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PloS One 2019Cells on the surface of the mesonephros give rise to replicating Gonadal Ridge Epithelial-Like (GREL) cells, the first somatic cells of the gonadal ridge. Later germ...
Cells on the surface of the mesonephros give rise to replicating Gonadal Ridge Epithelial-Like (GREL) cells, the first somatic cells of the gonadal ridge. Later germ cells associate with the GREL cells in the ovigerous cords, and the GREL cells subsequently give rise to the granulosa cells in follicles. To examine these events further, 27 bovine fetal ovaries of different gestational ages were collected and prepared for immunohistochemical localisation of collagen type I and Ki67 to identify regions of the ovary and cell proliferation, respectively. The non-stromal cortical areas (collagen-negative) containing GREL cells and germ cells and later in development, the follicles with oocytes and granulosa cells, were analysed morphometrically. Another set of ovaries (n = 17) were collected and the expression of genes associated with germ cell lineages and GREL/granulosa cells were quantitated by RT-PCR. The total volume of non-stromal areas in the cortex increased significantly and progressively with ovarian development, plateauing at the time the surface epithelium developed. However, the proportion of non-stromal areas in the cortex declined significantly and progressively throughout gestation, largely due to a cessation in growth of the non-stroma cells and the continued growth of stroma. The proliferation index in the non-stromal area was very high initially and then declined substantially at the time follicles formed. Thereafter, it remained low. The numerical density of the non-stromal cells was relatively constant throughout ovarian development. The expression levels of a number of genes across gestation either increased (AMH, FSHR, ESR1, INHBA), declined (CYP19A1, ESR2, ALDH1A1, DSG2, OCT4, LGR5) or showed no particular pattern (CCND2, CTNNB1, DAZL, FOXL2, GATA4, IGFBP3, KRT19, NR5A1, RARRES1, VASA, WNT2B). Many of the genes whose expression changed across gestation, were positively or negatively correlated with each other. The relationships between these genes may reflect their roles in the important events such as the transition of ovigerous cords to follicles, oogonia to oocytes or GREL cells to granulosa cells.
Topics: Animals; Cattle; Female; Gene Expression Regulation, Developmental; Germ Cells; Granulosa Cells; Mesonephros; Ovary
PubMed: 30901367
DOI: 10.1371/journal.pone.0214130 -
International Journal of Molecular... Oct 2019Cancer treatment, such as chemotherapy, induces early ovarian follicular depletion and subsequent infertility. In order to protect gametes from the gonadotoxic effects... (Review)
Review
Cancer treatment, such as chemotherapy, induces early ovarian follicular depletion and subsequent infertility. In order to protect gametes from the gonadotoxic effects of chemotherapy, several fertility preservation techniques-such as oocyte or embryo cryopreservation with or without ovarian stimulation, or cryopreservation of the ovarian cortex-should be considered. However, these methods may be difficult to perform, and the future use of cryopreserved germ cells remains uncertain. Therefore, improving the methods currently available and developing new strategies to preserve fertility represent major challenges in the area of oncofertility. Animal and ovarian culture models have been used to decipher the effects of different cytotoxic agents on ovarian function and several theories regarding chemotherapy gonadotoxicity have been raised. For example, cytotoxic agents might (i) have a direct detrimental effect on the DNA of primordial follicles constituting the ovarian reserve and induce apoptosis; (ii) induce a massive growth of dormant follicles, which are then destroyed; or (ii) induce vascular ovarian damage. Thanks to improvements in the understanding of the mechanisms involved, a large number of studies have been carried out to develop molecules limiting the negative impact of chemotherapy on the ovaries.
Topics: Animals; Antineoplastic Agents; Cryopreservation; Female; Fertility Preservation; Humans; Models, Animal; Ovary; Primary Ovarian Insufficiency
PubMed: 31717833
DOI: 10.3390/ijms20215342 -
Journal of Ovarian Research May 2018Fertility preservation by whole ovary cryopreservation and transplantation (WOCP&TP) with vascular anastomosis requires successful cryopreservation. In this study, we...
BACKGROUND
Fertility preservation by whole ovary cryopreservation and transplantation (WOCP&TP) with vascular anastomosis requires successful cryopreservation. In this study, we investigated the possibility of restoring ovarian function and natural fertility after WOCP&TP in a premature ovarian insufficiency (POI) rat model. The influence of cryopreservation on the offspring of rats following WOCP&TP was also explored.
METHOD
Rats aged 8-10 weeks were used as donors and recipients for allotransplantation. Fifteen rat whole ovaries were divided into three groups: the optimized group, the conventional group, and the fresh group. Different perfusion modes were used before cryopreservation and after thawing. Whole ovaries were observed by morphologic analysis, immunohistochemical staining, and transferase-mediated deoxyuridine triphosphate nick end-labeling assay. Ovarian function and fertility after WOCP&TP were then observed in 25 cyclophosphamide-induced POI rats for 8 months. Ovarian function was assessed by vaginal smears and blood hormone levels. Fertility restoration was quantified as the live birth rate after mating. The filial generation of rats was mated at 8-10 weeks of age. Offspring were observed for birth defect.
RESULTS
Histological evaluation demonstrated intact morphology of follicles in all groups, with 77.6% of the total number of follicles identified as intact in the optimized group. The apoptotic rates of ovarian cells in the optimized group were significantly lower than that in the conventional group. Of the 20 live POI rats, 14 (70%) began to recover ovarian function after 2 weeks of transplantation, with normal hormone levels achieved 4 weeks after transplantation. Four of 14 rats were pregnant and delivered live offspring. One rat had a second pregnancy and delivered a second litter of live offspring. When the offspring matured, they were mated, and second and third generations of rats were born. All offspring had no abnormalities in appearance.
CONCLUSIONS
High rates of restoration of ovarian function and natural fertility with multiple generations of offspring were obtained following WOCP&TP in a cyclophosphamide-induced POI rat model by utilizing optimized perfusion. Cryopreservation did not affect the viability of successive generations.
Topics: Animals; Cryopreservation; Cyclophosphamide; Disease Models, Animal; Female; Fertility; Fertility Preservation; Humans; Menopause, Premature; Ovarian Follicle; Ovary; Pregnancy; Primary Ovarian Insufficiency; Rats; Reproduction; Vitrification
PubMed: 29716634
DOI: 10.1186/s13048-018-0401-4 -
Arthropod Structure & Development Jul 2014Astigmatans are a large group of mites living in nearly every environment and exhibiting very diverse reproductive strategies. In spite of an uniform anatomical... (Review)
Review
Astigmatans are a large group of mites living in nearly every environment and exhibiting very diverse reproductive strategies. In spite of an uniform anatomical organization of their reproductive systems, gametogenesis in each sex is highly variable, leading to gamete formation showing many peculiar features and emphasizing the distinct position of Astigmata. This review summarizes the contemporary knowledge on the structure of ovaries and testes in astigmatic mites, the peculiarities of oogenesis and spermatogenesis, as well as provides new data on several species not studied previously. New questions are discussed and approaches for future studies are proposed.
Topics: Acari; Animals; Female; Male; Oogenesis; Ovary; Spermatogenesis; Testis
PubMed: 24791694
DOI: 10.1016/j.asd.2014.04.003 -
PloS One 2016Transplantation of ovarian tissue (OT) is currently the only clinical option to restore fertility with cryopreserved OT. However, follicle loss caused by ischemia and...
Transplantation of ovarian tissue (OT) is currently the only clinical option to restore fertility with cryopreserved OT. However, follicle loss caused by ischemia and slow revascularization occurs in transplanted OT. To shorten the ischemic period and promote angiogenesis, some angiogenic factors have been used. Angiopoietin-2 (Ang2) is one of the major angiogenic factors and has been reported to promote blood vessels and increase vascular permeability in ischemic and/or hypoxic environment. This study was performed to investigate the effects of Ang2 on follicle integrity and revascularization of transplanted mouse OT. Five-week-old B6D2F1 female mice were divided into a control group and two Ang2 groups, followed by ovary collection and vitrification. After warming, the ovaries were autotransplanted into kidney capsules with/without Ang2 injection (50 or 500 ng/kg), and then the mice were sacrificed at days 2, 7, 21, and 42 after transplantation. A total 2,437 follicles in OT grafts were assessed for follicular density, integrity, and classification by using hematoxylin and eosin staining. Apoptosis and revascularization were evaluated by using TUNEL assay and CD31 immunohistochemistry, respectively. Serum follicle-stimulating hormone (FSH) levels were measured by using enzyme-linked immunosorbent assay. Both Ang2 groups showed remarkable increase in morphologically intact follicle ratio across all grafting durations except D21. The numbers of CD31(+) vessels were significantly increased in both Ang2 groups compared with the control group at all durations, except in the 50 ng Ang2 group at D42. However, the mean numbers of follicles of the grafts, apoptosis ratios, and serum FSH levels showed no significant differences among the groups. Our results show that Ang2 treatment significantly increased the intact follicle ratios and the number of blood vessels of the mouse OT grafts. However, further studies performed with large animal or human OT are necessary before clinical application for fertility preservation in cancer patients, and the reliability of the systemic effects of Ang2 should be verified.
Topics: Animals; Apoptosis; Cryopreservation; Female; Fertility Preservation; Follicle Stimulating Hormone; Mice; Neovascularization, Physiologic; Ovarian Follicle; Ovary; Ribonuclease, Pancreatic; Transplantation, Autologous
PubMed: 27870915
DOI: 10.1371/journal.pone.0166782 -
Ultrasound in Medicine & Biology Jul 2021Timely angiogenesis and effective microcirculation perfusion are essential for the survival and functional recovery of transplanted ovaries. Ultrasound-targeted...
Timely angiogenesis and effective microcirculation perfusion are essential for the survival and functional recovery of transplanted ovaries. Ultrasound-targeted microbubble destruction (UTMD) can lead to angiogenesis and increase flow perfusion by causing transient inflammation. The purpose of this study was to evaluate the effects of UTMD on transplanted ovarian revascularization and survival. In vitro, for the criteria of cell viability and tube formation capability, the optimal exposure parameters were determined to be a microbubble concentration of 1 × 10/mL, mechanical index of 1 and exposure time of 30 s. After ovarian transplantation, 40 female Sprague Dawley rats were divided into four groups: transplantation alone, ultrasound alone, microbubbles alone and ultrasound and microbubbles (UTMD). At 7 d after transplantation, ovarian perfusion was assessed using qualitative and quantitative methods. The effect of angiogenesis was assessed by contrast-enhanced ultrasound, laser Doppler perfusion imaging and histologic analysis. The results, in which ovarian perfusion was highest in the UTMD group, suggest that UTMD can effectively improve ovarian perfusion. Compared with the other three groups, the number of follicles, microvascular density and rate of Ki-67-positive cells increased significantly in the UTMD group, while apoptosis decreased significantly (p < 0.05). The study indicates that UTMD promoted ovarian re-vascularization after ovarian transplantation and maintained follicular reserve.
Topics: Animals; Contrast Media; Female; Microbubbles; Neovascularization, Physiologic; Ovary; Rats; Rats, Sprague-Dawley; Ultrasonic Waves; Ultrasonography
PubMed: 33832825
DOI: 10.1016/j.ultrasmedbio.2021.02.025 -
Reproductive Biology and Endocrinology... May 2018Retinoids (retinol and its derivatives) are required for the development and maintenance of normal physiological functions of the ovary. However, the mechanisms...
BACKGROUND
Retinoids (retinol and its derivatives) are required for the development and maintenance of normal physiological functions of the ovary. However, the mechanisms underlying the regulation of ovarian retinoid homeostasis during follicular development remain unclear.
METHODS
The present study determined retinoid levels and the expression levels of genes involved in the retinol uptake and its metabolic pathway in the ovaries of follicle-stimulating hormone (FSH)-treated mice and in granulosa cells treated with FSH using ultra performance liquid chromatography (UPLC) combined with quadrupole time-of-flight high-sensitivity mass spectrometry (Q-TOF/HSMS) and real-time PCR analysis.
RESULTS
The levels of total retinoids and retinoic acid (RA) and expressions of retinol-oxidizing enzyme genes alcohol dehydrogenase 1 (Adh1) and aldehyde dehydrogenase (Aldh1a1) are increased in the ovaries of mice treated with FSH; in contrast, the retinyl ester levels and retinol-esterifying enzyme gene lecithin: retinol acyltransferase (Lrat) expression are diminished. In FSH-treated granulosa cells, the levels of retinyl esters, retinaldehyde, and total retinoids are augmented; and this is coupled with an increase in the expressions of stimulated by retinoic acid 6 (Stra6) and cellular retinol-binding protein 1 (Crbp1), genes in the retinol uptake pathway, and Adh1, Adh7, and Aldh1a1 as well as a diminution in Lrat expression.
CONCLUSIONS
These data suggest that FSH promotes retinol uptake and its conversion to RA through modulating the pathways of retinol uptake and metabolism in the mouse ovary. The present study provides a possible mechanism for the regulation of endogenous RA signaling in the developing follicles.
Topics: Animals; Cells, Cultured; Female; Follicle Stimulating Hormone; Gene Expression Regulation; Granulosa Cells; Metabolic Networks and Pathways; Mice; Mice, Inbred BALB C; Ovary; Vitamin A
PubMed: 29803227
DOI: 10.1186/s12958-018-0371-9 -
Reproductive Biology and Endocrinology... Feb 2021Melatonin has anti-inflammatory and antioxidative actions at the mitochondrial level. This indole-containing molecule may protect ovarian grafts during the process of...
BACKGROUD
Melatonin has anti-inflammatory and antioxidative actions at the mitochondrial level. This indole-containing molecule may protect ovarian grafts during the process of cryopreservation. Therefore, we aimed to determine whether melatonin pretreatment improves rat ovarian graft quality.
METHODS
Twenty-six female rats were allocated to two study groups of thirteen animals each: 1) control group: ovaries cryopreserved using the standard protocol; and 2) melatonin group: ovaries cryopreserved in a medium with melatonin. Ten rats of each group were submitted to 24-h freezing, and whole ovaries autologous and avascular transplantation with retroperitoneal placement. After postoperative (PO) day 15, daily vaginal smears were obtained for estrous cycle characterization. Between PO days 30 and 35, the animals were euthanized and ovarian grafts were recovered for histological and immunohistochemical (Ki-67, cleaved caspase-3, TUNEL, von Willebrand factor, estrogen, and progesterone receptors) analyses. The ovaries of the three remaining rats from each group were studied immediately after thawing to assess the effects of cryopreservation. ANOVA and Tukey's tests were used and the rejection level of the null hypothesis was set at 0.05 or 5% (p < 0.05).
RESULTS
Melatonin promoted faster restart of the estrous cycle and increased the expression of mature follicles, collagen type I, von Willebrand factor, Ki-67, and cleaved caspase-3 on corpora lutea and estrogen receptors in the ovaries as compared to control. There was a reduction in apoptosis by TUNEL on follicles, corpora lutea, and collagen type III.
CONCLUSION
Based on the evaluated parameters, melatonin may promote the quality of ovarian grafts. Reproductive function enhancement should be further studied.
Topics: Animals; Cryopreservation; Culture Media; Cytoprotection; Female; Melatonin; Ovary; Rats; Rats, Wistar; Time Factors
PubMed: 33536029
DOI: 10.1186/s12958-021-00705-4 -
Journal of Assisted Reproduction and... Jun 2020To develop a new protocol for whole-ovary decellularization for the production of a 3D bioscaffold suitable for in vitro/ex vivo studies and for the reconstruction of a...
PURPOSE
To develop a new protocol for whole-ovary decellularization for the production of a 3D bioscaffold suitable for in vitro/ex vivo studies and for the reconstruction of a bioengineered ovary.
METHODS
Porcine ovaries were subjected to the decellularization process (DECELL; n = 20) that involved a freeze-thaw cycle, followed by sequential incubations in 0.5% SDS for 3 h, 1% Triton X-100 for 9 h, and 2% deoxycholate for 12 h. Untreated ovaries were used as a control (CTR; n = 6). Both groups were analyzed to evaluate cell and DNA removal as well as ECM preservation. DECELL bioscaffolds were assessed for cytotoxicity and cell homing ability.
RESULTS
DECELL ovaries maintained shape and homogeneity without any deformation, while their color turned from red to white. Histological staining and DNA quantification confirmed a decrease of 98.11% in DNA content, compared with the native tissue (CTR). Histochemical assessments demonstrated the preservation of intact ECM microarchitecture after the decellularization process. This was also confirmed by quantitative analysis of collagen, elastin, and GAG contents. DECELL bioscaffold showed no cytotoxic effects in co-culture and, when re-seeded with homologous fibroblasts, encouraged a rapid cell adhesion and migration, with repopulating cells increasing in number and aggregating in cluster-like structures, consistent with its ability to sustain cell adherence, proliferation, and differentiation.
CONCLUSION
The protocol described allows for the generation of a 3D bioscaffold that may constitute a suitable model for ex vivo culture of ovarian cells and follicles, as well as a promising tool for the reconstruction of a bioengineered ovary.
Topics: Animals; Bioengineering; Extracellular Matrix; Female; Fibroblasts; Humans; Octoxynol; Ovary; Swine; Tissue Engineering; Tissue Scaffolds
PubMed: 32361917
DOI: 10.1007/s10815-020-01784-9 -
Biology of Reproduction Oct 2020Development and functions of the ovary rely on appropriate signaling and communication between various ovarian cell types. FOXL2, a transcription factor that plays a key...
Development and functions of the ovary rely on appropriate signaling and communication between various ovarian cell types. FOXL2, a transcription factor that plays a key role at different stages of ovarian development, is associated with primary ovarian insufficiency and ovarian cancer as a result of its loss-of-function or mutations. In this study, we investigated the impact of aberrant, constitutive expression of FOXL2 in somatic cells of the ovary. Overexpression of FOXL2 that started during fetal life resulted in defects in nest breakdown and consequent formation of polyovular follicles. Granulosa cell differentiation was impaired and recruitment and differentiation of steroidogenic theca cells was compromised. As a consequence, adult ovaries overexpressing FOXL2 exhibited defects in compartmentalization of granulosa and theca cells, significant decreased steroidogenesis and lack of ovulation. These findings demonstrate that fine-tuned expression of FOXL2 is required for proper folliculogenesis and fertility.
Topics: Animals; Cell Differentiation; Female; Forkhead Box Protein L2; Granulosa Cells; Mice; Mutation; Ovarian Follicle; Ovary; Theca Cells
PubMed: 32945847
DOI: 10.1093/biolre/ioaa146