Levels of MMP2, TIMP1 and TIMP2 in Follicular Fluids in Women Undergoing in Vitro Fertilization and Their Relationship to Oocyte Quality

2019 ◽  
Vol 12 (2) ◽  
pp. 100-107
Author(s):  
Nina P. Ayvazova ◽  
Lyubomira O. Ilieva ◽  
Emiliana I. Konova ◽  
Milena A. Atanasova
Keyword(s):  
Matrix Metalloproteinases ◽  
Ovarian Tissue ◽  
Follicular Development ◽  
Germinal Vesicle ◽  
Enzyme Linked Immunosorbent Assay ◽  
Oocyte Maturity ◽  
Vitro Fertilization ◽  
Follicular Fluids

Summary Recently, the important role of matrix metalloproteinases (MMPs) has been identified in follicular development and subsequent ovulation. Although the role of MMP in ovarian tissue remodeling during folliculogenesis has been well studied, the relationship between matrix protease activity and their inhibitors - Tisue inhibitors of matrix metalloproteinases (TIMP) and aging of the oocytes is still unclear. The present study aimed to establish the probable relationship between the expression levels of MMP-2 and TMP-1 and TIMP-2 in follicular fluid with the degree of oocyte maturity and quality. Follicular fluids from 20 women collected on the day of follicular puncture were tested for the presence of MMP-2, TIMP-1, and TIMP-2 using enzyme-linked immunosorbent assay (ELISA). The oocytes obtained were described in terms of maturity, morphology, and fertilization, as well as the embryo’s quality and rate of development. MMP-2 was significantly higher in follicular aspirates in the first prophase of meiosis - germinal vesicle (GV), compared to aspirates with first metaphase (MI) (p=0.011) and second metaphase (MII) of mature oocytes (p=0.010). The MMP-2/TIMP-1 ratio was significantly higher for GV compared to M1 (p=0.011), M2 (p=0.006) and atretic oocytes (p=0.032); (F(3, 71)=2.909, p=0.040). Based on our results, we can conclude that MMP-2 concentration in follicular fluids during the IVF / ICSI procedure had a significant relationship to oocyte maturation levels. It was significantly higher in the case of immature oocytes. On the other hand, oocytes with normal morphology were associated with a significantly higher MMP-2 concentration in follicular fluids.

198 KARYOPHERIN ALPHA EXPRESSION IN PORCINE OOCYTES AND EMBRYOS PRODUCED BY IN VITRO FERTILIZATION

2009 ◽  
Vol 21 (1) ◽  
pp. 197
Author(s):  
X. Wang ◽  
L. Magnani ◽  
R. Cabot
Keyword(s):  
Internal Control ◽  
Ovarian Tissue ◽  
Germinal Vesicle ◽  
Transcript Abundance ◽  
Cell Stage ◽  
Vitro Fertilization ◽  
Significance Level ◽  
Nuclear Localization Signals ◽  
Importin Α

Partitioning intracellular proteins between nuclear and cytoplasmic compartments is critically important for coordinating major cellular events involved in transcription and differentiation. Import of cytoplasmic proteins bearing classical nuclear localization signals (NLSs) into the nucleus is mediated by the importin α/β heterodimer. Importin α, also called karyopherin α (KPNA), serves to recognize the NLS-bearing cytoplasmic cargo. Six KPNA molecules have been characterized in human (KPNA1-6). Select KPNA molecules are known to be differently expressed in specific tissues; individual KPNA molecules also have specificity for unique NLS-bearing cargos. We hypothesized that transcripts for individual porcine KPNA molecules would be present at differing levels at specific stages of oocyte maturation and cleavage development, thereby reflecting the changing requirements for particular import pathways during this window of development. To test this hypothesis, we first identified the porcine orthologs of KPNA1-6. We also identified the open reading frame of a potentially novel KPNA, KPNA7. KPNA7 was highly represented in the porcine EST database from expressed sequence tags derived from oocytes and ovarian tissue. Transcript abundance of KPNA1-7 was determined in germinal vesicle (GV) and MII-stage (MII) porcine oocytes and 4-cell (4C) and blastocyst-stage (BL) porcine embryos using quantitative real-time PCR. mRNA was isolated from pools (50 200) of GV and MII oocytes and 4C and BL embryos produced by IVF. Transcripts for KPNA1-7 and YWHAG (internal control for transcript normalization) were amplified in duplicate across 3 to 5 replicates. Relative transcript abundance of these genes was measured using the comparative CT method; GV was taken as the calibrator stage. Data were analyzed using GLM procedures in SAS (SAS Institute Inc., Cary, NC, USA) with the significance level at 0.05, and differences were compared by Tukey’s post test. Our results showed that KPNA1 had a significant decrease in MII oocytes (4-fold, GV v. MII). Transcript abundance of KPNA2 was significantly higher in GV oocytes and 4C embryos than in MII oocytes (2-fold GV v. MII; 3-fold 4C v. MII); KPNA2 transcripts were not detectable in BL embryos. KPNA3 transcripts were reduced in BL embryos compared to GV oocytes (8-fold, BL v. GV). KPNA4 transcripts were increased at the 4-cell stage (18-fold, GV v. 4C). The transcripts of KPNA5 were detectable only in GV and MII oocytes. No significant changes in the amount of KPNA6 transcripts were detectable at the stages analyzed. Transcript levels of KPNA7 were reduced in BL embryos as compared to the GV oocytes (1165-fold, BL v. GV). Throughout all these stages examined, KPNA5 had the lowest transcript abundance and was not detectable in 4C and BL stages. Transcripts levels of KPNA7 were in higher abundance than KPNA1-6 in GV and MII oocytes. Results suggest that KPNA7 is a new member of the KPNA family. Our results also suggest that porcine oocytes and embryos, at discrete stages of development, have differing requirements for individual KPNA molecules.

Extrafollicular mediation of oocyte maturation by radial nerve factor in starfish Pisaster ochraceus

Zygote ◽  
2000 ◽  
Vol 8 (4) ◽  
pp. 359-368 ◽  
Author(s):  
Allen W. Schuetz
Keyword(s):  
Oocyte Maturation ◽  
Radial Nerve ◽  
Actinomycin D ◽  
Ovarian Tissue ◽  
Germinal Vesicle ◽  
Follicle Cells ◽  
Pisaster Ochraceus ◽  
Ovarian Wall

In starfish ovaries follicle cells that envelop each oocyte are thought to mediate the production of a maturation inducing substance (MIS), identified as 1-methyladenine, that induces maturation and spawning of oocytes after exposure to a gonadotropic substance secreted by the radial nerve (RNF). Studies were carried out to assess the possible role of extrafollicular cells within the ovarian wall in mediating this signal transduction process in the ovary of Pisaster ochraceus. Oocyte maturation and spawning occurred following the addition of RNF to intact ovarian tissue in vitro whereas no maturation occurred following the addition of RNF to germinal vesicle (GV) oocytes or GV oocytes surrounded by follicle cells. In contrast, oocyte maturation occurred when small ovarian wall fragments, lacking mature follicles, were incubated with GV oocytes and RNF. Neither actinomycin D nor cycloheximide altered RNF induction of oocyte maturation in the presence of the ovarian wall tissue whereas preheating (boiling water for 5 min) the tissue obliterated its response to RNF. Non-ovarian tissues failed to produce MIS in response to RNF. Results suggest that ovarian components other than the follicle cells that envelop fully grown immature oocyte are responsive to RNF and represent a significant and previously unrecognised intra-ovarian source of MIS.

Fibronectin production by chicken granulosa cells in vitro: effect of follicular development

1992 ◽  
Vol 127 (5) ◽  
pp. 466-470 ◽  
Author(s):  
Elikplimi K Asem ◽  
Jacqueline A Carnegie ◽  
Benjamin K Tsang
Keyword(s):  
Granulosa Cells ◽  
Follicular Development ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Ovarian Granulosa Cells ◽  
Preovulatory Follicles ◽  
Vitro Effect ◽  
Dose Dependent

In vitro studies were conducted to investigate the role of chicken ovarian granulosa cells in the production of fibronectin, a component of the basal lamina of ovarian follicles. Collagenase dispersed granulosa cells obtained from the first (F1; about 35 mm in diameter) and third (F3; 15–20 mm in diameter) largest preovulatory follicles, as well as from a pool of small yellow follicles (SF; 6–10 mm in diameter), were incubated in serum-free medium-199 for 24 to 96 h in the absence and presence of luteinizing hormone (LH) or forskolin. Fibronectin secreted in the medium was quantitated by enzyme linked immunosorbent assay. Basal fibronectin production (which increased with the duration of incubation) was significantly greater (p<0.001) in granulosa cells derived from mature follicle (F1) than in F3 or SF cells. Both LH and forskolin stimulated fibronectin production in SF and F3 cells in a dose-dependent manner; however, they were without effect in F1 cells. The magnitude of increase in fibronectin production elicited by LH or forskolin was greater in SF cells than in F3 cells. The cytoplasm of cultured granulosa cells taken at all stages of follicular development stained positively for fibronectin. These findings indicate that chicken granulosa cells produce fibronectin. This ability is acquired early in follicular development and the stimulatory effect of the gonadotropin (LH) diminished as the follicle approached ovulation.

scholarly journals Empty Zona Pellucida Only Case: A Critical Review of the Literature

2021 ◽  
Vol 18 (17) ◽  
pp. 9409
Author(s):  
Charalampos Siristatidis ◽  
Despoina Tzanakaki ◽  
Mara Simopoulou ◽  
Christina Vaitsopoulou ◽  
Petroula Tsioulou ◽  
...  
Keyword(s):  
Zona Pellucida ◽  
Critical Review ◽  
Treatment Options ◽  
Specific Treatment ◽  
Follicular Development ◽  
Oocyte Retrieval ◽  
Vitro Fertilization ◽  
Preimplantation Embryo Development

The presence of empty zona pellucida (EZP) in oocytes following oocyte retrieval (OR) during an in vitro fertilization (IVF) cycle presents a major clinical and laboratory challenge in assisted reproduction. It has been attributed to several factors such as the ovarian stimulation protocol employed, the damaging of the follicles during oocyte retrieval (OR) mainly through the high aspiration pressure, during the denudation technique, and the degeneration of oolemma within the zona pellucida (ZP) through apoptosis. The role of ZP is pivotal from the early stages of follicular development up to the preimplantation embryo development and embryo hatching. Polymorphisms or alterations on the genes that encode ZP proteins may contribute to EZP. We present a critical review of the published literature hitherto on EZP and available options when encountered with the phenomenon of EZP. Concerning the former, we found that there is rare data on this phenomenon that merits documentation. The latter includes technical, genetic, and pathophysiological perspectives, along with specific treatment options. In conclusion, we identify the lack of a definitive management proposal for couples presenting with this phenomenon, we underline the need for an algorithm, and indicate the questions raised that point towards our goal for a strategy when addressing a previous finding of EZP.

scholarly journals Zinc Chloride Supplemented during Ovarian Tissue Vitrification Improves In Vitro Follicle Development and Fertilization in Pigs

2021 ◽  
Vol 121 (2) ◽  
pp. 56-63
Author(s):  
Emma C. Hicks ◽  
Megan Martz ◽  
Haley A. Arena ◽  
Justin L. Rheubert ◽  
Brian D. Whitaker
Keyword(s):  
Embryonic Development ◽  
Zinc Chloride ◽  
Ovarian Tissue ◽  
Follicular Development ◽  
Vitro Fertilization ◽  
Antral Follicles ◽  
Mineral Supplementation ◽  
The Media ◽  
Pronuclear Formation

Ovarian tissue vitrification is a promising method to preserve follicles and gametes, but can be improved with mineral supplementation to the vitrification medium. The objective of this study was to determine the effects of supplementing 0.5 mg/mL zinc chloride to the media during cryopreservation of pig ovarian tissue. After thawing, the following criteria were evaluated: (1) follicular development and damage, (2) in vitro fertilization (IVF) characteristics, and (3) embryonic development. The number of damaged antral follicles (72.0 ± 3.8%) was less (p < 0.05) in ovarian tissue vitrified in media supplemented with zinc chloride compared to those not supplemented with zinc chloride (86.7 ± 4.1%). Oocytes obtained from the antral follicles on ovarian tissue vitrified in media supplemented with zinc chloride had less (p < 0.05) polyspermic penetration and higher (p < 0.05) male pronuclear formation during IVF than oocytes obtained from ovarian tissue not supplemented with zinc chloride. There were no statistical differences in embryonic development rates. Based on these results, supplementing zinc chloride during the vitrification protocol improves follicular development and subsequent IVF in pigs.

169 CHANGES OF PROTEIN PROFILES DURING FOLLICLE DEVELOPMENT AND IN VITRO OOCYTE MATURATION IN THE PIG

2011 ◽  
Vol 23 (1) ◽  
pp. 187 ◽  
Author(s):  
M. R. Ji ◽  
D. M. Jang ◽  
Y. S. Lee ◽  
H. T. Cheong ◽  
B. K. Yang ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Protein Extraction ◽  
Protein Profile ◽  
Follicular Development ◽  
Germinal Vesicle ◽  
Maldi Mass Spectrometry ◽  
Protein Patterns ◽  
Total Proteins ◽  
Follicular Fluids

When fully grown oocytes are removed from their follicles, they can resume meiosis and mature spontaneously under in vitro conditions. However, nuclear maturation under in vitro conditions is not accompanied by complete cytoplasmic maturation, which is essential for successful fertilization and the initiation of zygotic development. In the first study, to investigate protein patterns during oocyte maturation in vitro, immature oocytes (germinal vesicle stage; GV stage) as control and oocytes matured (M-II phase) in vitro were analysed. The porcine oocytes in the GV stage were put in culture with TCM-199 for 44 h for M-II-stage oocytes. Total proteins were extracted from 1200 oocytes for GV and M-II stages, separated on 2-D gels, and stained with silver. In general, the overall protein staining pattern between the 2 gels was remarkably similar for most protein spots. Analysis of the gels identified proteins that were up- or down-regulated between GV and M-II stages. Up-regulated proteins were identified as PDE4D, GPKOW, PGM5, HSP70, ZPG4, galK1, GST-β, PDX1, PDX2, and PDX3. In contrast, down-regulated proteins were identified as PRKAB1, GRP78, TD-pozl, ERP57, MPP1, DTNA, ZP3B, HSP90, HSP86, and HSP27. This study has identified a novel protein, named myomegalin, that interacts with cyclic nucleotide phosphodiesterase (PDE4D). The second study analysed changes in proteins in follicular fluids during porcine follicular development. Follicular fluids were collected from follicles of 1- to 2-, 2- to 6-, and 6- to 10-mm diameter from ovaries of slaughtered pigs. Total proteins were extracted from follicular fluids by M-PER Mammalian Protein Extraction Reagent. Differentially expressed proteins were analysed by MALDI mass spectrometry and searched on NCBInr. As a result, Spot No. 28 from the 2- to 6-mm follicle was Ig lambda chain C region, and Spot No. 32 and 33 from 6- to 10-mm were Apolipoprotein A-IV (APOA4). Increases of those proteins were correlated with follicular development. These results indicate that in vitro maturation changes the protein profile of porcine oocytes, which play important roles in the sequence of molecular events in porcine oocyte maturation and follicular development. This work was supported by Korean Research Foundation Grant funded by the Korean Government (2009-0071610).

Activation of bovine oocytes by protein synthesis inhibitors: new findings on the role of MPF/MAPKs†

2021 ◽  
Author(s):  
Cecilia Valencia ◽  
Felipe Alonso Pérez ◽  
Carola Matus ◽  
Ricardo Felmer ◽  
María Elena Arias
Keyword(s):  
Protein Synthesis ◽  
Kinase Activity ◽  
Cyclin B1 ◽  
Protein Synthesis Inhibitors ◽  
Vitro Fertilization ◽  
New Findings ◽  
Bovine Oocytes ◽  
Dependent Pathway

Abstract The present study evaluated the mechanism by which protein synthesis inhibitors activate bovine oocytes. The aim was to analyze the dynamics of MPF and MAPKs. MII oocytes were activated with ionomycin (Io), ionomycin+anisomycin (ANY) and ionomycin+cycloheximide (CHX) and by in vitro fertilization (IVF). The expression of cyclin B1, p-CDK1, p-ERK1/2, p-JNK, and p-P38 were evaluated by immunodetection and the kinase activity of ERK1/2 was measured by enzyme assay. Evaluations at 1, 4, and 15 hours postactivation (hpa) showed that the expression of cyclin B1 was not modified by the treatments. ANY inactivated MPF by p-CDK1Thr14-Tyr15 at 4 hpa (P &lt; 0.05), CHX increased pre-MPF (p-CDK1Thr161 and p-CDK1Thr14-Tyr15) at 1 hpa and IVF increased p-CDK1Thr14-Tyr15 at 17 hours postfertilization (hpf) (P &lt; 0.05). ANY and CHX reduced the levels of p-ERK1/2 at 4 hpa (P &lt; 0.05) and its activity at 4 and 1 hpa, respectively (P &lt; 0.05). Meanwhile, IVF increased p-ERK1/2 at 6 hpf (P &lt; 0.05); however, its kinase activity decreased at 6 hpf (P &lt; 0.05). p-JNK in ANY, CHX, and IVF oocytes decreased at 4 hpa (P &lt; 0.05). p-P38 was only observed at 1 hpa, with no differences between treatments. In conclusion, activation of bovine oocytes by ANY, CHX, and IVF inactivates MPF by CDK1-dependent specific phosphorylation without cyclin B1 degradation. ANY or CHX promoted this inactivation, which seemed to be more delayed in the physiological activation (IVF). Both inhibitors modulated MPF activity via an ERK1/2-independent pathway, whereas IVF activated the bovine oocytes via an ERK1/2-dependent pathway. Finally, ANY does not activate the JNK and P38 kinase pathways.

542 Objective Sleep Duration, Sleep Timing and Completion of In Vitro Fertilization Cycles; a Prospective Cohort Study

SLEEP ◽  
2021 ◽  
Vol 44 (Supplement_2) ◽  
pp. A214-A214
Author(s):  
Chawanont Pimolsri ◽  
Xiru Lyu ◽  
Cathy Goldstein ◽  
Chelsea Fortin ◽  
Sunni Mumford ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Sleep Duration ◽  
Medical Center ◽  
Working Women ◽  
Vitro Fertilization ◽  
Circadian Misalignment ◽  
Logistic Regression Models ◽  
Sleep Timing

Abstract Introduction Sleep duration and circadian misalignment have been linked to fertility and fecundability. However, sleep in women undergoing IVF has rarely been examined. This study investigated the role of sleep duration and timing with completion of an IVF cycle. Methods Prospective study of women undergoing IVF at a tertiary medical center between 2015 and 2017. Sleep was assessed by wrist-worn actigraphy 1–2 weeks prior to the initiation of their IVF cycle. Reproductive profile, IVF cycle details, demographic and health information were obtained from medical charts. Sleep duration, midpoint and bedtime were examined in relation to IVF cycle completion using logistic regression models, adjusted for age and anti-Müllerian hormone levels. A sub-analysis excluded women who worked non-day shifts to control for circadian misalignment. Results A total of 48 women were studied. Median age was 33y (range 25–42), with 29% of women older than 35 years. Ten women had an IVF cycle cancellation prior to embryo transfer. These women had shorter sleep duration, more nocturnal awakenings, lower sleep efficiency, and later sleep timing in comparison to those who completed their cycle. Twenty-minute increases in sleep duration were associated with lower odds of an uncompleted IVF cycle (OR = 0.88; 95% CI 0.78, 1.00). Women with later sleep midpoints and later bedtime had higher odds of an uncompleted cycle relative to those with earlier midpoints and earlier bedtime; OR=1.24; 95% CI 1.09, 1.40 and OR=1.33; 95% CI 1.17, 1.53 respectively, per 20-minute increments. These results were independent of age, levels of anti-Müllerian hormone, or sleep duration, and remained unchanged after exclusion of shift-working women. Conclusion This study demonstrated the influence of sleep duration and sleep timing on the odds of an uncompleted IVF cycle prior to embryo transfer. Sleep is a modifiable behavior that may contribute to IVF cycle success. Support (if any):

scholarly journals Age-related decline in the expression of GDF9 and BMP15 genes in follicle fluid and granulosa cells derived from poor ovarian responders

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Yan Gong ◽  
Jesse Li-Ling ◽  
Dongsheng Xiong ◽  
Jiajing Wei ◽  
Taiqing Zhong ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mrna Levels ◽  
Clinical Trial Registry ◽  
Altered Expression ◽  
Vitro Fertilization ◽  
Age Related ◽  
Tertiary Teaching ◽  
Poor Ovarian Responders

Abstract Background Growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) genes play important roles in folliculogenesis. Altered expression of the two have been found among patients with poor ovarian response (POR). In this prospective cohort study, we have determined the expression of the GDF9 and BMP15 genes in follicle fluid (FF) and granulosa cells (GCs) derived from poor ovarian responders grouped by age, and explored its correlation with the outcome of in vitro fertilization and embryo transfer (IVF-ET) treatment. Methods A total of 196 patients with POR were enrolled from a tertiary teaching hospital. The patients were diagnosed by the Bologna criteria and sub-divided into group A (< 35 year old), group B (35–40 year old), and group C (> 40 year old). A GnRH antagonist protocol was conducted for all patients, and FF and GCs were collected after oocyte retrieval. Expression of the GDF9 and BMP15 genes in the FF and GCs was determined with enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Results Compared with group C, groups A and B had significantly more two pronuclei (2PN) oocytes and transplantable embryos, in addition with higher rates of implantation and clinical pregnancy (P <  0.05). The expression level of GDF9 and BMP15 genes in the FF and GCs differed significantly among the three groups (P <  0.05), showing a trend of decline along with age. The ratio of GDF9/BMP15 mRNA levels were similar among the three groups (P > 0.05). The relative levels of GDF9 and BMP15 proteins in GCs have correlated with the relative mRNA levels in GCs and protein concentrations in FF (P <  0.05). Conclusions For poor ovarian responders, in particular those over 40, the expression of GDF9 and BMP15 is declined along with increased age and in accompany with poorer oocyte quality and IVF outcome, whilst the ratio of GDF9/BMP15 mRNA levels remained relatively constant. Trial registration Chinese Clinical Trial Registry Center (ChiCTR1800016107). Registered on 11 May 2018.

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