Dosimetric evaluation of ovaries and pelvic bones associated with clinical outcomes in patients receiving total body irradiation with ovarian shielding.

Total body irradiation (TBI) with ovarian shielding is expected to preserve fertility among hematopoietic stem cell transplant (HSCT) patients with myeloablative TBI-based regimens. However, the radiation dose to the ovaries that preserves ovarian function in TBI remains poorly understood. Furthermore, it is uncertain whether the dose to the shielded organs is associated with relapse risk. Here, we retrospectively evaluated the relationship between fertility and the dose to the ovaries, and between relapse risk and the dose to the pelvic bones. A total of 20 patients (median age, 23 years) with standard-risk hematologic diseases were included. Median follow-up duration was 31.9 months. The TBI prescribed dose was 12 Gy in six fractions for three days. Patients' ovaries were shielded with cylinder-type lead blocks. The dose-volume parameters (D98% and Dmean) in the ovaries and the pelvic bones were extracted from the dose-volume histogram (DVH). The mean ovary Dmean for all patients was 2.4 Gy, and 18 patients recovered menstruation (90%). The mean ovary Dmean for patients with menstrual recovery and without recovery were 2.4 Gy and 2.4 Gy, respectively, with no significant difference (P = 0.998). Hematological relapse was observed in five patients. The mean pelvis Dmean and pelvis D98% for relapse and non-relapse patients were 11.6 Gy and 11.7 Gy and 5.6 Gy and 5.3 Gy, respectively. Both parameters showed no significant difference (P = 0.827, 0.807). In conclusion, TBI with ovarian shielding reduced the radiation dose to the ovaries to 2.4 Gy, and preserved fertility without increasing the risk of relapse.

Ultrasound-guided follicle aspiration at time of laparotomy in a patient with Mayer-Rokitansky-Küster-Hauser syndrome.

OBJECTIVE: To demonstrate a unique case of direct ultrasound-guided follicle aspiration at time of a laparotomy in a patient with Mayer-Rokitansky-Küster-Hauser Syndrome. DESIGN: Video presentation. SETTING: Academic fertility center PATIENT(S): A 32-year-old gravida 0 with Mayer-Rokitansky-Küster-Hauser Syndrome and bilateral ovarian masses presented as a fertility preservation consult from gynecologic oncology due to the possibility of bilateral oophorectomy. Due to the appearance and size of the left ovary oncology planned to perform an exploratory laparotomy and left oophorectomy, with possible right oophorectomy. The patient and her partner desired embryo cryopreservation with plans for future use in a gestational carrier. She had previously undergone vaginal dilator therapy, however her ovaries were inaccessible transvaginally due to their cephalad location and small caliber of the vaginal pouch. The plan was made to proceed with controlled ovarian stimulation and concurrent ultrasound-guided follicle aspiration of the right ovary at the time of laparotomy following left oophorectomy. INTERVENTION(S): Ultrasound-guided follicle aspiration in vivo at time of laparotomy. MAIN OUTCOME MEASURE(S): Successful controlled ovarian stimulation, oocyte retrieval and embryo cryopreservation. RESULT(S): The patient underwent a long agonist protocol and received a total of 2,525 units of gonadotropin with a peak estradiol of 3,264 pg/ml. She required a total of 9 days of stimulation. The normal right ovary responded as expected, and the left ovary remained unchanged. Following laparotomy and left oophorectomy, direct application of the transvaginal ultrasound probe was used to aspirate all visible follicles on the right side in vivo. Twenty-four oocytes were retrieved, 15 were mature and 5 blastocysts were cryopreserved. Final pathology of left ovary returned as serous cystadenoma. The right ovary was examined by gynecologic oncology prior to and following retrieval and was thought to be normal and remained in situ. CONCLUSION(S): Although the approach described here is not feasible in most cases, this video demonstrates a unique and successful fertility preservation technique by direct ultrasound-guided follicle aspiration in vivo at the time of laparotomy in a Mayer-Rokitansky-Küster-Hauser Syndrome patient and to our knowledge, is the first description of its kind. This retrieval would have otherwise been limited by lack of access transvaginally and limited visualization transabdominally. This combined approach should be considered in future patients with müllerian anomalies and similar complicating factors necessitating laparotomy.

Using microscope for onco-testicular sperm extraction for bilateral testis tumors.

OBJECTIVE: To demonstrate a step-by-step approach to the use of the operating microscope for onco-testicular sperm extraction. DESIGN: Video presentation. SETTING: University hospital. PATIENT(S): A 34-year-old man (status post right orchiectomy at another institution for pT3 pure seminoma with negative preoperative tumor markers) was referred for contralateral orchiectomy for multifocal left testis mass and fertility preservation. A postoperative semen analysis for attempted cryopreservation of ejaculated semen identified azoospermia. INTERVENTION(S): Left radical orchiectomy, left microsurgical onco-testicular sperm extraction (TESE). MAIN OUTCOME MEASURE(S): Intraoperative technique with commentary highlighting tips for successful fertility preservation via microsurgical onco-TESE. Discussion of alternatives. RESULT(S): This video provides a step-by-step guide to microsurgical onco-TESE coordinated with radical orchiectomy for testis cancer as a means of fertility preservation in an azoospermic patient. Preoperative imaging with scrotal ultrasound can serve as a useful guide for targeting microdissection to areas of normal testicular parenchyma for extraction of seminiferous tubules likely to host normal spermatogenesis. This patient had successful recovery and cryopreservation of abundant testicular sperm following targeted ex-vivo testicular microdissection. CONCLUSION(S): Microsurgical onco-TESE may be offered to azoospermic patients when undergoing orchiectomy for testis cancer. Use of preoperative imaging and the surgical microscope guide surgical dissection and optimize sperm recovery.

Short-term preservation of Pecari tajacu ovarian preantral follicles using phosphate buffered saline (PBS) or powdered coconut water (ACP(r)) media / Preservação de folículos ovarianos pré-antrais de Pecari tajacu por curtos períodos utilizando meios à base de solução salina fosfatada tamponada (PBS) ou água de coco em pó (ACP(r))

We compare protocols for the short-term preservation of collared peccarie's ovarian preantral follicles (PFs) by using phosphate buffered saline- (PBS) or powdered coconut water- (ACP(r)) based medium. For morphology analysis each pair of ovaries collected from six females was divided into nine fragments. One fragment was destined for morphology analysis (histology and transmission electron microscopy - TEM), constituting the control group and the other fragments were placed in tubes with PBS or ACP(r), packed in 5 L Styrofoam boxes, stored for 4h, 12h, 24h, and 36h, and then analyzed. For viability analysis a pair of ovaries from two additional females was divided into nine fragments; one fragment was immediately destined for viability analysis (Trypan blue test) and the other fragments were stored as previously described, until 24h and then analyzed. After 4h storage in ACP(r) medium, the follicular integrity was similar to control (87.8% vs 94.4%, respectively); however, ultrastructural analyses revealed swollen mitochondria as the first signals of PF degeneration. It was observed that ACP(r) (66.7%) was more efficient than PBS (49.4%) to preserve the morphological integrity after 36h storage (P<0.05); however, no differences were observed on follicular viability (P>0.05). In conclusion, the use of the ACP(r) is recommended for the short-term preservation of Pecari tajacu preantral follicles...

Compararam-se protocolos para a preservação por curtos períodos de folículos ovarianos pré-antrais (PFs) de catetos, utilizando meios à base de solução salina tamponada (PBS) ou água de coco em pó (ACP(r)). Para a análise morfológica, cada par de ovários coletados de seis fêmeas foi dividido em nove fragmentos. Um fragmento foi destinado para a análise da morfologia (histologia e microscopia eletrônica de transmissão - MET), constituindo o grupo controle, e os demais fragmentos foram colocados em tubos contendo PBS ou ACP(r), acondicionados em caixas térmicas de poliestireno expandido de 5L, armazenados durante quatro, 12, 24 e 36 horas, e, então, analisados. Para a análise da viabilidade, pares de ovários de duas fêmeas adicionais foram divididos em nove fragmentos; um deles foi imediatamente destinado à análise da viabilidade (teste com azul de Trypan), os outros fragmentos foram armazenados como descrito previamente até 24h e, então, foram analisados. Após quatro horas de armazenamento em meio ACP(r), a integridade folicular foi similar ao grupo controle (87,8% vs. 94,4%, respectivamente); contudo, a análise ultraestrutural revelou mitocôndrias edemaciadas como os primeiros sinais de degeneração dos PFs. Foi observado que o ACP(r) (66,7%) foi mais eficiente do que o PBS (49.4%) em preservar a integridade morfológica após 36h (p<0,05); entretanto, nenhuma diferença foi observada para a viabilidade folicular (P>0,05). Em conclusão, o uso da ACP(r) é recomendado para a preservação por curtos períodos de folículos pré-antrais de Pecari tajacu...

First transplantation of isolated murine follicles in alginate.

AIM: Our aim is to develop an artificial ovary allowing survival and growth of isolated follicles and ovarian cells, to restore fertility in women diagnosed with pathologies at high risk of ovarian involvement. MATERIALS & METHODS: For this, alginate beads containing isolated preantral follicles and ovarian cells were autografted to immunocompetent mice. One week after grafting, the beads were invaded by proliferating murine cells (12.1%) and capillaries. RESULTS: The recovery rate of follicles per graft ranged from 0% to 35.5%. Of the analyzed follicles, 77% were Ki67-positive and 81%, TUNEL-negative. Three antral follicles were also identified, evidencing their ability to grow in the matrix. CONCLUSION: Our results suggest that an artificial ovary is now conceivable, opening new perspectives to restore fertility in women.

The use of a metal container for vitrification of mouse ovaries, as a clinical grade model for human ovarian tissue cryopreservation, after different times and temperatures of transport.

PURPOSE: Cryopreservation of ovarian tissue is paramount for fertility preservation, with important clinical applications, especially for women suffering from an oncological condition. Several cryopreservation methodologies have been tried in search of better outcomes, especially in terms of primor-dial and primary follicles integrity post-cryopreservation. Vitrification has successfully been applied to ovarian tissue using different carriers for tissue exposure to the liquid nitrogen (LN2). METHODS: We developed an enclosed metal vessel, which has the advantage of a faster heat transfer, when in contact with LN2 avoiding at the same time, the direct contact with tissue. Additionally, we assessed the effect of different times and temperatures of transport between the collection of mouse ovaries and the beginning of cryopreservation, on follicular morphology after vitrification. RESULTS: Our results suggest that 37 °C and R.T. help to maintain normal primordial and primary follicle morphology for up to 4 hrs after collection and beginning of vitrification in a metal container. CONCLUSION: These data show that the metal container is an appropriate carrier for mouse ovary vitrification. The rate of morphologically normal primordial follicles up to 4 hrs.