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PubMed2005

Effect of oviductal and cumulus cells on zona pellucida and cortical granules of porcine oocytes fertilized in vitro with epididymal spermatozoa.

The objective of this study was to evaluate the effects of porcine oviductal epithelial cell (POEC) monolayers and cumulus cells on the zona pellucida (ZP) and cortical granules (CG) of in vitro matured porcine oocytes. Denuded and cumulus-enclosed oocytes were exposed to POEC before or during in vitro fertilization (IVF). The functional effects of the co-culture system were the tested on the ZP resistance, measured by the time necessary to dissolve the ZP with 0.1% pronase, and the distribution and density of the cortical granules. CG density in the equator and cortex of each oocyte was evaluated by confocal microscopy after staining with fluorescein isothiocyanate-labelled peanut agglutinin (FITC-PNA). Both variables were assessed immediately after an in vitro maturation period (IVM group), 3 and 6h after culture with or without (Control) oviductal cells (Experiment 1) and 3h after insemination with frozen-thawed epididymal spermatozoa in the presence or absence (Control) of oviductal cells (Experiment 2). The time to dissolve the ZP of oocytes from IVM group was 440.4 +/- 61.7 s and no difference was observed among groups in Experiment 1. In contrast, the density of CG was affected; oocytes pre-incubated for 6h had a higher density than those pre-incubated for 3 h (P <0.001). Oocytes fertilized in vitro in the presence of POEC (Experiment 2) had a similar ZP digestion time as control oocytes 3 h after insemination. The presence of POEC during IVF as well as the presence of cumulus cells had no effect on the density and distribution of CG. However, a significant decrease in the density of CG was observed in the fertilized oocytes compared to in vitro matured oocytes (P <0.001). It is concluded that under the conditions employed the oviductal and cumulus cells in the perifertilization period had no effect on ZP hardening and CG density. However, an increase in CG density was observed when oocytes were maintained in culture. In addition, no hardening of ZP was observed after IVF, and denuded and cumulus-enclosed oocytes showed similar cortical reactions after insemination with epididymal spermatozoa regardless of the presence of POEC.

Animals↗
PubMed2006

Detection of ovine lentivirus in the cumulus cells, but not in the oocytes or follicular fluid, of naturally infected sheep.

The aim of this study was to examine the Maedi-Visna virus (MVV) infection status of oocytes, cumulus cells, and follicular fluid taken from 140 ewes from breeding flocks. MVV proviral-DNA and MVV RNA were detected using nested-PCR and RT-PCR MVV gene amplification, respectively in the gag gene. Nested-PCR analysis for MVV proviral-DNA was positive in peripheral blood mononuclear cells in 37.1% (52/140) of ewes and in 44.6% (125/280) of ovarian cortex samples. The examination of samples taken from ovarian follicles demonstrated that 8/280 batches of cumulus cells contained MVV proviral-DNA, whereas none of the 280 batches of oocytes taken from the same ovaries and whose cumulus cells has been removed, was found to be PCR positive. This was confirmed by RT-PCR analysis showing no MVV-viral RNA detection in all batches of oocytes without cumulus cells (0/280) and follicular fluid samples taken from the last 88 ovaries (0/88). The purity of the oocyte fraction and the efficacy of cumulus cell removal from oocytes was proved by absence of granulosa cell-specific mRNA in all batches of oocytes lacking the cumulus cells, using RT-PCR. This is the first demonstration that ewe cumulus cells harbor MVV genome and despite being in contact with these infected-cumulus cells, the oocytes and follicular fluid remain free from infection. In addition, the enzymatic and mechanical procedures we used to remove infected-cumulus cells surrounding the oocytes, are effective to generate MVV free-oocytes from MVV-infected ewes.

Animals↗
PubMed2003

Cumulus cells reduce the spermatozoa-zona binding inhibitory activity of human follicular fluid.

OBJECTIVE: To investigate the effects of human follicular fluid cultured with cumulus cells to inhibit the binding of spermatozoa to the zona pellucida of oocytes. DESIGN: Controlled experimental laboratory study. SETTING: University gynecology unit. PATIENT(S): Women undergoing assisted reproduction program and men visiting the subfertility clinics. INTERVENTION(S): Culture medium and human follicular fluid were used to culture cumulus cells in vitro for specified time periods. MAIN OUTCOME MEASURE(S): Zona binding capacity and motility of spermatozoa after incubation with cumulus cells treated culture medium or human follicular fluid. RESULT(S): Compared with the control medium, spent culture media after culturing cumulus cells for 3, 5, and 7 hours did not affect the motility and zona binding capacity of the treated spermatozoa. Significantly more spermatozoa treated with human follicular fluid that had been preincubated with cumulus cells for 5 and 7 hours bound onto hemizona in hemizona binding assay when compared with those preincubated in human follicular fluid without cumulus treatment. The hemizona index increased with the increase in the duration of cumulus cell treatment. Human follicular fluid with or without cumulus cells maintained sperm motility to similar extent for 3 hours. CONCLUSION(S): Cumulus cells reduced the inhibitory effect of human follicular fluid on spermatozoa-zona binding in vitro in a time-dependent manner.

Culture Media, Conditioned↗
PubMed1996

Complement-binding proteins are strongly expressed by human preimplantation blastocysts and cumulus cells as well as gametes.

Human preimplantation embryos, gametes and cumulus cells were studied for expression of the complement-binding proteins CD46 (membrane cofactor protein), CD55 (decay accelerating factor) and CD59 (membrane attack complex inhibitory factor) as well as complement receptors type 1 (CRI), type 2 (CR2) and type 3 (CR3). Both the CD55 and CD59 glycosyl phosphatidylinositol (GPI)-anchored proteins were expressed by the plasma membrane and zona pellucida of oocytes, early embryos and expanded preimplantation blastocysts; in contrast, CD46 was expressed only on the plasma membrane. Cumulus cells consistently expressed CD46 and, most strongly, CD59 whereas CD55 expression was variable. Monoclonal antibodies (mAbs) to CRI, CR2 and CR3 epitopes gave only occasional reactivity on oocytes and were unreactive with blastocysts, spermatozoa and cumulus cells. CD46 is expressed only on the spermatozoal inner acrosomal membrane, and CD59 on the plasma membrane; CD55 expression was confirmed on the plasma membrane as well as the inner acrosomal membrane. Control mAbs specific for factor H were usually unreactive with gametes, blastocysts and cumulus cells. These data support the concept that gametes and early embryonic cells are protected from complement-mediated attack by expression of CD46, CD55 and CD59, although these complement-binding proteins may have additional roles in reproductive events.

Acrosome↗
PubMed2003

Epidermal growth factor-receptor tyrosine kinase activity regulates expansion of porcine oocyte-cumulus cell complexes in vitro.

We have recently shown that epidermal growth factor (EGF) strongly stimulates expansion of porcine oocyte-cumulus complexes (OCCs) isolated from large follicles (>6 mm) and does not promote expansion of OCCs from small (3-4-mm) follicles. In order to elucidate the role of EGF in OCCs expansion, in the present study, we first examined the presence of EGF receptors (EGFRs) in cumulus cells isolated from follicles of different sizes. Surprisingly, immunoblotting showed that cumulus cells obtained from all follicular size categories contained similar amounts of EGFR protein. On the other hand, we found a dramatic difference in the pattern of protein tyrosine phosphorylation in a comparison of cumulus cells isolated from small and large follicles treated by EGF. Furthermore, tyrosine-phosphorylated EGFR was specifically immunoprecipitated with antiphosphotyrosine antibodies from EGF-treated cumulus cells isolated from the large follicles. This result strongly indicates that only OCCs from the large follicles contain mature EGFRs that are capable of becoming activated by EGF. Remarkably, preincubation of cumulus cells from small follicles (3-4 mm) with FSH strongly increased EGF-stimulated tyrosine phosphorylation to levels comparable with OCCs from large follicles. The FSH-dependent activation of EGFRs was beneficial for expansion of OCCs isolated from the small follicles since OCCs treated sequentially by FSH (3 h) and EGF (1 h) underwent expansion significantly better then OCCs cultured in FSH or EGF alone. We conclude that a FSH-dependent pathway has an important role in the maturation of the EGFR in cumulus cells and that activation of EGFR-dependent signaling is sufficient to induce expansion.

Animals↗
PubMed2004

Synergistic roles of BMP15 and GDF9 in the development and function of the oocyte-cumulus cell complex in mice: genetic evidence for an oocyte-granulosa cell regulatory loop.

Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-specific growth factors that appear to play key roles in granulosa cell development and fertility in most mammalian species. We have evaluated the role(s) of these paracrine factors in the development and function of both the cumulus cells and oocytes by assessing cumulus expansion, oocyte maturation, fertilization, and preimplantation embryogenesis in Gdf9+/-Bmp15-/- [hereafter, double mutant (DM)] mice. We found that cumulus expansion, as well as the expression of hyaluronon synthase 2 (Has2) mRNA was impaired in DM oocyte-cumulus cell complexes. This aberrant cumulus expansion was not remedied by coculture with normal wild-type (WT) oocytes, indicating that the development and/or differentiation of cumulus cells in the DM, up to the stage of the preovulatory luteinizing hormone (LH) surge, is impaired. In addition, DM oocytes failed to enable FSH to induce cumulus expansion in WT oocytectomized (OOX) cumulus. Moreover, LH-induced oocyte meiotic resumption was significantly delayed in vivo, and this delayed resumption of meiosis was correlated with the reduced activation of mitogen-activated protein kinase (MAPK) in the cumulus cells, thus suggesting that GDF9 and BMP15 also regulate the function of cumulus cells after the preovulatory LH surge. Although spontaneous in vitro oocyte maturation occurred normally, oocyte fertilization and preimplantation embryogenesis were significantly altered in the DM, suggesting that the full complement of both GDF9 and BMP15 are essential for the development and function of oocytes. Because receptors for GDF9 and BMP15 have not yet been identified in mouse oocytes, the effects of the mutations in the Bmp15 and Gdf9 genes on oocyte development and functions must be produced indirectly by first affecting the granulosa cells and then the oocyte. Therefore, this study provides further evidence for the existence and functioning of an oocyte-granulosa cell regulatory loop.

Animals↗
PubMed1993

Modulation of bovine oocyte-cumulus cell complex maturation and fertilization in vitro by glycosaminoglycans.

Oocyte-cumulus cell complex maturation and fertilization potential of 4700 bovine oocytes in the presence of various intrafollicular regulators were studied. Endpoints for oocyte maturation included cumulus cell expansion, retention of [3H]hyaluronic acid, elevated cyclic AMP concentrations, and germinal vesicle breakdown. Media supplemented with FSH and fetal bovine serum stimulated oocyte maturation and fertilization. Heparin and mixed glycosaminoglycans (40% heparan sulfate and 60% dermatan sulfate) stimulated cumulus cell expansion and increased [3H]hyaluronic acid but decreased cyclic AMP and reduced percentages of oocytes with germinal vesicle breakdown at 12 h. Oocytes cultured in heparin or mixed glycosaminoglycans were incapable of fertilization. Addition of serum to oocytes treated with heparin decreased cumulus cell expansion. Addition of FSH to heparin and serum cultures did not overcome the inhibitory effect of serum on heparin-induced expansion. Dermatan sulfate failed to stimulate expansion of cumuli but resulted in cells retaining [3H]hyaluronic acid and elevated cyclic AMP content of oocytes. Dermatan sulfate did not affect the percentage of germinal vesicle breakdown at 12 h. Nitrogen-desulfated heparin did not promote any signs of oocyte maturation. These results demonstrate that heparin and mixed glycosaminoglycans stimulate events related to oocyte maturation but were not capable of preparing oocytes for fertilization. The addition of serum or desulfation of heparin inhibited heparin effects on oocyte maturation.

Animals↗
PubMed2005

Presence of cumulus cells during in vitro fertilization protects the bovine oocyte against oxidative stress and improves first cleavage but does not affect further development.

The present study was conducted to evaluate the function of cumulus cells during bovine IVF Oocytes within cumulus-oocyte complexes (COCs) or denuded oocytes (DOs) were inseminated in control medium, or DOs were inseminated in cumulus cell conditioned medium (CCCM). DOs exhibited reduced cleavage and blastocyst formation rates when compared with intact COCs. The reduced blastocyst formation rate of DOs resulted from reduced first cleavage but subsequent embryo development was not changed. Live-dead staining and staining for apoptotic cells revealed no differences in blastocysts from oocytes fertilized as COC or DO. Fertilization of DOs in CCCM partially restored the cleavage rate, suggesting that factors secreted by cumulus cells are important for fertilization but that physical contact between oocytes and cumulus cells is required for optimal fertilization and first cleavage. Exposure of COCs to hydrogen peroxide shortly before fertilization reduced the cleavage rate, but did not lead to enhanced death of cumulus cells or oocyte death. Exposure of DOs to hydrogen peroxide, however, resulted in oocyte death and a complete block of first cleavage, suggesting that cumulus cells protect the oocyte against oxidative stress during fertilization.

Animals↗
PubMed2002

Luteinizing hormone receptor formation in cumulus cells surrounding porcine oocytes and its role during meiotic maturation of porcine oocytes.

We investigated the formation of LH receptor (LHR) in cumulus cells surrounding porcine oocytes and the role of LHR in meiotic maturation of oocytes. At least three splice variants of LHR mRNA were detected in cumulus cells, in addition to the full-length form. Low levels of three types of products were seen in cumulus cells from cumulus oocytes complexes (COCs), whereas the full-length form was significantly increased by 12-h cultivation with FSH. The addition of FSH also significantly increased the binding level of biotinylated hCG to COCs. The formation of LHR in FSH-stimulated cumulus cells was not affected by additional 0.5 mM phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX), and the oocytes were synchronized to the germinal vesicle (GV) II stage by exposure to 0.5 mM IBMX and FSH for 20 h. The binding of LH to its receptor induced a further increase in cAMP level and progesterone production and acceleration of meiotic progression to the metaphase I stage. The oocytes cultured with LH for 24 h following cultivation with FSH and IBMX were used for in vitro fertilization. At 6 days after in vitro fertilization, blastocyst rate in oocytes matured under these conditions was significantly higher than that of oocytes cultured in the absence of LH. Treatment of oocytes with FSH and 0.5 mM IBMX to express LH receptor in cumulus cells while holding oocytes at the GV II stage is a very beneficial way to produce in vitro-matured oocytes, which have high developmental competence.

1-Methyl-3-isobutylxanthine↗
PubMed1993

The enhancement by prostaglandin E2 of cumulus cell outgrowth in vitro.

Oocyte-cumulus cell complexes (OCCC) were aspirated from medium-sized (3-5 mm phi) porcine follicles to study quantitatively the process of forming monolayers. A monolayer index (MLI) was therefore developed. This index was evaluated by addition of graded quantities of fetal bovine serum. Furthermore, we observed that addition of prostaglandin E2 accelerated the monolayer formation (P < 0.001 at 20 hours versus 40 hours). The effect of PGE2 regarding both monolayer formation and cumulus cell expansion has been discussed.

Animals↗
PubMed2000

Apoptosis in human cumulus cells in relation to maturation stage and cleavage of the corresponding oocyte.

BACKGROUND: The purpose of this study was to determine the incidence of apoptosis in cumulus cells, and correlate these findings with the maturation stage, fertilization rate and embryo score of the corresponding oocyte, in couples undergoing ICSI due to a male factor. METHODS: The study group consisted of 21 couples where ICSI was performed. The total number of oocyte-cumulus complexes retrieved was 164. Sperm samples were assessed according to the WHO manual, morphology according to the strict criteria and for the presence of apoptosis. The degree of apoptotic DNA fragmentation was determined using the free 3'OH DNA termini in situ with chemically labeled and unlabeled nucleotides. The study was blinded for the technician involved in the assessments of apoptosis in the cumulus cells, apoptosis and morphology in spermatozoa. Ovarian hyperstimulation was carried out according to a long down regulation protocol using GnRH, recFSH and hCG. A maximum of 3 embryos were transferred on day 2 after ICSI. RESULTS: This study demonstrated that the incidence of apoptosis was significantly higher in cumulus cells from germinal vesicle and metaphase I oocytes compared to cumulus cells from metaphase II oocytes (p<0.0001). Non-fertilized metaphase II oocytes showed significantly higher incidence of apoptosis in cumulus cells compared to fertilized metaphase II oocytes (p=0.0082). Furthermore, apoptosis in spermatozoa had an impact on the embryo score (p=0.0087). CONCLUSION: Comparing apoptosis in cumulus cells with maturity of the corresponding oocytes, a significantly higher degree was found related to immature oocytes. Apoptosis in cumulus cells from human metaphase II oocytes impaired the fertilization rate. The degree of fragmentation in the embryo might be correlated to apoptosis in the spermatozoa.

Adult↗
PubMed1991

Importance of cumulus cells and insemination intervals for development of bovine oocytes into morulae and blastocysts in vitro.

Experiments were carried out to investigate influences of cumulus cells and insemination intervals on bovine in vitro fertilization (IVF) and early development. Cumulus-encased oocytes, aspirated from 2- to 5-mm ovarian follicles at slaughter, were incubated 24 hours for maturation in the presence of 20% proestrous (Day 20) cow serum and 100 microg LH/ml. Ova with mature (expanded) cumuli oophori were inseminated and removed from sperm-containing droplets (50 microl) after 6, 12, 24 or 48 hours. After maturation, some ova were removed from their surrounding cumulus cells and otherwise treated in the same way. The highest proportion of ova that cleaved (50/57, 87.7%) resulted from the 24-hour insemination group; this was significantly higher (P<0.05) than for 6-hour (35.1%) and 12-hour group (49.3%), but not for the 48-hour group (73.0%). A significantly (P<0.05) higher proportion of cleaved ova developed into morulae/blastocysts (28%) from the 24-hour group. Removal of cumulus cells before IVF resulted in lower cleavage rates; the morula/blastocyst stage was reached only when the denuded ova were with sperm for 48 hours. In additional experiments, cumulus cells recovered from follicular aspirates were cultured in HEPES-M 199 with 10% Day-20 serum, and 0, 10 and 100 microg LH/ml and resulting monolayers were used for zygote culture. The developmental stages reached after IVF were not altered by LH treatment of supporting cumulus cells. A 24-hour insemination interval with subsequent culture on a cumulus cell monolayer resulted in optimal in vitro development.

Journal Article↗
PubMed2001

Comet assay of cumulus cell DNA status and the relationship to oocyte fertilization via intracytoplasmic sperm injection.

This paper postulates that in the ovary, the close association between the cumulus cells and the oocytes permits the fertilizing ability of the oocytes to be determined indirectly through cumulus cell DNA status. The objective was to use a modified comet assay to analyse cumulus cell DNA and relate the data to oocyte fertilization after intracytoplasmic sperm injection (ICSI) procedures. Oocytes were retrieved (n = 15 cases) and free-floating cumulus cells were pooled and smeared on clear glass slides to dry. Meanwhile, the denuded oocytes were injected with spermatozoa and fertilization was assessed, followed by embryo transfer. The fixed cumulus cells were stained in acridine orange, coated with a mini-gel agarose layer, lysed in alkaline buffer and electrophoresis performed. Analyses of fluorescent cell images (n = 449) showed that the tail moment was positively correlated to the percentage of fertilization after ICSI (r = 0.567, P < 0.05). In contrast, there was no correlation between tail moment and number of oocytes retrieved, total ampoules used, endometrial thickness and age of patient. The results suggested that the competence of the oocytes was associated with the cumulus cell DNA status. A unique feature here was the comet assay for archived material with obvious advantages.

Adult↗
PubMed2002

The apoptotic profile of human cumulus cells changes with patient age and after exposure to sperm but not in relation to oocyte maturity.

OBJECTIVE: To determine the expression of apoptosis-associated molecules on cumulus cells removed from individual oocytes of different maturity, inseminated oocytes and to investigate the possibility of an age-dependent expression. DESIGN: Analysis of apoptosis in cumulus cells isolated from oocytes of different stages of maturity. SETTING: Assisted reproductive technology program of the Birmingham Women's Hospital, Birmingham, UK. PATIENT(S): Patients undergoing intracytoplasmic sperm injection or IVF cycles. MAIN OUTCOME MEASURE(S): Percentage of positive cumulus cells when assessed for nuclear DNA damage using the terminal deoxyuridine nucleotide end-labeling assay or stained with antibodies [Fas, Fas ligand, the antiapoptotic protein Bcl-xl, and the RNA-binding protein (TIAR)]. RESULT(S): Cumulus cells collected from mature oocytes showed no significant difference in the percentage of apoptotic markers compared to those recovered from immature oocytes, whereas those from patients >/=38 years differed significantly. When cumulus cells were exposed to sperm the levels of apoptotic markers altered significantly from those not exposed to sperm. CONCLUSION(S): The results show that the cumulus cells of human oocytes are equipped with a mechanism to undergo apoptosis and that patient age and the exposure of cumulus cells to sperm can alter their profiles of apoptotic markers.

Adult↗
PubMed1995

The influence of glucose, cumulus cells, and metabolic coupling on ATP levels and meiotic control in the isolated mouse oocyte.

The effects of glucose and cumulus cells on oocyte ATP levels and germinal vesicle breakdown (GVB) in isolated mouse oocytes have been examined. Oocyte-cumulus cell complexes or denuded oocytes (DO) from pregnant mare serum gonadotropin-primed immature mice were cultured in minimum essential medium containing 4 mM hypoxanthine and 1 mM pyruvate, in the absence or presence of 0.55 mM glucose. After 17-18 hr in the presence of glucose, ATP in both the oocyte-cumulus cell complexes and oocytes derived from such complexes (cumulus cell-enclosed oocytes; CEO) was elevated, and less than one-half of the oocytes had resumed maturation (48% GVB). Removal of glucose caused a decrease in ATP levels in complexes and CEO and reversed the meiotic arrest in CEO (98% GVB), and these effects were mimicked by iodoacetate treatment. Higher frequencies of GVB were observed in glucose-free medium after more than 6 hr of culture, while ATP levels were reduced within 3 hr. Glucose had no effect on ATP levels or the meiotic state of denuded oocytes. Interestingly, iodoacetate had a stimulatory effect on GVB in DO (86% GVB compared to 57% in controls), but did not effect ATP levels. Glycerrhetinic acid, a gap junction uncoupler, completely suppressed oocyte-cumulus cell coupling in cultured complexes (0.15% coupling compared to 16.6% in controls) and reversed the inhibitory effect of glucose on oocyte maturation (91 and 95% GVB at 10 and 25 microM compared to 42% GVB in controls). This agent also prevented follicle-stimulating hormone-induced meiotic maturation in dibutyryl cAMP-arrested CEO. These results thus implicate mediation by gap junctions of both inhibitory and stimulatory signals from the cumulus cells. Comparison of ATP levels in spontaneously maturing CEO in vitro with those from oocytes maturing in vivo in response to human chorionic gonadotropin revealed that a decrease in oocyte ATP preceded or accompanied GVB in spontaneously maturing oocytes but not in those maturing in vivo. The results of this study indicate that glucose-derived elevation of oocyte ATP contributes to meiotic arrest in cumulus cell-enclosed oocytes that is dependent on patient gap junctions. Furthermore, a model for reinitiation of oocyte maturation is supported in which spontaneous GVB results from cessation or interruption of inhibitory influences, while ligand-provoked GVB is brought about by the generation of stimulatory signals that override inhibitory input.

Adenosine Triphosphate↗
PubMed2003

Effect of roscovitine, a cdk1 inhibitor, and of the presence of oocyte on bovine cumulus cell expansion and cyclooxygenase-2 expression.

Oocyte maturation is accompanied by differentiation of surrounding cumulus cells. These cells produce hyaluronic acid (HA) and its storage in intercellular spaces results in expansion of the cells. The cumulus cells also accumulate cyclooxygenase-2 (cox-2) during maturation. Both expansion and cox-2 storage are regulated by FSH and EGF. The aim of this study was to determine whether oocyte meiotic resumption is involved in the regulation of cumulus differentiation or not. We investigated the effects of roscovitine, a reversible inhibitor of meiosis resumption of cattle oocytes on EGF induced expansion and cox-2 expression at the transcript and protein levels respectively (RT-PCR and Western blot), in cumulus oocyte complexes (COCs) and cumulus complexes alone (CCs). EGF induced expansion and cox-2 expression in both COCs and CCs. These effects were prevented by roscovitine, whether in the presence or in the absence of oocyte. However, the oocyte was essential for the reversibility of inhibition by roscovitine. In conclusion, our results indicate that i) oocyte secreted-factors are not essential for cumulus expansion, and ii) roscovitine mediated inhibition of meiotic resumption also respects the functionality of the surrounding somatic cells.

Animals↗
PubMed1997

Glutathione synthesis during in vitro maturation of bovine oocytes: role of cumulus cells.

Glutathione (GSH) synthesis during in vitro maturation (IVM) has been shown to play an important role in embryo development. The present study was carried out to evaluate the role of cumulus cells in GSH synthesis during IVM of bovine oocytes in the presence of cystine, cysteine, the cysteine analogue N-acetylcysteine, and cysteamine. For this purpose, cumulus-oocyte complexes (COCs), denuded oocytes (DOs), and DOs in coculture with a cumulus cell monolayer were used. An increase in GSH level stimulated by cystine was observed only in the presence of cumulus cells, either with COCs or in DOs matured on a coculture monolayer. Addition of cysteine and cysteamine to IVM medium increased GSH levels in COCs and DOs. N-Acetylcysteine increased GSH levels only in DOs. Moreover, cumulus cells contributed to the stimulatory effect exerted by cysteine and cysteamine on GSH synthesis in COCs. These results indicate that cumulus cells during IVM play an important role in oocyte GSH synthesis, allowing the oocytes to use cystine and contributing to the stimulatory effect exerted by cysteine and cysteamine. In addition, these results demonstrate that IVM medium supplemented with cysteine or cysteamine increased GSH content in oocytes without cumulus mass (DO) and in the absence of a cumulus cell monolayer. This may be useful to increase the efficacy of IVM of those oocytes having few cumulus cell layers, in a system without coculture.

Acetylcysteine↗
PubMed2002

Apoptosis in human cumulus cells in relation to zona pellucida thickness variation, maturation stage, and cleavage of the corresponding oocyte after intracytoplasmic sperm injection.

OBJECTIVE: To assess the degree of apoptosis in the cumulus cells and the variation of the zona pellucida and the maturity and fertilization of the corresponding oocyte. DESIGN: Retrospective study. SETTING: Private fertility clinic. PATIENT(S): Fifty couples undergoing ICSI. INTERVENTION(S): ICSI. MAIN OUTCOME MEASURE(S): Correlation between apoptosis in the cumulus cells and the zona pellucida thickness variation, maturation stage, fertilization rate, and embryo score. RESULT(S): This study demonstrated no correlation between apoptosis in cumulus cells and the thickness and variation of the zona pellucida in oocytes and embryos. The incidence of apoptosis was significantly higher in cumulus cells from empty zona pellucidas and germinal vesicle stage and metaphase I oocytes compared with metaphase II oocytes. Non-fertilized metaphase II oocytes showed significantly higher incidence of apoptosis compared with fertilized metaphase II oocytes. There was a correlation between embryo score and the zona pellucida thickness variation. CONCLUSION(S): Apoptosis in cumulus cells had no impact on the zona pellucida thickness and variation in oocytes and embryos. The zona pellucida thickness variation was positively correlated to good embryo score. A higher degree of apoptosis was seen in cumulus cells from immature oocytes compared with mature oocytes. Furthermore, apoptosis in cumulus cells impaired the fertilization rate of metaphase II oocytes after ICSI.

Adult↗