Follicular fluid supplementation during in vitro maturation promotes sperm penetration in bovine oocytes by enhancing cumulus expansion and increasing mitochondrial activity in oocytes

2012 ◽  
Vol 24 (5) ◽  
pp. 743 ◽  
Author(s):  
Tamás Somfai ◽  
Yasushi Inaba ◽  
Shinya Watanabe ◽  
Masaya Geshi ◽  
Takashi Nagai
Keyword(s):  
Embryo Development ◽  
Follicular Fluid ◽  
Calf Serum ◽  
In Vitro Fertilisation ◽  
Mitochondrial Activity ◽  
Atp Content ◽  
Promoting Effect ◽  
Cumulus Expansion ◽  
Sperm Penetration

The aim of this study was to examine the effects of bovine follicular fluid (bFF) on mitochondrial activity in in vitro-matured (IVM) oocytes and to assess its importance for fertilisation and embryo development. Bovine follicular oocytes were subjected to IVM in medium supplemented either with polyvinylpyrrolidone, bovine serum albumin, calf serum or bFF. Nuclear maturation, cumulus expansion, mitochondrial distribution and ATP content in oocytes were compared between groups along with subsequent in vitro fertilisation (IVF) and embryo development. Compared with other supplements, bFF generated significantly enhanced re-distribution of active mitochondria in oocytes and this effect was associated with elevated intracellular ATP content. Furthermore, bFF significantly improved cumulus expansion, which was associated with improved fertilisation rates when cumulus-enclosed oocytes were subjected to IVF; however, its promoting effect was neutralised when denuded oocytes were inseminated. Elevating ATP content in oocytes by bFF did not affect maturation or embryo development but promoted fertilisation when mitochondrial electron transport was blocked in oocytes before IVF by Rotenone. In conclusion, supplementation of IVM medium with bFF promotes sperm penetration both by the improvement of cumulus expansion and by enhancing ATP levels in oocytes, which maintains their ability to be fertilised after mitochondrial stress.

scholarly journals Antioxidant Nobiletin Enhances Oocyte Maturation and Subsequent Embryo Development and Quality

2020 ◽  
Vol 21 (15) ◽  
pp. 5340
Author(s):  
Yulia N. Cajas ◽  
Karina Cañón-Beltrán ◽  
Magdalena Ladrón de Guevara ◽  
María G. Millán de la Blanca ◽  
Priscila Ramos-Ibeas ◽  
...  
Keyword(s):  
Embryo Development ◽  
Biological Effects ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Developmental Competence ◽  
Mitochondrial Activity ◽  
Cortical Granules ◽  
Cleavage Rate ◽  
Cytoplasmic Maturation

Nobiletin is a polymethoxylated flavonoid isolated from citrus fruits with wide biological effects, including inhibition of reactive oxygen species (ROS) production and cell cycle regulation, important factors for oocyte in vitro maturation (IVM). Therefore, the objective of the present study was to evaluate the antioxidant activity of nobiletin during IVM on matured bovine oocyte quality (nuclear and cytoplasmic maturation; oocyte mitochondrial activity; intracellular ROS and glutathione (GSH) levels) and their developmental competence, steroidogenesis of granulosa cells after maturation, as well as quantitative changes of gene expression in matured oocytes, their cumulus cells, and resulting blastocysts. Bovine cumulus-oocyte complexes were in vitro matured in TCM-199 +10% fetal calf serum (FCS) and 10 ng/mL epidermal growth factor (EGF) (Control) supplemented with 10, 25, 50, or 100 μM of nobiletin (Nob10, Nob25, Nob50, and Nob100, respectively) or 0.1% dimethyl sulfoxide (CDMSO: vehicle for nobiletin dilution). A significantly higher percentage of matured oocytes in metaphase II was observed in Nob25 and Nob50 compared to other groups. Similarly, cleavage rate and cumulative blastocyst yield on Days 7 and 8 were significantly higher for Nob25 and Nob50 groups. Oocytes matured with 25 and 50 μM nobiletin showed a higher rate of migration of cortical granules and mitochondrial activity and a reduction in the ROS and GSH content in comparison with all other groups. This was linked to a modulation in the expression of genes related to metabolism (CYP51A1), communication (GJA1), apoptosis (BCL2), maturation (BMP15 and MAPK1), and oxidative stress (SOD2 and CLIC1). In conclusion, nobiletin offers a novel alternative for counteracting the effects of the increase in the production of ROS during IVM, improves oocyte nuclear and cytoplasmic maturation, and subsequent embryo development and quality in cattle.

Linoleic (LA) and linolenic (ALA) acid concentrations in follicular fluid of prepubertal goats and their effect on oocyte in vitro maturation and embryo development

2018 ◽  
Vol 30 (2) ◽  
pp. 286 ◽  
Author(s):  
Montserrat Roura ◽  
María G. Catalá ◽  
Sandra Soto-Heras ◽  
Sondes Hammami ◽  
Dolors Izquierdo ◽  
...  
Keyword(s):  
Embryo Development ◽  
Follicular Fluid ◽  
Dna Methyltransferase ◽  
Glucose Transporter ◽  
Mitochondrial Activity ◽  
Glucose Transporter 1 ◽  
Treatment Groups ◽  
Atp Concentration ◽  
Follicle Size

In this study we assessed the concentration of linoleic acid (LA) and linolenic acid (ALA) in follicular fluid of prepubertal goats according to follicle size (<3 mm or ≥3 mm) by gas chromatography and tested the addition of different LA and ALA (LA : ALA) concentration ratios (50 : 50, 100 : 50 and 200 : 50 µM) to the IVM medium on embryo development, mitochondrial activity, ATP concentration and relative gene expression (RPL19, ribosomal protein L19; SLC2A1, facilitated glucose transporter 1; ATF4, activating transcription factor 4; GPX1, glutathione peroxidase 1; HSPA5, heat-shock protein family A 70 kDa; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; DNMT1, DNA methyltransferase 1; GCLC, glutamate–cysteine ligase catalytic subunit; SOD1, superoxide dismutase 1). Oocytes were in vitro matured, fertilised or parthenogenetically activated and zygotes were cultured following conventional protocols. LA concentration ranged from 247 to 319 µM and ALA concentration from 8.39 to 41.19 µM without any effect of follicle size. Blastocyst production from the different groups was: control FCS (22.33%) and BSA (19.63%), treatments 50 : 50 (22.58%), 100 : 50 (21.01%) and 200 : 50 (9.60%). Oocytes from the 200 : 50 group presented higher polyspermy and mitochondrial activity compared with controls and the rest of the treatment groups. No differences were observed in ATP concentration or relative expression of the genes measured between treatment groups. In conclusion, the low number of blastocysts obtained in the 200 : 50 group was caused by a high number of polyspermic zygotes, which could suggest that high LA concentration impairs oocyte membranes.

206 THE IMPORTANCE OF CUMULUS EXPANSION FOR MONOSPERMIC FERTILIZATION OF PORCINE CUMULUS - OOCYTE COMPLEXES

2011 ◽  
Vol 23 (1) ◽  
pp. 202
Author(s):  
J. Beek ◽  
J. Bijttebier ◽  
D. Maes ◽  
H. Nauwynck ◽  
A. Van Soom
Keyword(s):  
Follicular Fluid ◽  
In Vitro Maturation ◽  
Indirect Evidence ◽  
Fertilization Rate ◽  
Control Group ◽  
Cumulus Expansion ◽  
Mean Diameter ◽  
Sperm Penetration ◽  
Epidermal Growth

In pigs, the exact role of the cumulus oöphurus during IVF still needs to be clarified. Indirect evidence exists that the rate of cumulus expansion is positively correlated with the defense against polyspermy. Epidermal growth factor (EGF) acts synergistically with FSH in the synthesis of hyaluronic acid, the deposition of which in the extracellular matrix is a prerequisite for cumulus expansion. Therefore, we aimed to evaluate the importance of cumulus expansion for fertilization results of porcine cumulus–oocyte complexes (COC) by using different EGF concentrations in the maturation medium. Cumulus–oocyte complexes were matured in vitro in NCSU23 medium supplemented with 10% follicular fluid (FF, obtained from 6- to 10-mm follicles) or 10% serum and 10, 20, or 50 ng mL–1 of EGF (n = 480 per experiment). In vitro maturation (IVM) in the presence of 10% FF and 10 ng mL–1 of EGF served as the control group. At 0, 22, 36, and 44 h of IVM, 20 COC of each group were selected for evaluation of cumulus expansion by measuring the maximum distance across the cumulus matrix (3 replications). Matured COC were co-incubated with frozen–thawed semen (6000 spermatozoa per oocyte) for 6 h. Subsequently, oocytes were cultured for 18 h. Zygotes were stained with 10 μg mL–1 of bis-benzimide (Hoechst) to assess the fertilization rate, polyspermy, and sperm penetration index (sp index, mean number of penetrated spermatozoa per fertilized oocyte; 2 replications). Differences in cumulus diameter were analyzed by one-way ANOVA. Fertilization parameters were analyzed by applying a logistic regression model to the results. Cumulus–oocyte complexes selected for IVM had a mean diameter of 240 μm. After 22 h of IVM in 10% FF, the mean diameter of COC was 336, 313, and 300 μm for 10, 20, and 50 ng mL–1 of EGF, respectively. After 44 h of IVM, these diameters had increased to 425, 388, and 397 μm. Twenty-two hours of IVM in 10% serum resulted in a COC diameter of 296, 305, and 276 μm for 10, 20, and 50 ng mL–1 of EGF. After 44 h of IVM, these diameters reached 330, 325, and 275 μm, respectively. Only 10% serum with 50 ng mL–1 of EGF proved unfavorable for cumulus expansion (P < 0.05). In vitro maturation for 44 h in 10% serum resulted in a smaller rate of cumulus expansion compared with IVM in 10% FF (P < 0.05), irrespective of EGF concentration. Penetration rate fluctuated between 84 and 100%, with no significant differences. Monospermic fertilization was lower in COC matured in the presence of 50 ng mL–1 of EGF compared with the control group (P < 0.05). The sp index increased in parallel with EGF concentrations and was higher after IVM in 10% serum than in 10% FF. Oocytes were penetrated by 3.1, 4.3, and 6.0 spermatozoa after IVM in serum with 10, 20, and 50 ng mL–1 of EGF, respectively. Results showed a tendency toward a lower rate of cumulus expansion concomitant with higher EGF concentrations. Follicular fluid was superior to serum in supporting cumulus expansion. Oocytes were penetrated by more spermatozoa when matured in 10% serum and 20 or 50 ng mL–1 of EGF compared with 10 ng mL–1 of EGF. Thus, the degree of cumulus expansion appears to be related to the sp index, confirming that the cumulus matrix may play a role in the polyspermy defense. This study was supported by Research Foundation-Flanders.

180 FATTY ACID COMPOSITION IN FOLLICULAR FLUID OF PREPUBERTAL GOAT OVARIES IN WINTER AND AUTUMN

2015 ◽  
Vol 27 (1) ◽  
pp. 181
Author(s):  
M. T. Paramio ◽  
M. Roura ◽  
S. Hammami ◽  
D. Izquierdo ◽  
M. G. Catalá
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Embryo Development ◽  
Follicular Fluid ◽  
Oocyte Quality ◽  
Blastocyst Stage ◽  
Omega 3 ◽  
Chromatography Analysis ◽  
Sperm Penetration

Fatty acids (FA) in follicular fluid (FF) play an important role on oocyte quality and embryo development (Fouladi-Nashta et al. 2007 Biol. Reprod. 77, 9–17). In our laboratory, we have shown in prepubertal goat differences in the percentage of blastocysts produced in vitro according to season. Thus, we have found in winter 15.8% and in autumn a decrease up to 4.7% of blastocysts that were produced from oocytes of 1 month old suckling Murciano-Granadina goat females and IVF with fresh semen. The aim of this study was to analyse composition of FF in order to find an explanation to seasonal changes in in vitro embryo production. Ovaries were recovered in winter and autumn from 1 month suckling goats (Murciano-Granadina) from a local slaughterhouse and the FF of all visible follicles was recovered using a sterile syringe. Each sample containing a pool of FF of different ovaries was frozen at –80°C until chromatography analysis. For the FA analysis, the Sukhija and Palmquist (1988 J. Agric. Food Chem. 36, 1202–1206) protocol with some adaptations was used. Briefly, 200 μL of FF sample was vortexed for 60 s with 250 μL of toluene and 1 mL of HCL (5%) and then warmed in a water bath for 1 h at 70°C. Subsequently 1.25 mL of K2CO3 (12%) and 500 μL of toluene was added, vortexed for 30 s and centrifuged for 5 min (3000 rpm). Finally the supernatant was recovered and dried with Na2SO4. The extracted samples were maintained in –20°C until gas chromatographic analysis (123–2362, Agilent Technologies Inc., Santa Clara, CA). The results in Table 1 express the mean of 3 replicates of follicular fluid pool as micromolar concentration of FA in FF. The FA profile in FF showed significant higher concentrations of α-linolenic (C18:3n3), eicosapentaenoic (EPA), docosahexaenoic (DHA), and omega-3 (n-3 polyunsaturated fatty acid; PUFA) in winter compared to autumn. This could be indicating that these PUFA have a positive effect on oocyte quality because of the higher embryo development of these oocytes during winter. Studies in our laboratory have shown that sperm penetration and normal zygotes were similar in both seasons even though the blastocyst yield was statistically higher in winter. We can speculate that fatty acids in the follicular environment are affecting the oocyte quality, increasing the possibility of reaching the blastocyst stage in prepubertal goat according to season. Further studies should be done to reach a more accurate conclusion. Table 1.Concentration (µM) of fatty acids in FF of prepubertal goat during winter and autumn (3 replicates)

scholarly journals Relationship of vascular perfusion of the wall of the preovulatory follicle to in vitro fertilisation and embryo development in heifers

Reproduction ◽  
2009 ◽  
Vol 137 (4) ◽  
pp. 689-697 ◽  
Author(s):  
M A R Siddiqui ◽  
E L Gastal ◽  
M O Gastal ◽  
M Almamun ◽  
M A Beg ◽  
...  
Keyword(s):  
Embryo Development ◽  
Follicular Fluid ◽  
Polar Body ◽  
Prostaglandin F2α ◽  
In Vitro Fertilisation ◽  
Preovulatory Follicle ◽  
Cleavage Rate ◽  
Vascular Perfusion ◽  
Cell Versus

The effect of the extent of vascular perfusion of the wall of the preovulatory follicle on in vitro cleavage rate of the recovered oocyte and embryo development to >8 cells was studied in 52 heifers. Heifers received a luteolytic dose of prostaglandin F2α (PGF2α) when the largest follicle was ≥11 mm. An ovulation-inducing injection of GnRH was given 36 h later (hour 0), and collection of follicular fluid and the oocyte was done at hour 26. Vascular perfusion of the follicular wall was assessed by colour Doppler ultrasonography at hours 0 and 26. Each of the recovered oocytes (41/52; 79%) was mature (extruded polar body). Cleavage and embryo development were assessed at 48 h and 120 h respectively, after in vitro fertilisation (IVF). The percentage of cleaved oocytes and >8 cell embryos was 80% (31/39) and 55% (17/31) respectively. Vascular perfusion of the follicular wall was greater (lower pulsatility index; P<0.001) for follicles that produced cleaved versus non-cleaved oocytes and greater (P<0.04) for follicles that produced >8 cell versus ≤8 cell embryos. Percentage of follicular wall with Doppler signals of blood flow was greater (P<0.001) for >8 cell versus ≤8 cell embryos. Follicular-fluid concentration of free IGF1 was lower for cleaved oocytes (P<0.001) and >8 cell embryos (P<0.05), and oestradiol was lower (P<0.05) for >8 cell embryos. Results supported the hypothesis that greater vascular perfusion of the wall of the preovulatory follicle was positively associated with IVF and embryo development.

scholarly journals Mitochondrial enrichment in infertile patients: a review of different mitochondrial replacement therapies

2021 ◽  
Vol 15 ◽  
pp. 263349412110235
Author(s):  
Cristina Rodríguez-Varela ◽  
Sonia Herraiz ◽  
Elena Labarta
Keyword(s):  
Stem Cells ◽  
Genetic Material ◽  
External Source ◽  
Oocyte Quality ◽  
Third Party ◽  
In Vitro Fertilisation ◽  
Mitochondrial Activity ◽  
Mitochondrial Transfer ◽  
Infertile Patients

Poor ovarian responders exhibit a quantitative reduction in their follicular pool, and most cases are also associated with poor oocyte quality due to patient’s age, which leads to impaired in vitro fertilisation outcomes. In particular, poor oocyte quality has been related to mitochondrial dysfunction and/or low mitochondrial count as these organelles are crucial in many essential oocyte processes. Therefore, mitochondrial enrichment has been proposed as a potential therapy option in infertile patients to improve oocyte quality and subsequent in vitro fertilisation outcomes. Nowadays, different options are available for mitochondrial enrichment treatments that are encompassed in two main approaches: heterologous and autologous. In the heterologous approach, mitochondria come from an external source, which is an oocyte donor. These techniques include transferring either a portion of the donor’s oocyte cytoplasm to the recipient oocyte or nuclear material from the patient to the donor’s oocyte. In any case, this approach entails many ethical and safety concerns that mainly arise from the uncertain degree of mitochondrial heteroplasmy deriving from it. Thus the autologous approach is considered a suitable potential tool to improve oocyte quality by overcoming the heteroplasmy issue. Autologous mitochondrial transfer, however, has not yielded as many beneficial outcomes as initially expected. Proposed mitochondrial autologous sources include immature oocytes, granulosa cells, germline stem cells, and adipose-derived stem cells. Presently, it would seem that these autologous techniques do not improve clinical outcomes in human infertile patients. However, further trials still need to be performed to confirm these results. Besides these two main categories, new strategies have arisen for oocyte rejuvenation by improving patient’s own mitochondrial function and avoiding the unknown consequences of third-party genetic material. This is the case of antioxidants, which may enhance mitochondrial activity by counteracting and/or preventing oxidative stress damage. Among others, coenzyme-Q10 and melatonin have shown promising results in low-prognosis infertile patients, although further randomised clinical trials are still necessary.

scholarly journals IProteomic study to identify factors in follicular fluid and/or serum involved in in vitro cumulus expansion of porcine oocytes

2020 ◽  
Author(s):  
J. Bijttebier ◽  
K. Tilleman ◽  
D. Deforce ◽  
M. Dhaenens ◽  
Soom A. Van ◽  
...  
Keyword(s):  
Follicular Fluid ◽  
Cumulus Expansion

scholarly journals Significance of disorders of proteomic profile of follicular fluid for predicting the effectiveness of in vitro fertilisation in women with infertility

2018 ◽  
Vol 99 (3) ◽  
pp. 496-503
Author(s):  
O S Zolotykh ◽  
S V Lomteva ◽  
K Yu Sagamonova
Keyword(s):  
Assisted Reproductive Technology ◽  
Follicular Fluid ◽  
Molecular Mechanisms ◽  
Reproductive Technologies ◽  
Reproductive Technology ◽  
In Vitro Fertilisation ◽  
Proteomic Profile ◽  
Technology Programs ◽  
Group 2

Aim. To study the proteomic profile of follicular fluid in patients with infertility in assisted reproductive technology programs. Methods. The study included women with infertility included in assisted reproductive technology programs: 15 women who had in vitro fertilisation which resulted in pregnancy (group 1) and 16 women with a negative result of this program (group 2). Fractionation of the follicular fluid samples was performed using the sets of special magnetic beads. Proteomic profiling was performed by tandem MALDI-mass-spectrometry. The anti-Müllerian hormone level was measured by ELISA. Results. The study revealed differences in the detectability of follicular fluid proteins with different regulatory properties in patients of groups 1 and 2. With the negative outcome of in vitro fertilisation, expression of a number of proteins involved in the processes of folliculogenesis, ovulation, selection of the dominant follicle, as well as proteins necessary for the development of the zygote and blastula was reduced in females' follicular fluid. Increased expression in women from group 2 was registered for proteins enhancing proteolytic reactions, cell apoptosis, including oocytes, which disrupt the positive action of activin and damage structural and functional state of mitochondria. A definite relationship was found between the level of anti-Müllerian hormone and rate of detection of a number of proteins, in particular protocadherin-2α, cystatin C, betaglycan, prostatic acid phosphatase, and dermicidin. Conclusion. The revealed changes in proteomic profile of the follicular fluid obviously play an important role in the molecular mechanisms that determine the effectiveness of assisted reproductive technologies; the identified differentially expressed proteins can serve as objective markers for predicting the outcomes of in vitro fertilisation.

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