperm

Objective: Although key roles for dietary vitamin E [VITE] and fatty acid [FA] in fertility have been confirmed, limited data are available on the effects of VITE alone, or a constant level of VITE supplemented by dietary omega-6 and omega-3 FAs in combination on male reproduction. Consequently in this paper, the effects of VITE, sunflower oil, fish oil and their combination on rat sperm were investigated

Materials and Methods: We divided 50 mature male Wistar rats into 5 groups [n=10] in a experimental completely randomized design for eight weeks: i. Control [CTR]: standard diet; ii. Vitamin E diet [VITE]: 2 times greater than recommendations; iii. Sunflower oil group [n-6] [gavaged with 0.5 ml/day/rat sunflower oil+VITE diet]; iv. Fish oil group [n-3]: [gavaged with 0.5 ml/day/rat fish oil+VITE diet] and v. n-3+n-6 group [gavaged with 0.3 ml fish oil/day/rat+0.2 ml sunflower oil/day/rat+VITE diet]. The sperm parameters were measured by computer assisted semen analyzer [CASA]. All data were analyzed with SPSS software

Results: Feed intake decreased in groups which were administered sunflower oil compared with the other groups [P<0.05]. The groups which received only VITE or fish oil+VITE had a significantly higher concentration of sperm compared with the n-6+n-3 and CTR group [P<0.05]. VITE and n-3 showed significant improved progressive motility compared to the CTR group, whereas the n-6 and n-6+n-3 groups were in the middle [P<0.05]. The highest sperm kinematic parameters were observed in the VITE only group. There was no strong correlation between sperm parameters and blood lipid profiles

Conclusion: Dietary VITE and fish oil+VITE can improve sperm quality. Our findings can be a focus for improvements in sperm quantity and motility in fertile animals using only dietary VITE

introduction to the royan human ovarian tissue bank

From December 2000 until 2010, the researchers at Royan Institute conducted a wide range of investigations on ovarian tissue cryopreservation with the intent to provide fertility preservation to cancer patients that were considered to be candidates for these services. In 2010, Royan Institute established the Royan Human Ovarian Tissue Bank as a subgroup of the Embryology Department. Since its inception, approximately 180 patients between the ages of 747 years have undergone consultations. Ovarian samples were cryopreserved from 47 patients [age: 7-35 years] diagnosed with cervical adenocarcinoma [n=9]; breast carcinoma [n=7], Ewing's sarcoma [n=7], opposite side ovarian tumor [n=7], endometrial adenocarcinoma [n=4], malignant colon tumors [n=3], as well as Hodgkin's lymphoma, major thalassemia and acute lymphoblastic leukemia [n=1-2 patients for each disease]. Additionally, two patients requested ovarian tissue transplantation after completion of their treatments

[Comparison of two different immature mouse ovarian vitrification methods using trypan blue staining]

Objective: Different cryoprotectants are used for cryopreservation of ovarian tissue in patients at risk of infertility. Ethylene glycol [EG], dimethyl sulfoxide [DMSO] and propanediol [PROH] have been chosen as the basic permeable cryoprotectants due to their decreased glass-formation characteristics compared to other cryoprotectants. In the present study, the effects of two different vitrification methods on whole mouse ovarian tissue by the use of a novel staining method [trypan blue] has been evaluated

Methods: Ovaries of 8 day-old NMRI mice were isolated and divided among the control, vitrification 1 [Vit1] and vitrification 2 [Vit2] groups. The Vit1 solution was composed of alpha-MEM+ 20% FBS + 15% EG + 15% DMSO. The Vit 2 solution was composed of alpha- MEM+ 15% FBS +20% EG + 20% PROH. Vit1 and Vit2 procedures were performed at 4°C and room temperature, respectively. Warming was performed in alpha-MEM+ 20% FBS supplemented with 1M sucrose in the Vit1 group and alpha-MEM+ 15% FBS with descending concentrations of sucrose [1, 0.5, 0.25 M] in the Vit2 group. Control and vitrified warmed ovaries were put in alpha-MEM supplemented by 0.4% trypan blue for 20 min, and then stained ovaries were fixed in Bouin's fixative, serially sectioned in paraffin wax and finally quantitatively evaluated under a light microscope

Results: The highest percentage of primordial follicles was observed in the control group. There was a significant difference between the control and Vit1 groups, and between the Vit1 and Vit2 groups [p<0.05]. No significant difference was observed in primary and preantral follicles between the control and vitrification groups

Conclusion: Vitrification with EG and PROH are more suitable for preservation of follicle reserves in ovaries. Trypan blue staining is a faster and easier method for evaluation of ovarian tissue

Maternal-effect gene expression in cultured preantral follicles derived from vitrified-warmed mouse ovary

This study was conducted to assess survival of follicles, their oocyte maturation and fertilization potential as well as expression of early embryo developmental genes in in vitro cultured pre-antral follicles derived from vitrified-warmed mouse ovary. In this experimental study, ovaries of 12-day old Naval Medical Research Institute [NMRI] female mice were placed into non-vitrified and vitrified-warmed groups. Isolated preantral follicles from experimental groups were cultured in vitro for 12 days. On the 12[th] day of culture, oocyte maturation was induced and then matured oocytes were in vitro fertilized. The rates of oocyte maturation and two-cell stage embryo formation were assessed. Relative expression of Mater and Zar1 was evaluated on days 1, 6, 10 and 12 of culture. Data analysis was performed by t test and two-way ANOVA [P<0.05]. Our data showed no significant difference between the control and vitrification groups in the rate of follicular survival, oocyte maturation and two-cell stage embryo formation. The level of gene expression was higher on the 6[th] and 10[th] days of culture for Mater and Zar1 in vitrified-warmed group compared with non-vitrified group, however, there was no significant difference between the two groups. It seems that the applied vitrification method did not reveal any negative effect on maturation and developmental competence of oocytes surrounded in preantral follicles and therefore could preserve follicular reserves efficiently

[ review on the activity of hormones and growth factors after ovarian tissue transplantation]

The majority of cancer treatments are invasive. Gonadal injuries cause reductions in fertility which results in lack of hope for conception in cancer patients and frustration for their partners. Fortunately, current advancements in cryopreservation and transplantation sciences regarding fertility preservation lead to cryostorage of gonads and preservation prior to the onset of chemo- and radiotherapy treatments. Accordingly in women, the main goal of ovarian cryopreservation is establishment of fertility and hormonal cycle restoration after auto-transplantation. Although the history of ovarian transplantation dates back to the 19[th] century, there are reports of live human births following ovarian tissue cryopreservation and transplantation since the past 100 years. Despite this success and additional research in the field of ovarian cryopreservation and transplantation, numerous questions remain unanswered. Among these questions, growth factors and hormonal changes because of their effects on follicular function appear to be more important during ovarian tissue transplantation. This review attempts to address hormones and growth factor functions with the specifics of ovarian cryopreservation and auto-transplantation

Effect of different thawing rates on post-thaw viability, kinematic parameters and chromatin structure of buffalo [Bubalus bubalis] spermatozoa

The aim of the present study was to evaluate three thawing rates on the post thaw motility, viability and chromatin structure of buffalo semen frozen in 0.5-ml straws. In this experimental study semen was collected with artificial vagina [42[degree sign]C] from four buffalo bulls. Split pooled ejaculates [n=4] were extended at 37[degree sign]C with a Bioxcell[registered sign] extender. Semen was cooled to 4[degree sign]C within 2 hours, equilibrated at 4[degree sign]C for 4 hours, then filled in 0.5 ml French straws, and frozen in programmable cell freezer before plunging into liquid nitrogen. Straws were thawed at water bath temperatures of 37, 50 or 70[degree sign]C for 30, 15 and 6 seconds, respectively. Semen was incubated at 37[degree sign]C for 2 hours and evaluated for post thaw motility, viability, acrosomal and DNA integrity of spermatozoa. Analysis of variance [ANOVA] was used for comparisons of means. When the ANOVA test showed statistical differences, the mean of the treatments were compared using Duncan's multiple range tests. The initial postthaw motility [0 hour] averaged 62.7 +/- 7.2%, 73.1 +/- 9.77%, and 74.9 +/- 8.58% for the three thaw rates, respectively. Kinematic parameters such as average path velocity, linearity and beat/cross frequency in the thaw rate of 70?C for 6 seconds were superior to other rates studied [p<0.05]. After 2 hours of incubation, proportions of progressive motility and Kinematic parameters decreased in all groups [p>0.05]. A positive correlation was detected between sperm motility and thawing rate after two hours incubation times. The percentage of viable spermatozoa and spermatozoa with an intact acrosome and plasma membrane integrity were not different between the groups of samples thawed at different temperatures [p>0.05]. The percentage of spermatozoa with chromatin dispersion for the thaw rate of 70[degree sign]C for 6 seconds was significantly higher than for the to other rates studied [p< 0.05]. In contrast with motility and viability, the DNA integrity of post thaw spermatozoa remained unaffected during 2 hours incubation. The post thaw motility and kinematic parameters of buffalo spermatozoa were significantly improved immediately after thawing by increasing the thawing rate from 37[degree sign]C in 30 seconds to70[degree sign]C in 6 seconds. However, this relative advantage had disappeared after incubation in a water bath at 37[degree sign]C for two hours. A thaw rate of 70[degree sign]C for 6 seconds was associated with higher chromatin dispersion than the other thaw rates studied. Sperm thawing over at 50 degrees could be safely used to improve motility recovery after sperm cryopreservation in buffalo bulls

[Temperature reduction process - a way for fertility preservation in cancer patients]

More than the half of cancer patients undergo cancer treatments of chemotherapy and/or radiotherapy. Unfortunately treatment with invasive methods occasionally lead to severe side effects. Patients who undergo chemotherapy can be affected by premature ovarian failure, an important cause of infertility. Ovarian tissue cryopreservation is suggested as the only way for preservation of sex cells and fertility preservation in cases of prepubertal girls and women with sterility attributed to chemotherapy, radiotherapy, genetic disorders or specific diseases